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7 Reproductive and Developmental Effects This chapter summarizes the scientiï¬c literature published since Veterans and Agent Orange: Update 2004, hereafter referred to as Update 2004 (IOM, 2005), on the association between exposure to herbicides and adverse reproduc- tive or developmental effects. (Analogous shortened names are used to refer to the updates for 1996, 1998, 2000, and 2002 [IOM, 1996, 1999, 2001, 2003].) The categories of association and the approach to categorizing the health outcomes are discussed in Chapters 1 and 2. The literature discussed includes papers that describe environmental, occupational, and Vietnam-veteran studies that evalu- ate herbicide exposure and the risk of birth defects, declines in sperm quality and fertility, spontaneous abortion, stillbirth, neonatal and infant mortality, low birth weight and preterm birth, and childhood cancer. In addition to studies of herbicides and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), studies of popula- tions exposed to polychlorinated biphenyls (PCBs) were reviewed when relevant because TCDD is sometimes a contaminant of PCBs. For new studies that report only a single reproductive health outcome and that are not revisiting a previously studied population, design information is summarized here with the results; de- sign information on all other new studies can be found in Chapter 4. This chapterâs primary emphasis is on the potential adverse reproductive effects of herbicide exposure in men because the vast majority of Vietnam veter- ans are men. About 8,000 women served in Vietnam (H. Kang, US Department of Veterans Affairs, personal communication, December 14, 2000), so ï¬ndings relevant to female reproductive health are also included. Studies that investigated the potential reproductive consequences of exposure of either parent were consid- ered; whenever the information was available, an attempt was made to evaluate the effects of maternal and paternal exposure separately. 517
518 VETERANS AND AGENT ORANGE: UPDATE 2006 FERTILITY Male reproductive function is under the control of several components whose proper coordination is important for normal fertility. Several of the components and some endpoints related to male fertility, including reproductive hormones and sperm characteristics, can be studied as indicators of fertility. The reproductive neuroendocrine axis involves the central nervous system, the anterior pituitary gland, and the testis. The hypothalamus integrates neural inputs from the cen- tral and peripheral nervous systems and regulates the gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH). Both are secreted into the circulation in episodic bursts by the anterior pituitary gland and are neces- sary for normal spermatogenesis. In the testis, LH interacts with receptors on Leydig cells, where it stimulates increased testosterone synthesis. FSH and the testosterone from the Leydig cells interact with the Sertoli cells in the seminifer- ous tubule epithelium to regulate spermatogenesis. More detailed reviews of the male reproductive hormones can be found elsewhere (Knobil et al., 1994; Yen and Jaffe, 1991). Several agents, such as lead and dibromochloropropane, affect the neuroendocrine system and spermatogenesis (for reviews, see Bonde and Giwercman, 1995; Tas et al., 1996). Whereas many studies have investigated the relationship between chemi- cals and male fertility, studies among women are sparse. Some chemicals may disrupt the female hormonal balance necessary for proper functioning. Normal menstrual-cycle functioning is also important in the risk of hormonally related diseases, such as osteopenia, breast cancer, and cardiovascular disease. Chemicals can have multiple effects on the female system, including modulation of hormone concentrations, such menstrual- or ovarian-cycle irregularities as changes in men- arche and menopause, and impairment of fertility (Bretveld et al., 2006a,b). In this chapter, we discuss studies that have focused on menstrual-cycle characteris- tics and age of menarche or age of menopause. Studies of the association between the chemicals of interest and endometriosis are reviewed in Chapter 9. Conclusions from VAO and Updates The committee responsible for Veterans and Agent Orange, hereafter re- ferred to as VAO (IOM, 1994), concluded that there was inadequate or insufï¬- cient evidence of an association between exposure to 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), TCDD, picloram, or cacodylic acid and altered sperm characteristics or infertility. Additional in- formation available to the committees responsible for Update 1996, Update 1998, Update 2000, Update 2002, and Update 2004 did not change that ï¬nding. Reviews of the relevant studies are presented in the earlier reports. Table 7-1 summarizes the studies.
