National Academies Press: OpenBook

Veterans and Agent Orange: Update 2002 (2003)

Chapter: 3. Toxicology

« Previous: 2. Considerations in Evaluating the Evidence
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

3

Toxicology

As in Veterans and Agent Orange: Health Effects of Herbicides Used in Vietnam (hearafter referred to as VAO; IOM, 1994), Veterans and Agent Orange: Update 1996 (hearafter Update 1996; IOM, 1996), Veterans and Agent Orange: Update 1998 (hearafter Update 1998; IOM, 1999), and Veterans and Agent Orange: Update 2000 (hearafter Update 2000; IOM, 2001), this review summarizes the recent experimental data that serve as a scientific basis of assessment of the biologic plausibility of health outcomes reported in epidemiologic studies. Efforts to establish the biologic plausibility of effects of herbicide exposure in the laboratory strengthen the evidence of the herbicide effects suspected to occur in humans. Toxic outcomes are influenced by differences in dosage (magnitude and frequency of administration); by exposure to other chemicals, including chemicals other than herbicides; by pre-existing health status; by genetic factors; and by the route and rate of absorption, distribution, metabolism, and excretion. Any attempt to extrapolate from experimental studies to human exposure must therefore carefully consider such variables before conclusions are made.

Multiple chemicals were used for various purposes in Vietnam. The chemical nature of the substances themselves is discussed in more detail in Chapter 6 of VAO. Four herbicides documented in military records were of particular concern and are addressed here: 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), 4-amino-3,5,6-trichloropicolinic acid (picloram), and cacodylic acid (dimethylarsenic acid, DMA). In addition, this chapter focuses to a large extent on a contaminant of 2,4,5-T, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, or dioxin) because its potential toxicity is of concern and considerably more information is available on it than on the herbicides. Most of the

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

experimental studies of those chemicals, unless otherwise noted, were conducted with pure chemicals, in contrast with the epidemiologic studies discussed in later chapters, in which exposures were often to mixtures of chemicals.

This chapter begins with a brief summary of major conclusions presented in previous Veterans and Agent Orange reports regarding the toxicology of the compounds of interest. That summary is followed by what makes up the majority of the chapter, overviews and discussions of the relevant experimental studies that have been published on 2,4-D, 2,4,5-T, picloram, cacodylic acid, and TCDD since Update 2000. Within the update for each of the chemicals, the experimental studies investigating the toxicokinetics, mechanisms of action, and disease outcomes of exposure to the chemical are discussed. Where appropriate, the mechanisms of action are discussed as they relate to a particular endpoint. Estimating potential human health risks on the basis of the animal data is then discussed.

HIGHLIGHTS OF PREVIOUS REPORTS

Chapter 4 of VAO and Chapter 3 of Update 1996, Update 1998, and Update 2000 review the results of animal and in vitro studies published through 2000 that investigate the toxicokinetics, mechanism of action, and disease outcomes of the herbicides used in Vietnam, and the contaminant TCDD. The toxicity of the four herbicides has not been studied extensively, but in general they are not considered particularly toxic because high concentrations are usually required to modulate cellular and biochemical processes. In contrast, the toxicity of TCDD has been studied extensively. On the basis of the experimental data reviewed in previous Agent Orange reports, the committees concluded that TCDD elicits a diverse spectrum of sex-, strain-, age-, and species-specific effects, including carcinogenesis, immunotoxicity, reproductive and developmental toxicity, hepatotoxicity, neurotoxicity, chloracne, and loss of body weight. The scientific consensus is that TCDD is not directly genotoxic and that its ability to influence the carcinogenic process is mediated by epigenetic events, such as enzyme induction, cell proliferation, apoptosis, and intracellular communication. Most, if not all, of TCDD's effects are mediated through the aryl hydrocarbon receptor (AhR), which interacts with other proteins, binds to DNA and results in biochemical effects, including enzyme induction.

TOXICITY PROFILE UPDATE OF 2,4-D

Toxicokinetics

Toxicokinetics (also referred to as pharmacokinetics) pertains to the routes and rates of uptake, tissue distribution, transformation, and elimination of a toxicant. Those processes, in part, determine the amount of a particular chemical that reaches potential target organs or cells and thereby influences toxicity to organs

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

or cells. Understanding the toxicokinetics of a compound is important for valid reconstruction of exposure to it.

Since Update 2000, several studies have examined the pharmacokinetics and metabolism of 2,4-D in animal species. Recent data support the conclusions of previous updates that metabolism and elimination of 2,4-D are relatively rapid and that tissue uptake is small. Kim et al. (2001) constructed a physiologically based pharmacokinetic (PBPK) model to describe and predict the kinetic behavior of 2,4-D in rats after long-term exposures to low doses. The model was tested with experimental data from rats that were given 2,4-D at 1 or 10 mg/kg body weight per day by subcutaneous infusion for 7, 14, and 28 days. In general, the experimental data fell within the range of 2,4-D concentrations predicted by the PBPK model for the blood and different brain regions. The model supports the concept that uptake of the chemical into brain was limited primarily by the membrane components of the blood–brain barrier. In another study (Barnekow et al., 2001) the elimination and metabolism of 2,4-D following oral administration were evaluated in laying hens dosed with 2,4-D at 18 mg/kg body weight for 7 days and in lactating goats dosed with 2,4-D at 483 mg/kg body weight for 3 days. More than 90% of the total dose was eliminated within 24 h of the final dose. Individual tissue residues accounted for less than 0.1% of the dose. The most abundant residue was 2,4-D; a minor metabolite, 2,4-dichlorophenol, was also present. Overall, those studies suggest that in the species used 2,4-D is eliminated relatively rapidly and that uptake and metabolism by most tissues are low.

A study by Dickow et al. (2000) attempted to correlate plasma concentrations with observed clinical effects in dogs after a dose of twice the reported LD50 (the lowest dose that kills half the animals that receive it), 2,4-D at 100 mg/kg body weight. All dogs survived, but vomiting and diarrhea were observed. The mean total and unbound plasma 2,4-D concentrations were 511 mg/L and 129 mg/L, respectively.

As discussed in previous updates, studies suggest that although 2,4-D is relatively nontoxic, the developing nervous system might be a target after exposure to high concentrations. Sturtz et al. (2000) therefore investigated the lactational transfer of 2,4-D by measuring it in tissues of rats whose dams received 2,4-D at 50, 70, and 100 mg/kg body weight during nursing. 2,4-D residues in tissues depended on dose and exposure time. At the highest dose, there was impaired body growth, low tissue weights, and diminished stomach contents of the offspring. The analysis of tissues indicated that 2,4-D was transferred to the neonates during nursing and that, at least at the highest maternal dose, the toxicity might be explained by diminished milk intake or direct toxic effects on the neonate. When 2,4-D treatment was discontinued, the residues remained in the stomach contents of the neonates for at least a week.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Mechanisms of Toxic Action
Mechanisms Related to Genotoxic Effects

Several studies reviewed in Update 2000 indicate a relatively weak or no genotoxic potential of 2,4-D. Two studies published since then are consistent with a lack of genotoxicity. A study by Venkov et al. (2000) demonstrated a lack of mutagenic action of 2,4-D by using tests in yeast, transformed hematopoietic cells, and mouse bone marrow cells. Charles et al. (2000) also demonstated a lack of genotoxicity after exposure to 4-(2,4-dichlorophenoxy) butyric acid, of which 2,4-D is a metabolite, by looking at gene mutation in bacteria and cultured mammalian cells, cytogenetic abnormalities in mammalian cells, and induction of DNA damage and repair in rat hepatocytes. A study by Amer and Aly (2001), however, observed increased genotoxity after oral exposure to 2,4-D at 3.3 mg/kg body weight for 3 and 5 consecutive days; a significant increase in the percentage of chromosomal aberrations in bone marrow and spermatocytes was observed with both regimens. The genotoxic effects of 2,4-dichlorophenol, a metabolite of 2,4-D, were also investigated in that study and were much weaker. Only the highest concentration tested, 2,4-D at 180 mg/kg body weight, induced a significant percentage of effects after intraperitoneal injection (Amer and Aly, 2001).

Mechanisms Related to Effects on Energy Metabolism or Mitochondrial Function

Several reports cited in previous updates suggest that the toxicity of 2,4-D might be related, at least in part, to its effect at relatively high concentrations on calcium homeostasis and energy metabolism. Those actions might be mediated by a direct action on mitochondria. A study discussed in Update 2000 indicated that the mitochondrial effects of some herbicide preparations, including those containing 2,4-D, might be due primarily to the surfactant in the formulations and not to 2,4-D itself. A similar study by Oakes and Pollak (2000) confirmed that as much as 50% of the effects of several formulations, including Agent Orange, on oxidative functions of submitochondrial particles is due to “inert” components. A molecular study by Di Paolo et al. (2001) isolated a single protein contained in rat liver mitochondria to which radiolabeled 2,4-D or one of its metabolites was covalently bound. Although the identity of the protein is not known, the investigators suggest that the alteration of its function may be related to known alterations in mitochondrial function produced by 2,4-D.

Previous updates noted that 2,4-D is a peroxisome proliferator, that is, it causes an increase in the number and size of peroxisomes in several tissues of susceptible species. Such chemicals are nonmutagenic carcinogens in the livers of rodents. Humans and hamsters are considered to be relatively resistant to the effects of peroxisome proliferators. A study by Ozaki et al. (2001) observed distinct morphologic changes in the kidneys of rats and mice chronically exposed

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

to 2,4-D and WY-14643 (a known peroxisome proliferator) for up to 3 months. The changes were characterized by alteration in tubule structures, long brush borders of tubule cells, and reduced volume and number of mitochondria. Those changes were not observed in hamsters. The authors indicate that although 2,4-D is considered a weak peroxisome proliferator in the rodent liver, it appeared to be more effective in inducing renal changes. Kaioumova et al. (2001) determined that the dimethylammonium salt of 2,4-D (up to 3 mM) caused concentration-and time-dependent apoptosis in peripheral lymphocytes of healthy people and in vitro in Jurkat T cells. Further examination of the mechanism indicated that those effects were mediated by direct action of the chemical on mitochondria. Hepatocyte ultrastructural changes were observed in rats whose mothers received the sodium salt of 2,4-D in drinking water (at a daily dose of 2,4-D at 250 mg/kg body weight) before fertilization and during pregnancy and lactation; the changes were consistent with effects of 2,4-D on mitochondria and energy metabolism (Pilat-Marcinkiewicz et al., 2000).

Mechanisms Related to Effects on Thyroid Hormones

Effects of 2,4-D on serum concentrations of thyroid hormones, particularly decreases in thyroxine, were noted in previous updates. A recent report by Kobal et al. (2000) likewise observed decreased serum concentrations of thyroxine and triiodothyronine after oral exposure of male and female rats to 2,4-D at 11 and 110 mg/kg body weight per day for 10 days. Chemical-induced alterations in thyroid homeostasis can adversely affect the development of many organ systems including the nervous and reproductive systems. Most of these effects are caused by lack of thyroid hormone alone rather than by increases in TSH.

Mechanisms Related to Effects on Cell Stress Responses

Stress proteins (for example, heat-shock proteins) are most often induced in a variety of cells in response to environmental and chemical stressors and have been proposed as markers of the presence of stressors. Two studies examined the ability of 2,4-D to increase heat-shock proteins in bacteria and a human cell line. 2,4-D exposure induced several heat-shock proteins in bacteria (Cho et al., 2000), but did not induce the hsp70 promoter in a HeLa cell line (Ait-Aissa et al., 2000). An additional study determined that a single exposure to 1 mM 2,4-D diminished growth and total protein in all E. coli strains tested; successive exposures to 0.01 mM 2,4-D also had a toxic effect on cell growth (Balague et al., 2001).

Disease Outcomes

Studies of disease outcomes published since Update 2000 are consistent with the previous conclusion that 2,4-D is relatively nontoxic and has weak oncogenic

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

potential. Also as previously indicated, the developing fetus appears to be most sensitive to the effects of 2,4-D for a number of toxic end points. One recent investigation yielded no evidence that paternal exposure to a herbicide formulation containing 2,4-D and picloram caused birth defects or any other adverse reproductive outcome (Oakes et al., 2002a). Recent animal studies of disease outcomes of 2,4-D exposure are discussed below.

Neurotoxicity

Bortolozzi et al. (2001) studied the effects of nonphysiologic, direct, intracebral administration of 2,4-D (2,4-D at 50 or 100 µg/rat) on behavior and neurochemical alterations in the rat brain. 2,4-D induced a regionally specific neurotoxicity in the basal ganglia, but the neurotoxic effects depended on the location of injection, the dose, and the length of time since the injection. Those data suggest that 2,4-D has the ability to produce direct effects on the brain if high enough concentrations can be achieved. In another study, 2–4 mM 2,4-D directly affected the viability of isolated frog sciatic nerve (Kouri and Theophilidis, 2002). Garabrant and Philbert (2002) reviewed the scientific evidence relevant to neurologic effects of 2,4-D. Although high doses in experimental animals have been found to produce myotonia and alterations in gait and behavioral indexes, there is no evidence of effects on the neurologic system at doses in the microgram-per-kilogram-per-day range. That information is consistent with the conclusion of this and previous Agent Orange updates.

Reproductive and Developmental Toxicity

Several studies have examined the developmental toxicity of 2,4-D. Charles et al. (2001) examine the potential for 2,4-D and its salts and esters to induce developmental toxicity in rats and rabbits. In both species, effects on maternal body weight manifest with 2,4-D at 30 mg/kg maternal body weight per day. At higher doses, body weights and feed consumption were more severely affected. The no-observed-adverse-effect level (NOAEL) for maternal toxicity was about 10 mg 2,4-D/kg body weight per day. Significantly decreased fetal body weights and fetal variations were seen in rats only at doses greater than 90 mg 2,4-D/kg body weight per day. At maternally toxic doses in rabbits, embryonal and fetal development were unaffected. Those data suggest that those end points in the developing rat and rabbit are not uniquely sensitive to 2,4-D or its salt and ester forms. Postnatal measures were not examined in that study. A study by Fofana et al. (2000) examined maternal and fetal toxicity after exposure of pregnant dams to 2,4-D at daily doses of 50, 70, 110, or 150 mg/kg maternal body weight on gestational days 6–10, 6–15, or 11–15. There was significant maternal weight loss in all experimental groups and a dose-related embryolethality. Kidney and urogenital malformations were found in the fetuses. A later study by Fofana et al.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

(2002) reported similar findings except that impaired growth of the unaffected offspring was not observed.

Two studies examined brain development in rats and concluded that exposure to 2,4-D may produce alterations in brain development. Ferri et al. (2000) injected dams with 2,4-D (at 100 mg/kg body weight per day) or vehicle during lactation (on postnatal day 9–15 or 9–25). No overt signs of toxicity were observed in the dams, but significant differences were observed in the development of the brain monoaminergic system of neonates exposed to 2,4-D through mother's milk. There was an increase in 5-hydroxyindolacetic acid and serotonin in brains of 25-day-old pups. Alterations in serotonin, dopamine, and norepinephrine were also seen in several brain areas. Rosso et al. (2000) exposed neonatal rats to 2,4-D at 100 mg/kg body weight per day on postnatal days 7–25 or 2,4-D at 70 mg/kg body weight per day on postnatal days 12–25. Decreased body and brain weights were noted only at the higher dose regimen, but both regimens decreased the amount of brain myelin gangliosides and myelin deposition. Alterations in muscular force and motor activity were also seen. Another study investigated whether 2,4-D alters sensitivity to amphetamine by altering the number of D2-like receptors, a subtype of dopamine receptors in the brain (Bortolozzi et al., 2002). Rats exposed to 2,4-D at 70 mg/kg body weight per day from gestational day 16 to postnatal day 23 and acutely challenged with amphetamine exhibited increased sensitivity to amphetamine and an increase in D2-like receptor density. The increased density depended more on the particular brain region and the sex of the animal than on the timing of the 2,4-D exposure. A reversal to basal density of D2-like receptors did not occur after cessation of 2,4-D exposure.

A recent study investigated the male-mediated reproductive toxicity of a mixture of 2,4-D and picloram similar to Agent White called Tordon 75D® (Oakes et al., 2002b). Male rats were exposed by gavage (5 days/week for 9 weeks) to Tordon 75D® (2.5%, 5%, and 10% solutions, corresponding to approximate Tordon 75D® doses of 37, 75, and 150 mg/kg body weight per day) and then mated with untreated females at various times during the treatment and after an 11-week recovery period. On gestational day 20, pregnant females were killed, and fetuses were weighed and examined for malformations. The positive control, cyclophosphamide, increased postimplantation fetal death, but no effects on fetal survival or malformations were observed in the herbicide-treated groups.

Garabrant and Philbert (2002) reviewed the scientific evidence relevant to reproductive risks posed by 2,4-D exposure. They conclude that there is a lack of reproductive and developmental toxicity by any route of administration at 2,4-D doses that do not exceed 50 mg/kg body weight, a dose that saturates renal clearance mechanisms, and that offspring of treated pregnant animals show mild to moderate alterations in skeletal development only in the presence of overt maternal toxicity. Those conclusions are consistent with the data presented in this and previous updates.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Immunotoxicity

Lee et al. (2001) examined the effect of exposure to a commercial 2,4-D formulation during gestation on the immune response in mice. Pregnant mice were given the formulation in drinking water (0–1.0%, equivalent to 2,4-D at 0– 650 mg/kg body weight per day) on gestational days 6–16. Immune function in the offspring was evaluated 7 weeks after birth. Decreased body weights and minor reductions in kidney weights were seen in the two highest-dose groups (0.1 and 1.0%). Immune alterations were observed only in the highest-dose group. Suppression of the lymphocyte response to mitogens, an increase in relative B-cell counts, and reduction in the number of cytotoxic and suppressor T cells were seen. The humoral immune response, as measured by antibody production against sheep red blood cells, and peritoneal macrophage phagocytic function were not altered. The authors conclude that the effect on human and animal immune function would probably be minimal when 2,4-D is encountered after normal application in the environment. Garabrant and Philbert (2002) reviewed the scientific evidence relevant to possible effects of 2,4-D on the immune system and concluded that there is little evidence of a significant effect at any dose. That conclusion is consistent with the conclusion of this and previous updates,which note that 2,4-D has at most a weak effect on the immune system.

Carcinogenicity

Using a protocal similar to that discussed above, Lee et al. (2000) examined the effect of exposure to a commercial 2,4-D formulation during gestation on urethan-induced lung adenoma in mice. Female offspring of dams exposed to 2,4-D (0–1.0%) on gestational days 6 –16 were given urethan (1.5 mg/g) at the age of 7 weeks to induce pulmonary adenoma. Offspring were examined at the age of 12 weeks for formation of pulmonary adenomas. Gestational 2,4-D exposure did not affect the number of tumors produced, but it did reduce the mean tumor diameter in the highest-dose group. The authors concluded that gestational 2,4-D exposure had no persistent effect on immune cells involved in cell-mediated immunosurveillance mechanisms. Garabrant and Philbert (2002) reviewed the scientific evidence relevant to cancer risks posed by 2,4-D exposure and concluded that there was no experimental evidence that 2,4-D or any of its salts or esters damages DNA and that studies in experimental animals had demonstrated a lack of carcinogenic effects of 2,4-D. Those conclusions are consistent with the conclusions of the present and previous updates.

TOXICITY PROFILE UPDATE OF 2,4,5-T

No relevant studies on the toxicokinetics of 2,4,5-T or the disease outcomes seen in experimental animals after exposure to 2,4,5-T have been published since Update 2000.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Previous updates reviewed several possible mechanisms by which 2,4,5-T may affect biologic systems. Much of the available information suggests that 2,4,5-T may disrupt cellular pathways involving acetylcoenzyme A. Several reports suggested that 2,4,5-T has only weak mutagenic potential but that it may alter the profile of enzymes involved in the metabolism of procarcinogens.

Two recent studies have investigated the mechanisms underlying the cellular effects of 2,4,5-T. A study by Kaya et al. (2000) examined the ability of several herbicides, including 2,4,5-T, to produce genotoxicity in the wing-spot test of Drosophila melanogaster. It was found to increase slightly the frequency of small single spots but not other types of mutant clones. Furthermore, the slight effect was observed only in a particular type of cross. Those data are consistent with a weak mutagenic potential of 2,4,5-T. A study by Yamanoshita et al. (2001) investigated whether low concentrations of 2,4,5-T affect apoptosis in PC12 cells, a cell line of rat pheochromocytoma cells. Exposure to 2,4,5-T concentrations as low as 10–12 g/L increased cell viability and inhibited DNA fragmentation induced by serum deprivation. The authors concluded that because the physiologic mechanisms leading to cell death are necessary for the normal development of tissues, the inhibitory effect of 2,4,5-T on those mechanisms might cause damage by interrupting normal cell homeostasis and differentiation.

TOXICITY PROFILE UPDATE OF CACODYLIC ACID

Cacodylic acid was present (at 4.7%) in a herbicide that was used in Vietnam in defoliation and crop-destruction missions. The active ingredient in cacodylic acid is dimethylarsinic acid (DMA), which is a metabolite of inorganic arsenic in humans; inorganic arsenic is known to cause cancers in humans. Because of possible concerns that the health effects seen following exposure to inorganic arsenic might be seen after exposure to cacodylic acid, the committee discussed whether studies of inorganic arsenic are relevant to its conclusions. Dimethylarsinic acid is resistant to hydrolysis, and is not demethylated to inorganic arsenic. Although dimethylarsinic acid is formed and is an active metobolite in humans following inorganic arsenic exposure, as discussed in Chapter 2, it has not been established and cannot be inferred that the effects seen following exposure to inorganic arsenic occur following exposure to cacodylic acid. Therefore, in general, the literature on inorganic arsenic is not considered in this report. The reader is referred to Arsenic in Drinking Water (NRC, 1999) and Arsenic in Drinking Water: 2001 Update (NRC, 2001) for further details on the effects of inorganic arsenic. The toxicokinetics of inorganic arsenic as they relate to cacodylic acid formation are discussed below.

Toxicokinetics

Arsenic forms reactive metabolites that affect cellular respiration in nearly every organ system in the body. It was thought for years that methylation of

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

inorganic arsenic was a detoxification process, but recent studies have disputed that theory. The initial methylation of arsenic yields pentavalent monomethylarsonic acid (MMAv), which is reduced to trivalent monomethylarsonous acid (MMAIII) and further methylated to pentavalent dimethylarsinic acid (DMA v). DMA is further reduced to dimethylarsinous acid (DMAIII), which is methylated to form trimethylarsine oxide (Styblo et al., 2000). The route of excretion is primarily the urinary system. As discussed in Arsenic in Drinking Water (NRC 1999), in most animals the DMA that is formed is rapidly excreted in the urine, but in rats DMA accumulates in the red cells and tissues. The pentavalent arsenic species (MMAv and DMAv) are less toxic than the trivalent ones. MMAIII is about 4 times more toxic than inorganic arsenic following acute exposure; the toxicity of DMAIII is similar to that of arsenic III (NRC, 2001).

Mechanisms of Toxic Action

A primary mechanism of the acute toxicity of arsenic is interference with cellular respiration, but recent attention has been devoted mostly to understanding the carcinogenic properties and pathways of arsenic. Inorganic arsenic, a known human carcinogen, does not induce neoplasia in laboratory animals, but cancer has been induced in the urinary bladder, kidneys, liver, thyroid glands, and lungs of laboratory animals by exposure to high concentrations of the metabolite DMA (IOM, 2001; Kenyon and Hughes, 2001; NRC, 2001). The mechanisms responsible for those neoplasms remain unknown. Recent studies have suggested that DMA may act through induction of oxidative damage (Yamanaka et al., 2001) or damage to DNA (Kenyon and Hughes, 2001; Mass et al., 2001; Noda et al., 2002; Sordo et al., 2001). Another recent study demonstrated that DMA caused necrosis of the epithelium of the urinary bladder followed by regenerative hyperplasia (Cohen et al., 2001).

Disease Outcomes

Few animal studies are available on the noncancer health effects of cacodylic acid. Previous reports indicate that cacodylic acid is fetotoxic and teratogenic in rats and mice but only at high, maternally toxic doses (Kenyon and Hughes, 2001). Cacodylic acid acts as a tumor promotor in several organ systems. In a recent initiation-promotion study, however, cacodylic acid given in the drinking water at 220 ppm for 29 weeks did not act as a promotor of kidney tumors in male NCI-Black Reiter rats initiated with N-ethyl-N-hydroxyethylnitrosamine (Vijayaraghavan et al., 2000). In another study, a dose-dependent increase in the incidence of transitional-cell carcinoma occurred in the urinary bladder of male rats given cacodylic acid at 50 or 200 ppm in the drinking water for 104 weeks starting at the age of 10 weeks (Wei et al., 1999). The authors conclude that cacodylic acid is a weak carcinogen. In another study by Seike et al. (2002), oral

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

administration of cacodylic acid at 400 ppm did not exert promoting effects in the lungs of male F344 rats initiated with N-bis (2-hydroxypropyl) nitrosamine.

TOXICITY PROFILE UPDATE OF PICLORAM

Picloram and 2,4-D are components of Agent White, a herbicide formulation used in Vietnam. Studies reviewed in previous updates and in VAO reported a fairly rapid elimination of picloram and suggest carcinogenic and some neurologic effects of exposure but only at extremely high doses. Some cellular abnormalities in liver and inconsistent developmental effects have also been reported.

Two relevant studies of picloram have been published since Update 2000, both focusing on its potential reproductive effects. A study by Oakes et al. (2002b) investigated the possibility of male-mediated reproductive toxicity of a mixture of 2,4-D and picloram similar to Agent White, called Tordon 75D®. As discussed earlier, male rats were exposed to Tordon 75D® by gavage for 5 days per week for 9 weeks at 37 (low dose), 75 (medium dose), or 150 mg/kg of body weight per day (high dose). The 9-week treatment caused a reduction in testicular weight in some animals treated with the highest dose. The small testes had shrunken tubules and germ-cell depletion that was still evident in some rats after a 21-week recovery period. There were no significant differences in the serum concentration of testosterone between control animals and treated animals. In a related study by the same investigators (Oakes et al., 2002a), each of the males exposed to the three doses of herbicide was mated with two untreated females during weeks 2 and 3, 4 and 5, and 8 and 9 of treatment and with four untreated females after an 11-week recovery period. Negative control males were treated with distilled water, and positive controls with cyclophosphamide. On day 20 of gestation, litter size, fetal weight, and fetal malformation rate were all unaffected by herbicide treatment. The positive and negative controls showed the expected results. The results of those studies suggest that exposure to herbicide formulation containing 2,4-D and picloram can cause male-mediated birth defects or other adverse reproductive outcomes.

TOXICITY PROFILE UPDATE OF TCDD

Toxicokinetics

The distribution of TCDD and other chlorodibenzo-p-dioxin congeners has been examined extensively in animal models and to a smaller extent in humans over the last two decades. Similar planar halogenated aromatic hydrocarbons (PHAHs), especially the polychlorinated dibenzofurans and non-ortho-polychlorinated biphenyls, have also been examined extensively. As discussed in numerous papers reviewed in previous reports (VAO and Updates 1996, 1998, 2000), those chemicals are hydrophobic and tend to be readily absorbed across cell membranes.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Properties of the chemicals, properties of the organs and cells, and the route of exposure affect the partitioning, absorption, and accumulation of chemicals. TCDD is distributed to all compartments of the body in amounts that differ from organ to organ. Lipid content is a major factor in the accumulation of TCDD and other PHAHs in different organs and in the body as a whole. Biologic processes, especially metabolism, can affect the distribution and elimination of the chemicals. The concentration of a chemical in a given organ or tissue therefore depends on the dose, absorption, lipid content, and metabolism in the organ of concern. TCDD metabolism can be influenced by processes in other organs. For example, the capacity of the binding protein cytochrome P450 1A2 (CYP1A2), which is prominently expressed in the liver, can alter the accumulation of TCDD in extrahepatic organs.

Since Update 2000, several refinements have been made in the understanding of processes that affect the distribution and elimination of TCDD, including efforts to refine PBPK models in animals and humans. Those models are important in exposure assessment because they can be used to extrapolate from measured concentrations, usually in serum, to estimate initial doses in humans.

Animal Studies

Studies in rodent models continue to support the value of PBPK models to predict the disposition of TCDD. Wang et al. (2000) showed that a PBPK model developed for female Sprague-Dawley rats could accurately predict the disposition of TCDD in male Sprague-Dawley rats and in different strains of rats and mice. Evans and Andersen (2000) refined a “steady-state” PBPK model that successfully described the dose-dependent distribution of TCDD in adipose and liver tissues of rats. The analyses showed further that the induction of CYP1A2 affects the maximal accumulation in the liver but that an inflection point in the curve describing the accumulation was affected by conditions associated with TCDD binding to the AhR and to the binding of the activated receptor complex to DNA.

Diliberto et al. (2001) described the distribution of TCDD in mice after a subchronic low-dose exposure. They used repeated dosing at multiple times and doses to examine the disposition of TCDD and used the induction of CYP1A1 as a marker of an effective dose. The dosing regimen was thought to emulate the most likely mode of exposure in humans, by diet. The dosing regimens resulted in nearly steady-state concentrations of TCDD. Hepatic CYP1A1 activity was measurably induced by TCDD at 0.15 ng/kg body weight per day (a body burden of TCDD of 2.8 ng/kg of body weight). The study emphasized the cumulative effects of daily dosing and the importance of determining tissue dosimetry or body burden for chemicals with long half-lives such as TCDD.

Hurst et al. (2000) examined the distribution of TCDD in exposed dams and rat pups exposed to TCDD in utero by using a low-dose subchronic exposure; the regimen achieved the desired steady-state distribution. The objective of the study

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

was to obtain information to aid in developing models to assess the potential risk associated with such exposure in humans. The authors determined that low-dose maternal exposure results in low concentrations of TCDD in the fetus and that the subchronic exposure produced concentrations in the fetus similar to those seen after a single higher dose during gestation. The study also adds to the information about relationships between maternal and fetal TCDD; individual embryo concentrations of TCDD were 1.6, 7, and 16 pg/g of body weight after maternal exposure to TCDD at 1, 10, and 30 ng/kg body weight per day. On day 16, TCDD concentrations were similar in fetal liver, urogenital tract, head, and all other parts of the body together; this suggests that there was no preferential sequestration in the fetal liver. There was evidence, however, of sequestration in liver as soon as 4 days after birth.

Chen CY et al. (2001) further examined the disposition of polychlorinated dibenzo-p-dioxin (PCDD), polychlorinated biphenyl (PCB), and polychlorinated dibenzofuran (PCDF) congeners in female rats and their offspring. Mixtures of nine dioxin-like compounds were given to female Long-Evans rats at various doses to determine transfer to the placenta, fetuses, and pups. Pharmacokinetic differences among the chemicals influenced the transfer of TCDD from dam to offspring. Those differences were the result of preferential sequestration of planar dioxins and furans by CYP1A2. The authors emphasize, however, that uncertainties remain regarding the sequestration of such compounds by CYP1A2 in humans and that the uncertainties need to be clarified for human risk assessment.

A study of rats exposed during gestation and through lactation (Iba and Fung, 2001) showed that the effects of TCDD, as indicated by CYP1A induction, in the offspring persisted longer than might be expected on the basis of the half-life of TCDD in rats; the effects were more prolonged in lung than in liver. The study also revealed a pronounced induction of CYP1A2 protein in lung, supporting earlier suggestions that CYP1A2 can be induced in some extrahepatic organs, especially the lungs of rodents. Whether CYP1A2 is induced in human lung is not clear, although there have been reports of CYP1A2 transcript in human lung (Mace et al., 1998). There is also evidence suggesting CYP1A2 expression in human bronchial mucosa and peripheral lung tissue (Wei et al., 2001).

Studies modeling the disposition and effects of TCDD in rodents provide evidence that supports the development and use of PBPK models in humans. Comparisons of humans and laboratory animals, however, may not be as accurate as comparisons among rodent species. Van Birgelen and van den Berg (2000) point out that the well-known differences in pharmacokinetic behavior of TCDD in humans and rodents (principally the rates of metabolism, which is slower in humans, and the possible differences in the role of CYP1A2) indicate that a lower daily intake would be needed in humans to reach tissue burdens similar to those in rats. The authors emphasize that estimation of daily intake will be important in applying PBPK models to humans.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Human Studies

Human populations of concern in assessing exposure and risk include those directly exposed to TCDD at high concentrations by accident, in the workplace, or in Vietnam. Comparison populations exposed at background concentrations are also important for assessing the risk posed by higher exposures. In a study by Arfi et al. (2001) of 16 people in France exposed only to background TCDD, the TCDD in benign lipomas of patients was evaluated in relation to their mean daily dietary TCDD intake. The mean daily dietary TCDD intake in France in TCDD equivalents (TEQs) is 1–4 pg TEQ/kg body weight per day—similar to the intake in other industrialized countries. The authors did not find a correlation between the daily dietary dioxin intake and TCDD in the lipomas.

The possibility that TCDD residues might be transferred from more highly exposed to less highly exposed people was examined in a study by Manz et al. (2001) of 14 workers in Germany exposed to TCDD occupationally and their female partners who were not occupationally exposed. Analysis of blood and fatty tissue showed that the female partners accumulated substantial TCDD—as much as 10% of the concentrations in the tissues of their male partners. The underlying mechanism or pathway of transfer reflected in those results is not clear. The authors did not report on TCDD in semen, a possible route of transfer to the female partners. Earlier studies have measured TCDD in semen samples from Vietnam veterans (Schecter et al., 1996); the findings suggest a possible mechanism of male-mediated adverse reproductive outcomes after Agent Orange or other dioxin exposure.

The elimination of TCDD from highly exposed people has been examined in a study of two patients who were exposed to and had accumulated extremely high concentrations of TCDD—at 144,000 pg/g of blood fat in one patient and 26,000 pg/g of blood fat in the other (Geusau et al., 2001). A route of elimination seldom studied is through the skin, and cutaneous elimination of TCDD was examined in those patients. Elimination via skin was assessed in one of the patients by comparing TCDD concentrations in material collected at various times from the skin surface and in cerumen and epithelial cysts with blood concentrations. The authors conclude that elimination via the skin, most probably in cellular material, accounted for 1–2% of the daily elimination. But it is possible that skin is one avenue by which nonexposed partners of highly exposed people could be exposed to TCDD concentrations somewhat greater than in the general population.

Various attempts to accelerate the elimination of TCDD were made in the two patients of Geusau et al. (2002), principally by continuous administration of Olestra. Olestra had been shown to enhance TCDD elimination (discussed in Update 2000). It was given at three different doses; at the highest dose, 66 g/day, fecal excretion of TCDD was increased by factors of 10 and 8 for the more and less severely contaminated patients, respectively. At Olestra treatments of 33 g/ day, fecal excretion was increased by factors of 4.9 and 6.7, respectively. Efforts

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

were made to remove lipids from the blood (LDL-apheresis) of the more severely contaminated patient. Although the binding of TCDD to VLDL (see Dalton et al., 2001) suggests that such a treatment might be effective, it did not have a major impact on TCDD body burden.

The relationship between concentrations of chemical residues in blood to those in other tissues is important for assessing tissue dose and elimination rates based on blood concentrations. Elimination of TCDD residues is generally through fecal excretion, which may include residues not absorbed and residues eliminated in bile. The concentrations of 20 dioxin-like compounds (seven PCDDs, 10 PCDFs, and three PCBs) in bile were compared with concentrations in blood and liver in 27 autopsy cases (Kitamura et al., 2001). Total TEQs were the same in bile and blood but were higher in liver. Correlation between blood and bile TEQs was high: a correlation coefficient of 0.89 among the 27 autopsy cases. The study estimated that the rate of accumulation of dioxins, in TEQs, was 0.99, 0.70, and 1.91 pg/g of lipid per year in bile, blood, and liver, respectively.

Half-Life Studies

A number of recent studies are revealing substantial variation in the half-life of TCDD in humans that is associated with a number of variables. Dose and time after exposure have been identified as important. A summary of half-lives estimated in studies in humans and animals is presented in Table 3-1.

Miniero et al. (2001) reviewed data concerning the half-life of TCDD and how it correlates with body weight. The major determinants of TCDD half-life are thought to be lipophilicity, metabolism, and sequestration in the liver, but it seems to correlate empirically with the body weight of mammals. To evaluate that correlation, Miniero et al. (2001) regressed half-life measures with body weight and found a significant correlation between them. Other factors did not seem to influence the dispersion of points about the regression line. That suggests that although uncertainties about the role of CYP1A2 in TCDD distributions in humans remain, metabolism might not affect the half-life substantially. The data may be relevant to a minimal physiologic toxicokinetic model being developed by Salvan and colleagues to describe the long-term behavior of TCDD at the individual level (Bortot et al., 2002; Salvan et al., 2001). The model principally describes the variations of TCDD in serum lipids in terms of body mass, not tissue distribution or sequestration of TCDD in the liver, and it has been used for exposure assessment in studies to identify TCDD effects (Schnoor et al., 2001).

The two highly exposed patients studied by Geusau et al. (2001) showed overall TCDD half-lives of 1.5 and 2.9 years in the more and less severely contaminated patients, respectively. Those half-lives are considerably shorter than values commonly reported: 7.2 years in the Boehringer cohort (Flesch-Janys et al., 1996) and 6.9 and 9.8 years in men and women, respectively, in the Seveso cohort (Michalek et al., 2002) (see below and Table 3-1).

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

TABLE 3-1 Estimates of TCDD Half-Life in Humans and Animals

Reference

Half-lifea

Confidence Interval

Comment

Human Studies

Pirkle et al., 1989

7.1 yr

5.8–9.6 yr

Adult males, Ranch Hands,

9–23 yr PE

Michalek et al., 2002

7.5 yr

 

Adult males, Ranch Hands

9–33 yr PE

Flesch-Janys et al., 1996

7.2 yr

 

Adult males, Boehringer cohort

Needham et al., 1994

7.8 yr

7.2–9.7 yr

Adults, Seveso cohort

Michalek et al., 2002

6.9 yr

 

Adult males, Seveso cohort,

3–16 yr PE

 

9.8 yr

Adult females, Seveso cohort,

3–16 yr PE

 

0.34 yrb

Adult males, Seveso cohort,

0–3 mo PE

Geusau et al., 2002

1.5 yrb

 

Adult female, severe exposure

0–3 yr PE

 

2.9 yrb

Adult female, severe exposure

0–3 yr PE

Animal Studies

Viluksela et al., 1996

20.2 days

 

Rats, Long-Evans TurkuAB strain

28.9 daysc

 

Rats, Long-Evans Charles River strain

Weber et al., 1993

16.3 ± 3.0 days

 

Rats, male Sprague-Dawley

Pohjanvirta et al., 1990

21.9 days

 

Rats, male Han/Wistar resistant strain

Neubert et al., 1990

73.7 days

60.9–93.8

Monkey, Marmoset, single injection

aHalf-lives of TCDD in humans based on measurement of TCDD in serum samples.

bShorter half-lives measured in humans during first months after exposure or in severely contaminated persons consistent with nonlinear elimination predicted by PBPK modeling (e.g., by Carrier et al., 1995). Greater half-life in females attributed to greater body mass index.

cAttributed to differences in dilution due to different growth rates.

ABBREVIATION: PE, postexposure.

The toxicokinetics of TCDD were examined in adults exposed to TCDD in the Seveso accident and were compared with data on TCDD elimination in the veterans of Operation Ranch Hand (Michalek et al., 2002). Serum TCDD concentrations in the Seveso cohort were available from samples obtained within days of exposure, providing a measured initial dose, whereas the earliest serum measurements in the Ranch Hand population were obtained in 1982 and at 5-year intervals thereafter to 1997, or 9–33 years after initial exposure. The mean half-life in the Seveso males during the first 3 months after exposure was 0.34 year. The mean half-life in males during the period from 3 to just over 16 years after exposure was 6.9 years. The mean half-life in the Ranch Hands 9–33 years after

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

exposure was 7.5 years, which is barely significantly different from the latter rate in Seveso males. In the Seveso females, the half-life during the slower phase of elimination, 9.8 years, was longer than that in males. The authors point out that the fast initial elimination in the Seveso cohort is consistent with the pattern expected on the basis of a two-compartment model and with temporal changes in whole-body elimination observed in rodents. The slower elimination in females has been observed in other studies as well (see Update 1998 and Update 2000 for discussion of studies).

In a related study, Jackson and Michalek (2001) presented follow-up temporal changes in TCDD concentrations in the Vietnam-era veterans who were not occupationally exposed to herbicides and from whom serum samples were available from 1987, 1992, and 1997. Among those veterans, serum TCDD concentrations decreased by 0.25 parts per trillion (ppt) per year. The decrease in the comparison veteran group from 1987 to 1992 was similar to the decline in serum TCDD observed in Germany from 1989 to 1994 and is considered to reflect a decline in background TCDD.

Van der Molen et al. (2000) used a pharmacokinetic model described in an earlier study (Van der Molen et al., 1996) to assess elimination in longitudinal and cross-sectional studies on the basis of previously published data (Flesch-Janys et al., 1996; Schrey et al., 1993). In the Van der Molen model, body composition, body weight, and intake rate are assumed to depend on age, and elimination rate is assumed to depend on body composition. The model also treats background intake rates as an input, avoiding the subtraction of background concentrations from observed concentration in persons temporarily exposed to higher doses. It does not consider sequestration in the liver. The model 's prediction that elimination rate must change with age is consistent with other observations. Differences in half-life could be due to differences in a number of variables, including body mass index, weight, initial dose, time after exposure, and age. Differing half-lives and a biphasic elimination of TCDD could confound the extrapolation from serum measurements to initial exposure. Whether a biphasic curve like that which might be inferred from the Seveso cohort could be used in exposure reconstruction is not certain, but the possibility should be tested.

There have been continuing efforts to identify substances that can aid in the elimination of TCDD and similar compounds from the body. Morita et al. (2001) determined that chlorophyll can inhibit absorption of dioxin and dibenzofuran in rats and enhance their excretion; fecal excretion of seven PCDD congeners and 10 PCDF congeners was increased with increasing dietary chlorophyll. In a related study, Morita and Nakano (2002) observed that seaweed, common in the Japanese diet, also accelerated the elimination of PCDD and PCDF congeners. But they observed that the elimination was principally of metabolites and that the amount of parent compound eliminated was less than 10% of the total; in humans, parent compounds of the congeners constituted 37–90% of the total eliminated. Morita and Nakano (2002) did see an increased elimination of parent

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

compound in rats when fed seaweed, but whether seaweed would have a similar effect in humans is not known.

To summarize the information on toxicokinetics, there is now considerable understanding of the toxicokinetic behavior of TCDD. The data continue to show that body composition (that is, percentage of fat) is a key determinant of disposition and half-life at low body burdens (background exposures). At higher exposures, CYP1A2 binding holds the dioxins in the liver and results in faster elimination. Therefore, there is both dose-dependent tissue distribution, with the amount in liver relative to that in fat increasing as the dose goes up, and dose-dependent elimination. New data on individuals exposed to very high concentrations of TCDD have shown that at such high doses, the half-life can be quite short relative to that in individuals exposed to background or only moderate concentrations. This is graphically illustrated in the two highly exposed Viennese women (Geusau et al., 2001) and in the new analysis of Seveso exposure data (Michalek et al., 2002). Despite the substantial knowledge, however, it still is quite difficult to reconstruct initial exposure levels from blood levels taken much later.

Mechanisms of Toxic Action

Studies published since Update 2000 are consistent with the hypothesis that TCDD produces its biologic and toxic effects by binding to a gene regulatory protein, the aryl hydrocarbon receptor (AhR). The mechanistic model indicates that binding of TCDD to the AhR, dimerization of the AhR with a nuclear protein (AhR nuclear transport protein, or Arnt), and interaction of this complex with specific DNA sequences (Ah-responsive elements, or AhREs, and dioxin-responsive elements, or DREs) present in the 5'-promoter regions of responsive genes lead to the inappropriate modulation of gene expression. Those molecular changes are the initial steps in a series of biochemical, cellular, and tissue changes that result in the toxicity observed. That hypothesis is supported by numerous studies that have evaluated structure-activity relationships of various chemicals that bind to the AhR, the genetics of mutant genes that express the AhR, AhR-deficient mice, and the molecular events contributing to and regulating AhR expression and its activity. Additional details of the events have been uncovered since Update 2000. The exact relationships between the modulated expression of known regulated or modulated genes (Table 3-2) and the diversity of toxic effects elicited by TCDD in humans and numerous animal species, however, have yet to be uncovered.

The finding that many AhR-regulated genes are modulated in a species-, cell-, and developmental stage-specific pattern suggests that the molecular and cellular pathways leading to a particular toxic event are complex. Many of the data are consistent with the notion that cellular processes involving growth, maturation, and differentiation are most sensitive to TCDD-induced modulation as mediated by the AhR. The findings in animals continue to indicate that reproduc-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

TABLE 3-2 Genes and Proteins Known to Be Modulated by TCDD and/or Dioxin-like Chemicals

Reference

Genes

Genes and Proteins Directly Regulated by AhR

Poland and Knutson, 1982

CYP1A1

Tukey and Nebert, 1984

CYP1A2

Sutter et al., 1994

CYP1B1

Pimental et al., 1993

glutathione-S-transferase Ya

Takimoto et al., 1992

aldehyde dehydrogenase 4

Favreau and Pickett, 1991

NAD(P)H-menadione oxidoreductase 1

Lamb et al., 1994

UDP glucuronosyltransferase1

Krishnan et al., 1995

cathepsin D (inhibition); Sp1 (inhibition)

Gillesby et al., 1997

pS2 (inhibition)

Porter et al., 2001

heat shock protein 27 (inhibition)

Gaido and Maness, 1994

plasminogen activator inhibitor-2

Jeon and Esser, 2000

interleukin-2

Kraemer et al., 1996

cyclooxygenase-2

Gao et al., 1998

ecto-ATPase

Genes and Proteins Suspected to Be Directly Regulated by AhR

Rivera et al., 2002

CYP2S1

Masten and Shiverick, 1995

BSAP

Lai et al., 1996

transforming growth factor-beta (TGF-β)

Matikainen et al., 2002

Bax

Lai et al., 1996

interleukin-6; interferon-gamma

Kim et al., 2000

c-myc

Sugawara et al., 2001

steroidogenic acute regulatory protein

Ogi et al., 2001

polκ

Ohbayashi et al., 2001

DIF-3

Genes and Proteins Modulated by Posttranscriptional Mechanisms

Gaido et al., 1992

transforming growth factor-alpha (TGF-α); urokinase plasminogen activator

Dong et al., 1997

MHC Q1

Puga et al., 1992

c-fos; c-jun

Genes and Proteins Reported to Be Altered by AhR Ligand Exposure

Vogel and Abel, 1995

tumor necrosis factor-alpha (TNF-α)

Shridhar et al., 2001

corticotrophin-releasing hormone

Park and Lee, 2002

Hrk; interleukin-3 receptor-beta (IL-3β)

Svensson and Lundberg, 2001

adseverin

Kolluri et al., 1999

p27Kip1

Ohsako et al., 2001

5-alpha reductase 2

Nukaya et al., 2001

low molecular weight prekinonogen

Nishimura et al., 2001

metallothionein

Ma et al., 2001

poly(ADP-ribose) polymerase

Kakeyama et al., 2001

NMDA receptor

Yang et al., 2001

carboxylesterase

Oikawa et al., 2002

IgE-dependent histamine-releasing factor

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Reference

Genes

Ishimura et al., 2002

glucose transporter 3

Tian et al., 1998

estrogen receptor

Roth et al., 1988

malic enzyme

Sewall et al., 1995

epidermal growth factor receptor (EGFR)

Poland and Glover, 1973

ALA synthetase

Sparrow et al., 1994

pyruvate carboxylase

Kolluri et al., 2001

N-myristoyltransferase 2

Mathieu et al., 2001

multidrug resistance 1

Sugihara et al., 2001

xanthine oxidase/dehydrogenase

Sutter et al., 1991

interleukin-1beta (IL-1β)

Krig and Rice, 2000

transglutaminase

Puga et al., 2000a

guanine nucleotide exchange factor; Ki-ras2 proto-oncogene; semaphorin; inositol 1,4,5-triphosphate receptor; MEK5; advillin; casein kinase 1 delta; RAY1; SHP-2 tyrosine phosphatase; MST2 serine/threonine kinase; phosphoinositide 3-kinase; phospholipase A2; calcium-modulating cyclophilin; calmodulin; neurogranin; phosphatidic acid phosphatase; S100 calcium-binding protein A4, A7, and A12; FRK, fyn-related kinase; calcineurin A; visinin-like factor 1; phospholamban; calbindin 1; spindle pole body protein; p56 Lck; FLT1, fms-related tyrosine kinase; protein kinase C-beta and zeta; IkB-alpha; PKA regulatory subunit; phosphatidylinositol 4-kinase; serine/threonine kinase 2; Thy-1 cell surface antigen; A-kinase anchor protein; CD3E; E74-like Ets-domain transcription factor; Erg-1 and Erg-2 transcription factors; MAD; MADS box transcription enhancer; frizzled homolog; FREAC-2 forkhead-like protein; troponin C; inducible NO synthase; G protein-coupled endothelin; endothelial NO synthase; follistatin; FGL2, prothombinase; arginine vasopressin receptor 1A; 5-lipoxygenase-activating protein; midkine; very low density lipoprotein receptor; coagulation factor XII; neuropeptide Y receptor Y1; vascular endothelium growth factor; multiple exostoses 2; dermatan sulphate proteoglycan; decorin; thrombomodulin; granzyme A; cyclin B2; human RACH1; matrilin 2; lamin B receptor; caspase-1 and 4; PDCD2; P107, RB-related protein; glycogen synthase kinase 3 beta; tumor necrosis factor members 3, 6, 8, 9 and 10; CC3; growth arrest-specific GAS-1; anti-mullerian hormone receptor; heat-shock protein hsp-40; NEK-2 serine/threonine kinase; breakpoint cluster region; matrilin 2; LIM domain; alpha (1,3) fucosyltransferase; integrin beta 1 and 3; CD63 antigen; translation initiation factor 4; homeobox HB9 and Pax 3; bagpipe homeobox homolog; epimorphin; CD47 antigen; phosphatidylinositol glycan, class C, H, and K;

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Reference

Genes

Puga et al., 2000a

dystrophin-associated glycoprotein 1; mannosidase alpha type II; VAMP 8; VAMP-associated protein; ankyrin 2, neuronal; gamma-butyrobetaine hydroxylase; epimorphin; membrane fatty acid desaturase; sulfotransferase 2B family member; N-methylpurine-DNA glycosylase; thioredoxin peroxidase; 3-hydroxybutyrate dehydrogenase; suppressor of Ty homolog; estrogen sulfotransferase; mammalian mutS homolog; endonuclease G; cytochrome c-1

Frueh et al., 2001

G protein-coupled receptor HM74; agrin precursor-like protein; enhancer of filamentation; XMP; cytochrome b5; DNA-binding protein inhibitor ID-2; aquaporin 3-like protein; mannose-binding protein C precursor; coagulation factor XI; arylacetamide deacetylase; cytochrome P450 subfamily XIX; calponin; endothelial actin-binding protein; phospholipase A2, membrane-associated precursor; keratin 17; apolipoprotein C-1 precursor; lectin galactdoside-binding soluble 3 (galectin 3); phospholipase D; ATP synthase lipid-binding protein P1 precursor; glutaminyl-tRNA synthetase; hybrid receptor gp250 precursor; glutamate-cysteine ligase regulatory subunit; inwardly rectifying potassium channel Kir3.2; aminopeptidase N; aminoacylase-1; B2-bradykinin receptor, 3; histidine ammonia-lyase; L-myc-1-proto-oncogene protein; YL-1 protein; fibrogen alpha chain precursor; thyroxin-binding globulin and globulin precursor; SPARC/osteonectin; alcohol dehydrogenase 1 alpha polypeptide; fibrinogen gamma-B chain

Kurachi et al., 2002

RAB11a and RAB3D (members of RAS oncogene family); Ral-A protein; interferon-inducible GTPase; insulin-like growth factor binding protein 3 and ALS; calcium binding protein A11; regucalcin; S100 calcium binding protein A1; cyclin-dependent kinase inhibitor 1A; proteasome 26S subunit; proprotein convertase subtilison/kexin type 6; metaxin 2; ubiquitin-like 1; tyrosine 3-monooxygenase activation protein; Sin3-associated polypeptide; eukaryotic translation initiation factor s 2 and 3; eukaryotic translation elongation factors 1 and 2; nucleobindin; heterogeneous nuclear ribonucleoproteins C and K; high mobility group protein 1; histone gene complex 1; YY1 transcription factor; zinc-finger protein 207; upstream transcription factor 2; histone deacetylase 5; nuclear RNA export factor 1 homolog; homeodomain interacting protein kinase 1; RNA polymerase 1-3; basic transcription factor 3; ribosomal proteins S6, L9, S8, L7a, S3; RNase A family 4;

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Reference

Genes

Kurachi et al., 2002

splicing factor, arginine/serine-rich 2 (SC-35); endothelial monocyte activating polypeptide 2; protein that interacts with C kinase 1; GrpE-like 1, mitochondrial; chaperonin subunits 3 and 4; enoyl coenzyme A hydratase, short chain 1; phosphatidylcholine transfer protein; organic anion transporter member 10; FXYD domain-containing ion transport regulator 1; ATPase-like vacuolar proton channel;ATP-binding cassette, sub-family G; neurophilin; tenomodulin; CD82 antigen; poliovirus sensitivity; LDL receptor related protein, associated protein 1; integral membrane protein 3; natural killer tumor recognition sequence; syndecan 4; pigment epithelium-derived factor; argininosuccinate synthetase 1; cytochrome P450, 2d10; protein phosphatase 1, catalytic subunit; tryptophan-2,3-dioxygenase; sialyltransferase 9; acetyl-coenzyme A dehydrogenase, long chain; ornithine decarboxylase; ketohexokinase; plasmin inhibitor alpha 2; phosphoenolpyruvate carboxykinase 1; stearoyl-coenzyme A desaturase 1; sorbitol dehydrogenase 1; phosphoglycerate kinase; ornithine transcarbamylase; galactokinase; lactate deyhydrogenase 1, A chain; lysosomal acid lipase 1; carbonyl reductase 1; carbonic anhydrase 3 and 5; cytochrome c oxidase, subunits Va and VIIb; glucose regulated protein, 58 kDa; heat shock protein cognate 70; cell death-inducing DNA fragmentation factor; heat shock 10kDa protein 1 (chaperonin 10); clusterin; fatty acid binding protein 1, liver; presenilin 2; GM2 ganglioside activator protein; peroxisome biogenesis factor 16; male enhanced antigen 1; mucolipin 1; gene trap ankyrin repeat; apolipoproteins E and CI; serum albumin variant;major urinary protein; alpha-2-HS-glycoprotein; ferritin heavy chain; serine protease inhibitors 1-3; beta-2 microglobulin;hepcidin antimicrobrial peptide; complement components 1, 3, and 4; plasminogen; transthyretin; haptoglobin; alpha-2-glycoprotein 1, zinc; kininogen; alpha-2-macroglobulin; glutathione peroxidase; profilin 1; melanoma X-actin; destrin; gelsolin; keratin complex 2, basic gene 1; prefoldin 2

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

tive, developmental, and oncogenic end points are very sensitive to TCDD. The data support the biologic plausibility of similar end points of toxicity in exposed humans. However, many of the responses are tissue- and species-specific, and the exact mechanistic basis of the differences is not known.

The conclusions indicated above are similar to those in Update 2000. Since that update, many interactions of the AhR at the cell and molecular levels have been reported. However, in many cases it is not clear how these might be related to a particular toxic end point. Therefore, although the text below references all related work published since Update 2000 that was identified by the committee, closer attention is given only to studies that added substantial new information, particularly as it might be relevant to the exposure of veterans in Vietnam. As discussed in Update 2000, it is important to consider exposure and species sensitivity when discussing animal data and their relevance to humans.

Structural and Functional Aspects of the AhR

The AhR Gene and Protein Several studies have characterized AhR structure, expression, and function in different animal species, including zebrafish (Andreasen et al., 2002a,b), rainbow trout (Pollenz et al., 2002), Atlantic killifish (Karchner et al., 2002), chick embryos (Walker MK et al., 2000), and guinea pigs (Korkalainen et al., 2001). In general, the published data are consistent with the conservation of AhR structure and function among species. Zebrafish, however, express two AhR-like molecules; one is not responsive to TCDD, and the other appears to be active in mediating cardiovascular toxicity in developing animals (Andreasen et al., 2002b). The closest homologue of the human AhR is the guinea pig receptor (Korkalainen et al., 2001); this is of interest because the guinea pig is one of the species most susceptible to TCDD-induced lethality.