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 519 TABLE 7-1 Selected Epidemiologic StudiesâFertility (altered hormone concentrations, decreased sperm counts or quality, subfertility, or infertility) Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a OCCUPATIONAL New Studies Farr et al., 2006 Age of menopause women who self-reported pesticide exposure 8,038 0.9 (0.8â1.0) Oh et al., 2005 Male fertilityâdioxin exposure with air monitoring 31 1.4* Farr et al., 2004 Menstrual cycle characteristics of premenopausal women in AHS aged 21â40 1,754 Short menstrual cycle 0.8 (0.6â1.0) Long menstrual cycle 1.4 (0.9â2.1) Irregular 0.6 (0.4â0.8) Missed Period 1.6 (1.3â2.0) Intermenstrual bleeding 1.1 (0.9â1.4) Studies Reviewed in Update 2000 Abell et al., 2000 Female greenhouse workers in Denmark (maternal exposure) 20 hours manual contact per week 220 0.7 (0.5â1.0)b Never used gloves 156 0.7 (0.5â1.0)b High exposure 202 0.6 (0.5â0.9)b Larsen et al., 1998 Danish farmers who used any potentially spermatotoxic pesticides, including 2,4-D Farmers using pesticides vs. organic farmers 523 1.0 (0.8â1.4)b Used three or more pesticides 0.9 (0.7â1.2)b Used manual sprayer for pesticides 0.8 (0.6â1.1)b Studies Reviewed in Update 1998 Heacock et al., Workers at sawmills using chlorophenates 1998 Standardized fertility ratio 18,016 0.7 (0.7â0.8)c (births) Mantel-Haenszel rate ratio estimator 18,016 0.9 (0.8-0.9)c (births) Cumulative exposure (hours) 120â1,999 7,139 0.8 (0.8â0.9)c 2,000â3,999 4,582 0.9 (0.8â1.0)c 4,000â9,999 4,145 1.0 (0.9â1.1)c 10,000 1,300 1.1 (1.0â1.2)c continued
520 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-1 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Lerda and Rizzi, Argentinean farmers exposed to 2,4-D 32 1991 Sperm count (millions/ml) exposed: 49.0 vs. control: 101.6 Motility (%) exposed: 24.8 vs. control: 70.4 Sperm death (%) exposed: 82.9 vs. control: 37.1d Anomalies (%) exposed: 72.9 vs. control: 33.4 (p 0.01 overall) ENVIRONMENTAL New Studies Eskanazi et al., Seveso cohort-serum dioxin concentrations 2005 and age of menopause 616 Premenopause 260 43.6 (0.2â0.9) Natural Menopause 169 45.8 (0.3â1.0) Surgical menopause 83 43.4 (0.3â1.0) Impending menopause 13 43.8 (0.2â0.9) Perimenopause 33 36.5 (0.2â0.9) Other 58 39.6 (0.2â0.9) Warner et al., 2004 Age of menarche at time of exposure 282 1.0 (0.8â1.1) Greenlee et al., Women from Wisconsin, US infertility 2003 (maternal exposure) Mixed or applied herbicides 21 2.3 (0.9â6.1) Used 2,4,5-T 9 9 cases (2.7%) 11controls (3.4%) Used 2,4-D 4 4 cases (1.2%) 4 controls (1.2%) Swan et al., 2003 Men from Missouri, US low sperm quality Elevated urinary metabolite marker for 2,4-D 5 0.8 (0.2â3.0) Studies Reviewed in Update 2002 Staessen et al., Adolescents in communities close to industrial 2001 sources of heavy metals, PCBs, VOCs, and PAHsâdelays in sexual maturity In Hoboken, Belgium 8 4.0 (*) In Wilrik, Belgium 15 1.7 (*)
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 521 TABLE 7-1 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a VIETNAM VETERANS Studies Reviewed in Update 1996 Henriksen et al., Effects on speciï¬c hormone levels or sperm 1996 count in Ranch Hands Low testosterone High dioxin (1992) 18 1.6 (0.9â2.7) High dioxin (1987) 3 0.7 (0.2â2.3) Low dioxin (1992) 10 0.9 (0.5â1.8) Low dioxin (1987) 10 2.3 (1.1â4.9) Background (1992) 9 0.5 (0.3â1.1) High FSH High dioxin (1992) 8 1.0 (0.5â2.1) Low dioxin (1992) 12 1.6 (0.8â3.0) Background (1992) 16 1.3 (0.7â2.4) High LH High dioxin (1992) 5 0.8 (0.3â1.9) Low dioxin (1992) 5 0.8 (0.5â3.3) Background (1992) 8 0.8 (0.4â1.8) Low sperm count High dioxin 49 0.9 (0.7â1.2) Low dioxin 43 0.8 (0.6â1.0) Background 66 0.9 (0.7â1.2) Studies Reviewed in VAO CDC, 1989b Vietnam Experience Study Lower sperm concentration 42 2.3 (1.2â4.3) Proportion of abnormal sperm 51 1.6 (0.9â2.8) Reduced sperm motility 83 1.2 (0.8â1.8) Stellman et al., American Legionnaires who served in 1988 Southeast Asia Difï¬culty having children 349 1.3 (p 0.01) Unless otherwise indicated, studies show paternal exposure. a Given when available. b For this study, relative risk has been replaced with the fecundability ratio, for which a value less than 1.0 indicates an adverse effect. c For this study, relative risk has been replaced with the standardized fertility ratio, for which a value less than 1.0 indicates an adverse effect. d Table 1 in the reference reverses these ï¬guresâcontrol: 82.9%; exposed: 37.1%âbut the text (âThe percentages of asthenospermia, mobility, necrosperma and teratospermia were greater in the exposed group than in controlsâ¦â) suggests that this is a typographic error. * Information not provided by study authors.