There are many data on the ability of various regions of the AhR protein to function in ligand binding, DNA binding, nuclear localization, and interaction with other proteins, including Arnt. New information has added to our understanding of those regions (Kronenberg et al., 2000; Andreasen et al., 2002c; Berg and Pongratz, 2001; Elbi et al., 2002; Ikuta et al., 2002; Jones and Whitlock, 2001; Kumar et al., 2001; Levine et al., 2000). Exposure of cells and animals to relatively high TCDD concentrations has been shown to stimulate pathways that mediate the degradation of the AhR, leading to a subsequent decrease in AhR-mediated gene alterations. Additional data have also improved our understanding of those pathways (Ma and Baldwin, 2000; Pollenz and Barbour, 2000; Santiago-Josefat et al., 2001). Notably, recent data indicate that environmentally relevant concentrations do not appear to change AhR concentrations in the rat (Franc et al., 2001). It is of interest that nuclear localization and transcriptional activation of the AhR have been found to occur in the absence of an exogenous ligand, such as TCDD (Richter et al., 2001). That finding might indicate the existence of an endogenous ligand for the AhR whose identification would greatly increase our

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

knowledge of AhR function and of how the inappropriate stimulation of the AhR by TCDD may lead to toxic effects.

A recent publication characterizes the regulatory regions of the Ahr gene (Garrison and Denison, 2000). The data suggest that the expression of the gene depends on acetylation of nuclear histone components (Garrison and Denison, 2000; Garrison et al., 2000). Three polymorphisms in the human Ahr gene have been reported at codons 517, 554, and 570 in exon 10 (Smart and Daly, 2000; Wong et al., 2001a,b). Exon 10 is the major region that is responsible for the transactivation of other genes. In vitro assays using expressed variant forms of the AhR indicate that none of those individual polymorphisms has an important effect on the ability of the AhR to induce the CYP1A1 gene in a TCDD-dependent manner (Wong et al., 2001b). A combination of the lysine554 + isoleucine570 variant and the lysine554 + isoleucine570 + serine517 variant, however, was unable to support induction of CYP1A1 by TCDD. That suggests that people with that combined polymorphism might be less susceptible to some of the effects of TCDD. Cauchi et al. (2001) identified several human Ahr polymorphisms, but none was found to be associated with altered CYP1A1 inducibility or with susceptibility to lung cancer. Roberts et al. (2000) reported that TCDD failed to induce the CYP genes in a human hepatoma cell line and that this was probably because of the presence of a defective AhR protein.

Interaction of the AhR with Other Proteins The function and regulation of the AhR depends on the presence of several other intracellular proteins. Additional studies have been published on the interactions of the AhR with a protein known as X-associated protein 2 (XAP2; also called ARA9 and AhR-interacting protein, or AIP) (Bell and Poland, 2000; Kazlauskas et al., 2000, 2002; LaPres et al., 2000; Petrulis et al., 2000), with 90-kilodalton (90-kDa) heat-shock protein (hsp90) (Heid et al., 2000; Kazlauskas et al., 2001), and with p23 (Cox and Miller, 2002). Those proteins have been shown to function in stabilizing the AhR in cells and in regulating the intracellular localization of the AhR. An additional study indicates that tyrosine phosphorylation of the AhR is required for its DNA-binding activity (Park et al., 2000). Interactions of the AhR with the retinoblastoma protein (Elferink et al., 2001), tyrosine kinases (Dieter et al., 2001), the orphan receptor COUP-TF (Klinge et al., 2000), the short heterodimer partner (SHP) orphan nuclear receptor (Klinge et al., 2001), myb-binding protein 1a (Jones et al., 2002), the Brahms/SWI2 gene 1 protein (Wang and Hankinson, 2002), the silencing mediator of retinoic acid and thyroid hormone receptors (SMRT) (Rushing and Denison, 2002), and the NcoA/SRC-1/p160 family of transcriptional coactivator proteins (Beischlag et al., 2002) may also modulate the ability of the AhR to regulate genes.

Update 2000 noted the identification of an AhR repressor (AhRR) protein that inhibits AhR function by competing with the AhR for dimerization with Arnt. Additional data on this protein from human, mouse, and killifish (Karchner

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

et al., 2002) are consistent with previous information. They also indicate that AhRR is evolutionarily conserved. Notably, recent data indicate the association of an AhRR polymorphism (proline to leucine at the 185 position) with micropenis in humans (Fujita et al., 2002); this is particularly relevant because TCDD exposure to animals is known to affect the development of reproductive tissue.

Chemicals Other Than TCDD That Affect AhR Function As indicated in previous updates, data on the ability of various dioxin-like chemicals to bind to the AhR and cause toxicity support a role of the AhR in mediating the toxicity of those chemicals; newer information is consistent with that role (Simanainen et al., 2002). Recent data, however, also indicate that relative potency values derived from one end point of toxicity, in this case lethality, are not necessarily valid for other end points of toxicity.

Since Update 2000, a number of chemicals have been found to modulate AhR function by directly binding to the AhR. Several of those chemicals—such as indirubin and indigo (Adachi et al., 2001), some prostaglandins (Seidel et al., 2001), and 7-ketocholesterol (Savouret et al., 2001)—appear to act as AhR agonists. Notably, these molecules are found in human and animal tissues. Other chemicals, such as the naturally occurring flavonoids (Ashida et al., 2000a; Ciolino et al., 1999; Quadri et al., 2000) and synthetic flavonoids (Nazarenko et al., 2001), also bind to the AhR, but some of them act as antagonists. The ability of some PCBs to have agonist or antagonist activity appears to depend on their relative affinity for the AhR and properties related to their intrinsic efficacy, that is, their ability to produce a response once bound to the AhR (Hestermann et al., 2000). The ability of PCBs to bind to the AhR and elicit a conformation that would bind to DREs in vitro and induce CYP1A1 in whole cells has been used to detect AhR agonist and antagonist activity (Petrulis and Bunce, 2000). The work by Pohjanvirta et al. (2002) emphasizes the point that although a chemical like indolo[3,2-b]carbazole can bind to the AhR and act as an AhR agonist, it might not elicit the same toxic responses as TCDD. That is probably because of the rapid metabolism of the chemical, and it further suggests that prolonged occupation of the AhR and persistent modulated gene expression may be necessary for some toxic end points. Additional work has been done to understand the molecular constraints responsible for the ability of the dioxin-like chemicals to bind to the AhR (Arulmozhiraja et al., 2000; Mhin et al., 2002).

Other chemicals—such as resveratrol (Lee and Safe, 2001), some heavy metals (Maier et al., 2000), and bisphenol A (Jeong et al., 2000)—appear to block TCDD- and AhR-dependent gene transcription by mechanisms not related to their ability to bind the AhR.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
AhR-Mediated Alterations of Gene Expression

Much of our current understanding of the mechanism of TCDD action is based on analysis of the induction of particular genes. Several genes known to be modulated by TCDD and dioxin-like chemicals in a variety of biologic systems, including human cells, are listed in Table 3-2, which includes several genes published since Update 2000. Genes in which either mRNA or protein concentrations have been shown to be altered are included, but enzymes or proteins whose biologic activities are altered by some other mechanisms are not. Several genes are known to be modulated by direct interaction of the AhR –Arnt complex with DREs in the promoter region; recent data indicate that the induction of some of these genes failed to occur when mice in which the expression of Arnt was conditionally disrupted were treated with TCDD (Tomita et al., 2000). Other genes are suspected, but not yet proved, to be induced by this mechanism. The expression of several genes—such as cathepsin D, Sp1, pS2, and heat-shock protein 27—has been shown to be inhibited by the ability of the AhR to bind to DREs near the DNA-binding sites for the estrogen receptor. The expression of other genes, including transforming growth factor-alpha (TGF-α) and MHC Q1, is thought to be altered by posttranscriptional mechanisms. Finally, Table 3-2 includes genes modulated in a variety of cells or tissues in several species, including humans, but of which the mechanisms of alteration are not yet understood; it is quite likely that the induction or repression of many of these genes might be secondary to the ability of the AhR–Arnt complex to act directly on other genes. The size of the latter category emphasizes the ability of the ligand-bound AhR to initiate a cascade of molecular and biochemical events that eventually leads to cell and tissue alterations. That these events are known to be tissue-, species-, and developmental stage-specific also emphasizes the complex nature of the biochemical events that lead to some particular toxic response. Several recent investigations of those gene changes have been carried out in human cells (Frueh et al., 2001; Puga et al., 2000a).

Data on several individual genes are discussed below in the context of particular tissue systems or toxic end points that might be affected by TCDD. However, this should not be interpreted to indicate that the ability of TCDD to modulate the expression of a particular gene is limited to that particular tissue; although the effects of TCDD are very tissue- and cell-specific, genes or biochemical pathways modulated in one tissue are often found to be modulated in several other tissues.

Mechanisms Related to Particular Toxic End Points

As indicated in previous updates, an accumulation of studies in experimental animals indicates that TCDD affects a variety of tissues, and the type of effect observed is often tissue-specific. In addition, effects are most often dose-depen-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

dent, that is, some toxic end points appear to be most sensitive to low exposures, and others may occur only at high concentrations. Furthermore, effects have been found to depend on the species examined and often on the age and sex of the animal. There is no reason to suspect that humans would be different in that respect. Findings in animals suggest that reproductive, developmental, and oncogenic end points are the most sensitive to TCDD and are consistent with the notion that growth, maturation, and differentiation are the most sensitive cellular processes. Those data support the biologic plausibility of similar toxic end points in humans. Although the exact biochemical mechanisms of those end points and the observed differences are not yet understood, recent data have emphasized the possibility that at least some of the effects are mediated by TCDD's ability, through the AhR, to modulate cell-cycle control, signaling pathways that lead to cell death, hormones and growth factors and the responses to them, or the biochemical pathways that lead to oxidative stress (Barouki and Morel, 2001; Nebert et al., 2000). Those mechanisms are implicated in many of the toxic end points discussed below.

Mechanisms Related to Wasting Syndrome Exposure of most animal species to relatively high doses of TCDD elicits a wasting syndrome characterized by decreased food consumption and loss of body weight. The biochemical pathways affected by TCDD that lead to the syndrome have not been identified. Tuomisto et al. (2000) observed that TCDD treatment alters food intake and food selection in rats. A study by Dunlap et al. (2002) demonstrated that the absence of c-src protein kinase expression affects the development of several TCDD-elicited toxic end points—such as decreased body-weight gain, adipose tissue to liver weight ratio, decreased weight of pancreas, glycogen depletion, and phosphoenolpyruvate carboxykinase downregulation —that are related to the syndrome. Previous updates discuss the effect of TCDD on vitamin stores in animals. Two additional studies suggest that altered retinoid homeostasis, particularly an increased mobilization of vitamin A from storage sites mainly in the liver, leading to increased serum and kidney retinoic acid may contribute to the syndrome (Fletcher et al., 2001; Kelley et al., 2000; Nilsson et al., 2000). Glover et al. (2000) observed that rats treated with TCDD had increased sensitivity to endotoxin, which resulted in increased nitric oxide. Previous studies had suggested that endotoxin is a contributing factor in TCDD-induced wasting syndrome, possibly through its effects on the stimulation of cytokine production, which can suppress appetite.

Mechanisms Related to Effects on Skin and Adipose Tissue Skin lesions, including chloracne, are often reported in animals and humans after exposure to TCDD and related chemicals. Loertscher et al. (2001a) observed that TCDD exposure reduces the number of normal human keratinocytes grown under culture conditions, but not because of increased cellular apoptosis. Using an immortalized human keratinocyte line, those investigators also observed that TCDD

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

caused alterations in the pattern of terminal differentiation without increased apoptosis or altered cellular proliferation (Loertscher et al., 2001b). Krig et al. (2002) investigated the mechanism whereby TCDD suppresses retinoid induction of the transglutaminase gene in human keratinocytes. They found that the suppressive action of TCDD is at the transcriptional level but that it occurs indirectly through a DNA site outside a 5-kilobase region of the TGM2 promoter and does not directly interfere with retinoid action or at the retinoid response element in this gene.

As indicated in previous updates, TCDD has been shown to inhibit the differentiation of some preadipocyte cell lines to adipocytes, and this process is AhR-dependent. Several research groups have examined the mechanism of that effect because it may be relevant to how TCDD acts in various tissues. Shimba et al. (2001) demonstrated that overexpression of the AhR in fibroblast cells in the absence of TCDD exposure suppresses morphologic differentiation and induction of adipocyte-related genes, whereas underexpression of the AhR induces differentiation and expression of the genes. The authors conclude that the AhR is a negative regulator of adipose differentiation and that regulation can occur independently of TCDD exposure. Liu et al. (2002) present data suggesting that the antiadipogenic action of TCDD is related to the expression of C/EBPα, a factor believed to coordinate genes involved in lipogenesis. Kern et al. (2002a) observed a decrease in catalase activity in the adipose tissue of TCDD-treated rats. Nagashima and Matsumura (2002) established that TCDD causes downregulation of glucose uptake activity in preadipocytes. Kern et al. (2002b) observed a decrease in glucose transport in a TCDD-treated adipocyte line; in addition, TCDD stimulated secretion of TNF and decreased lipoprotein lipase activity. The authors conclude that this may indicate a physiologic mechanism for epidemiologic studies linking dioxin exposure to diabetes.

Mechanisms Related to Effects on Bone and Teeth Possible effects of TCDD on bone have not been thoroughly investigated. Jamsa et al. (2001) studied effects of 19 weekly TCDD treatments amounting to total TCDD doses of 0.17– 170 µg/kg body weight in young adult rats. Tibial growth was inhibited in a dose-dependent manner, but there was no effect on bone mineral density. Breaking force and stiffness were reduced by TCDD at 17 µg/kg body weight. Resistance of the H/W rat strain to those effects was associated with an altered transcription domain of the AhR. Singh et al. (2000) observed that TCDD inhibits osteogenesis in a chicken periosteal osteogenesis model and that this effect is restricted primarily to the osteoblastic differentiation phase. Collagen type I, osteopontin, bone sialoprotein, and alkaline phosphatase mRNA content were decreased. Similar results were observed with a rat stromal bone cell line. And TCDD induced a reduction in bone mineralization. All those effects were antagonized by cotreatment with resveratrol, which is found at high concentrations in red wine. The authors postulated that the high concentrations of AhR ligands in cigarette smoke,

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

might be important in linking smoking to osteoporosis and periodontal disease. Partridge et al. (2000) noted that low concentrations of TCDD and estrogen affected synthesis or secretion of parathyroid-stimulated collagenase-3 in a rat osteoblast osteosarcoma cell line.

Previous reports have suggested that defects in children's first molars may be associated with environmental exposure to dioxins. Render et al. (2000) observed proliferation of periodontal squamous epithelium in mink fed TCDD at 5 ppb for 6 months. Sahlberg et al. (2002) reported that AhR and Arnt are coexpressed in developing mouse tooth buds, especially in secretory odontoblasts and ameloblasts. Allen and Leamy (2001) found that in utero exposure to TCDD did not affect the fluctuating asymmetry of mandibles in mice, but did decrease mandible size and affect their overall shape.

Mechanisms Related to Cardiovascular Toxicity There is a paucity of information on the potential for TCDD to exert toxic effects on the mammalian cardiovascular system. However, a recent study by Riecke et al. (2002) noted a dose-dependent increase in the incidence of myocardial fibrosis in marmosets 2 or 4 weeks after a single dose of TCDD at 1, 10, or 100 ng/kg body weight. Further analysis indicated increased transforming growth factor β1 (TGFβ1) and TGF-β receptor type I in heart tissue. The authors suggested a relation to the myocardial fibrosis in that this factor is known to cause fibrosis in many tissues of experimental animals. Additional studies in chicken embryos, which appear to be very susceptible to cardiac effects after TCDD exposure, noted that the activation of the AhR signaling pathway correlates with the ability of various dioxin-like chemicals to induce cardiotoxicity (Heid et al., 2001) and that TCDD-induced apoptosis may contribute to the observed changes in myocyte proliferation, coronary development, and structural malformations (Ivnitski et al., 2001).

Mechanisms Related to Pulmonary Toxicity Iba and Fung (2001) observed that gestational and lactational exposure of rats to TCDD caused long-lasting and sex-dependent induction of CYP1A1 and CYP1A2 protein in the lungs. Wei et al. (2001) observed both CYP1A1 and CYP1A2 in lung biopsy specimens from human subjects.

Genter et al. (2001) indicated that TCDD can induce many metabolic enzymes—including CYP1A1, CYP1A2, CYP2B1, CYP2C11, and epoxide hydrolase —in rat olfactory mucosa and that the pattern of the changes was different from that in liver. The increase in nasal mucosal enzymes resulted in enhanced metabolism of lidocaine. The authors concluded that environmental exposure to TCDD may affect drugs administered nasally.

Mechanisms Related to Hepatotoxicity The liver is a primary target organ of TCDD and related chemicals, but the severity of effects can vary considerably

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

among species. The liver and its cells are often used to study the effects of TCDD on biochemical pathways that may be responsible for toxic end points.

Porphyria cutanea tarda, the most common clinical form of porphyria, has been reported in humans exposed to hexachlorobenzene and TCDD. A study in CYP1A2-null mice by Smith et al. (2001) demonstrated that the presence and inducibility of this gene are necessary for the uroporphyrinogenic effects of TCDD and contribute to TCDD-induced hepatocellular injury. Injury probably occurs through the metabolism of uroporphyrinogen III to uroporphyrin III, which cannot be used for heme synthesis. Data published by Robinson et al. (2002), however, suggest that a gene in addition to Ahr may modulate hepatic porphyria and injury caused by TCDD in mice.

Since the last update, the ability of TCDD or dioxin-like compounds to elicit changes in the expression of several genes or change the activity of proteins has been observed in liver cells from a number of species. They include the multidrug-resistance gene (Mathieu et al., 2001), the N-myristoyltransferase 2 gene (Kolluri et al., 2001), carboxylesterase (Yang et al., 2001), the low-molecular-weight prekininogen gene (Nukaya et al., 2001), poly (ADP-ribose) polymerase (Ma et al., 2001), and CYP2S1 (Rivera et al., 2002). Additional data on the abilty of TCDD to modulate the activity of the transcription factors AP-1, c-myc, and NF-κB (Ashida et al., 2000b; Puga et al., 2000b) and to induce CYP1A1 (Kono et al., 2001; Korner et al., 2002), CYP1B1 (Shehin et al., 2000), and an ecto-ATPase gene (Gao and Whitlock, 2001) have been published since Update 2000. Santini et al. (2001) observed that the induction of CYP1A1 in mouse hepatoma cells depends heavily on the stage in the cell cycle, and is markedly suppressed in G2/ M cells. However, Muller et al. (2000) noted that TGF-β1 inhibits TCDD-induced CYP1 activities in primary rat hepatocytes. Ashida et al. (2000b) suggested that changes in activity of c-myc and AP-1 may be affected by TCDD-induced alterations in protein kinase and phosphatase activities. Shimba et al. (2000) also reported that TCDD treatment may upregulate the urokinase-type plasminogen activator gene by inducing binding of a 50-kDa protein to the mRNA. Several recent publications reported the use of advancing technologies, including microarrays, to analyze TCDD-induced gene changes in mouse (Kurachi et al., 2002) and human liver-tumor cells (Frueh et al., 2001; Puga et al., 2000a). Holman et al. (2000) used synchrotron infrared spectromicroscopy to determine that TCDD exposure to human liver-tumor cells results in a relative increase in the number of methyl–methylene groups in single cells; this suggests an increase in DNA methylation and altered gene expression. A publication by Kohn et al. (2001) described a model to characterize the relationships between rat liver concentrations of TCDD and enzyme induction.

Several research groups have suggested the induction of cellular oxidative stress as a mechanism by which TCDD could elicit damage via the AhR and lead to many of the toxic end points observed, including liver injury. Since Update 2000, several publications have examined the mechanism whereby that may

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

occur. Subchronic or acute exposure of rats and mice to TCDD has been shown to increase production of liver superoxide anion, thiobarbituric acid-reactive substances (evidence of lipid peroxidation), and DNA single-strand breaks (Bagchi et al., 2002; Hassoun et al., 2001, 2002; Slezak et al., 2000). Several other investigations suggested that changes in tissue catalase and glutathione peroxidase activities (Kern et al., 2002a), suppression of carbonic anhydrase III (Ikeda et al., 2000), or induction of xanthine oxidase–dehydrogenase activity (Sugihara et al., 2001) may contribute to the oxidative damage observed after TCDD exposure. TCDD treatment of mice was found to increase reactive-oxygen production by liver mitochondria, and liver ATP concentrations were significantly decreased at the peak times of reactive-oxygen production (Senft et al., 2002). In contrast, the induction of metallothionein by TCDD may play a protective role in TCDD-elicited oxidative stress responses (Nishimura et al., 2001). Notably, Arnt protein is also required by hypoxia-inducible factor-1α (HIF-1α) to enhance expression of various genes in response to hypoxia. The data provided by Nie et al. (2001) suggest that exposure to TCDD, via the recruitment of Arnt by the AhR, may repress HIF-1α-inducible responses. Hirai et al. (2002) found that α-tocopherol and L-dehydroascorbic acid, but not vitamin C, protect human cells in culture from toxicity induced by very high concentrations of TCDD.

Update 2000 reported that TCDD, via the AhR, blocks many estrogen-induced responses (see “Mechanisms Related to Effects on the Mammary Gland” below). Stanton et al. (2001a,b) demonstrated that TCDD treatment strongly blocked several responses of male chickens to estrogen, including hepatic lipid synthesis and metabolism and weight gain.

Mechanisms Related to Gastrointestinal Tract Effects Sterling and Cutroneo (2002) observed that TCDD and benzo[a]pryene induce CYP1A1 in rat small intestinal epithelial cells and human colon carcinoma cells. It cannot be concluded from those data alone, however, that human colon cells are responsive to TCDD; the authors noted that induction depended heavily on the presence of extracellular matrix and the differentiation state of those cells.

Mechanisms Related to Neurotoxicity Few studies have examined the possibility of nervous system damage in adult animals exposed to TCDD; the developing brain appears to be more sensitive (see “Developmental Toxicity” below). However, observed behavioral alterations —such as anorexia, weight loss, changes in circadian rhythm, and altered reproductive behavior—suggest that the nervous system may be affected even in adult animals. Huang et al. (2000) demonstrated that the AhR and Arnt are expressed in rat brain and pituitary and that CYP1A1 is induced in these tissues after TCDD exposure. Chronic exposure of Cynomolgus monkeys to TCDD was found to increase corticotropin-releasing hormone mRNA in the paraventricular nucleus of the hypothalamus (Shridhar et al., 2001). As in the liver, exposure to TCDD was shown to increase brain

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

oxidative stress as determined by increased superoxide anion, lipid peroxidation, and DNA single-strand breaks (Bagchi et al., 2002; Hassoun et al., 2001, 2002). The AhR was expressed and CYP1A1 was induced by TCDD in a rat glial cell line. In addition, cyclic-AMP-induced differentiation, as measured by extension of astrocyte processes and the induction of glial fibrillary acidic protein, was inhibited by TCDD; TCDD's inhibiting effect was blocked by an AhR antagonist (Takanaga et al., 2001). The AhR and Arnt have been found to be widely expressed throughout the brain and brainstem (Petersen et al., 2000).

Mechanisms Related to Immunotoxicity The animal immune system is highly sensitive to the toxic effects of TCDD, but the primary cell targets remain unclear. Multiple cell types make up this system, and many undergo rapid proliferation and differentiation in response to stress or a foreign antigen. It is possible, and indeed highly likely, that many cell types at different stages of development might be affected by TCDD.

In a recent review, Kerkvliet (2002) suggested that TCDD may cause the inappropriate activation of cells and lead to anergy or cell death and the premature termination of the immune response. Recent data on T-cell responses (Camacho et al., 2001; Fujimaki et al., 2002; Nohara et al., 2002a) and dendritic cell function (Vorderstrasse and Kerkvliet, 2001) in mice after TCDD exposure are consistent with that hypothesis. Hematopoietic stem cells also have been suggested as sensitive targets for TCDD (Murante and Gasiewicz, 2000). That effect in particular may contribute to a previously demonstrated reduction in the capacity of bone marrow from TCDD-treated mice to generate pro-T lymphocytes associated with elicited thymic atrophy. TCDD may also target thymocytes directly by inducing cell-cycle arrest (Lai et al., 2000).

The exact genes or biochemical alterations induced by TCDD leading to those effects are not known. Since Update 2000, TCDD has been shown to increase the production of TNF-1 by peripheral lymphocytes from exposed rhesus monkeys (Rier et al., 2001a), inhibit AP-1 activity in activated B cells (Suh et al., 2002), induce the expression of adseverin in mouse thymus (Svensson and Lundberg, 2001; Svensson et al., 2002), and induce the expression of the genes encoding Hrk and interleukin-3 receptor in Jurkat T cells (Park and Lee, 2002). TCDD has been shown not to affect the complement system in the guinea pig (Wagner et al., 2001). The role of the AhR in TCDD-induced immunotoxicity has been described in the previous updates, and additional data consistent with a role of the AhR have been published recently (Vorderstrasse et al., 2001).

Mechanisms Related to Carcinogenesis As indicated in previous updates, TCDD has been demonstrated to be a potent tumor promoter in several model systems. Its ability to induce cell proliferation and to alter differentiation is believed to be an important factor in the mechanism of TCDD-induced carcinogenesis. A study by Thornton et al. (2001) used Big Blue® lacI transgenic rats to

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

assess the mutagenicity of TCDD. After 6 weeks of exposure to TCDD at 2 µg/kg body weight, there was no increase in mutation frequency or change in mutation spectrum; this is consistent with many previous reports indicating the low mutagenic potential of this chemical. Also consistent with earlier data is the observation by Ramakrishna et al. (2002) of an increase in tumor multiplicity in mouse N-nitrosodimethlyamine-initiated lung tumors after treatment with a single dose of TCDD at 1.6 µg/kg body weight. The authors observed a significant decrease in membrane-associated K-ras protein p21 but an increase in raf-1; they hypothesized that these biochemical alterations constitute one mechanism by which TCDD may promote tumors.

Chronic bioassays have shown TCDD to increase the incidence of hepatic tumors in female, but not male, rats. A recent study by Wyde et al. (2002) indicated that this may be explained by the weaker potency of tumor promotion and DNA damage in male rats compared with females and lack of induction of cell replication in females. The investigators also found that hepatic 8-oxodeoxyguanosine adduct formation, a measure of oxidative DNA damage, was significantly higher in TCDD-treated female rats and depended on the presence of 17-β estradiol (Wyde et al., 2001a). However, when diethylnitrosamine-treated ovariectomized rats were treated for 20 or 30 weeks with TCDD in the presence and absence of 17-β estradiol supplementation, estradiol supplementation did not appear to increase hepatotoxicity, as determined histopathologically and with analysis of serum characteristics (Wyde et al., 2000).

Several of the enzymes induced by TCDD, including CYP1A1 and CYP1A2, are responsible for the metabolic activation of many promutagens, so the activation of the AhR is considered to be important for the carcinogenic activity of many chemicals. Machala et al. (2001) determined that the ability of several polycyclic aromatic hydrocarbons to induce AhR-mediated gene expression could contribute to their mutagenic potential. Similarly, Jeffrey et al. (2002) observed that both activation of the AhR pathway and metabolism of benzo[a]pyrene are necessary for the transcriptional repression of the BRCA-1 gene by benzo[a]pyrene. Repression of this gene may be a predisposing event in the onset of sporadic breast cancer. However, Uno et al. (2001) observed that although the lack of the CYP1A1 gene in CYP1A1-knockout mice protected the animal from benzo[a]pyrene-mediated liver toxicity and death by decreasing the formation of large amounts of toxic metabolites, there was a paradoxic increase in liver benzo[a]pyrene–DNA adduct formation because of the slower metabolic clearance of this chemical.

All available data suggest that the AhR plays a role in TCDD-induced tumor promotion. An interesting study by Andersson et al. (2002) found that transgenic mice that had a constitutively active AhR had reduced life span and induced tumors of the glandular part of the stomach. Up to the age of 6 months, the animals also had decreased thymic weights and increased liver weights, which were consistent with a simulation of low-dose exposure to TCDD. Notably, sev-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

eral species of animals treated with AhR ligands have previously been reported to develop lesions of the glandular stomach. A recent study showed that TCDD and four AhR antagonists inhibited the growth of human pancreatic cell lines in a dose-dependent manner (Koliopanos et al., 2002).

Greenlee et al. (2001) observed that the proliferation of tumor cells and their ability to invade normal tissue are inhibited in the presence of TCDD. The authors suggest that that might be the basis of some rodent bioassays that show a decrease in the incidence of mammary tumors after TCDD exposure. On the other hand, there is data cited in this and earlier updates to suggest that under some conditions TCDD exposure may increase the risk for the development of breast tumors (see below and “Biologic Plausibility” for Breast Cancer in Chapter 6). Clearly, additional work is needed to determine how different exposure factors may affect breast tumor development.

Mechanisms Related to Effects on the Testis Many effects of TCDD in male rodents have been reported previously, including decreases in the size of accessory sex organs and in daily sperm production. Other investigations suggest that TCDD may cause tissue damage by induction of oxidative stress. Those findings are consistent with results of studies by Latchoumycandane et al. (2002a,b) in rat testis. Subchronic treatment of adult male rats with TCDD at 1–100 ng/kg of body weight per day resulted in a dose-dependent decrease in epididymal sperm counts and a decline in the activities of superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase. There was also an increase in hydrogen peroxide and lipid peroxidation in testis. Similar changes were seen in the mitochondrial and microsomal fractions of testis. AhR activation was found by Ogi et al. (2001) to enhance the expression of the mouse Polκ gene. That gene is a low-fidelity DNA polymerase that appears to be developmentally regulated in the mouse testis. TCDD was also found to induce expression of DIF-3, a gene implicated in spermatogenesis in mouse embryonic stem cells (Ohbayashi et al., 2001). TCDD treatment had no effect on the production of testosterone by isolated mouse testicular cells, primary cultures of rat Leydig cells, or mouse Leydig tumor cells (Mandal et al., 2001; Uchida et al., 2002), although a single dose of TCDD at 50 µg/kg body weight resulted in a 75% decrease in serum testosterone in adult male rats. Mandal et al. (2001) concluded that the effect of TCDD on testosterone in rats may not be a direct result of decreased capacity of Leydig cells to synthesize steroids.

Mechanisms Related to Effects on the Prostate TCDD has been shown to induce CYP1A1 in three human prostate cancer cell lines (Jana et al., 2000; Schaufler et al., 2002). In the study by Jana et al. (2000), although expression of the CYP1B1 and PA1-2 genes were induced by TCDD in breast and uterine human cell lines, they were not induced in LNCaP prostate cells. TCDD also inhibited testosterone-induced reporter gene activation in all three cell lines,

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

suggesting that TCDD and the target steroid hormones negatively regulate each other's activity. No effects on cell-cycle distribution and cell growth were observed. CYP1B1 was induced in the PC3 and DU145 human prostate cell lines (Schaufler et al., 2002). Using human prostate-carcinoma cells, Schrader and Cooke (2000) observed that TCDD partially inhibited 5α-dihydrotestosterone-induced gene induction.

Mechanisms Related to Effects on the Ovary Immature female rats treated with TCDD before gonadotropin-induced follicular development and ovulation have been shown to produce significantly fewer ova than untreated control animals. A review by Petroff et al. (2001) indicated that a blockade of ovulation by TCDD probably involves actions at several target tissues and on multiple intracellular pathways. For example, they found an effect on matrix metalloproteinase in the follicular wall, altered hypothalamohypophyseal mechanisms responsible for the ovulatory luteinizing hormone surge, and effects on the expression of several genes including those that encode for cyclooxygenase-2, plasminogen activator inhibitor-2, and the progesterone receptor.

Data from Roby (2001) indicate that TCDD treatment alters the ability of the ovary to respond to gonadotropin and results in the abnormal development of follicles as assessed by lower gonadotropin binding, lower estradiol production, and lower concentrations of cAMP. Treatment of rats with gonadotropin-releasing hormone was able to partially restore ovulation inhibited by TCDD. The authors interpreted those data as confirming a direct effect of TCDD on the hypothalamic–pituitary axis and the ovary (Gao et al., 2000). A later study confirmed that TCDD decreased the responsiveness of the hypothalamus to estrogen as a feedback inducer of gonadotropin secretion (Gao et al., 2001). Blockade of ovulation by TCDD was alleviated by treatment with estradiol, although the systemic toxicity, as determined by weight loss, was increased (Petroff et al., 2000). A study by Petroff et al. (2002) suggested that inhibin, a member of the TGF superfamily, did not have a role in TCDD-elicited effects on the ovary. Recent investigations have suggested that the AhR-dependent alteration of Bax expression may have a role in developing fetal ovarian germ cells (Matikainen et al., 2002).

The AhR and Arnt were found to be expressed in rat ovary and liver, but the relative expression depended on the estrous cycle (Chaffin et al., 2000). Human uterine tissue and ovarian tissue were also found to express the AhR and Arnt (Khorram et al., 2002), but the relative expression was altered in some pathologic conditions, such as endometriosis and uterine leiomyoma.

TCDD has been reported to have a number of biochemical effects on cultured granulosa cells, including CYP1A1 and CYP1B1 induction and an increase in estrogen receptor-β mRNA (Dasmahapatra et al., 2001), reduction in expression of mRNA for P450scc and P450arom involved in the steroid biosynthetic pathway (Dasmahapatra et al., 2000), increases in luteinizing hormone receptor

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

protein and mRNA (Hirakawa et al., 2000), and alerations in the secretion of estradiol and progesterone (Pieklo et al., 2000). Gregoraszczuk et al. (2001) also observed that TCDD treatment decreased progesterone secretion of luteal cells. Studies by Rogers and Denison (2002) suggest that TCDD induces a factor that inhibits estrogen-dependent gene expression in human ovarian-carcinoma cells.

Mechanisms Related to Effects on the Uterus TCDD has been shown to decrease uterine weights in rodents and alter endometrial structure. Buchanan et al. (2000) examined the relative contributions of uterine stromal and epithelial cells to TCDD-induced effects. TCDD inhibited estradiol-induced epithelial-cell mitogenic and secretory activity, but the effects appeared to be mediated indirectly through the AhR in stromal cells. The authors postulated that TCDD impairs uterine epithelial function by altering normal stromal–epithelial interactions. Additional work by Buchanan et al. (2002) indicated that changes in epithelial-cell mitogenesis might involve changes in cell cycle and in cyclin and TGF-β expression.

Bulun et al. (2000) found transcripts of the AhR, Arnt, and several AhR target genes—including CYP1A1, CYP1A2, and CYP1B1—to be expressed in human endometriotic tissues. Pitt et al. (2001) found that expression of AhR mRNA in cultured human endometrial explants increases with age and that Arnt mRNA expression is tissue-phase- and age-dependent. The presence of endometriosis did not change the expression of the AhR and Arnt. Both CYP1A1 and CYP1B1 were induced by TCDD in those explant cultures (Bofinger et al., 2001). Hasan and Fischer (2001), however, observed that a change in AhR localization in rabbit uterus occurred in pregnancy and shortly before the expected time of implantation. Those data suggest that maternal steroid hormones may regulate AhR expression in this tissue. Neither estrogen nor progesterone inhibited induction of CYP1A1 activity by TCDD in human Ishikawa endometrial-cancer cells, although TCDD blocked estradiol-induced responses. However, estrogen was shown to inhibit induction of CYP1A1-dependent enzyme activity in ECC-1 endometrial cells (Wormke et al., 2000a). Both CYP1A1 and plasminogen activator inhibitor-2 were induced in the human RL95-2 uterine cell line (Jana et al., 2000). Coexpression of the estrogen receptor increased responsiveness of these cells to TCDD.

Some concern exists about the possibility that TCDD increases the prevalence of endometriosis in humans. Some of the evidence of such an effect comes from animal studies that have been summarized by Birnbaum and Cummings (2002). Although the mechanisms remain unclear, the authors suggest that effects of the dioxin-like compounds on growth factors, cytokines, and hormones may mediate the promotion of endometriosis.

Mechanisms Related to Effects on the Mammary Gland Human breast-cancer cells have been useful for investigating the mechanisms of AhR signaling

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

and the effects of TCDD on hormonally induced responses, especially responses to estrogen. Update 2000 reported that TCDD has been shown to block many estrogen-induced responses in human breast-cancer cells. Wang et al. (2001) reported additional data on the mechanism whereby TCDD inhibits estradiolmediated induction of the cathepsin D gene via the AhR. TCDD was shown by Porter et al. (2001) to inhibit estradiol-induced heat-shock protein 27 (hsp27) gene by a similar mechanism, that is, the binding of the AhR to a DRE present in the gene-promoter region. Using suppression subtractive hybridization, Chen I et al. (2001) identified 33 genes in human breast-cancer MCF-7 cells that are induced by estradiol and inhibited by AhR agonists and that may be important for mediating the antiestrogenic activity of TCDD. Additional work by Wormke et al. (2000b) also suggested that TCDD-induced activation of proteosomes and later increased degradation of estrogen receptor-α may contribute to the noted antiestrogenic activity of TCDD. Notably, expression of a constitutively active AhR in human MCF-7 cells enhanced expression of CYP1A1, inhibited estrogen-dependent cathepsin D expression, and inhibited growth of these cells (Kohle et al., 2002).

Angus et al. (2000) found that exposure of human breast epithelial cells to TCDD results in a time-dependent increase in membrane protein kinase erbB2 and erbB3 and leads to cell proliferation. The authors suggest that TCDD might facilitate the transition of breast cells from being estrogen-dependent to being estrogen-independent. That has been shown to be a key step in the progression of breast cancer and the overexpression of erbB2 and erbB3, which are negative prognostic indicators for survival. Studies by Davis et al. (2001) suggest that TCDD inhibits apoptosis in a human mammary epithelial cell line by stimulating the production of TGF-α, which results in the activation of the epidermal growth-factor receptor pathway. Kim et al. (2000) found that TCDD was able to induce the c-myc promoter in MCF-10F cells by a mechanism that required the interaction of the AhR with RelA and the binding to NF-κB elements in the promoter; the author suggested this may be a novel mechanism by which the AhR can stimulate proliferation and tumorigenesis of mammary cells.

TCDD was shown to induce the expression of CYP1A1, CYP1B1, and plasminogen activator inhibitor-2 in MCF-7 cells (Jana et al., 2000). Guo et al. (2001) found that the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) suppresses TCDD-induced CYP1A1 through a protein kinase C-dependent process in MCF-10A cells; alteration of TGF-β activity may also contribute to this suppression.

Mechanisms Related to Other Endocrine Effects As indicated in previous updates, TCDD has been shown to affect the thyroid and thyroid hormones in several animal species. TCDD treatment of rats increased thyroid-stimulating hormone concentrations in the anterior pituitary, changed thyroid structure in a way consistent with accelerated biosynthesis of T4 in the thyroid, but signifi-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

cantly decreased both serum T4 and free T4 concentrations. The UDP-glucuronosyltransferase-1 gene was induced in the liver at TCDD doses as low as 1 µg/kg body weight. The authors suggest that induction of UGT1 is the main event that triggers the alterations in thyroid-hormone homeostasis. Additional studies by Craft et al. (2002) suggest that species differences in the response to chemicals that induce alterations in thyroid-hormone status may be due to differential induction of hepatic UGT enzymes.

Work by Pitt et al. (2000) indicates that a single exposure to TCDD during pregnancy does not appear sufficient to induce maternally mediated developmental, reproductive, and behavioral toxicity through an effect on the hypothalamic– pituitary–adrenal axis.

Mechanisms Related to Developmental Toxicity Extensive data from studies in experimental animals suggest that developing tissues are highly sensitive to the toxic effects of TCDD as mediated by the AhR and that tissue growth and differentiation processes are affected.

Matthews et al. (2001) examined the effect of TCDD treatment of mouse embryos in culture on the number of embryos developing from the morula to the blastocyst stage and the number of apoptotic blastomeres; no effects of 3 µM TCDD were observed. And Wu et al. (2002) did not observe CYP1A1 mRNA in mouse embryos at the one-, two-, and eight-cell stages after exposure to TCDD, whereas CYP1A1 mRNA was significantly increased at the blastocyst stage. TCDD exposure had no effect on the rate of development of preimplantation embryos, the cell number of blastocyst embryos, or apoptotic indexes.

Bryant et al. (2001a) examined the effects of loss of EGF and TGF-α expression on the incidence of hydronephrosis and cleft palate in developing mice. Animals expressing no EGF did not develop cleft palate after maternal TCDD exposure. The lack of the EGF or TGF-α genes, however, increased the incidence and severity of TCDD-induced hydronephrosis. Previous studies have found that TCDD produces hydronephrosis by altering the differentiation and proliferation of ureteric epithelial cells in fetal mouse urinary tract. Bryant et al. (2001b) found that late-gestational ureteric cells in vitro respond to TCDD by a stimulation of epithelial growth and differentiation, but that effect was not observed in the presence of EGF over longer culture periods. Notably, immunostimulation was found to decrease the incidence of cleft palate in mice significantly in response to TCDD; the decrease is thought to be due to a modulation of growth factor or cytokine production in developing tissues (Holladay et al., 2000). TCDD was also found to retard lower jaw development and circulation in the zebrafish embryo; however, the reduction in perfusion rate occurred well after the inhibition of jaw development (Teraoka et al., 2002).

Update 2000 cited several reports indicating that development of the male reproductive system is exceptionally sensitive to in utero and lactational TCDD exposure. Impaired prostate growth has been shown consistently, and recent

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

studies confirm it. Ohsako et al. (2001) reported that low-dose administration of TCDD in rats affected the development of the external genital organs and ventral prostate more than the testis and other internal genital organs. They also suggested that the effect of TCDD on the prostate may be due to decreased responsiveness of the prostate to androgen and decreased expression of androgen receptor. Theobald et al. (2000) found that the ability of the ventral prostate to form 5α-dihydrotestosterone was not significantly altered in rats after in utero and lactational exposure. However, the androgen-induced expression of prostate-binding protein subunit C3 was transiently decreased, and the formation of androgen-responsive luminal epithelial cells was inhibited. Altered prostate epithelial-cell differentiation was hypothesized as the major effect leading to impaired prostate growth. Timms et al. (2002) demonstrated that intrauterine position and individual differences in gonadal steroid concentrations influenced the response of the developing prostate to TCDD and that these effects may be mediated by a decrease in serum estradiol. Additional studies by Ohsako et al. (2002) indicate that there is a critical and narrow window of exposure—about gestation day 15 in the rat—during which the fetus is sensitive to these TCDD effects. Using Ahr null-allele mice, Lin et al. (2002) demonstrated that effects depend on the AhR. Notably, they also found that development of several tissues—including liver, heart, spleen, thymus, lung, submandibular gland, testis, epididymis, and kidney—are affected by absence of the AhR (Lin et al., 2001). Haavisto et al. (2001) reported that exposure of male rats in utero caused a stimulatory effect on testicular testosterone synthesis and increased circulating testosterone; these effects were caused by stimulation of pituitary luteinizing-hormone production and enhanced sensitivity of the fetal testis to luteinizing hormone. Slezak et al. (2002) reported that male rat pups exposed perinatally to TCDD showed increased production of reactive oxygen species in the liver, although no alteration in lipid peroxidation or total glutathione was observed.

Lewis et al. (2001) reported that in utero and lactational exposure of female fetuses to TCDD impairs mammary gland differentiation as determined by the distribution of terminal ductal structures and increased expression of estrogen receptor-α. The ability of those tissues to differentiate in response to estrogen, however, was not affected. That result is consistent with data published by Fenton et al. (2002), who also observed reduced lactation and milk production by the offspring when they reach adulthood. Exposure at gestation day 15 appeared to be critical for the inhibition of breast epithelial development.

As indicated in the previous updates, several reports of studies in animals and exposed humans suggest that perinatal exposure to TCDD or dioxin-like chemicals may impair brain development. Recent publications by Markowski et al. (2002) and Hojo et al. (2002) indicate that low-dose prenatal TCDD exposure (less than 0.54 µg/kg body weight) of rats results in subtle behavioral effects, including altered sexually dimorphic behavior and impaired ability to inhibit or delay voluntary behavior. Several publications have documented specific bio-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

chemical changes in the brains of animals after transplacental exposure to TCDD, including altered hippocampal astroglia–neuronal gap junction communication (Legare et al., 2000), changes in NMDA receptor subunit mRNA expression (Kakeyama et al., 2001), alterations in fetal brain aromatase activity (Ikeda et al., 2002), a decrease in serotonin-immunoreactive neurons in raphe nuclei of male offspring that lasted up to the age of 42 days (Kuchiiwa et al., 2002), and altered Sp1 DNA binding in the developing cerebral cortex and cerebellum (Nayyar et al., 2002).

Ishimura et al. (2002) observed that treatment of pregnant rats with a single oral dose of TCDD at 0.8 or 1.6 µg/kg body weight on gestation day 15 increased glycogen content and glucose transporter 3 mRNA. The authors hypothesized that altered glucose kinetics in the placenta contribute to TCDD-induced fetal toxicity.

Disease Outcomes
Lethality and Defects

An unusual characteristic of TCDD is the large variation among species in susceptibility to its lethal effects. Both intraspecies variations and interspecies variation can occur; some of the difference in sensitivity is related to differences in the expression and primary amino acid sequences of the AhR protein. Regardless, most species develop a wasting syndrome when exposed to acute toxic doses of TCDD.

Mink in particular appear to be highly sensitive to the toxic effects of TCDD. In a recent study (Hochstein et al., 2001), mink fed diets containing TCDD at 0.016–1.40 ppb exhibited several dose-dependent adverse health effects, including the wasting syndrome. Theirs was the first report of thickened, deformed, and elongated toenails in TCDD-exposed mink. The effects of TCDD on toenails in mink are reminiscent of metaplasias seen in nail beds of rhesus macaques treated with PCB mixtures (McNulty, 1985) and pigmented or deformed nails in children exposed to PCBs in utero following a poisoning in Taiwan from PCB contaminated cooking oil (Rogan et al., 1988). In another study (Render et al., 2001), loose and displaced incisor teeth were observed in mink kits fed TCDD at 2.4 ppb; radiographs of the jaw showed maxillary and mandibular osteolysis with lysis of the lamina dura.

Kattainen et al. (2001) exposed three lines of F10 generation rats from Hans/ Wistar and Long/Evans crossings to TCDD in utero and via lactation; TCDD at 1 µg/kg body weight prevented the development of the third lower molars in the most sensitive lines. The authors acknowledge that tooth development appears to be a highly sensitive end point of TCDD-induced toxicity. A large proportion of the pups from mother rats exposed 1 day after birth to TCDD at 1,000 µg/kg body weight lacked their third molars (Lukinmaa et al., 2001). However, in contrast

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

with prevention of development of the lower third molars after in utero exposure, the molars missing after neonatal (lactational) exposure were most frequently the upper third molars. It has been proposed that coexpression of the AhR and ARNT during early tooth development and during the formation and mineralization of the dental matrix suggests that the AhR–ARNT pathway is a mediator of dental toxicity of TCDD (Sahlberg et al., 2002).

Cardiovascular Toxicity

There is a paucity of information on the potential cardiovascular toxicity of TCDD. In a study in which TCDD in corn oil was injected into chicken eggs at 0.40 pmol/g egg, TCDD reduced the size and number of coronary arteries (Ivinitski et al., 2001). The effect was associated with a reduction in proliferation of monocytes that was preceded by an increase in apoptosis in structures in which cell death normally occurs. Others have suggested that TCDD induces developmental defects in the cardiovascular system—that the cells that line the blood vessels could be a target of TCDD toxicity (IOM, 1999). A study by Dong et al. (2002) demonstrated that a decrease in blood flow in the mesencephalic vein is one of the earliest adverse effects of TCDD in the zebrafish embryo; they hypothesized that the decrease is a result of endothelial damage caused by oxidative stress associated with CYP1A induction. Dalton et al. (2001) observed that daily treatments of mice with 5 µg of TCDD for 3 days resulted in an increase in urinary excretion of vasoactive eicosanoids and serum triglycerides and an increase in mean tail-cuff blood pressure.

Subchronic treatment of hyperlipidemic apolipoprotein E (ApoE)-null mice with TCDD at 150 ng/kg body weight three times a week for 7 or 26 weeks caused a trend toward earlier onset and greater severity of atherosclerotic lesions than in vehicle-treated mice. Notably, ApoE-deficient mice have a lipoprotein profile similar to that of humans with type III hyperlipoproteinemia, and they develop extensive aortic and coronary atherosclerosis with lesions that are similar to those observed in humans. In a recent study (Riecke et al., 2002), male marmosets treated with a single subcutaneous injection of TCDD at 100 ng/kg body weight showed no overt signs of toxicity or effects on heart weights. Histologic examination, however, revealed an increase in picrosirius red-positive areas in the hearts of the TCDD-treated marmosets compared with controls. Western blotting confirmed an increase in collagen, fibronectin, and laminin in the hearts of the TCDD-treated marmosets.

Although the cardiovascular system appears to be affected predominantly in developing animals, there is only minimal evidence that it is a primary target of TCDD toxicity in adult animals. Further research is needed to determine whether exposure to TCDD might exacerbate lesions in blood vessels and be a risk factor for atherosclerosis.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Pulmonary Toxicity

Very little information is available to incriminate TCDD as a pulmonary toxicant in acute exposures. When rats were treated with TCDD at 125 ng/kg body weight for 60 weeks, there was a significant increase in alveolar–bronchiolar metaplasia (Tritscher et al., 2000). In the same study, when rats were initiated with diethylnitrosamine and then exposed to TCDD at 125 ng/kg body weight for 60 weeks, bronchiolar epithelial hyperplasia was noted. But the lesions were not observed in rats treated with TCDD for 30 weeks and then with corn oil for 30 weeks; continuous TCDD exposure might be required for their continued development. The study suggests that chronic oral exposure to TCDD can induce metaplasia and proliferative changes in the lung.

Hepatotoxicity

The liver is a primary target organ of halogenated aromatic hydrocarbons, but the severity of lesions varies considerably among species (IOM, 2001). It has been reported that a single oral dose of TCDD can inhibit normal hepatic accumulation of dietary vitamin A (Kelley et al., 1998); this effect has recently been confirmed in four species: hamster (TCDD at 0.9 µg/kg body weight), guinea pigs (0.1 µg/kg body weight), rats (1.1 µg/kg body weight), and mice (3.6 µg/kg body weight) (Fletcher et al., 2001).

Neurotoxicity

Some studies have implicated TCDD as a neurotoxin, but others have been unable to show neurotoxic activity (IOM, 1999, 2001). Hans/Wistar rats given TCDD intraperitoneally at 1,000 µg/kg body weight exhibited weight loss but no neurologic impairment (IOM, 2001). However, male and female Sprague-Dawley rats exposed in utero to TCDD at 1 µg/kg maternal body weight showed a deficit in learning in the spatial discrimination-reversal learning (RL) task, whereas the male progeny also showed a facilitation of task-specific spatial learning and memory (IOM, 2001). A study in primates showed that prenatal exposure to TCDD facilitated some spatial tasks but impaired visual RL tasks (IOM, 2001). In a more recent study in which Holtzman rats were exposed to TCDD at 20–180 ng/kg body weight on gestation day 18, the offspring had dose-dependent reductions in the number of earned opportunities to run on specifically designed running wheels, in lever response rates, and in total number of revolutions on the wheel (Markowski et al., 2001). Another study by the same group (Markowski et al., 2002) showed that female Holtzman rats exposed in utero to a single dose of TCDD at 0.18 µg/kg maternal body weight on gestation day 15 had significantly less accuracy and committed more errors in lever chambers than did the nonexposed controls. It was shown recently that GABA neurons in the brain are

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

targets of TCDD (Hays et al., 2002). In 3-day-old pups exposed in utero to TCDD at 1 µg/kg maternal body weight on gestation day 15, virtually all GABA neurons expressed the AhR gene. The noted effects of in utero exposure to TCDD have not been observed when animals have been exposed postnatally. In some species, the neurobehavorial development of the fetus seems to be sensitive to the toxic effects of TCDD.

Reproductive and Developmental Toxicity

It has been discussed in previous updates that low doses of TCDD can affect reproductive development and fertility of progeny. In Update 2000, one study demonstrated decreased sperm production and an increased number of abnormal sperm in male offspring of female rats treated with TCDD from before mating through lactation. All TCDD-exposed males, however, were able to impregnate females and produce viable fetuses in them. Other reproductive indexes evaluated in the study were also unaffected by TCDD.

TCDD has been reported to decrease seminal vesicle growth and prostate weight in male rats (IOM, 2001). Recent studies have confirmed that in utero exposure to TCDD impairs prostate growth. One study suggested that low-dose (TCDD at 800 ng/kg body weight) administration of TCDD to pregnant Holtzman dams had a greater effect on development of the external genital organs and ventral prostate than on development of the testis and other internal genital organs in the male offspring (Ohsako et al., 2001). A continuation of those studies (Ohsako et al., 2002) showed that TCDD exposure at 1 µg/kg maternal body weight on gestation day 18 resulted in significant decreases in the urogenital complex and ventral prostate weight and in urogenital–glans penis length (the length between the anterior end of the urethra and the glans penis) of male offspring. In utero TCDD exposure (1 µg/kg body weight) in Holtzman rats impaired prostate growth and androgen responsiveness by inhibiting prostate epithelial-cell differentiation (Theobald et al., 2000).

Administration of TCDD at 1.0 µg/kg maternal body weight on gestation day 13.5 resulted in increased pituitary luteinizing hormone concentrations and testicular testosterone synthesis in Han/Wistar fetuses but not in Long/Evans fetuses (Haavisto et al., 2001). Testosterone concentrations, however, were not affected in offspring of Holtzman rats given a single oral dose of TCDD at 1 µg/kg body weight or when primary murine testicular cells were cultured with TCDD (Timms et al., 2002; Uchida et al., 2002). Male Sprague-Dawley rats treated with TCDD at 10 µg/kg body weight had a reduced adrenocorticotropin-to-corticosterone ratio, suggesting that TCDD disturbs the hypothalamic–pituitary–adrenal axis (Pitt et al., 2000); this effect, however, was not detected in the pregnant females. It has been suggested that dioxin inducible factor-3 may be a target gene for TCDD during spermatogenesis (Ohbayashi et al., 2001).

TCDD exposure can also affect female reproduction. Female Sprague-Dawley rats given TCDD at 32 µg/kg body weight had reduced numbers of ova in

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

their oviducts, which affected ovulation (Gao et al., 2000). It was also noted that gonadotropin-releasing hormone induced surges in luteinizing hormone and follicle stimulating hormone, but the increases only partially restored the inhibitory effects of TCDD on ovulation. The female fetuses and offspring of pregnant Long-Evans rats treated with TCDD at 1 µg/kg maternal body weight on gestation day 15 were evaluated for developmental effects (Fenton et al., 2002; Hurst et al., 2002). In the fetus, the mesenchyme that separates the Müllerian ducts was widened, and the zone of unfused ducts was increased (Hurst et al., 2002). The postnatal effects included delayed vaginal opening and persistent vaginal threads but no altered estrous cycle (Fenton et al., 2002). There was also a reduction of the primary branches, decreased epithelial elongation, and fewer alveolar buds and lateral branches in the mammary glands of the TCDD-exposed rats.

Nonhuman primates (rhesus monkeys) had increased serum TCDD and endometriosis 13 years after treatment with TCDD at 25 ng/kg body weight (Rier et al., 2001b). They also exhibited an increase in TNF-α secretion from peripheral-blood mononuclear cells and a decrease in cytolytic activity of natural killer cells (Rier et al., 2001a). In a study in which cynomolgus macaques were treated with a single oral dose of TCDD at 4 µg/kg body weight, the menstrual cycle was eliminated and mean follicle-stimulating hormone concentrations were increased (Moran et al., 2001); the endometria of the noncycling monkeys were inactive. In another study in which cynomolgus monkeys were treated with TCDD at 25 ng/ kg body weight, progesterone concentrations and menstruation data indicated that TCDD did not interfere with ovulation (Shridhar et al., 2001). TCDD stimulated production of the corticotropin-releasing hormone, however, suggesting that the hypothalamic–pituitary–adrenal pathway is stimulated. TCDD (4 µg/kg body weight) has also been shown to induce epithelial transdifferentiation in the cynomolgus cervix 1 –2 years after treatment (Scott et al., 2001). When human endometrial explants were cultured in medium containing estradiol or estradiol and progesterone, TCDD significantly increased the expression of the AhR. TCDD did not affect ARNT mRNA or endometriosis (Bofinger et al., 2001; Pitt et al., 2001).