522 VETERANS AND AGENT ORANGE: UPDATE 2006 Update of the Epidemiologic Literature Occupational Studies After exclusion of women who were pregnant, were nursing, were taking oral contraceptives, had extreme body-mass indexes, or had missing values, Farr et al. (2004) reported on the menstrual-cycle characteristics of 3,103 premeno- pausal women in the Agricultural Health Study (AHS) who were 21â40 years old when they completed a female health and family health questionnaire. They examined the association between pesticide mixing or applying and menstrual characteristics of short cycles, long cycles, irregular cycles, missed periods, and bleeding or spotting between periods in the preceding 12 months. Women who had never mixed or applied pesticides were considered the control group. The investigators reported a signiï¬cant relationship between increased cycle length and ever mixing or applying any type of pesticide (p 0.02) and increased reports of missed periods (OR 1.6). There was a trend toward increased odds of long cycles (p 0.08) and missed periods (p 0.001) with increasing days of pesticide exposure. Although using hormonally active pesticides was found to be associ- ated with increased cycle length and increased frequency of missed cycles, the pesticides with this observed association did not include any of the chemicals of interest to the present review committee. The study used self-reported informa- tion on menstrual cycle that may have been unreliable, and no hormonal conï¬r- mation of menstrual dysfunction was available. Overall, there was no indication of an association with menstrual-cycle characteristics and the speciï¬c chemicals of interest in this review. There also has been a report from the AHS (Farr et al., 2006) concerning age at menopause in 8,038 women who were 35â55 years old at the time of enroll- ment. Women were classiï¬ed according to their self-reported pesticide exposure. Overall, women who ever mixed or applied pesticides were found to have a higher age at menopause (hazard ratio [HR] by Cox proportional hazard analysis 0.87, 95% CI 0.78â0.97) that translates into a delay of about 3 months. The estimate did not vary much when restricted to herbicides (HR 0.88, 95% CI 0.74â1.05) or to phenoxy herbicides (HR 0.85, 95% CI 0.65â1.11). One study of male fertility outcomes has been published since the last update. Oh et al. (2005) studied a group of 31 male incinerator workers and 84 controls in Seoul, South Korea. They measured dioxin exposure with air monitor- ing in the facility and found that levels were 100 times higher than those reported for the general Seoul area (31.17 ng TEQ/m3 compared with 0.32 ng TEQ/m3). Sperm characteristics were analyzed for eight controls and six workers. No sta- tistically signiï¬cant differences were observed in the number of sperm (p = 0.05) or sperm mobility (p 0.35). The fractions of sperm with DNA damage in waste- incineration workers and control subjects were measured at 1.40% 0.08% and 1.26% 0.03%, respectively (p 0.001).