In a teratogenic study in which pregnant female mice were treated with TCDD at 24 µg/kg body weight on gestation day 12, cleft palate and hydronephrosis were induced in the offspring (Bryant et al., 2001a). An evaluation of the role of epidermal growth factor (EGF) using wild-type mice and knockout mice (mice that do not express EGF) revealed that EGF influences the induction of cleft palate by TCDD but that EGF is not required for formation of hydronephrosis.

Endocrine Effects

TCDD has been reported to affect concentrations of thyroid hormone, but contrasting results confuse interpretation. In a recent study (van der Plas et al., 2001), Sprague-Dawley rats exposed to TCDD at 1 µg/kg body weight per week

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

exhibited a decrease in total thyroid hormone but an increase in plasma retinal. Most TCDD investigations, however, have focused on assessing the effects of hormonal changes on reproduction. Most of those effects are mentioned in the preceding section. Estradiol produced by mature ovarian follicles triggers increased secretion of gonadotropin-releasing hormone. A recent study reported that TCDD at 32 µg/kg body weight decreased the responsiveness of the hypothalamus to estradiol, which normally acts as a feedback inducer of preovulatory gonadotropin secretion in Sprague-Dawley rats (Gao et al., 2001). In another study, it was determined that estradiol was not associated with the hepatotoxicity commonly observed in TCDD-treated rats (Wyde et al., 2000).

Immunotoxicity

The immune system of animals is highly sensitive to the toxic effects of TCDD and is a primary target of TCDD toxicity. For that reason, many investigations have focused on the immunotoxic effects of TCDD and the mechanisms responsible for them. TCDD is a potent immunosuppressant in laboratory animals at extremely low doses (0.1 µg TCDD/kg body weight or lower). There are, however, considerable differences in the immune response between rats and mice and strain differences within those species. Both resistant and sensitive strains have been identified for the numerous immunotoxic end points of TCDD toxicity.

TCDD has been shown to alter host resistance to infectious disease. It was reported that a single dose of TCDD increased the mortality of mice infected with influenza A (Burleson et al., 1996). That effect, however, was not confirmed in a more recent study using a similar protocol in four strains of mice (B6C3F1, BALB/c, C57Bl.6N, and DBA2) (Nohara et al., 2002b). The reasons for the discrepancy between the studies are not clear. Another group reported that in mice treated with TCDD (10 µg/kg body weight) and influenza virus there was no increase in the pulmonary virus burden; that suggested that TCDD impaired viral replication in lung epithelial cells (Lawrence et al., 2000). In another disease-susceptibility investigation, brain lesions and number of brain cysts after Toxoplasma gondi infection were not altered in C57Bl/6 mice dosed with TCDD at 50 µg/kg body weight (King et al., 2000). But in male A/J mice given TCDD intraperitoneally at 5 µg/kg body weight followed by three weekly doses at 1.4 µg/kg body weight, an increase in mortality from myocardial coxsackievirus B3 infection was seen 7 days after virus inoculation; no effect on the inflammatory lesions in the myocardium was seen (Funseth et al., 2002). When Brown Norway rats were exposed concurrently to TCDD (30 µg/kg bw) and house mites, TCDD exposure suppressed, rather than enhanced, immune response, as measured by immunoglobulin E (IgE) synthesis, and decreased immune-mediated lung disease (Luebke et al., 2001). TCDD has also been shown to stimulate the expression of IgE-dependent histamine-releasing factor mRNA via the AhR-dependent pathway (Oikawa et al., 2002).

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

In thymocytes obtained from C57BL/6N mice and treated in culture, TCDD skewed differentiation of thymocytes toward CD8 T cells, which require activation of the extracellular signal-related kinase pathway (Tsukumo et al., 2002). NC/Nga mice given TCDD intraperitoneally at 5.0 or 20 µg/kg body weight markedly suppressed the concentrations of interleukin-4 (IL-4) and interleukin-5 (IL-5) in culture supernatants of spleen cells (Fujimaki et al., 2002). TCDD exposure also reduced anti-ova antibody and total IgE antibody titers in the plasma of those mice. A recent study indicates that the maturing B220+ B cells are not the direct target for TCDD-induced toxicity but that hematopoietic progenitor cells are most likely a direct target for TCDD-mediated effects (Wyman et al., 2002).

Dendritic cells play a major role in activation of naive T cells. When dendritic cells from C57Bl/6 mice treated with TCDD at 15 µ g/kg body weight were cocultured with allogeneic T cells, the proliferative response and production of IL-2 and interferon-δ were increased, as was the production of IL-12 (Vorderstrasse and Kerkvliet, 2001). The total number of dendritic cells recovered from the TCDD-treated mice, however, was significantly decreased. It was suggested that TCDD leads to premature deletion of dendritic cells. TCDD (15 µg/kg bw) has also been shown to interfere with survival and differentiation of OVA-specific T-helper cells; that interference could prevent expansion and differentiation of those cells into effector T-helper cells (Shephard et al., 2000).

In utero exposure of rats on gestation day 15 to TCDD at 12.5–800 ng/kg maternal body weight had no significant effect on the thymic or splenic weights of the offspring (Nohara et al., 2000). A dose-dependent induction of CYP1A1 mRNA occurred in the thymus of the offspring, and a very weak induction occurred in the spleen of the offspring. In contrast, there was no effect of TCDD on cell numbers or populations (CD4 and CD8 markers) in the thymus, but splenocyte numbers decreased in a dose-dependent manner with no effect on splenic cellularity.

The complement system affords a defense against microbial infections. Administration of TCDD to guinea pigs at 0.5 µg/kg of body weight did not induce any significant change in complement activity (Wagner et al., 2001). That suggests that the complement system is resistant to the toxic effects of low doses of TCDD.

Carcinogenicity

TCDD is a known hepatocarcinogen in rats and mice and is considered to be a carcinogen in humans. It is not genotoxic but acts as a promoter involving multiple pathways in regulatory cell proliferation and differentiation. TCDD is known to promote hepatic neoplasia in laboratory rats at doses as low as 0.01 ng/ kg body weight per day (IOM, 2001). Recent multistage carcinogen models have used a variety of initiators —including diethylnitrosamine, azaserine, and methylnitrosourea—to investigate the tumor-promoting activities of TCDD (Desaulniers

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

et al., 2001; Oztas, 2000; Wyde et al., 2001a,b). They showed that TCDD promoted hepatocellular foci formation, pancreatic acinar cell foci, and mammarytumor development. In another experiment using diethylnitrosamine as an initiator, TCDD (1.75 µg/kg bw) administered continuously (biweekly) for 60 weeks, but not 30 weeks, to Sprague-Dawley rats promoted development of hepatocellular adenomas and carcinomas (Walker NJ et al., 2000). In fact, there were considerably fewer hepatocellular adenomas and carcinomas combined in the 30-week TCDD-treated group (17%) than in the vehicle control group (55%). Those data indicate that liver promotion by TCDD in female rats depends on long-term continuous exposure to TCDD. Another feature of that study is that the mean focus volume of preneoplastic altered hepatic foci (AHF) continued to increase after the 30-week treatment with TCDD.

Conclusions

TCDD is one of the most highly toxic chemicals known to affect animals, but there is an extreme range (of a factor of 1,000) in lethal effects among species and even within strains of animals. Oral intake is the primary route of exposure, and doses in nanogram amounts can elicit toxic effects in the most sensitive strains and species. The most sensitive time of exposure to TCDD is exposure of the fetus during pregnancy; toxicity results primarily in the nervous, immune, and reproductive systems. Structural abnormalities can also be a result of prenatal exposure to TCDD. TCDD is a potent promoter of hepatocarcinogenesis and is a hepatotoxin in animals. The immune system of animals is particularly sensitive to the toxic effects of TCDD; TCDD is immunosuppressive at doses lower than 0.1 µg/kg body weight. The endocrine system also appears to be a sensitive target of TCDD. Endocrine effects on the male and female reproductive tract have been reported at extremely low doses of TCDD. As mentioned above, the most toxic effects occur when animals are exposed in utero, but TCDD also interferes with reproduction when sexually mature animals are exposed. The cardiovascular and pulmonary systems appear more resistant to the toxicity of TCDD than the aforementioned organ systems.

SUMMARY OF TOXICITY PROFILES

This section synthesizes the experimental data on 2,4-D, 2,4,5-T, picloram, cacodylic acid, and TCDD reviewed in this and previous VAO reports, with a focus on recent data.

2,4-D

The majority of the studies of 2,4-D have found it to be relatively nontoxic, with health effects in animals observed only at high doses. Three studies pub-

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

lished since Update 2000 support earlier findings demonstrating that tissue uptake of 2,4-D is poor and metabolism fairly rapid, which could partially explain its low toxicity.

Earlier studies demonstrated the potential of high doses of 2,4-D to cause behavioral effects, muscle weakness, and incoordination in animals. Recent studies have demonstrated the ability of 2,4-D to damage nerve cells and produce neurotoxicity, but they tested high concentrations of the chemical after in vitro exposure or direct injection into the brain and therefore might not be relevant to physiologic exposures. The reproductive and developmental effects of 2,4-D have been examined recently. No developmental toxicity was seen in one study, but recent studies suggest that neonatal exposure to 2,4-D (100 mg/kg body weight per day) can result in alterations in brain development. A study of the effects of paternal exposure to 2,4-D did not demonstrate any effects on fetal survival or malformations. The effects of in utero exposure to 2,4-D on immune function in the offspring demonstrated effects only at the highest dose tested (650 mg/kg body weight per day). Carcinogenicity tests of 2,4-D have generally been negative; a recent study looking at the effects of 2,4-D on urethan-induced lung adenomas did not show an increase in adenomas with 2,4-D treatment, and thus supported 2,4-D's lack of carcinogenicity. Previous studies have suggested that 2,4-D might affect thyroid hormones (more specifically serum thyroxine). A recent study found that both thyroxine and triiodothyronine concentrations decreased after oral exposure to 2,4-D.

Genotoxicity tests and mechanistic studies have also been conducted on 2,4-D. It is either nongenotoxic or only weakly mutagenic in the many assays used. 2,4-D has been shown to have a number of effects on cells or biochemical measures, including effects on some hormones, on cellular components involved in the development and functioning of brain cells, and on some enzymes and transporters. Effects on calcium metabolism and energy metabolism, possibly through direct effects on mitochondrial function, have also been seen in response to 2,4-D treatment, as have effects on stress proteins. The relationship of any of those effects to any disease outcomes in animals or humans, however, is unknown.

Taken all together, the experimental data reviewed in this and previous reports indicate that 2,4-D is relatively nontoxic, with neurodevelopmental effects after neonatal exposure (at 100 mg/kg body weight per day) being one of the few effects seen.

2,4,5-T

Although not a great deal of research has been conducted recently on 2,4,5-T, the available data indicate that 2,4,5-T itself is relatively nontoxic. Studies indicate that 2,4,5-T is absorbed into the body after oral exposure, but absorption after dermal exposure is much slower. No recent toxicokinetic studies have been conducted. Studies of the reproductive effects of 2,4,5-T have demonstrated that

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

it can be fetotoxic in rodents at doses greater than 20 mg/kg body weight per day on days 6-15 of pregnancy, retarding growth and causing increased embryolethality and cleft palate. No such effects were seen in rabbits, sheep, or monkeys, and evidence suggests that TCDD contamination of the 2,4,5-T might underlie the reproductive effects seen in rodents. The carcinogenicity of 2,4,5-T has also been investigated; no indications of carcinogenicity were seen. Studies of the genotoxicity of 2,4,5-T show it to have weak genotoxic potential. Little is known regarding the cellular effects of 2,4,5-T, but it has been shown to alter cellular metabolism (for example, on the acetylcoenzyme A system), affect cholinergic transmission and the tyrosine kinase receptor, and disrupt apoptosis. As in the case of 2,4-D, the relevance of those effects to human diseases is not known, and the data consistently indicate that 2,4,5-T is relatively nontoxic.

Cacodylic Acid

Cacodylic acid, or DMA, is a metabolite of inorganic arsenic. As discussed in Chapter 2, because the relevance of studies of inorganic-arsenic exposure for evaluating effects of exposure to cacodylic acid has not been established and cannot be inferred, the literature on inorganic arsenic is not considered in this report. Methylation of inorganic arsenic to DMA was long thought to be a detoxification pathway. More recently, however, the trivalent methylated forms of arsenic, DMAIII and MMAIII, have been shown to be toxic—following acute exposure MMAIII is about 4 times more toxic than inorganic arsenic and DMAIII has similar toxicity to that of arsenicIII (NRC, 2001). Urinary excretion of DMA appears to be species-dependent; rapid excretion occurs in many animals. Rats, however, accumulate DMA in red cells and tissues.

Few animal studies are available on the noncancer health effects of cacodylic acid, but previous reports indicate that high, maternally toxic doses are fetotoxic and teratogenic in rats and mice. With respect to carcinogenicity, there is evidence that cacodylic acid can promote skin tumorigenesis in animals that are initiated chemically or with UVB radiation. Evidence of cacodylic acid's pulmonary and bladder carcinogenic activity has also been seen in mice and rats, respectively. In other studies, however, cacodylic acid did not promote kidney tumors or lung tumors in nitrosamine-initiated rats.

A primary mechanism of the acute toxicity of arsenic is interference of cellular respiration, but the mechanisms underlying the effects of cacodylic acid are not well understood. Some data indicate that cacodylic acid acts through induction of oxidative damage or damage to DNA, and it has been shown to affect microtubule networks at particular points in mitosis. A recent study demonstrated that cacodylic acid causes necrosis of the epithelium of the urinary bladder followed by regenerative hyperplasia, and other studies have found that cacodylic acid is a potent inducer of apoptosis (or programed cell death).

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Picloram

Few studies have been conducted on the toxicity of picloram, but those done indicate that it is relatively nontoxic. Two of three carcinogenicity studies reviewed in VAO indicate that pilcoram is not carcinogenic; a third was positive for liver tumors, but on review of the data, an Environmental Protection Agency committee concluded that the tumors were due to contamination with hexachlorobenzene (HCB). The VAO committee did note, however, that because the study was carried out with technical picloram, the compound used in Vietnam most likely contained similar amounts of HCB. Although the data on reproductive effects are not extensive, no effects considered to be treatment-related have been seen. Notably, a study of the male-mediated reproductive toxicity of Tordon 75D® (a commercial mixture of 2,4-D and picloram) found no effects on fetal survival or malformations. Another commercial mixture of 2,4-D and picloram, Tordon 202C®, had immunotoxic effects, reducing antibody production in mice in response to sheep red-cell inoculation at concentrations only marginally above those expected to be encountered after recommended application of the herbicide. Once again, however, the relevance of the few effects seen to human health outcomes is not known; taken together, the data indicate that picloram is relatively nontoxic.

TCDD

In contrast with the effects of the herbicides themselves, the effects of TCDD, a contaminant of 2,4,5-T, have been studied extensively. TCDD is hydrophobic and therefore is absorbed well across membranes, distributes to all compartments of the body, and partitions with lipids. Data also indicate that TCDD is transferred across the placenta to the fetus and is transferred to neonates through lactation. The enzyme cytochrome P450 1A2 (CYP1A2) plays an important role in the distribution of TCDD. Studies of TCDD in Ranch Hand Vietnam veterans indicate that it has a mean half-life of 7.6 years. Recent studies in two people exposed to very high amounts of TCDD, however, showed an elimination half-life of 1.5 and 2.9 years in the more and less exposed people, respectively, indicating that the half-life depends on body burden. Recent data from Seveso also indicate that the half-life is shorter in the first 3 months after exposure than from 3 to 16 years after exposure. Those data on half-life are consistent with a two-compartment toxicokinetic model for TCDD. Olestra somewhat increased the excretion of TCDD in the two heavily exposed patients, and this is consistent with earlier studies that indicate that the diet can affect the toxicokinetics of TCDD. A recent study in rats demonstrated that dietary seaweed can increase TCDD excretion, but whether the increase would occur in humans is not known. Evidence also suggests that the half-life is correlated with body weight. TCDD concentrations are often measured in blood, and previous and recent autopsy

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

studies indicate that blood concentrations correlate with tissue concentrations. Studies have also been conducted to validate physiologically based pharmacokinetic models to estimate the distribution and tissue concentrations of TCDD. Such models appear to be useful for toxicokinetic predictions.

Many effects have been observed in animals after exposure to TCDD, and TCDD is considered more toxic than the active ingredients of the herbicides used in Vietnam. Sensitivity to the lethal effects of TCDD varies among species and strains, but most species studied develop a wasting syndrome after acutely toxic doses that is characterized by a loss of body weight and fatty tissue. One target of TCDD is the liver, where lethal doses of TCDD cause necrosis, but the effect depends on the species exposed. Effects on the structure and function of the liver are also seen at lower doses. A recent study demonstrated that TCDD inhibits the ability of the liver to accumulate vitamin A.

TCDD may affect, directly or indirectly, many organs of the endocrine system in a species-specific manner; for example, thyroid-hormone concentrations have been shown to be affected. But some of the results of studies of thyroid hormones are contradictory, and this makes their interpretation difficult.

The adult nervous system has been shown to be sensitive to TCDD only at high doses. After in utero exposure, however, even the effects at high doses are not straightforward: in utero TCDD exposure decreases performance in some learning and memory tasks but improves performance in other tasks.

In animals, one of the most sensitive systems to TCDD is the immune system. Recent studies have demonstrated that TCDD can alter the numbers of immune cells, the measured activity of the cells, and the ability of animals to fight off infection. Effects on the immune system, however, appear to depend on the species, strain, and developmental stage of the animal studied.

Recent studies have further investigated whether TCDD is involved in endometriosis. TCDD did not affect surgically induced endometrial lesions in rats, although earlier studies demonstrated that prenatal and postnatal exposure of mice to TCDD increased sensitivity to endometrial-lesion growth.

Reproductive and developmental effects have been seen in animals exposed to TCDD, such as effects of developmental exposure to TCDD on sperm counts, sperm production, and seminal vesicle weights in male offspring and effects on the reproductive system in female offspring. In some recent studies, however, the effects on the reproductive system were not accompanied by effects on reproductive outcomes. Effects on the developing cardiovascular system have also been seen after TCDD exposure. The developing nervous system is potentially very sensitive to the effects of TCDD.

TCDD is carcinogenic and an extremely potent promoter of neoplasia in laboratory rats. Liver cancers have been seen consistently after TCDD treatment, and increases in skin cancer, lung cancer, and cancers of the thyroid and adrenal glands have been seen in some studies. A decrease in cancers of the uterus, the pancreas, and the pituitary, mammary, and adrenal glands has also been seen, but

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

most of those tumors showed decreases only at the high dose and were associated with the decrease in body weight gain, and the decrease in mammary tumors was only seen in one study. In a recent study, there was an increase in hepatic foci at TCDD doses as low as 0.01 ng/kg body weight per day—the lowest dose of TCDD known to promote tumors. Recent data also suggest that promotion of liver tumors by TCDD in female rats depends on continuous exposure.

Data published since Update 2000 are consistent with the hypothesis that TCDD produces most, if not all, of its effects by binding to a protein that regulates gene expression, the aryl hydrocarbon receptor (AhR). The binding of TCDD to the AhR and interaction of the complex with other proteins is followed by its binding to DNA, which triggers a number of cellular events, including induction of numerous proteins. Research in animals that have been engineered not to express the AhR and in animals with slightly different forms of it provides evidence that the AhR is necessary for the toxicity of TCDD. Modulation of genes by the AhR appears to have developmental-stage-, species-, and cell-specific patterns, which suggest that the molecular and cellular pathways that lead to any particular toxic event are complex.

Additional research has demonstrated that the outcomes of TCDD exposure can be modulated by numerous other proteins with which the AhR interacts. It is plausible, therefore, that the AhR could divert proteins and transcription factors from other signaling pathways; the disruption of the other pathways could have serious consequences for cellular and tissue processes.

Despite the large amount of research conducted on the cellular effects of TCDD, details of the mechanism(s) underlying its effects are not yet determined. Possible mechanisms underlying different effects have been discussed in this chapter and include effects on protein kinase expression, effects on vitamin stores, effects on cellular differentiation and the cell cycle, and oxidative stress. Although the mechanism underlying the carcinogenic effects of TCDD remains unknown, available data indicate that TCDD does not act directly on the genetic material; most genotoxic assays have negative results. Effects on enzymes or hormones could be involved in the carcinogenicity of TCDD.

RELEVANCE TO HUMAN HEALTH

As indicated above, exposure to TCDD has been associated with both cancer and noncancer end points in animals, and most TCDD effects are mediated through the AhR. Although structural differences in the AhR have been identified, it operates in a similar manner in animals and humans, and a connection between TCDD exposure and human health effects is, in general, considered biologically plausible. Animal research indicates that TCDD can cause cancers and benign tumors, and it can increase the incidence of some cancers or tumors in the presence of known carcinogens. However, experimental animals differ greatly in their susceptibility to TCDD-induced effects, and the sites at which tumors are

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

induced vary from species to species. Noncancer health effects also vary according to dose, time, and the animal exposed. Whether the effects of TCDD and other exposures are threshold-dependent—that is, whether some exposures may be too low to induce any effect—is still controversial. The relationship between mechanism and the shape of the dose-response curve, whether it be linear or nonlinear, is complex, not well understood, and may be different for different end points.

Little information is available on the biologic plausibility of causation of health effects by Agent Orange through chemicals other than TCDD. Although concerns have been raised about nondioxin contaminants of herbicides, far too little is known about their distribution and concentration in the formulations used in Vietnam to permit conclusions concerning their impact.

Considerable uncertainty remains about how to apply mechanistic information from nonhuman studies to an evaluation of the potential health effects of herbicide or dioxin exposure in Vietnam veterans. While the data specific to humans is inadequate to demonstrate strong relationships between exposure and disease conditions or pathologies, the growing and abundant evidence from experimental animals and wildlife strongly suggests that similar adverse effects are likely in human populations, the issues of sensitivity and dose-response perhaps being paramount in species differences. It is hoped that as the cellular mechanisms of these compounds are discovered, future VAO updates will have better information on which to base conclusions, including better information on the relevance of experimental data to effects in humans.

REFERENCES

Adachi J, Mori Y, Matsui S, Takigami H, Fujino J, Kitagawa H, Miller CA III, Kato T, Saeki K, Matsuda T. 2001. Indirubin and indigo are potent aryl hydrocarbon receptor ligands present in human urine. The Journal of Biological Chemistry 276:31475–31478.

Ait-Aissa S, Porcher JM, Arrigo AP, Lambre C. 2000. Activation of the hsp70 promoter by environmental inorganic and organic chemicals: relationships with cytotoxicity and lipophilicity. Toxicology 145:147–157.

Allen DE, Leamy LJ. 2001. 2,3,7,8-tetrachlorodibenzo-p-dioxin affects size and shape, but not asymmetry, of mandibles in mice. Ecotoxicology 10(3):167–176.

Amer SM, Aly FAE. 2001. Genotoxic effect of 2,4-dichlorophenoxy acetic acid and its metabolite 2,4-dichlorophenol in mouse. Mutation Research 494:1–12.

Andersson P, McGuire J, Rubio C, Gradin K, Whitelaw ML, Pettersson S, Hanberg A, Poellinger L. 2002. A constitutively active dioxin/aryl hydrocarbon receptor induces stomach tumors. Proceedings of the National Academy of Sciences of the United States of America 99:9990–9995.

Andreasen EA, Hahn ME, Heideman W, Peterson RE, Tanguay RL. 2002a. The zebrafish (Danio rerio) aryl hydrocarbon receptor type 1 is a novel vertebrate receptor. Molecular Pharmacology 62:234–249.

Andreasen EA, Spitsbergen JM, Tanguay RL, Stegeman JJ, Heideman W, Peterson RE. 2002b. Tissue-specific expression of AHR2, ARNT2, and CYP1A in zebrafish embryos and larvae: Effects of developmental stage and 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure. Toxicological Sciences 68:403–419.

Andreasen EA, Tanguay RL, Peterson RE, Heideman W. 2002c. Identification of a critical amino acid in the aryl hydrocarbon receptor The Journal of Biological Chemistry 277:13210–13218.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Angus WGR, Larsen MC, Jefcoate CR. 2000. TCDD elevates erbB2 expression and signaling in T47D cells by reversing serum potentiation of estrogen receptor activity, independent of estrogen levels and enhanced ER down-regulation. Molecular and Cellular Endocrinology 170:1– 13.

Arfi C, Seta N, Fraisse D, Revel A, Escande JP, Momas I. 2001. Dioxins in adipose tissue of non-occupationally exposed persons in France: correlation with individual food exposure. Chemosphere 44(6):1347–1352.

Arulmozhiraja S, Fujii T, Tokiwa H. 2000. Electron affinity for the most toxic 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): A density functional theory study. Journal of Physical Chemistry A 104:7068–7072.

Ashida H, Fakuda I, Yamashita T, Kanazawa K. 2000a. Flavones and flavonols at dietary levels inhibit a transformation of aryl hydrocarbon receptor induced by dioxin. FEBS Letters 476:213– 217.

Ashida H, Nagy S, Matsumura F. 2000b. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)-induced changes in activities of nuclear protein kinases and phosphatases affecting DNA binding activity of c-myc and AP-1 in the livers of guinea pigs. Biochemical Pharmacology 59:741–751.

Bagchi D, Balmoori J, Bagchi M, Ye X, Williams CB, Stohs SJ. 2002. Comparative effects of TCDD, endrin, naphthalene and chromium (VI) on oxidative stress and tissue damage in the liver and brain tissues of mice. Toxicology 175:73–82.

Balague C, Sturtz N, Duffard R, de Duffard AME. 2001. Effect of 2,4-dichlorophenoxyacetic acid herbicide on Escherichia coli growth, chemical composition, and cellular envelope. Environmental Toxicology 16:43–53.

Barnekow DE, Hamburg AW, Puvanesarajah V, Guo M. 2001. Metabolism of 2,4-dichlorophenoxyacetic acid in laying hens and lactating goats. Journal of Agricultural and Food Chemistry 49:156–163.

Barouki R, Morel Y. 2001. Repression of cytochrome P450 1A1 gene expression by oxidative stress: mechanisms and biological implications. Biochemical Pharmacology 61:511–516.

Beischlag TV, Wang S, Rose DW, Torchia J, Reisz-Porszasz S, Muhammad K, Nelson WE, Probst MR, Rosenfeld MG, Hankinson O. 2002. Recruitment of the NcoA/SRC-1/p160 family of transcriptional coactivators by the aryl hydrocarbon receptor nuclear complex. Molecular and Cellular Biology 22:4319–4333.

Bell DR, Poland A. 2000. Binding of aryl hydrocarbon receptor (AhR) to AhR-interacting protein. The role of hsp 90. The Journal of Biological Chemistry 275:36407–36414.

Berg P, Pongratz I. 2001. Differential usage of nuclear export sequences regulates intracellular localization of the dioxin (aryl hydrocarbon) receptor. The Journal of Biological Chemistry 276:43231–43238.

Birnbaum LS, Cummings AM. 2002. Dioxins and endometriosis: a plausible hypothesis. Environmental Health Perspectives 110:15–21.

Bofinger DP, Feng L, Chi L-H, Love J, Stephen FD, Sutter TR, Osteen KG, Costich TG, Batt RE, Koury ST, Olson JR. 2001. Effect of TCDD exposure on CYP1A1 and CYP1B1 expression in explant cultures of human endometrium. Toxicological Sciences 62:299–314.

Bortolozzi A, de Duffard AME, Dajas F, Duffard R, Silveira R. 2001. Intracebral administration of 2,4-dichlorophenoxyacetic acid induces behavioral and neurochemical alterations in the rat brain. Neurotoxicology 22(2):221–232.

Bortolozzi A, Dufard R, Antonelli M, de Duffard AME. 2002. Increased sensitivity in dopamine D2-like brain receptors from 2,4-dichlorophenoxyacetic acid (2,4-D)-exposed and amphetamine-challenged rats. Annals of the New York Academy of Sciences 965:314–323.

Bortot P, Thomaseth K, Salvan A. 2002. Population toxicokinetic analysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin using Bayesian techniques. Statistics in Medicine 21:533–547.