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 523 Environmental Studies The committee reviewed two reports from the Seveso Womenâs Health Study (SWHS) published since the last update that focused on age at menarche and age at menopause in the Seveso population, which was exposed to high concentrations of TCDD as the result of an industrial explosion in 1976. Warner et al. (2004) examined age at menarche in 282 women who were premenarcheal at the time of the explosion. TCDD was measured in archived blood samples. Subjects had a mean age of 6.9 years at the time of the explosion. The median serum TCDD concentration was 140.3 ppt for all premenarcheal women. Serum TCDD did not vary with self-reported age at menarche in all subjects or in a group that were less than 8 years old at the time of the explosion. A major limi- tation of the study was that age at menarche was based on recall, and the time between onset of menarche and study interview ranged from 5 to 19 years. The ï¬nding of no association between age at menarche and exposure of young girls to TCDD may be related to the possibility that susceptibility is greater in utero than during childhood. The committee reviewed a second SWHS paper by Eskenazi et al. (2005) on serum dioxin concentrations and age at menopause in the Seveso cohort. The study included 616 women who were premenopausal at the time of the explo- sion and were older than 35 years at the time of the interview. The median lipid- adjusted serum TCCD concentration was 43.7 ppt and did not vary signiï¬cantly among the menopausal categories of premenstrual, natural menopause, surgical menopause, impending menopause, and perimenopause. The HRs of the serum TCDD quintiles (1.0, 1.1, 1.4, 1.6, and 1.1) suggested a trend between TCDD exposure (up to about 100 ppt) and earlier onset of natural menopause but also suggested that women with the highest serum TCDD did not have the earliest onset of menopause. Age at which the subjects of this study were exposed rep- resents an appropriate match for the experience of female Vietnam veterans. The literature suggests, however, that ovarian follicles are most susceptible to effects in the prepubertal period. A publication by Swan (2006) only reiterated the ï¬ndings in Swan et al. (2003), which were considered in Update 2004. Vietnam-Veteran Studies No new Vietnam-veteran studies concerning exposure to the compounds of interest and fertility were published since Update 2004. Biologic Plausibility There is little evidence that 2,4-D or 2,4,5-T has substantial effects on re- productive organs or fertility. One recent study has demonstrated that 2,4,5-T
524 VETERANS AND AGENT ORANGE: UPDATE 2006 competes with 17 -estradiol for binding to estrogen receptor and can function as an antiestrogen in cell culture (Lemaire et al., 2006), suggesting 2,4,5-T may have the potential to disrupt female reproductive function. In contrast with the lack of evidence on 2,4-D and 2,4,5-T, many diverse laboratory studies provide evidence that TCDD can affect reproductive organ function and reduce fertility in both men and women. TCDD exposure can re- duce fertility in male rats and is associated with histologic changes in the testis (Chahoud et al., 1989). More recent studies of TCDDâs effects on the testis have shown that it can induce signiï¬cant changes in gene expression (Kuroda et al., 2005; Lai et al., 2005a; Volz et al., 2005; Yamano et al., 2005), leading to modiï¬cation of steroidogenesis in particular (Lai et al., 2005b). Those changes are associated with disruption or complete inhibition of spermatogenesis (Fisher et al., 2005; Simanainen et al., 2004; Volz et al., 2005). Furthermore, the TCDD- induced reduction in spermatogenesis has been associated with reduced erectile function in one study (Moon et al., 2004) and reduced serum testosterone in another (Simanainen et al., 2004). In women, TCDD has been shown to reduce reproductive success, and this reduction could be mediated by alterations in the ovaries, uterus, and placenta. TCDD has been shown to disrupt ovarian steroidogenesis, impair ovulation, re- duce circulating progesterone and estradiol, and decrease fertility (Li et al., 2006; Petroff and Mizinga, 2003; Ushinohama et al., 2001). Recent studies demonstrate that TCDD at low concentrations suppresses gene expression essential to ovarian function and downregulates estrogen-dependent signaling (Hombach-Klonisch et al., 2006; Miyamoto, 2004). TCDD-induced reduction in fertility in women could also be mediated by changes in the uterus. TCDD has antiestrogenic activ- ity on the uterus, causing impairment of uterine epithelial function (Buchanan et al., 2000) that may contribute to TCDD-induced reduction in the survival of implanted embryos early in gestation (Kitajima et al., 2004). TCDD-induced reduction in reproductive success may also be mediated by altered placental function, which can lead to fetal death. TCDD alters gene expression in the pla- centa, suppresses placental vascular remodeling, and induces placental hypoxia (Ishimura et al., 2002, 2006; Mizutani et al., 2004). The biologic plausibility of reproductive effects in general arising from ex- posure to the chemicals of interest is discussed at the end of this chapter. Synthesis Although there is much evidence of the biologic plausibility of disruption of male and female fertility by exposure to the chemicals of interest, there continues to be a lack of substantive epidemiologic data that demonstrate any association in human populations.