Bryant PL, Reid LM, Schmid JE, Buckalew AR, Abbott BD. 2001a. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on fetal mouse urinary tract epithelium in vitro. Toxicology 162:23– 34.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Bryant PL, Schmid JE, Fenton SE, Buckalew AR, Abbott BD. 2001b. Teratogenicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in mice lacking the expression of EGF and/or TGF-α. Toxicological Sciences 62:103–114.

Buchanan DL, Sato T, Peterson RE, Cooke PS. 2000. Antiestrogenic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin in mouse uterus: critical role of the aryl hydrocarbon receptor in stromal tissue. Toxicological Sciences 57:302–311.

Buchanan DL, Ohsako S, Tohyama C, Cooke PS, Iguchi T. 2002. Dioxin inhibition of estrogen-induced mouse uterine epithelial mitogenesis involves changes in cyclin and transforming growth factor-β expression Toxicological Sciences 66:62–68.

Bulun SE, Zeitoun KM, Kilic G. 2000. Expression of dioxin-related transactivating factors and target genes in human eutopic endometrial and endometriotic tissues. American Journal of Obstetrics and Gynecology 182:767–775.

Burleson GR, Lebrec H, Yang YG, Ibanes JD, Pennington KN, Birnbaum LS. 1996. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on influenza virus host resistance in mice. Fundamental and Applied Toxicology 29:40–47.

Camacho IA, Hassuneh MR, Nagarkatti M, Nagarkatti PS. 2001. Enhanced activation-induced cell death as a mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced immunotoxicity in peripheral T cells. Toxicology 165:51–63.

Carrier G, Brunet RC, Brodeur J. 1995. Modeling of the toxicokinetics of polychlorinated dibenzo-p-dioxins and dibenzofurans in mammalians, including humans. I. Nonlinear distribution of PCDD/PCDF body burden between liver and adipose tissue Toxicology and Applied Pharmacology 131:253–266.

Cauchi S, Stucker I, Solas C, Laurent-Puig P, Cenee S, Hemon D, Jacquet M, Kremers P, Beaune P, Massaad-Massade L. 2001. Polymorphisms of human aryl hydrocarbon receptor (AhR) gene in a French population: relationship with CYP1A1 inducibility and lung cancer. Carcinogenesis 22:1819–1824.

Chaffin CL, Trewin AL, Hutz RJ. 2000. Estrous-dependent changes in the expression of aromatic hydrocarbon receptor (AHR) and AHR-nuclear translocator (ARNT) mRNAs in the rat ovary and liver. Chemico-Biological Interactions 124:205–216.

Charles JM, Cifone MA, Lawlor T, Murli H, Young RR, Leeming NM. 2000. Evaluation of the invitro genetic toxicity of 4-(2,4-dichlorophenoxy)butyric acid. Mutation Research 472:75–83.

Charles JM, Hanley TR Jr, Wilson RD, van Ravenzwaay B, Bus JS. 2001. Developmental toxicity studies in rats and rabbits on 2,4-dichlorophenoxyacetic acid and its forms. Toxicological Sciences 60:121–131.

Chen CY, Hamm JT, Hass R, Birnbaum LS. 2001. Disposition of polychlorinated dibenzo-p-dioxins, dibenzofurans and non-ortho polychlorinated biphenyls in pregnant Long Evans rats and the transfer to offspring. Toxicology and Applied Pharmacology 173: 65–88.

Chen I, Hsieh T, Thomas T, Safe S. 2001. Identification of estrogen-induced genes downregulated by AhR agonists in MCF-7 breast cancer cells using suppression subtractive hybridization Gene 262:207–214.

Cho Y-S, Park S-H, Kim C-K, Oh K-H. 2000. Induction of stress shock proteins DnaK and GroEL by phenoxyherbicide 2,4-D in Burkholderia sp. YK-2 isolated from rice field. Current Microbiology 41:33–38.

Ciolino H, Daschner P, Yeh G. 1999. Dietary flavonols quercetin and kaempferol are ligands of the aryl hydrocarbon receptor that affect CYP1A1 transcription differentially The Biochemical Journal 340:715–722.

Cohen SM, Yamamoto S, Cano M, Arnold LL. 2001. Urothelial cytotoxicity and regeneration induced by dimethylarsinic acid in rats. Toxicological Sciences 59:68–74.

Cox MB, Miller CA III. 2002. The p23 co-chaperone facilitates dioxin receptor signaling in a yeast model system. Toxicology Letters 129:13–21.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Craft ES, DeVito MJ, Crofton KM. 2002. Comparative responsiveness of hypothyroxinemia and hepatic enzyme induction in Long-Evans rats versus C57BL/6 mice exposed to TCDD-like and phenobarbital-like polychlorinated biphenyl congeners. Toxicological Sciences 68:372–380.

Dalton TP, Kerzee JK, Wang B, Miller M, Dieter MZ, Lorenz JN, Shertzer HG, Nerbert DW, Puga A. 2001. Dioxin exposure is an environmental risk factor for ischemic heart disease. Cardiovascular Toxicolology 1:285–298.

Dasmahapatra AK, Wimpee BAB, Trewin AL, Wimpee CF, Ghorai JK, Hutz RJ. 2000. Demonstration of 2,3,7,8-tetrachlorodibenzo-p-dioxin attenuation of P450 steroidogenic enzyme mRNAs in rat granulosa cells in vitro by competitive reverse transcriptase-polymerase chain reaction assay. Molecular and Cellular Endocrinology 164:5–18.

Dasmahapatra AK, Wimpee BAB, Trewin AL, Hutz RJ. 2001. 2,3,7,8-Tetrachlorodibenzo-p-dioxin increases steady-state estrogen receptor–β mRNA levels after CYP1A1 and CYP1B1 induction in rat granulosa cells in vitro. Molecular and Cellular Endocrinology 182:39–48.

Davis JW III, Lauer FT, Burdick AD, Hudson LG, Burchiel SW. 2001. Prevention of apoptosis by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the MCF-10A cell line: correlation with increased transforming growth factor α production. Cancer Research 61:3314–3320.

Desaulniers D, Leingartner K, Russo J, Perkins G, Chittim BG, Archer MC, Wade M, Yang J. 2001. Modulatory effects of neonatal exposure to TCDD or a mixture of PCBs, p,p′-DDT, and p-p′-DDE on methylnitrosourea-induced mammary tumor development in the rat. Environmental Health Perspectives 109:739–747.

Dickow LM, Podell M, Gerken DF. 2000. Clinical effects and plasma concentration determination after 2,4-dichlorophenoxyacetic acid 200 mg/kg administration in the dog. Clinical Toxicology 38(7):747–753.

Dieter MZ, Freshwater SL, Solis WA, Nebert DW, Dalton TP. 2001. Tryphostin AG879, a tyrosine kinase inhibitor: prevention of transcriptional activation of the electrophile and the aromatic hydrocarbon response elements. Biochemical Pharmacology 61:215–225.

Diliberto JJ, Devito MJ, Ross DG, Birnbaum LS. 2001. Subchronic exposure of [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in female B6C3F1 mice: relationship of steady-state levels to disposition and metabolism. Toxicological Sciences 61:241–255.

Di Paolo O, de Duffard AME, Duffard R. 2001. In vivo and in vitro binding of 2,4-dichlorophenoxyacetic acid to a rat liver mitochondrial protein. Chemico-Biological Interactions 137:229–241.

Dong L, Ma Q, Whitlock JP Jr. 1997. Down-regulation of major histocompatibility complex Q1b gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin. The Journal of Biological Chemistry 272:29614–29619.

Dong W, Teraoka H, Yamazake K, Tsukiyama S, Imani S, Imagawa T, Stegeman JJ, Peterson RE, Hiraga T. 2002. 2,3,7,8-Tetrachlorodibenzo-p-dioxin toxicity in zebrafish embryo: local circulation failure in the dorsal midbrain is associated with increased apoptosis. Toxicological Sciences 69:191–201.

Dunlap DY, Ikeda I, Nagashima H, Vogel CFA, Matsumura F. 2002. Effects of src-deficiency on the expression of in vivo toxicity of TCDD in a strain of c-src knockout mice procured through six generations of backcrossings to C57BL/6 mice. Toxicology 172:125–141.

Elbi C, Misteli T, Hager GL. 2002. Recruitment of dioxin receptor to active transcription sites. Molecular Biology of the Cell 13:2001–2015.

Elferink CJ, Ge N-L, Levine A. 2001. Maximal aryl hydrocarbon receptor activity depends on an interaction with the retinoblastoma protein. Molecular Pharmacology 59:664–673.

Evans MV, Andersen ME. 2000. Sensitivity analysis of a physiological model for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): Assessing the impact of specific model parameters on sequestration in liver and fat in the rat. Toxicological Sciences 54:71–80.

Favreau LV, Pickett CB. 1991. Transcriptional regulation of the rat NAD(P)H:quinone reductase gene. Identification of regulatory elements controlling basal level expression and inducible expression by planar aromatic compounds and phenolic antioxidants. The Journal of Biological Chemistry 266:4556–4561.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Fenton SE, Hamm JT, Birnbaum LS, Youngblood GL. 2002. Persistent abnormalities in the rat mammary gland following gestational and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Toxicological Sciences 67:63–74.

Ferri A, Bortolozzi A, Duffard R, de Duffard AME. 2000. Monoamine levels in neonate rats lactationally exposed to 2,4-dichlorophenoxyacetic acid. Biogenic Amines 16:73–100.

Flesch-Janys D, Becher H, Gurn P, Jung D, Konietzko J, Manz A, Papke O. 1996. Elimination of polychlorinated dibenzo-p-dioxins and dibenzofurans in occupationally exposed persons. Journal of Toxicology and Environmental Health 47: 363–378.

Fletcher N, Hanberg A, Hakansson H. 2001. Hepatic vitamin A depletion is a sensitive marker of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure in four rodent species. Toxicological Sciences 62:166–175.

Fofana D, Kobae H, Oku S, Nishi J-I, Miyata K. 2000. Prenatal developmental effects of pure 2,4-dichlorophenoxyacetic acid (2,4-D) on the rat. Congenital Anomalies 40:287–296.

Fofana D, Kobae H, Sameshima K, Miyata K. 2002. Postnatal survival of rat offspring prenatally exposed to pure 2,4-dichlorophenoxyacetic acid (2,4-D). Congenital Anomalies 42:32–35.

Franc M-A, Pohjanvirta R, Tuomisto J, Okey AB. 2001. Persistent, low-dose 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure: Effect on aryl hydrocarbon receptor expression in a dioxin-resistance model. Toxicology and Applied Pharmacology 175:43–53.

Frueh FW, Hayashibara KC, Brown PO, Whitlock JP Jr. 2001. Use of cDNA microarrays to analyze dioxin-induced changes in human liver gene expression. Toxicology Letters 122:189–203.

Fujimaki H, Nohara K, Kobayashi T, Suzuki K, Eguchi-Kasai K, Tsukumo S, Kijima M, Tohyama C. 2002. Effect of a single oral dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin on immune function in male NC/Nga mice. Toxicological Sciences 66:117–124.

Fujita H, Kosaki R, Yoshihashi H, Ogata T, Tomita M, Hasegawa T, Takahashi T, Matsuo N, Kosaki K. 2002. Characterization of the aryl hydrocarbon receptor repressor gene and association with its Pro185Ala polymorphism with micropenis. Teratology 65:10–18.

Funseth E, Wesslen L, Lindh U, Friman G Ilback NG. 2002. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on trace elements, inflammation and viral clearance in the myocardium during coxsackievirus B3 infection in mice. The Science of the Total Environment 284:135–147.

Gaido KW, Maness SC. 1994. Regulation of gene expression and acceleration of differentiation in human keratinocytes by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicology and Applied Pharmacology 127:199–208.

Gaido KW, Maness SC, Leonard LS, Greenlee WF. 1992. 2,3,7,8-Tetrachlorodibenzo-p-dioxin-dependent regulation of transforming growth factors-α and -β2 expression in a human keratinocyte cell line involves both transcriptional and post-transcriptional control. The Journal of Biological Chemistry 267:24591–24595.

Gao L, Whitlock JP Jr. 2001. Acessibility and activity of the promoter for a dioxin-inducible ecto-ATPase gene. Archives of Biochemistry and Biophysics 392:270–278.

Gao L, Dong L, Whitlock JP Jr. 1998. A novel response to dioxin. Induction of ecto-ATPase gene expression The Journal of Biological Chemistry 273:15358–15365.

Gao X, Petroff BK, Rozman KK, Terranova PF. 2000. Gonadotropin-releasing hormone (GnRH) partially reverses the inhibitory effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on ovulation in the immature gonadotropin-treated rat. Toxicology 147:15–22.

Gao X, Mizuyachi K, Terranova PF, Rozman KK. 2001. 2,3,7,8-Tetrachlorodibenzo-p-dioxin decreases responsiveness of the hypothalamus to estradiol as a feedback inducer of preovulatory gonadotropin secretion in the immature gonadotropin-primed rat. Toxicology and Applied Pharmacology 170:181–190.

Garabrant DH, Philbert MA. 2002. Review of 2,4-dichlorophenoxyacetic acid (2,4-D) epidemiology and toxicology. Critical Reviews in Toxicology 32:233–257.

Garrison PM, Denison MS. 2000. Analysis of the murine AhR gene promoter. Journal of Biochemical and Molecular Toxicology 14:1–10.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Garrison PM, Rogers JM, Brackney WR, Denison MS. 2000. Effects of histone deacetylase inhibitors on the Ah receptor gene promoter. Archives of Biochemistry and Biophysics 374:161–171.

Genter MB, Apparaju S, Desai PB. 2001. Induction of olfactory mucosal and liver metabolism of lidocaine by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Journal of Biochemical and Molecular Toxicology 16(3):128–134.

Geusau A, Tschachler K, Meixner M, Papke O, Stingl G, McLachlan M. 2001. Cutaneous elimination of 2,3,7,8-tetrachlorodibenzo-p-dioxin. British Journal of Dermatology 145:938–943.

Geusau A, Schmaldienst S, Derfler K, Papke O, Abraham K. 2002. Severe 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) intoxication: kinetics and trials to enhance elimination in two patients. Archives of Toxicology 76:316–325.

Gillesby B, Santostefano M, Porter W, Wu ZF, Safe S, Zacharewski T. 1997. Identification of a motif within the 5'-regulatory region on pS2 which is responsible for Ap1 binding and TCDD-mediated suppression. Biochemistry 36:6080–6089.

Glover RE, Germolec DR, Patterson R, Walker NJ, Lucier GW, Mason RP. 2000. Endotoxin (lipopolysaccharide)-induced nitric oxide production in 2,3,7,8-tetrachlorodibenzo-p-dioxintreated Fisher rats: detection of nitrosyl hemoproteins by EPR spectroscopy. Chemical Research in Toxicology 13:1051–1055.

Greenlee WF, Hushka LJ, Hushka DR. 2001. Molecular basis of dioxin actions: evidence supporting chemoprotection Toxicologic Pathology 29:6–7.

Gregoraszczuk EL, Zabielny E, Ochwat D. 2001. Aryl hydrocarbon receptor (AhR)-linked inhibition of luteal cell progesterone secretion in 2,3,7,8-tetrachlorodibenzo-p-dioxin treated cells. Journal of Physiology and Pharmacology 52:303–311.

Guo M, Joiakim A, Dudley DT, Reiners JJ Jr. 2001. Suppression of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-mediated CYP1A1 and CYP1B1 induction by 12-O-tetradecanoylphorbol-13-acetate: role of transforming growth factor β and mitogen-activated protein kinases. Biochemical Pharmacology 62:1449–1457.

Haavisto T, Nurmela K, Pohjanvirta R, Huuskonen H, El-Gahani F, Paranko J. 2001. Prenatal testosterone and luteinizing hormone levels in male rats exposed during pregnancy to 2,3,7,8-tetrachlorodibenzo-p-dioxin and diethylstilbestrol. Molecular and Cellular Endocrinology 178: 169–179.

Hasan A, Fischer B. 2001. Hormonal control of arylhydrocarbon receptor (AhR) expression in the preimplantation rabbit uterus. Anatomy and Embryology 204:189–196.

Hassoun EA, Li F, Abushaban A, Stohs SJ. 2001. Production of superoxide anion, lipid peroxidation and DNA damage in the hepatic and brain tissues of rats after subchronic exposure to mixtures of TCDD and its congeners. Journal of Applied Toxicology 21:211–219.

Hassoun EA, Wang H, Abushaban A, Stohs SJ. 2002. Induction of oxidative stress in the tissues of rats after chronic exposure to TCDD, 2,3,4,7,8-pentachlorodibenzofuran, and 3,3',4,4 ',5-pentachlorobiphenyl. Journal of Toxicology and Environmental Health, PartA 65:825–842.

Hays LE, Carpenter CD, Petersen SL. 2002. Evidence that GABAergic neurons in the preoptic area of the rat brain are target of 2,3,7,8-tetrachlorodibenzo-p-dioxin during development. Environmental Health Perspectives 110:369–376.

Heid SE, Pollenz RS, Swanson HI. 2000. Role of heat shock protein 90 dissociation in mediating agonist-induced activation of the aryl hydrocarbon receptor. Molecular Pharmacology 57:82– 92.

Heid SE, Walker MK, Swanson HI. 2001. Correlation of cardiotoxicity mediated by halogenated aromatic hydrocarbons to aryl hydrocarbon receptor activation. Toxicological Sciences 61:187– 196.

Hestermann EV, Stegeman JJ, Hahn ME. 2000. Relative contributions of affinity and intrinsic efficacy to aryl hydrocarbon receptor ligand potency. Toxicology and Applied Pharmacology 168:160–172.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Hirai K-I, Pan J-H, Shui Y-B, Simamura E, Shimada H, Kanamaru T, Koyama J. 2002. Alphatocopherol protects cultured human cells from the acute lethal cytotoxicity of dioxin. International Journal of Vitamin and Nutrition Research 72:147–153.

Hirakawa T, Minegishi T, Abe K, Kishi H, Ibuki Y, Miyamoto K. 2000. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the expression of luteinizing hormone receptors during cell differentiation in cultured granulosa cells. Archives of Biochemistry and Biophysics 375:371–376.

Hochstein JR, Render JA, Bursian SJ, Aulerich RJ. 2001. Chronic toxicity of dietary 2,3,7,8-tetrachlorodibenzo-p-dioxin to mink. Veterinary and Human Toxicology 43:134–139.

Hojo R, Stern S, Zareba G, Markowski VP, Cox C, Kost JT, Weiss B. 2002. Sexually dimorphic behavioral responses to prenatal dioxin exposure Environmental Health Perspectives 110:247– 254.

Holladay SD, Sharova L, Smith BJ, Gogal RM Jr, Ward DL, Blaylock BL. 2000. Nonspecific stimulation of the maternal immune system. I. Effects on teratogen-induced fetal malformations. Teratology 62:413–419.

Holman H-Y N, Goth-Goldstein R, Martin MC, Russell ML, McKinney WR. 2000. Low-dose responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin in single living human cells measured by synchrotron infrared spectromicroscopy. Environmental Science and Technology 34:2513–2517.

Huang P, Rannug A, Ahlbom E, Hakansson H, Ceccatelli S. 2000. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the expression of cytochrome P450 1A1, the aryl hydrocarbon receptor, and aryl hydrocarbon receptor nuclear translocator in rat brain and pituitary. Toxicology and Applied Pharmacology 169:159–167.

Hurst CH, Abborr B, Schmid JE, Birnbaum LS. 2002. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) disrupts early morphogenetic events that form the lower reproductive tract in female rat fetuses. Toxicological Sciences 65:87–98.

Hurst CH, DeVito MJ, Birnbaum, LS. 2000. Tissue distribution of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in maternal and developing Long-Evans rats following subchronic exposure. Toxicological Sciences 57:275–283.

Iba MM, Fung J. 2001. Pulmonary CYP1A1 and CYP1A2 levels and activities in adult male and female offspring of rats exposed during gestation and lactation to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Biochemical Pharmacology 62:617–626.

Ikeda M, Ishii Y, Kato H, Akazawa D, Hatsumura M, Ishida T, Matsusue K, Yamada H, Oguri K. 2000. Suppression of carbonic anhydrase III in rat liver by a dioxin-related toxic compound, coplanar polychlorinated biphenyl 3,3',4,4'-pentachlorobiphenyl Archives of Biochemistry and Biophysics 380:159–164.

Ikeda M, Inukai N, Mitsui T, Sone H, Yonemoto J, Tohyama C, Tomita T. 2002. Changes in fetal brain aromatase activity following in utero 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure in rats. Environmental Toxicology and Pharmacology 11:1–7.

Ikuta T, Watanabe J, Kawajiri K. 2002. Characterization of the LxxLL motif in the aryl hydrocarbon receptor: Effects on subcellular localization and transcriptional activity Journal of Biochemistry 131:79–85.

IOM (Institute of Medicine). 1994. Veterans and Agent Orange: Health Effects of Herbicides Used in Vietnam Washington, DC: National Academy Press.

IOM. 1996. Veterans and Agent Orange: Update 1996. Washington, DC: National Academy Press.

IOM. 1999. Veterans and Agent Orange: Update 1998. Washington, DC: National Academy Press.

IOM. 2001. Veterans and Agent Orange: Update 2000. Washington, DC: National Academy Press.

Ishimura R, Ohsako S, Miyabara Y, Sakaue M, Kawakami T, Aoki Y, Yonemoto J, Tohyama C. 2002. Increased glycogen content and glucose transporter 3 mRNA level in the placenta of Holtzman rats after exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicology and Applied Pharmacology 178:161–171.

Ivnitski I, Elmaqued R, Walker MK. 2001. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) inhibition of coronary development is preceded by a decrease in myocyte proliferation and an increase in cardiac apoptosis Teratology 64:201–212.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Jackson WG Jr, Michalek JE. 2001. Temporal changes in TCDD levels in 1410 Air Force Vietnam-era veterans not occupationally exposed to herbicides. Journal of Exposure Analysis and Environmental Epidemiology 11:50–55.

Jamsa T, Viluksela M, Tuomisto JT, Tuomisto J, Tuukkanen J. 2001. Effects 2,3,7,8-tetrachlorodibenzo-p-dioxin on bone in two rat strains with different aryl hydrocarbon receptor structures. Journal of Bone and Mineral Research 16:1812–1820.

Jana NR, Sarkar S, Ishizuka M, Yonemoto J, Tohyama C, Sone H. 2000. Comparative effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on MCF-7, RL95-2 and LNCaP cells: role of target steroid hormones in cellular responsiveness to CYP1A1 induction. Molecular Cell Biology Research Communications 4:174–180.

Jeffrey BD, Chirnomas RB, Chen EJ, Gudas JM, Romagnolo DF. 2002. Activation of the aromatic hydrocarbon receptor pathway is not sufficient for transcriptional repression of BRCA-1: requirements for metabolism of benzo[a]pyrene to 7r8t-Dihydroxy-9t,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene Cancer Research 62:113–121.

Jeon M-S, Esser C. 2000. The murine IL-2 promoter contains distal regulatory elements responsive to the Ah receptor, a member of the evolutionarily conserved bHLH-PAS transcription factor family. Journal of Immunology 165:6975–6983.

Jeong HG, Kim JY, Choi CY. 2000. Down-regulation of murine Cyp1a1 in mouse hepatoma Hepa-1c1c7 cells by bisphenol A. Biochemical and Biophysical Research Communications 277:594– 598.

Jones LC, Whitlock JP Jr. 2001. Dioxin-inducible transactivation in a chromosomal setting. Analysis of the acidic domain of the Ah receptor. The Journal of Biological Chemistry 276:25037– 25042.

Jones LC, Okino ST, Gonda TJ, Whitlock JP Jr. 2002. Myb-binding protein 1a augments AhR-dependent gene expression. The Journal of Biological Chemistry 277:22515–22519.

Kaioumova D, Susal C, Opelz G. 2001. Induction of apoptosis in human lymphocytes by the herbicide 2,4-dichlorophenoxyacetic acid. Human Immunology 62:64–74.

Kakeyama M, Sone H, Tohyama C. 2001. Changes in expression of NMDA receptor subunit mRNA by perinatal exposure to dioxin. Neuropharmacology and Neurotoxicology NeuroReport 12: 4009–4012.

Karchner SI, Franks DG, Powell WH, Hahn ME. 2002. Regulatory interactions among three members of the vertebrate aryl hydrocarbon receptor family: AHR repressor, AHR1, and AHR2. The Journal of Biological Chemistry 277:6949–6959.

Kattainen H, Tuukkanen J, Simanainen U, Tusmisto JT, Kovero O, Lukinmaa PL, Alaluusua S, Tuomisto J, Viluksela M. 2001. In utero/lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure impairs molar tooth development in rats. Toxicology and Applied Pharmacology 174:216– 224.

Kaya B, Creus A, Yanikoglu A, Cabre O, Marcos R. 2000. Use of the Drosophila wing spot test in the genotoxicity testing of different herbicides. Environmental and Molecular Mutagenesis 36: 40–46.

Kazlauskas A, Poellinger L, Pongratz I. 2000. The immunophilin-like protein XAP2 regulates ubiuitination and subcellular localization of the dioxin receptor. The Journal of Biological Chemistry 275:41317–41324.

Kazlauskas A, Sundstrom S, Poellinger L, Pongratz I. 2001. The hsp90 chaperone complex regulates intracellular localization of the dioxin receptor. Molecular and Cellular Biology 21:2594–2607.

Kazlauskas A, Poellinger L, Pongratz I. 2002. Two distinct regions of the immunophilin-like protein XAP2 regulate dioxin receptor function and interaction with hsp90. The Journal of Biological Chemistry 277:11795–11801.

Kelley SK, Nilsson CB, Green MH, Green JB, Hakansson H. 1998. Use of model-based compartmental analysis to study effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on vitamin A kinetics in rats. Toxicological Sciences 44:1–13.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Kelley SK, Nilsson CB, Green MH, Green JB, Hakansson H. 2000. Mobilization of vitamin A stores in rats after administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin: a kinetic analysis. Toxicological Sciences 55:478–484.

Kenyon EM, Hughes MF. 2001. A concise review of the toxicity and carcingenicity of demethylarsinic acid. Toxicology 160:227–236.

Kerkvliet NI. 2002. Recent advances in understanding the mechanisms of TCDD immunoxicity International Immunopharmacology 2:277–291.

Kern PA, Fishman RB, Song W, Dicker-Brown A, Fonseca VA. 2002a. The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on oxidative enzymes in adipocytes and liver. Toxicology 171:117–125.

Kern PA, Dicker-Brown A, Said ST, Kennedy R, Fonseca VA. 2002b. The stimulation of tumor necrosis factor and inhibition of glucose transport and lipoprotein lipase in adipose cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Metabolism 51:65–68.

Khorram O, Garthwaite M, Golos T. 2002. Uterine and ovarian aryl hydrocarbon receptor (AHR) and aryl hydrocarbon receptor nuclear translocator (ARNT) mRNA expression in benign and malignant gynaecological conditions. Molecular Human Reproduction 8:75–80.

Kim CS, Sandberg JA, Slikker W Jr, Binienda Z, Schlosser PM, Patterson TA. 2001. Quantitative exposure assessment: application of physiologically-based pharmacokinetic (PBPK) modeling of low-dose, long-term exposures of organic acid toxicant in the brain. Environmental Toxicology and Pharmacology 9:153–160.

Kim DW, Gazourian L, Quadri SA, Romieu-Mourez R, Sherr DH, Sonenshein GE. 2000. The RelA NF-κB subunit and the aryl hydrocarbon receptor (AhR) cooperate to transactivate the c-myc promoter in mammary cells. Oncogene 19:5498–5506.

King MD, Lindasy, DS, Ehrich MF, Nagarkatti M. 2000. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on T cell subpopulations in the thymus and spleen of mice with chronic Toxoplasma gondi infection. International Journal of Toxicology 19:323–329.

Kitamura K, Nagao M, Yamada T, Sunaga M, Hata J, Watanabe S. 2001. Dioxins in bile in relation to those in human liver and blood. Journal of Toxicological Science 26:327–336.

Klinge CM, Kaur K, Swanson HI. 2000. The aryl hydrocarbon receptor interacts with estrogen receptor alpha and orphan receptors COUP-TF1 and ERRa1. Archives of Biochemistry and Biophysics 378:163–174.