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 525 Conclusions On the basis of its evaluation of the evidence reviewed here and in previous VAO reports, the committee concludes that there is inadequate or insufï¬cient evidence of an association between exposure to the compounds of interest and altered hormone concentrations, menstrual-cycle abnormalities, decreased sperm counts or sperm quality, subfertility, or infertility. SPONTANEOUS ABORTION Spontaneous abortion is the expulsion of a nonviable fetus, generally before 20 weeks of gestation, that is not induced by physical or pharmacologic means. The background risk of recognized spontaneous abortion is generally 7â15 per- cent (Hertz-Picciotto and Samuels, 1988), but it is established that many more pregnancies terminate before women become aware of them (Wilcox et al., 1988)âthese terminations are known as subclinical pregnancy losses and gener- ally are not included in studies of spontaneous abortion. Estimates of the risk of recognized spontaneous abortion vary with the design and method of analysis. Study designs include cohorts of women asked retrospectively about pregnancy history, cohorts of pregnant women (usually those receiving prenatal care), and cohorts of women who are monitored for future pregnancies. Retrospective re- ports can be limited by memory loss, particularly of spontaneous abortions that took place a long time before. Studies that enroll women who appear for prenatal care require the use of life tables and specialized statistical techniques to account for differences in the times at which women seek medical care during pregnancy. Enrollment of women before pregnancy provides the theoretically most valid estimate of risk, but it can attract non-representative study groups because pro- tocols are demanding. Conclusions from VAO and Updates The committee responsible for VAO concluded that there was inadequate or insufï¬cient evidence of an association between exposure to 2,4-D, 2,4,5-T, TCDD, picloram, or cacodylic acid and spontaneous abortion. Additional infor- mation available to the committees responsible for Update 1996, Update 1998, and Update 2000 did not change that conclusion. Information available to the committee responsible for Update 2002, however, led to the conclusion that there was suggestive evidence that paternal exposure to TCDD is not associated with the risk of spontaneous abortion, but that there was insufï¬cient information to determine whether an association exists between maternal exposure to TCDD and the risk of spontaneous abortion or between maternal or paternal exposure to 2,4-D, 2,4,5-T, picloram, or cacodylic acid and the risk of spontaneous abortion. The additional information reviewed by the committee responsible for Update
526 VETERANS AND AGENT ORANGE: UPDATE 2006 2004 did not change this conclusion. The relevant studies are reviewed in the earlier reports. Table 7-2 summarizes them. Update of the Epidemiologic Literature Environmental Studies Tango et al. (2004) studied the distribution of several birth outcomes around Japanese municipal-waste incinerators with elevated dioxin emissions. They found fetal death after the 12th week of gestation (with or without congenital malformations) was not associated with the distance the mother lived from an incinerator at the time of birth or whether her residence was in the area known to have the highest dioxin soil concentrations. No new occupational or Vietnam-veteran studies concerning exposure to the compounds of interest and spontaneous abortion were published since Update 2004. Biologic Plausibility Laboratory animal studies demonstrate that TCDD exposure during preg- nancy can alter circulating steroid hormone concentrations (Simanainen et al., 2004) and disrupt placental development and function (Ishimura et al., 2006; Mizutani et al., 2004) and thus contribute to a reduction in survival of implanted embryos and to fetal death (Kitajima et al., 2004). However, the reproductive signiï¬cance of those effects and the risk of recognized pregnancy loss before 20 weeks of gestation in humans are not clear. There is no evidence of a relationship between paternal or maternal exposure to TCDD and spontaneous abortion. Ex- posure to 2,4-D or 2,4,5-T causes fetal toxicity and death after maternal exposure in experimental animals. However, that effect occurs only at high doses and in the presence of maternal toxicity. No fetal toxicity or death has been reported to occur after paternal exposure to 2,4-D. The biologic plausibility of reproductive effects in general arising from ex- posure to the chemicals of interest is discussed at the end of this chapter. Synthesis No additional information was available to the committee responsible for Update 2006 to motivate changing the assessment of the last two committees. Given the age of the Vietnam-veteran cohort, it is highly unlikely that additional information on this outcome among the population will appear.
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 527 TABLE 7-2 Selected Epidemiologic StudiesâSpontaneous Abortion Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a OCCUPATIONAL Studies Reviewed in Update 2002 Schnorr et al., Wives and partners of men in the NIOSH 2001 cohort Estimated paternal TCDD serum level at the time of conception 20 ppt 29 0.8 (0.5â1.2) 20 to 255 ppt 11 0.8 (0.4â1.6) 255 to 1,120 11 0.7 (0.3â1.6) 1,120 ppt 8 1.0 (0.4â2.2) Studies Reviewed in Update 2000 Driscoll et al., Women employed by the US Forest 1998 Serviceâmiscarriages (maternal exposure) 141 2.0 (1.1â3.5) Studies Reviewed in VAO Moses et al., 1984 Follow-up of 2,4,5-T production workers 14 0.9 (0.4â1.8) Suskind and Hertzberg, 1984 Follow-up of 2,4,5-T production workers 69 0.9 (0.6â1.2) Smith et al., Follow-up of 2,4,5-T sprayers vs 1982 non-sprayers 43 0.9 (0.6â1.3)** Townsend et al., Wives of men employed involved in 1982 chlorophenol processing at Dow Chemical Company 85 1.0 (0.8â1.4) Carmelli et al., Wives of men occupationally exposed to 1981 2,4-D All reported work exposure to herbicides (high and medium) 63 0.8 (0.6â1.1)** Farm exposure 32 0.7 (0.4â1.5) Forest and commercial exposure 31 0.9 (0.6â1.4) Exposure during conception period Farm exposure 15 1.0 (0.5â1.8) Forest and commercial exposure 16 1.6 (0.9â1.8) All exposures, father aged 18â25 years Forest and commercial exposure 8 3.1 (1.2â7.8) Exposure during conception period Father aged 31â35 years, farm exposure 10 2.9 (0.8â10.9) ENVIRONMENTAL New Studies Eskenazi et al., Seveso (Italy) Womenâs Health Study 2003 participants living in exposure Zones A and B in 1976 (maternal exposure) Pregnancies 1976â1998 97 0.8 (0.6â1.2) Pregnancies 1976â1984 44 1.0 (0.6â1.6) continued
528 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-2 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Studies Reviewed in Update 2002 Arbuckle et al., Ontario farm families (maternal and paternal 2001 exposure) Phenoxyacetic acid herbicide exposure in the preconception period & spontaneous abortion risk 48 1.5 (1.1â2.1) Revich et al., Residents of the Samara Region, Russia 2001 (maternal and paternal exposure) Chapaevsk 24.4% (20.0â29.5%)b Samara * 15.2% (14.3â16.1%)b Toliatti * 10.6% (9.8â11.5%)b Syzran * 15.6% (13.4â18.1%)b Novokuibyshevsk * 16.9% (14.0â20.3%)b Other small towns * 11.3% (9.4â13.8%)b Tuyet and Vietnamese women who were or whose (*) [anecdotal reports Johansson, husbands were exposed to herbicides sprayed of miscarriage in 2001 during the Vietnam war * pilot study] Studies Reviewed in Update 2000 Axmon et al., Wives of Swedish ï¬shermen 2000 Before gestation week 12 0.5 (0.3â1.0) East coast residents 12 West coast residents 54 Petrelli et al., Wives of pesticide appliers 26 3.8 (1.2â12.0) 2000 VIETNAM VETERANS Studies Reviewed in Update 2002 Kang et al., Female Vietnam-era veterans (maternal 2000 exposure) 1.0 (0.82â1.21) Vietnam veterans (1,665 pregnancies) 278 (*) Vietnam-era veterans who did not serve in Vietnam (1,912 pregnancies) 317 (*) Studies Reviewed in Update 2000 Schwartz, 1998 Female Vietnam veterans (maternal exposure) Women who served in Vietnam 113 (*) Women who did not serve in the war zone 124 (*) Civilian women 86 (*) Studies Reviewed in Update 1996 Wolfe et al., Air Force Ranch Hand veterans 157 1995 Background 57 1.1 (0.8â1.5) Low-level exposure 56 1.3 (1.0â1.7) High-level exposure 44 1.0 (0.7â1.3) Studies Reviewed in VAO Aschengrau and Wives of Vietnam veterans presenting at Monson, 1989 Boston Hospital for Women 27 weeks gestation 10 0.9 (0.4â1.9) 13 weeks gestation * 1.2 (0.6â2.8)
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 529 TABLE 7-2 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a CDC, 1989b Vietnam Experience Study Overall 1,566 1.3 (1.2â1.4) Self-reported low exposure 489 1.2 (1.0â1.4) Self-reported medium exposure 406 1.4 (1.2â1.6) Self-reported high exposure 113 1.7 (1.3â2.1) Field and Kerr, Follow-up of Australian Vietnam veterans 199 1.6 (1.3â2.0) 1988 Stellman et al., American Legionnaires with service 1988 1961â1975 Vietnam-era veterans vs Vietnam veterans All Vietnam veterans 231 1.4 (1.1â1.6) Low exposure 72 1.3 (1.0â1.7) Medium exposure 53 1.5 (1.1â2.1) High exposure 58 1.7 (1.2â2.4) Vietnam-era veterans vs herbicide handlers 