Klinge CM, Jernigan SC, Risinger KE, Lee JE, Tyulmenkov VV, Falkner KC, Prough RA. 2001. Short heterodimer partner (SHP) orphan nuclear receptor inhibits the transcriptional activity of aryl hydrocarbon receptor (AHR)/AHR nuclear translocator (ARNT). Archives of Biochemistry and Biophysics 390:64–70.

Kobal S, Cebulj-Kadunc N, Cestnik V. 2000. Serum T3 and T4 concentrations in adult rats treated with herbicide 2,4-dichlorophenoxyacetic acid. European Journal of Physiology 440(suppl): R171–R172.

Kohle C, Hassepass I, Bock-Hennig BS, Bock KW, Poellinger L, McGuire J. 2002. Conditional expression of a constitutively active aryl hydrocarbon receptor in MCF-7 human breast cancer cells. Archives of Biochemistry and Biophysics 402:172–179.

Kohn MC, Walker NJ, Kim AH, Portier CJ. 2001. Physiological modeling of a proposed mechanism of enzyme induction by TCDD. Toxicology 162:193–208.

Koliopanos A, Kleef J, Xiao Y, Safe S, Zimmerman A, Buchler MW, Friess H. 2002. Increased arylhydrocarbon receptor expression offers a potential therapeutic target for pancreatic cancer. Oncogene 21(39):6059–6070.

Kolluri SK, Weiss C, Koff A, Gottlicher M. 1999. P27kip1 induction and inhibition of proliferation by the intracellular Ah receptor in developing thymus and hepatoma cells. Genes and Development 13:1742–1753.

Kolluri SK, Balduf C, Hofmann M, Gottlicher M. 2001. Nover target genes of the Ah (dioxin) receptor: transcriptional induction of N-myristoyltransferase 2. Cancer Research 61:8534–8539.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Kono Y, Okada S, Tazawa Y, Kanzaki S, Mura T, Ueta E, Nanba I, Otsuka Y. 2001. Effect of lactational exposure to 1,2,3,4-tetrachlorodibenzo-p-dioxin on cytochrome P-450 1A1 mRNA in the neonatal rat liver: Quantitative analysis by the competitive RT-PCR method. Pediatrics International 43:458–464.

Korkalainen M, Tuomisto J, Pohjanvirta R. 2001. The AH receptor of the most dioxin-sensitive species, guinea pig, is highly homologous to the human AH receptor. Biochemical and Biophysical Research Communications 285:1121–1129.

Korner W, Golor G, Schulz T, Wiesmuller T, Hagenmaier H, Neubert D. 2002. Tissue concentrations and induction of hepatic monooxygenase in male Wistar rats after repeated doses of defined polychlorinated dibenzo-p-dioxin and dibenzofuran (PCDDs and PCDFs) mixtures. Archives of Toxicology 75:653–664.

Kouri G, Theophilidis G. 2002. The action of the herbicide 2,4-dichlorophenoxyacetic acid on the isolated sciatic nerve of the frog (Rana Ridibunda). Neurotoxicity Research 4:25–32.

Kraemer SA, Arthur KA, Denison MS, Smith WL, DeWitt DL. 1996. Regulation of prostaglandin endoperoxide H synthase-2 expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Archives of Biochemistry and Biophysics 330:319–328.

Krig SR, Rice RH. 2000. TCDD suppression of tissue transglutaminase stimulation by retinoids in malignant human keratinocytes. Toxicological Sciences 56:357–364.

Krig SR, Chandraratna RAS, Chang MMJ, Wu R, Rice RH. 2002. Gene-specific TCDD suppression of RARα- and RXR-mediated induction of tissue transglutaminase. Toxicological Sciences 68:102–108.

Krishnan V, Porter W, Santostefano M, Wang X, Safe S. 1995. Molecular mechanism of inhibition of estrogen-induced cathepsin D gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in MCF-7 cells. Molecular and Cellular Biology 15:6710–6719.

Kronenberg S, Esser C, Carlberg C. 2000. An aryl hydrocarbon receptor conformation acts as the functional core of nuclear dioxin signaling. Nucleic Acids Research 28:2286–2291.

Kuchiiwa S, Cheng S-B, Nagatomo I, Akasaki Y, Uchida M, Tominaga M, Hashiguchi W, Kuchiiwa T. 2002. In utero and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin decreases serotonin-immunoreactive neurons in raphe nuclei of male mouse offspring. Neuroscience Letters 317:73–76.

Kumar MB, Ramadoss P, Reen RK, Vanden Heuvel JP, Perdew GH. 2001. The Q-rich subdomain of the human Ah receptor transactivation domain is required for dioxin-mediated transcriptional activity. The Journal of Biological Chemistry 276:42302–42310.

Kurachi M, Hashimoto S-I, Obata A, Nagai S, Nagahata T, Inadera H, Sone H, Tohyama C, Kaneko S, Kobayashi K-I, Matsushima K. 2002. Identification of 2,3,7,8-tetrachlorodibenzo-p-dioxin-responsive genes in mouse liver by serial analysis of gene expression. Biochemical and Biophysical Research Communications 292:368–377.

Lai Z-W, Pineau T, Esser C. 1996. Identification of dioxin-responsive elements (DREs) in the 5'-region of putative dioxin inducible genes. Chemical-Biological Interactions 100:97–112.

Lai Z-W, Fiore NC, Hahn PJ, Gasiewicz TA, Silverstone AE. 2000. Differential effects of diethylstilbestrol and 2,3,7,8-tetrachlorodibenzo-p-dioxin on thymocyte differentiation, proliferation, and apoptosis in blc-2 transgenic mouse fetal thymus organ culture. Toxicology and Applied Pharmacology 168:15–24.

Lamb JG, Straub P, Tukey RH. 1994. Cloning and characterization of cDNAs encoding mouse Ugt 1.6 and rabbit UGT 1.6: differential induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Biochemistry 33:10513–10520.

LaPres JJ, Glover E, Dunham EE, Bunger MK, Bradfield CA. 2000. ARA9 modifies agonist signaling through an increase in cytosolic aryl hydrocarbon receptor. The Journal of Biological Chemistry 275:6153–6159.

Latchoumycandane C, Chitra KC, Mathur PP. 2002a. Induction of oxidative stress in rat epididymal sperm after exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Archives of Toxicology 76:113– 118.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Latchoumycandane C, Chitra KC, Mathur PP. 2002b. The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the antioxidant system in mitochondrial and microsomal fractions of rat testis. Toxicology 171:127–135.

Lawrence BP, Warren TK, Luong G. 2000. Fewer T lymphocytes and decreased pulmonary influenza virus burden in mice exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Journal of Toxicology and Environmental Health 61:39–53.

Lee JE, Safe S. 2001. Involvement of a post-transcriptional mechanism in the inhibition of CYP1A1 expression by resveratrol in breast cancer cells. Biochemical Pharmacology 62:1113–1124.

Lee K, Johnson VJ, Blakley BR. 2001. The effect of exposure to a commercial 2,4-D formulation during gestation on the immune response in CD-1 mice. Toxicology 165:39–49.

Lee K, Johnson VJ, Blakley BR. 2000. The effect of exposure to a commerical 2,4-D herbicide formulation during gestation on urethan-induced lung adenoma formation in CD-1 mice. Veterinary and Human Toxicology 42(3):129–132.

Legare ME, Hanneman WH, Barhouni R, Burghardt RC, Tiffany-Castiglioni E. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin alters hippocampal astroglia-neuronal gap junction communication NeuroToxicology 21:1109–1116.

Levine SL, Petrulis JR, Dubil A, Perdew GH. 2000. A tetratricopeptide repeat half-site in the aryl hydrocarbon receptor is important for DNA binding and trans-activation potential. Molecular Pharmacology 58:1517–1524.

Lewis BC, Hudgins S, Lewis A, Schorr K, Sommer R, Peterson RE, Flaws JA, Furth PA. 2001. In utero and lactational treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin impairs mammary gland differentiation but does not block the response to exogenous estrogen in the postpubertal female rat Toxicological Sciences 62:46–53.

Lin T-M, Ko K, Moore, RW, Buchanan DL, Cooke PS, Peterson RE. 2001. Role of the aryl hydrocarbon receptor in the development of control and 2,3,7,8-tetrachlorodibenzo-p-dioxin-exposed male mice. Journal of Toxicology and Environmental Health, Part A 64:327–342.

Lin T-M, Ko K, Moore RW, Simanainen U, Oberly TD, Peterson RE. 2002. Effects of aryl hydrocarbon receptor null mutation and in utero and lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure on prostate and seminal vesicle development in C57BL/6 mice. Toxicological Sciences 68:479–487.

Liu PCC, Moreno-Aliaga MJ, Dunlap DY, Hu XM, Denison MS, Matsumura F. 2002. Correlation between the high expression of C/EBPb protein in F442A cells and their relative resistance to antiadipogenic action of TCDD in comparison to 3T3-L1 cells. Journal of Biochemical and Molecular Toxicology 16:70–83.

Loertscher JA, Sadek CS, Allen-Hoffmann BL. 2001a. Treatment of normal human keratinocytes with 2,3,7,8-tetrachlorodibenzo-p-dioxin causes a reduction in cell number, but no increase in apoptosis Toxicology and Applied Pharmacology 175:114–120.

Loertscher JA, Sattler CA, Allen-Hoffmann BL. 2001b. 2,3,7,8-Tetrachlorodibenzo-p-dioxin alters the differentiation pattern of human keratinocytes in organotypic culture. Toxicology and Applied Pharmacology 175:121–129.

Luebke RW, Copeland CB, Daniels M, Lambert AL, Gilmour MI. 2001 Suppression of allergic immune responses to house dust mite (HDM) in rats exposed to 2,3,7,8-TCDD. Toxicolological Sciences 62:71–79.

Lukinmaa PL. Sahlberg C, Leppäniemi A, Partanen AM, Kovero O, Pohjanvirta R, Tuomisto J, Alaluusua S. 2001. Arrest of rat molar tooth development by lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin 173:38–47.

Ma Q, Baldwin KT. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced degradation of aryl hydrocarbon receptor (AhR) by the ubiquitin-proteasome pathway. The Journal of Biological Chemistry. 275:8432–8438.

Ma Q, Baldwin KT, Renzelli AJ, McDaniel A, Dong L. 2001. TCDD-inducible poly (ADP-ribose) polymerase: a novel response to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Biochemical and Biophysical Research Communications 289:499–506.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Mace K, Bowman ED, Vautravers P, Shields PG, Harris CC, Pfeifer AMA. 1998. Characterization of xenobiotic metabolizing enzyme expression in human bronchial mucosa and peripheral lung tissues. European Journal of Cancer 34:914–920.

Machala M, Vondracek J, Blaha L, Ciganek M, Neca J. 2001. Aryl hydrocarbon receptor-mediated activity of mutagenic polycyclic aromatic hydrocarbons determined using in vitro reporter gene assay Mutation Research 497:49–62.

Maier A, Dalton TP, Puga A. 2000. Disruption of dioxin-inducible phase I and phase II gene expression patterns by cadmium, chromium, and arsenic. Molecular Carcinogenesis 28:225–235.

Mandal PK, McDaniel LR, Prough RA, Clark BJ. 2001. 7,12-dimethylbenz[a]anthracene inhibition of steroid production in MA-10 mouse Leydig tumor cells is not directly linked to induction of CYP1B1. Toxicology and Applied Pharmacology 175:200–208.

Manz A, Papke O, Baur X. 2001. Transfer at home of 2,3,7,8-tetrachlorodibenzo-p-dioxin and β-hexachlorocyclohexane. Gesundheitswesen 63:398–403.

Markowski VP, Zareba G, Stern S, Cox, C, Weiss B. 2001. Altered operant responding for motor reinforcement and the determination of benchmark doses following perinatal exposure to lowlevel 2,3,7,8-tetrachlorodibenzo-p-dioxin. Environmental Health Perspectives 109:621–627.

Markowski VP, Cox C, Preston R, Weiss B. 2002. Impaired cued delayed alteration behavior in adult rat offspring following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin on gestation day 15. Neurotoxicology and Teratology 24:209–218.

Mass MJ, Tennant A, Roop BC, Cullen WR, Styblo M, Thomas DJ, Kligerman AD. 2001. Methylated trivalent arsenic species are genotoxic. Chemical Research in Toxicology 14:355–361.

Masten SA, Shiverick KT. 1995. The Ah receptor recognizes DNA binding sites for the B cell transcription factor, BSAP: A possible mechanism for dioxin-mediated alteration of CD19 gene expression in human B lymphocytes. Biochemical and Biophysical Research Communications 212:27–34.

Mathieu M-C, Lapierre I, Brault K, Raymond, M. 2001. Aromatic hydrocarbon receptor (AhR)-AhR nuclear translocator- and p53-mediated induction of the murine multidrug resistance mdr1 gene by 3-methylcholanthrene and benzo(a)pyrene in hepatoma cells. The Journal of Biological Chemistry 276:4819–4827.

Matikainen TM, Moriyama T, Morita Y, Perez GI, Korsmeyer SJ, Sherr DH, Tilly JL. 2002. Ligand activation of the aromatic hydrocarbon receptor transcription factor drives Bax-dependent apoptosis in developing fetal ovarian germ cells. Endocrinology 143:615–620.

Matthews M, Heimler I, Fahy M, Radwanska E, Hutz R, Trewin A, Rawlins R. 2001. Effects of dioxin, an environmental pollutant, on mouse blastocyst development and apoptosis. Fertility and Sterility 75:1159–1162.

McNulty WP. 1985. Toxicity and fetotoxicity of TCDD, TCDF and PCB isomers in rhesus macaques (Macaca mulatta). Environmental Health Perspectives 60:77–88.

Mhin BJ, Lee JE, Choi W. 2002. Understanding the congener-specific toxicity in polychlorinated dibenzo-p-dioxins: Chlorination pattern and molecular quadrupole moment. Journal of the American Chemical Society 124:144–148.

Michalek JE, Pirkle JL, Needham LL, Patterson DG Jr, Caudill SP, Tripathi RC, Mocarelli P. 2002. Pharmacokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin in Seveso adults and veterans of operation Ranch Hand. Journal of Exposure Analysis and Environmental Epidemiology 12:44– 53.

Miniero R, De Felip E, Ferri F, di Domenico A. 2001. An overview of TCDD half-life and its correlation to body weight. Chemosphere 43:839–844.

Moran FM, Chen TJ, Santos S, Cheney A, Overstreet JW, Lasley BL. 2001. Effect of dioxin on ovarian function in the cynomogus macaque (M. fascicularis). Reproductive Toxicology 15: 377–383.

Morita K, Nakano T. 2002. Seaweed accelerates the excretion of dioxin stored in rats. Journal of Agricultural and Food Chemistry 50:910–917.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Morita, K, Ogata M, Hasegawa T. 2001. Chlorophyll derived from Chlorella inhibits dioxin absorption from the gastrointestinal tract and accelerates dioxin excretion in rats Environmental Health Perspectives 109:289–294.

Muller GF, Dohr O, El-Bahay C, Kahl R, Abel J. 2000. Effect of transforming growth factor-beta1 on cytochrome P450 expression inhibition of CYP1 mRNA and protein expression in primary rat hepatocytes Archives of Toxicology 74:145–152.

Murante FG, Gasiewicz TA. 2000. Hemopoietic progenitor cells are sensitive targets of 2,3,7,8-tetrachlorodibenzo-p-dioxin in C57BL/6J mice. Toxicological Sciences 54:374–383.

Nagashima H, Matsumura F. 2002. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induced downregulation of glucose transporting activities in mouse 3T3-L1 preadipocyte. Journal of Environmental Science and Health B37:1–14.

Nayyar T, Zawia NH, Hood DB. 2002. Transplacental effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on the temporal modulation of Sp1 DNA binding in the developing cerebral cortex and cerebellum. Experimental Toxicologic Pathology 53:461–468.

Nazarenko DA, Dertinger SD, Gasiewicz TA. 2001. In vivo antagonism of AhR-mediated gene induction by 3'-methoxy-4'-nitroflavone in TCDD-responsive lacZ mice. Toxicological Sciences 61:256–264.

Nebert DW, Roe AL, Dieter MZ, Solis WA, Yang Y, Dalton TP. 2000. Role of the aromatic hydrocarbon receptor and [Ah] gene battery in the oxidative stress response, cell cycle control, and apoptosis Biochemical Pharmacology 59:65–85.

Needham LL, Gerthoux PM, Patterson DG Jr, Brambilla P, Pirkle JL, Tramacere PL, Turner WE, Beretha C, Sampson EJ, Mocarelli P. 1994. Half-life of 2,3,7,8-tetrachlorodibenzo-p-dioxin in serum of Seveso adults. Interim report. Organohalogen Compounds 21:81–85.

Neubert D, Wiesmuller T, Abraham K, Krowke R, Hagenmaier H. 1990. Persistence of various polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDDs and PCDFs) in hepatic and adipose tissue of marmoset monkeys. Archives of Toxicology 64:431–442.

Nie M, Blankenship AL, Giesy JP. 2001. Interactions between aryl hydrocarbon receptor (AhR) and hypoxia signaling pathways. Environmental Toxicology and Pharmacology 10:17–27.

Nilsson CB, Hoegberg P, Trossvik C, Azais-Braesco V, Blaner WS, Fex G, Harrison EH, Nau H, Schmidt CK, van Bennekum AM, Hakansson H. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin increases in serum and kidney retinoic acid levels and kidney retinal esterification in the rat. Toxicology and Applied Pharmacology 169:121–131.

Nishimura N, Miyabara Y, Suzuki JS, Sato M, Aoki Y, Satoh M, Yonemoto J, Tohyama C. 2001. Induction of metallothionein in the livers of female Sprague Dawley rats treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin. Life Sciences 69:1291–1303.

Noda Y, Suzuki T, Kohara A, Hasegawa A Yotsuyanagi T, Hayashi M, Sofuni T, Yamanaka K, Okada S. 2002. In vivo genotoxicity evaluation of dimethylarsinic acid in Muta TM Mouse. Mutation Research 513:205–212.

Nohara K, Fujimaki H, Tsukumo S, Ushio H, Miyabara Y, Kijima M, Tohyama C, Yonemoto J. 2000. The effects of perinatal exposure to low doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin on immune organs in rats. Toxicology 154:123–133.

Nohara K, Fujimaki H, Tsukumo S-I, Inouye K, Sone H, Tohyama C. 2002a. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on T cell-derived cytokine production in ovalbumin (OVA)-immunized C57Bl/6 mice. Toxicology 172:49–58.

Nohara K, Izumi H, Tamura SI, Nagata R, Tohyama C. 2002b. Effect of low-dose 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on influenza A virus-induced mortality in mice. Toxicology 170:131–138.

NRC (National Research Council). 1999. Arsenic in Drinking Water. Washington, DC: National Academy Press.

NRC. 2001. Arsenic in Drinking Water: Update 2001. Washington, DC: National Academy Press.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Nukaya M, Takahashi Y, Gonzalez FJ, Kamataki T. 2001. Aryl hydrocarbon receptor-mediated suppression of expression of the low-molecular weight prekininogen gene in mice. Biochemical and Biophysical Research Communications 287:301–304.

Oakes DJ, Pollak JK. 2000. The in vitro evaluation of the toxicities of three related herbicide formulations containing ester derivatives of 2,4,5-T and 2,4-D using sub-mitochondrial particles. Toxicology 151:1–9.

Oakes DJ, Webster WS, Brown-Woodman PDC, Ritchie HE. 2002a. A study of the potential for a herbicide formulation containing 2,4-D and picloram to cause male-mediated developmental toxicity in rats Toxicological Sciences 68:200–206.

Oakes DJ, Webster WS, Brown-Woodman PD, Ritchie HE. 2002b. Testicular changes induced by chronic exposure to the herbicide formulation, Tordon 75D (2,4-dichlorophenoxyacetic acid and picloram) in rats Reproductive Toxicology 16:281–289.

Ogi T, Mimura J, Hikida M, Fujimoto H, Fujii-Kuriyama Y, Ohmori H. 2001. Expression of human and mouse genes encoding polkappa: testis-specific developmental regulation and AhR-dependent inducible transcription Cells to Genes 6:943–953.

Ohbayashi T, Oikawa K, Iwata R, Kamea A, Evine K, Isobe T, Matsuda Y, Mimura J, Fujii-Kuriyama Y, Kuroda M, Mukai K. 2001. Dioxin induces a novel nuclear factor, DIF-3, that is implicated in spermatogenesis. FEBS Letters 508:341–344.

Ohsako S, Miyahara Y, Nishimura N, Kurosawa S, Sakaue M, Ishimura R, Sato M, Takeda K, Aoki Y, Sone H, Tohyama C, Yonemoto J. 2001. Maternal exposure to a low dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) suppressed the development of reproductive organs of male rats: Dose dependent increase of mRNA levels of 5alpha-reductase type 2 in contrast to decrease of androgen receptor in the pubertal ventral prostate. Toxicological Sciences 60:132– 143.

Ohsako S, Miyabara Y, Sakaue M, Ishimura R, Kakeyame M, Izumi H, Yonemoto J, Tohyama C. 2002. Developmental stage-specific effects of perinatal 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure on reproductive organs of male rat offspring. Toxicological Sciences 66:283–292.

Oikawa K, Ohbayashi T, Mimura J, Fujii-Kuriyama Y, Teshima S, Rokutan K, Mukai K, Kuroda M. 2002. Dioxin stimulates synthesis and secretion of IgE-dependent histamine-releasing factor. Biochemical and Biophysical Research Communications 290:984–987.

Ozaki K, Mahler JF, Haseman JK, Moomaw CR, Nicolette ML, Nyska A. 2001. Unique renal tubule changes induced in rats and mice by the peroxisome proliferators 2,4-dichlorophenoxy acetic acid (2,4-D) and WY-14643 Toxicologic Pathology 29(4):440–450.

Oztas H. 2000. Effects of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)) on the early stages of pancreatic carcinogenesis induced by azaserine in the rat pancreas. Turkish Journal of Medical Sciences 30:29–34.

Park J-H, Lee S-W. 2002. Up-regulated expression of genes encoding Hrk and IL-3R beta subunit by TCDD in vivo and in vitro. Toxicology Letters 129:1–11.

Park S, Henry EC, Gasiewicz TA. 2000. Regulation of DNA binding activity of the ligand-activated aryl hydrocarbon receptor by tyrosine phosphorylation. Archives of Biochemistry and Biophysics 381:302–312.

Partridge NC, Fiacco GJ, Walling HW, Barmina OY, Jeffrey JJ, Ruh MF. 2000. Effects of dioxin and estrogen on collagenase-3 in UMR 106-01 osteocarcoma cells. Archives of Biochemistry and Biophysics 382:182–188.

Petersen SL, Curran MA, Marconi SA, Carpenter CD, Lubbers LS, McAbee MD. 2000. Distribution of mRNAs encoding the arylhydrocarbon receptor, arylhydrocarbon receptor nuclear translocator, and arylhydrocarbon receptor nuclear translocator-2 in the rat brain and brainstem. The Journal of Comparative Neurology 427:428–439.

Petroff BK, Gao X, Rozman KK, Terranova PF. 2000. Interaction of estradiol and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in an ovulation model: evidence for systemic potentiation and local ovarian effects. Reproductive Toxicology 14:247–255.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Petroff BK, Roby KF, Gao X, Son D-S, Williams S, Johnson D, Rozman KK, Terranova PF. 2001. A review of mechanisms controlling ovulation with implications for the anovulatory effects of polychlorinated dibenzo-p-dioxins in rodents. Toxicology 158:91–107.

Petroff BK, Gao X, Ohshima K-I, Shi FX, Son D-S, Roby KF, Rozman KK, Watanabe G, Taya K, Terranova PF. 2002. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on serum inhibin concentrations and inhibin immunostaining during follicular development in female Sprague-Dawley rats. Reproductive Toxicology 16:97–105.

Petrulis JR, Bunce NJ. 2000. Competitive behavior in the interactive toxicology of halogenated aromatic compounds. Journal of Biochemical and Molecular Toxicology 14:73–81.

Petrulis JR, Hord NG, Perdew GH. 2000. Subcellular localization of the aryl hydrocarbon receptor is modulated by the immunophilin homolog hepatitis B virus X-associated protein 2. The Journal of Biological Chemistry 275:37448–37453.

Pieklo Z, Grochowalski A, Gregoraszczuk EL. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin alters follicular steroidogenesis in a time- and cell-specific manner. Experimental and Clinical Endocrinology and Diabetes 108:299–304.

Pilat-Marcinkiewicz B, Sulik M, Szynaka B, Jablonska E, Andrzejewicz A. 2000. Fetotoxic action of 2,4-dichlorophenolyacetic acid (2,4-D). II. Ultrastructural changes in rat hepatocytes. Acta Poloniae Toxicologica 8:71–79.

Pimental RA, Liang B, Yee GK, Wilhelmsson A, Poellinger L, Paulson KE. 1993. Dioxin receptor and C/EBP regulate the function of the glutathione S-transferase Ya gene xenobiotic response element. Molecular and Cellular Biology 13:4365–4373.

Pirkle JL, Wolfe WH, Patterson DG, Needham LL, Michalek JE, Miner JC, Peterson MR, Phillips DL. 1989. Estimates of the half-life of 2,3,7,8-tetrachlorodibenzo-p-dioxin in Vietnam veterans of Operation Ranch Hand. Journal of Toxicology and Environmental Health 27(2):165–171.

Pitt JA, Buckalew AR, House DE, Abbott BD. 2000. Adrenocorticotropin (ACTH) and corticosterone secretion by perfused pituitary and adrenal glands from rodents exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Toxicology 151:25–35.

Pitt JA, Feng L, Abbott BD, Schmid J, Batt RE, Costich TG, Koury ST, Bofinger DP. 2001. Expression of AhR and ARNT mRNA in cultured human endometrial explants exposed to TCDD. Toxicological Sciences 62:289–298.

Pohjanvirta R, Vartiainen T, Uusi-Rauva A, Monkkonen J, Tuomisto J. 1990. Tissue distribution, metabolism, and excretion of 14C-TCDD in a TCDD-susceptible and a TCDD-resistant rat strain. Pharmacology and Toxicology 66:93–100.

Pohjanvirta R, Korkalainen M, McGuire J, Simanainen U, Juvonen R, Tuomisto JT, Unkila M, Viluksela M, Bergman J, Poellinger L, Tuomisto J. 2002. Comparison of acute toxicities of indolo[3,2-b]carbazole (ICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in TCDD-sensitive rats. Food and Chemical Toxicology 40:1023–1032.

Poland A, Glover E. 1973. Chlorinated dibenzo-p-dioxins: potent inducers of δ-aminolevulinic acid synthetase and aryl hydrocarbon hydroxylase. II. Study of the struture-activity relationship. Molecular Pharmacology 9:736–747.

Poland A, Knutson JC. 1982. 2,3,7,8-Tetrachlorodibenzo-p-dioxin and related halogenated aromatic hydrocarbons: examination of the mechanism of toxicity. Annual Review of Pharmacology and Toxicology 22:517–554.

Pollenz RS, Barbour ER. 2000. Analysis of the complex relationship between nuclear export and aryl hydrocarbon receptor-mediated gene regulation. Molecular and Cellular Biology 20:6095–6104.

Pollenz RS, Necela B, Marks-Sojka K. 2002. Analysis of rainbow trout Ah receptor isoforms in cell culture reveals conservation of function of Ah receptor-mediated signal transduction Biochemical Pharmacology 64:49–60.

Porter W, Wang F, Duan R, Qin C, Castro-Rivera E, Kim K, Safe S. 2001. Transcriptional activation of heat shock protein 27 gene expression by 17β-estradiol and modulation by antiestrogens and aryl hydrocarbon receptor agonists. Journal of Molecular Endocrinology 26:31–42.

Puga A, Nebert DW, Carrier F. 1992. Dioxin induces expression of c-fos and c-jun proto-oncogenes and a large increase in transcription factor AP-1. DNA Cell Biology 11:269–281.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Puga A, Maier A, Medvedovic M. 2000a. The transcriptional signature of dioxin in human hepatoma HepG2 cells Biochemical Pharmacology 60:1129–1142.

Puga A, Barnes SJ, Chang C-y, Zhu H, Nephew KP, Khan SA, Shertzer HG. 2000b. Activation of transcription factors activator protein-1 and nuclear factor-κB by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Biochemical Pharmacology 59:997–1005.

Quadri SA, Qadri AN, Hahn ME, Mann KK, Sherr DH. 2000. The bioflavonoid galangin blocks aryl hydrocarbon receptor activation and polycyclic aromatic hydrocarbon-induced pre-B cell apoptosis Molecular Pharmacology 58:515–525.