9 1.6 (0.7â3.3) Vietnam veterans with medium or high exposure vs Vietnam veterans with low exposure Medium exposure 53 1.2 (0.8â1.7) High exposure 58 1.4 (0.9â1.9) ABBREVIATIONS: CDC, Centers for Disease Control and Prevention; CI, conï¬dence interval; NIOSH, National Institute for Occupational Safety and Health. Unless otherwise indicated, studies show paternal exposure. a Given when available. b Spontaneous abortion rate per 100 full-term pregnancies for the years 1991â1997. * Information not provided by study authors. ** 90% conï¬dence interval. Conclusions The present committee concurs with the overall conclusion of the previ- ous committees that paternal exposure to TCDD is not associated with risk of spontaneous abortion and that insufï¬cient information is available to determine whether an association exists between the risk of spontaneous abortion and mater- nal exposure to TCDD or either maternal or paternal exposure to 2,4-D, 2,4,5-T, picloram, or cacodylic acid. STILLBIRTH, NEONATAL DEATH, AND INFANT DEATH Stillbirth or late fetal death typically refers to the delivery at or after 20 weeks of gestation of a fetus that shows no signs of life, including fetuses that
530 VETERANS AND AGENT ORANGE: UPDATE 2006 weigh more than 500 g regardless of gestational age (Kline et al., 1989). Neonatal death refers to the death of a liveborn infant within 28 days of birth, while infant death includes deaths occurring before the ï¬rst birthday. Because the causes of stillbirth and early neonatal death overlap considerably, they are commonly analyzed together in a category referred to as perinatal mor- tality (Kallen, 1988). Stillbirths make up less than 1 percent of all births (CDC, 2000). The most common causes of perinatal mortality (Kallen, 1988) among low-birth-weight (500 g to 2,500 g) liveborn and stillborn infants are placental and delivery complicationsâabruptio placenta, placenta previa, malpresentation, and umbilical-cord complications. Among infants weighing more than 2,500 g at birth, the most common causes of perinatal death are complications of the cord, placenta, and membranes and congenital malformations (Kallen, 1988). Conclusions from VAO and Updates The committee responsible for VAO concluded that there was inadequate or insufï¬cient evidence of an association between exposure to 2,4-D, 2,4,5-T, TCDD, picloram, or cacodylic acid and stillbirth, neonatal death, or infant death. Additional information available to the committees responsible for Update 1996, Update 1998, Update 2000, Update 2002, and Update 2004 did not change that conclusion. Reviews of the relevant studies are presented in the earlier reports. Update of the Epidemiologic Literature The only relevant study published since the last update was a large study of multiple pregnancy outcomes in Japan (Tango et al., 2004). The study found no associations between proximity of the motherâs residence at the time of birth (deï¬ned in terms of 1-km bands around the incinerators) and rates infant deaths ( 1 yr, 1 mo, 1 wk with and without congenital malformations). Analyses were conducted for a âpeak-declineâ relationship with âpeakâ dioxin soil concentra- tions (known to occur about 2 km from an incinerator); only death within the ï¬rst year of life, overall (p 0.023) or with congenital malformations (p 0.047), showed a signiï¬cant result. This study had several methodologic weaknesses, including the lack of individual-level information on other risk factors and of in- dividual exposure data. In addition, the birth-outcome effects might be associated with socioeconomic differences in residents in the different zones. No new occupational or Vietnam-veteran studies concerning exposure to the compounds of interest and stillbirth, neonatal death, or infant death were published since Update 2004. Biologic Plausibility Laboratory studies of maternal TCDD exposure during pregnancy have demonstrated the induction of fetal death; neonatal death, however, is only rarely