Ramakrishna G, Perella C, Birely L, Diwan BA, Fornwald LW, Anderson LM. 2002. Decrease in K-ras p21 and increase in Raf1 and activated Erk 1 and 2 in murine lung tumors initiated by N-nitrosodimethylamine and promoted by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicology and Applied Pharmacology 179:21–34.

Render JA, Hochstein JR, Aulerich RJ, Bursian SJ. 2000. Proliferation of periodontal squamous epithelium in mink fed 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Veterinary and Human Toxicology 42:85–86.

Render JA, Bursian SJ, Rosenstein DS, Aulerich FJ. 2001. Squamous epithelial proliferation in the jaws of mink fed diets containing 3,3',4,4',5-pentachlorobiphenyl (PCB 126) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Veterinary and Human Toxicology 43:22–26.

Richter CA, Tillitt DE, Hannink M. 2001. Regulation of subcellular localization of the aryl hydrocarbon receptor (AhR). Archives of Biochemistry and Biophysics 389:207–217.

Riecke K, Grimm D, Shakibaei M, Kossmehl P, Schulze-Tanzil G, Paul M, Stahlmann R. 2002. Low doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin increase transforming growth factor β and cause myocardial fibrosis in marmosets (Callithrix jacchus). Archives of Toxicology 76:360–366.

Rier SE, Coe CL, Lemieux AM, Martin DC, Morris R, Lucier GW, Clark GC. 2001a. Increased tumor necrosis factor-α production by peripheral blood leukocytes from TCDD-exposed Rhesus monkeys. Toxicological Sciences 60:327–337.

Rier SE, Turner WE, Martin DC, Morris R, Lucier GW, Clark GC. 2001b. Serum levels of TCDD and dioxin-like chemicals in Rhesus monkeys chronically exposed to dioxin: correlation of increased serum PCB levels with endometriosis. Toxicological Sciences 59:147–159.

Rivera SP, Saarikoski ST, Hankinson O. 2002. Identification of a novel dioxin-inducible cytochrome P450. Molecular Pharmacology 61:255–259.

Roberts EA, Harper PA, Wong JMY, Wang Y, Yang S. 2000. Failure of Ah receptor to mediate induction of cytochromes P450 in the CYP1 family in the human hepatoma line SK-Hep-1. Archives of Biochemistry and Biophysics 384:190–198.

Robinson SW, Clothier B, Akhtar RA, Yang AL, Latour I, Van Ijperen C, Festing MFW, Smith AG. 2002. Non-Ahr gene susceptibility loci for porphyria and liver injury induced by the interaction of “dioxin” with iron overload in mice. Molecular Pharmacology 61:674–681.

Roby KF. 2001. Alterations in follicle development, steroidogenesis, and gonadotropin receptor binding in a model of ovulatory blockade. Endocrinology 142:2328–2335.

Rogan W, Gladen B, Hung K, Koong S, Shih L, Taylor J, Wu Y, Yang D, Ragan N, Hsu C. 1988. Congenital poisoning by polychlorinated biphenyls and their contaminants in Taiwan. Science 241:334-336.

Rogers JM, Denison MS. 2002. Analysis of the antiestrogen activity of 2,3,7,8-tetrachlorodibenzo-p-dioxin in human ovarian carcinoma BG-1 cells. Molecular Pharmacology 61:1393–1403.

Rosso SB, Garcia GB, Madariaga MJ, de Duffard AME, Duffard RO. 2000. 2,4-Dichlorophenoxyacetic acid in developing rats alters behavior, myelination and regions in brain gangliosides pattern. Neurotoxicology 21(1–2):155–164.

Roth W, Voorman R, Aust SD. 1988. Activity of thyroid hormone-inducible enzymes following treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicology and Applied Pharmacology 92:65–74.

Rushing SR, Denison MS. 2002. The silencing mediator of retinoic acid and thyroid hormone receptors can interact with the aryl hydrocarbon (Ah) receptor but fails to repress Ah receptordependent gene expression. Archives of Biochemistry and Biophysics 403:189–201.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Sahlberg C, Pohjanvirta R, Gao Y, Aluluusua S, Tuomisto J, Lukinmaa P-L. 2002. Expression of the mediators of dioxin toxicity, aryl hydrocarbon receptor (AhR) and the AhR nuclear translocator (ARNT) is developmentally regulated in mouse teeth. International Journal of Developmental Biology 46:295–300.

Salvan A, Thomaseth K, Bortot P, Sartori N. 2001. Use of a toxicokinetic model in the analysis of cancer mortality in relation to the estimated absorbed dose of dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD). Science of the Total Environment 274:21–35.

Santiago-Josefat B, Pozo-Guisado E, Mulero-Navarro S, Fernandez-Salguero PM. 2001. Proteasome inhibition induces nuclear translocation and transcriptional activation of the dioxin receptor in mouse embryo primary fibroblasts in the absence of xenobiotics. Molecular and Cellular Biology 21:1700–1709.

Santini RP, Myrand S, Elferink C, Reiners JJ Jr. 2001. Regulation of CYP1A1 induction by dioxin as a function of cell cycle Journal of Pharmacology and Experimental Therapeutics 299:718–728.

Savouret JF, Antenos M, Quesne M, Xu J, Milgrom E, Casper RF. 2001. 7-Ketocholesterol is an endogenous modulator for the arylhydrocarbon receptor. The Journal of Biological Chemistry 276(5):3054–3059.

Schaufler K, Haslmayer P, Jager W, Pec M, Thalhammer T. 2002. The environmental toxin 2,3,7,8-tetrachlorodibenzo-p-dioxin induces cytochrome P450 activity in high passage PC 3 and DU 145 human prostate cancer cell lines. International Journal of Molecular Medicine 9:411–416.

Schecter A, McGee H, Stanley JS, Boggess K, Brandt-Rauf P. 1996. Dioxins and dioxin-like chemicals in blood and semen of American Vietnam veterans from the state of Michigan. American Journal of Industrial Medicine. 30:647–654.

Schnoor TM, Lawson CC, Whelan EA, Dankovic DA, Deddens JA, Piacitelli LA, Reefhuis J, Sweeney MH, Connally B, Fingerhut, MA. 2001. Spontaneous abortion, sex ratio, and paternal occupational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Environmental Health Perspectives 109(11):1127–1132.

Schrader TJ, Cooke GM. 2000. Examination of selected food additives and organochlorine food contaminants for androgenic activity in vitro. Toxicological Sciences 53:278–288.

Schrey P, Wittsiepe J, Ewers U, Exner M, Selenka F. 1993. Polychlorinated dibenzo-p-dioxins and dibenzofurans in human blood. Bundesgesundheitsblatt 11:455–463.

Scott MA, Tarara RP, Hendrichs AG, Benirschke K, Overstreet JW, Lasley BL. 2001. Exposure to the dioxin 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces squamous metaplasia in the endocervix of cynomolgus macaques. Journal of Medical Primatology 30:156–160.

Seidel SD, Winters GM, Rogers WJ, Ziccardi MH, Li V, Keser B, Denison MS. 2001. Activation of the Ah receptor signaling pathway by prostglandins. Journal of Biochemical and Molecular Toxicology 15:187–196.

Seike N, Wanibuchi H, Morimura K, Nishikawa T, Kishida H, Nakae D, Hirata K, Fukushima S. 2002. Lack of promoting effect due to oral administration of dimethylarsinic acid on rat lung carcinogenesis initiated with N-bis(2-hydroxypropyl)nitrosamine Cancer Letters 175:113–119.

Senft AP, Dalton TP, Nebert DW, Genter MB, Hutchinson RJ, Shertzer HG. 2002. Dioxin increases reactive oxygen production in mouse liver mitochondria Toxicology and Applied Pharmacology 178:15–21.

Sewall CH, Clark GC, Lucier GW. 1995. TCDD reduces rat hepatic epidermal growth factor receptor: comparison of binding, immunodetection and autophosphorylation. Toxicology and Applied Pharmacology 132:263–272.

Shehin SE, Stephenson RO, Greenlee WF. 2000. Transcriptional regulation of the human CYP1B1 gene. Evidence for involvement of an aryl hydrocarbon receptor response element in constitutive expression. Journal of Biological Chemistry 275:6770–6776.

Shepherd DM, Dearstyne EA, Kerkvliet NI. 2000. The effects of TCDD on the activation of ovalbumin (OVA)-specific DO11.10 transgenic CD4(+) T cells in adoptively transferred mice Toxicological Sciences 56:340–350.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Shimba S, Hayashi M, Sone H, Yonemoto J, Tezuka M. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces binding of a 50 kDa protein on the 3' untranslated region of urokinase-type plasminogen activator mRNA Biochemical and Biophysical Research Communications 272: 441–448.

Shimba S, Wada T, Tezuka M. 2001. Arylhydrocarbon receptor (AhR) is involved in negative regulation of adipose differentiation in 3T3-L1 cells: AhR inhibits adipose differentiation independently of dioxin. Journal of Cell Science 114:2809–2817.

Shridhar S, Farley A, Reid FL, Foster WG, Van Vugt DA. 2001. The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on corticotrophin-releasing hormone, arginine vasopressin, and pro-opiomelanocortin mRNA levels in the hypothalamus of the cynomolgus monkey. Toxicological Sciences 63:181–188.

Simanainen U, Tuomisto JT, Tuomisto J, Viluksela M. 2002. Structure-activity relationships and dose responses of polychlorinated dibenzo-p-dioxins for short-term effects in 2,3,7,8-tetrachlorodibenzo-p-dioxin-resistant and -sensitive rat strains. Toxicology and Applied Pharmacology 181:38–47.

Singh SU, Casper RF, Fritz PC, Sukhu B, Ganss B, Girard B Jr, Savouret JF, Tenenbaum HC. 2000. Inhibition of dioxin effects on bone formation in vitro by a newly described aryl hydrocarbon receptor antagonist, resveratrol. Journal of Endocrinology 167:183–195.

Slezak BP, Hatch GE, DeVito MJ, Diliberto JJ, Slade R, Crissman K, Hassoun E, Birnbaum LS. 2000. Oxidative stress in female B6C3F1 mice following acute and subchronic exposure to 2,3,7,8-tetrachlodibenzo-p-dioxin (TCDD). Toxicological Sciences 54:390–398.

Slezak BP, Hamm JT, Reyna J, Hurst CH, Birnbaum LS. 2002. TCDD-mediated oxidative stress in male rat pups following perinatal exposure. Journal of Biochemical and Molecular Toxicology 16:49–52.

Smart J, Daly AK. 2000. Variation in induced CYP1A1 levels: Relationship to CYP1A1, Ah receptor and CSTM1 polymorphisms. Pharmacogenetics 10:11–24.

Smith AG, Clothier B, Carthew P, Childs NL, Sinclair PR, Nebert DW, Dalton TP. 2001. Protection of the CYP1A2(-/-) null mouse against uroporphyria and hepatic injury following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicology and Applied Pharmacology 173:89–98.

Sordo M, Herrera LA, Ostrosky-Wegmand P, Rojas E. 2001. Cytotoxic and genotoxic effects of As, MMA, and DMA on leukocytes and stimulated human lymphocytes. Teratogenesis Carcinogenesis and Mutagenesis 21:249–260.

Sparrow BR, Thompson CS, Ryu B-W, Selevonchick DP, Schaup HW. 1994. 2,3,7,8-Tetrachlorodibenzo-p-dioxin induced alterations of pyruvate carboxylase levels and lactate dehydrogenase isozyme shifts in C57BL/6J male mice. Journal of Biochemical Toxicology 9:329–335.

Stanton BJ, El-Sabeawy F, Yang XF, Enan E, Lasley BL. 2001a. Interaction of estrogen and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in immature male chickens (Gallus domesticus). Comparative Biochemistry and Physiology Part C 129:35–47.

Stanton B, Watkins S, German JB, Lasley B. 2001b. Interaction of estrogen and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) with hepatic fatty acid synthesis and metabolism of male chickens (Gallus domesticus). Comparative Biochemistry and Physiology Part C 129:137–150.

Sterling KM Jr, Cutroneo KR. 2002. Differentiation-dependent induction of CYP1A1 in cultured rat small intestinal epithelial cells, colonocytes, and human colon carcinoma cells; basement membrane-mediated apoptosis. Journal of Cellular Biochemistry 86:440–450.

Sturtz N, Evangelista de Duffard AM, Duffard R. 2000. Detection of 2,4-dichlorophenoxyacetic acid (2,4-D) residues in neonates breast-fed by 2,4-D exposed dams. Neurotoxicology 21(1-2):147– 154.

Styblo M, Del Razo LM, Vega L, Germolec DR, LeCluyse EL, Hamilton GA, Reed W, Wang C, Cullen WR, Thomas DJ. 2000. Comparative toxicity of trivalent and pentavalent inorganic and methylated arsenicals in rat and human cells. Archives of Toxicology 74:289–299.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Sugawara T, Nomura E, Sakuragi N, Fujimoto S. 2001. The effect of the arylhydrocarbon receptor on the human steroidogenic acute regulatory gene promoter activity. Journal of Steroid Biochemistry and Molecular Biology 78:253–260.

Sugihara K, Kitamura S, Yamada T, Ohta S, Yamashita K, Yasuda M, Fujii-Kuriyama Y. 2001. Aryl hydrocarbon receptor (AhR)-mediated induction of xanthine oxidase/xanthine dehydrogenase activity by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Biochemical and Biophysical Research Communications 281:1093–1099.

Suh J, Jeon YJ, Kim HM, Kang JS, Kaminski NE, Yang K-H. 2002. Aryl hydrocarbon receptor-dependent inhibition of AP-1 activity by 2,3,7,8-tetrachlorodibenzo-p-dioxin in activated B cells. Toxicology and Applied Pharmacology 181:116–123.

Sutter TR, Guzman K, Dold KM, Greenlee WF. 1991. Targets for dioxin: genes for plasminogen activator inhibitor-2 and interleukin-1 beta. Science 254: 415–418.

Sutter TR, Tang YM, Hayes CL, Wo Y-Y, Jabs EW, Li X, Yin H, Cody CW, Greenlee WF. 1994. Complete cDNA sequence of a human dioxin-inducible mRNA identifies a new gene subfamily of cytochrome P450 that maps to chromosome 2 Journal of Biological Chemistry 269: 13092–13099.

Svensson C, Lundberg K. 2001. Immune-specific up-regulation of adseverin gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin Molecular Pharmacology 60:135–142.

Svensson C, Silverstone AE, Lai Z-W, Lundberg K. 2002. Dioxin-induced adseverin expression in the mouse thymus is strictly regulated and dependent on the aryl hydrocarbon receptor. Biochemical and Biophysical Research Communications 291:1194–1200.

Takanaga H, Kunimoto M, Adachi T, Tohyama C, Aoki Y. 2001. Inhibitory effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on camp-induced differentiation of rat C6 glial cell line Journal of Neuroscience Research 64:402–409.

Takimoto K, Lindahl R, Pitot HC. 1992. Regulation of 2,3,7,8-tetrachlorodibenzo-p-dioxin-inducible expression of aldehyde dehydrogenase in hepatoma cells. Archives of Biochemistry and Biophysics 298(2):493–497.

Teraoka H, Dong W, Ogawa S, Tsukiyama S, Okuhara Y, Niiyama M, Ueno N, Peterson RE, Higaga T. 2002. 2,3,7,8-Tetrachlorodibenzo-p-dioxin toxicity in the zebrafish embryo: altered regional blood flow and impaired lower jaw development. Toxicological Sciences 65:192–199.

Theobald HM, Roman BL, Lin TM, Ohtani S, Chen SW, Peterson RE. 2000. 2,3,7,8-Tetrachlorodibenzo-p-dioxin inhibits luminal cell differentiation and androgen responsiveness of the ventral prostate without inhibiting prostatic 5alpha-dihydrotestosterone formation or testicular androgen production in rat offspring. Toxicological Sciences 58:324–338.

Thornton AS, Oda Y, Stuart GR, Glickman BW, de Boer JG. 2001. Mutagenicity of TCDD in Big Blue® transgenic rats. Mutation Research 478:45–50.

Tian Y, Ke S, Thomas T, Meeker RJ, Gallo MA. 1998. Transcriptional suppression of estrogen receptor gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Journal of Steroid Biochemistry and Molecular Biology 67:17–24.

Timms BG, Peterson RE, vom Saal FS. 2002. 2,3,7,8-Tetrachlorodibenzo-p-dioxin interacts with endogenous estradiol to disrupt prostate gland morphogenesis in male rat fetuses. Toxicological Sciences 67:264–274.

Tomita S, Sinal CJ, Yim SH, Gonzalez FJ. 2000. Conditional disruption of the aryl hydrocarbon receptor nuclear translocator (Arnt) gene leads to loss of target gene induction by the aryl hydrocarbon receptor and hypoxia-inducible factor 1α. Molecular Endocrinology 14:1674– 1681.

Tritscher AM, Mahler J, Portier CJ, Lucier GW, Walker NJ. 2000. Induction of lung lesions in female rats following chronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicologic Pathology 28:761–769.

Tsukumo SI, Iwata M, Tohyama C, Nohara K. 2002. Skewed differentiation of thymocytes toward CD8 T cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin requires activation of extracellular signal-related kinase pathway. Archives of Toxicology 76:335–343.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Tukey RH, Nebert DW. 1984. Regulation of mouse cytochrome P3-450 by the Ah receptor. Studies with a P3-450 cDNA clone. Biochemistry 23: 6003–6008.

Tuomisto JT, Viluksela M, Pohjanvirta R, Tuomisto J. 2000. Changes in food intake and food selection in rats after 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure. Pharmacology Biochemistry and Behavior 65:381–387.

Uchida T, Yoshida S, Inui Y, Takeda K. 2002. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on testosterone production in isolated murine testicular cells. Journal of Health Science 48:292– 295.

Uno S, Dalton TP, Shertzer HG, Genter MB, Warshawsky D, Talaska G, Nebert DW. 2001. Benzo[a]pyrene-induced toxicity: paradoxical protection in CYP1A1 (-/-) knockout mice having increased hepatic BaP-DNA adduct levels Biochemical and Biophysical Research Communications 289:1049–1056.

Van Birgelen APJM, van den Berg M. 2000. Toxicokinetics. Food Additives and Contaminants 17:267–273.

Van der Molen GW, Kooijman SALM, Slob W. 1996. A generic toxicokinetic model for persistent lipophilic compounds in humans: an application to TCDD. Fundamental and Applied Toxicology 31:83–94.

Van der Molen GW, Kooijman SALM, Wittsiepe J, Schrey P, Flesch-Janys D, Slob W. 2000. Estimation of dioxin and furan elimination rates with a pharmacokinetic model. Journal of Exposure Analysis and Environmental Epidemiology 10:579–585.

van der Plas SA, Lutkeschipholt I, Spenkelink B, Brouwer A. 2001. Effects of subchronic exposure to complex mixtures of dioxin-like and non-dioxin-like polyhalogenated aromatic compounds on thyroid hormone and vitamin A levels in female Sprague-Dawley rats. Toxicological Sciences 59:92–100.

Venkov P, Topashka-Ancheva M, Georgieva M, Alexieva V, Karanov E. 2000. Genotoxic effect of substituted phenoxyacetic acids. Archives of Toxicology 74:560–566.

Vijayaraghavan M, Wanibuchi H, Yamamoto S, Hakoi K, Nakae D, Konishi Y, Fukushima S. 2000. Lack of promoting potential of dimethylarsinic acid in kidney of male NCI-Black Reiter rats. Journal of Toxicologic Pathology 13:87–91.

Viluksela M, Duong TV, Stahl BU, Li X, Tuomisto J, Rozman KK. 1996. Toxicokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in two substrains of male Long-Evans rats after intravenous injection. Fundamental and Applied Toxicology 31:184–191.

Vogel C, Abel J. 1995. Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on growth factor expression in the human breast cancer cell line MCF-7. Archives of Toxicology 16:259–265.

Vorderstrasse BA, Kerkvliet NI. 2001. 2,3,7,8-Tetrachlorodibenzo-p-dioxin affects the number and function of murine splenic dendritic cells and their expression of accessory molecules. Toxicology and Applied Pharmacology 171:117–125.

Vorderstrasse BA, Steppan LB, Silverstone AE, Kerkvliet NI. 2001. Aryl hydrocarbon receptor-deficient mice generate normal immune responses to model antigens and are resistant to TCDD-induced immune suppression Toxicology and Applied Pharmacology 171:157–164.

Wagner E, Frank MM, Smialowicz RJ. 2001. 2,3,7,8-Tetrachlorodibenzo-p-dioxin and natural immunity: lack of an effect on the complement system in a guinea pig model. Toxicology 159:107– 113.

Walker MK, Heid SE, Smith SM, Swanson HI. 2000. Molecular characterization and developmental expression of the aryl hydrocarbon receptor from the chick embryo. Comparative Biochemistsry and Physiology Part C 126:305–319.

Walker NJ, Tritscher AM, Sills RC, Lucier GW, Portier CJ. 2000. Hepatocarcinogenesis in female Sprague-Dawley rats following discontinuous treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin. Toxicological Sciences 54:330–337.

Wang F, Samudio I, Safe S. 2001. Transcriptional activation of cathepsin D gene expression by 17β-estradiol: mechanism of aryl hydrocarbon receptor-mediated inhibition Molecular and Cellular Endocrinology 172:91–103.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×

Wang S, Hankinson O. 2002. Functional involvement of the Brahms/SWI2-related gene 1 protein in cytochrome P4501A1 transcription mediated by the aryl hydrocarbon receptor complex. Journal of Biological Chemistry 277:11821–11827.

Wang X, Santostefano MJ, DeVito MJ, Birnbaum LS. 2000. Extrapolation of a PBPK model for dioxins across dosage regimen, gender, strain and species. Toxicological Sciences 56:49–60.

Weber LW, Ernst SW, Stahl BU, Rozman K. 1993. Tissue distribution and toxicokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin in rats after intravenous injection. Fundamental and Applied Toxicology 21:523–534.

Wei M, Eanibuchi H, Yamamoto S, Li W, Fukushima S. 1999. Urinary bladder carcinogenicity of dimethylarsinic acid in male F344 rats. Carcinogenesis 20:1873–1876.

Wei C, Cacavale RJ, Kehoe JJ, Thomas PE, Iba MM. 2001. CYP1A2 is expressed along with CYP1A1 in the human lung. Cancer Letters 171:113–120.

Wong JMY, Harper PA, Meyer UA, Bock KW, Morike K, Lagueux J, Ayotte P, Tyndale RF, Sellers EM, Manchester DK, Okey AB. 2001a. Ethnic variability in the allelic distribution of human aryl hydrocarbon receptor codon 554 and assessment of variant receptor function in vitro. Pharmacogenetics 11:85–94.

Wong JMY, Okey AB, Harper PA. 2001b. Human aryl hydrocarbon receptor polymorphisms that result in loss of CYP1A1 induction. Biochemical and Biophysical Research Communications. 288:990–996.

Wormke M, Castro-Rivera E, Chen I, Safe S. 2000a. Estrogen and aryl hydrocarbon receptor expression and crosstalk in human Ishikawa endometrial cancer cells. Journal of Steroid Biochemistry and Molecular Biology 72:197–207.

Wormke M, Stoner M, Saville B, Safe S. 2000b. Crosstalk between estrogen receptor alpha and the aryl hydrocarbon receptor in breast cancer cells involves unidirectional activation of proteasomes. FEBS Letters 478:109–112.

Wu Q, Ohsako S, Baba T, Miyamoto K, Tohyama C. 2002. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on preimplantation mouse embryos. Toxicology 174:119–129.

Wyde ME, Seely J, Lucier GW, Walker NJ. 2000. Toxicity of chronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin in diethylnitrosamine-initiated ovariectomized rats implanted with subcutaneous 17β-estradiol pellets. Toxicological Sciences 54:493–499.

Wyde ME, Wong VA, Kim AH, Lucier GW, Walker NJ. 2001a. Induction of hepatic 8-oxy-deoxyguanosine adducts by 2,3,7,8-tetrachlorodibenzo-p-dioxin in Sprague-Dawley rats is female-specific and estrogen-dependent Chemical Research in Toxicology 14:849–855.

Wyde ME, Eldridge SR, Lucier GW, Walker NJ. 2001b. Regulation of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced tumor promotion by 17β-estradiol in female Sprague-Dawley rats. Toxicology and Applied Pharmacology 173:7–17.

Wyde ME, Cambre T, Lebetkin M, Eldridge SR, Walker NJ. 2002. Promotion of altered hepatic foci by 2,3,7,8-tetrachlorodibenzo-p-dioxin and 17β-estradiol in male Sprague-Dawley rats. Toxicology and Applied Pharmacology 68:295–303.

Wyman A, Lavin AL, Wilding GE, Gasiewicz TA. 2002. 2,3,7,8-Tetracholodibenzo-p-dioxin does not directly alter the phenotype of maturing B cells in a murine coculture system. Toxicology and Applied Pharmacology 180:164–177.

Yamanaka K, Takabayashi F, Mutsumi M, An Y, Hasegawa A, Okada S. 2001. Oral exposure of dimethylarsinic acid, a main metabolite of inorganic arsenics, in mice leads to an increase in 8-oxo-2'-deoxyguanosine level, specifically in the target organs for arsenic carcinogenisis Biochemical and Biophysical Research Communications 287:66–70.

Yamonoshita O, Saito T, Takahashi K, Hosokawa T, Okabe M, Ito K, Kurasaki M. 2001. 2,4,5-Trichlorophenoxyacetic acid inhibits apoptosis in PC12 cells Life Sciences 69:403–408.

Yang D, Li Y, Yuan X, Matoney L, Yan B. 2001. Regulation of rat carboxylesterase expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD): A dose-dependent decrease in mRNA levels but a biphasic change in protein levels and activity. Toxicological Sciences 64:20–27.

Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 30
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 31
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 32
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 33
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 34
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 35
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 36
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 37
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 38
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 39
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 40
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 41
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 42
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 43
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 44
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 45
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 46
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 47
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 48
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 49
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 50
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 51
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 52
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 53
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 54
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 55
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 56
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 57
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 58
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 59
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 60
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 61
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 62
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 63
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 64
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 65
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 66
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 67
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 68
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 69
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 70
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 71
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 72
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 73
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 74
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 75
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 76
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 77
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 78
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 79
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 80
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 81
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 82
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 83
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 84
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 85
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 86
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 87
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 88
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 89
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 90
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 91
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 92
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 93
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 94
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 95
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 96
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 97
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 98
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 99
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 100
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 101
Suggested Citation:"3. Toxicology." Institute of Medicine. 2003. Veterans and Agent Orange: Update 2002. Washington, DC: The National Academies Press. doi: 10.17226/10603.
×
Page 102
Next: 4. Overview of Epidemiologic Studies »
Veterans and Agent Orange: Update 2002 Get This Book
×
Buy Hardback | $107.00 Buy Ebook | $84.99
MyNAP members save 10% online.
Login or Register to save!
Download Free PDF

This book updates and evaluates the available scientific evidence regarding statistical associations between diseases and exposure to dioxin and other chemical compounds in herbicides used in Vietnam, focusing on new scientific studies and literature.

  1. ×

    Welcome to OpenBook!

    You're looking at OpenBook, NAP.edu's online reading room since 1999. Based on feedback from you, our users, we've made some improvements that make it easier than ever to read thousands of publications on our website.

    Do you want to take a quick tour of the OpenBook's features?

    No Thanks Take a Tour »
  2. ×

    Show this book's table of contents, where you can jump to any chapter by name.

    « Back Next »
  3. ×

    ...or use these buttons to go back to the previous chapter or skip to the next one.

    « Back Next »
  4. ×

    Jump up to the previous page or down to the next one. Also, you can type in a page number and press Enter to go directly to that page in the book.

    « Back Next »
  5. ×

    Switch between the Original Pages, where you can read the report as it appeared in print, and Text Pages for the web version, where you can highlight and search the text.

    « Back Next »
  6. ×

    To search the entire text of this book, type in your search term here and press Enter.

    « Back Next »
  7. ×

    Share a link to this book page on your preferred social network or via email.

    « Back Next »
  8. ×

    View our suggested citation for this chapter.

    « Back Next »
  9. ×

    Ready to take your reading offline? Click here to buy this book in print or download it as a free PDF, if available.

    « Back Next »
Stay Connected!