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 531 observed and is usually the result of cleft palate leading to an inability to nurse. Studies addressing the potential for perinatal death as a result of paternal expo- sure to TCDD or herbicides are too limited to support conclusions. The biologic plausibility of reproductive effects in general arising from ex- posure to the chemicals of interest is discussed at the end of this chapter. Synthesis The study reviewed for this update did not ï¬nd signiï¬cant associations be- tween the relevant exposures and rates of infant or fetal deaths. The study was limited in that exposure was based on environmental concentrations of dioxin and individual exposure data were not obtained. Furthermore, several risk factors that could confound associations between exposure and outcome were not assessed. Conclusions On the basis of its evaluation of the evidence reviewed here and in previous VAO reports, the committee concludes that there is inadequate or insufï¬cient evidence of an association between exposure to the compounds of interest and stillbirth, neonatal death, or infant death. BIRTH WEIGHT AND PRETERM DELIVERY The World Health Organization recommends 2,500 g as the threshold for designation of low birth weight (Alberman, 1984). Low infant weight at birth is among the important predictors of neonatal death and morbidity in the United States. The concept of low birth weight actually encompasses two causal path- ways, often treated as a single entity: low birth weight secondary to intrauterine growth retardation (IUGR), in which case a fetus or baby is referred to as small for gestational age, and low birth weight secondary to preterm delivery (PTD), which can have other long-term consequences. The concept of IUGR represents birth weight adjusted for gestational age. The current deï¬nition of PTD is de- livery at less than 259 days, or 37 completed weeks, of gestation, calculated on the basis of the date of the ï¬rst day of the last menstrual period (Bryce, 1991). About 7 percent of live births are low birth weight. The incidence of IUGR is much more difï¬cult to quantify because there are no standards for distribut- ing birth weight by gestational age. When no distinction is made between the causes of low birth weight (IUGR or PTD), the factors most strongly associated with it are maternal tobacco use during pregnancy, multiple births, and race or ethnicity. Other potential risk factors are socioeconomic status (SES), maternal weight, birth order, maternal complications during pregnancy (such as severe pre- eclampsia) and obstetric history, job stress, and cocaine or caffeine use during pregnancy (Kallen, 1988). Established risk factors for PTD include race (black), marital status (single), low SES, previous low birth weight or PTD, multiple ges-
532 VETERANS AND AGENT ORANGE: UPDATE 2006 tations, tobacco use, and cervical, uterine, or placental abnormalities (Berkowitz and Papiernik, 1993). Conclusions from VAO and Updates The committee responsible for VAO concluded that there was inadequate or insufï¬cient evidence of an association between exposure to the compounds of interest and low birth weight or preterm delivery. Additional information avail- able to the committees responsible for Update 1996, Update 1998, Update 2000, Update 2002, and Update 2004 did not change that conclusion. Reviews of the relevant studies are presented in the earlier reports. Update of the Epidemiologic Literature Update 2000, Update 2002, and Update 2004 discussed change in the sex ratio at birth from the usual ratio of male-to-female newborns, which is approxi- mately 1.02 as a possible adverse reproductive outcome. If the herbicides used in Vietnam did alter this endpoint in humans, it would not represent an outcome for which an individual veteran could be compensated. Such an alteration would, however, be indicative of some impact on reproductive function, as mentioned in the section on biologic plausibility at the end of this chapter. Of the new articles reporting reproductive effects, two environmental studies that presented ï¬ndings on birth weight and gestational age also assessed possible changes in the sex ratio at birth; their ï¬ndings for this additional endpoint are included below. Occupational Studies Lawson et al. (2004) conducted a follow-on study of PTD and birth weight among the children of men in the National Institute for Occupational Safety and Health (NIOSH) cohort in comparison to age- and race-matched neighborhood referents. The analysis of birth weight considered 1,117 singleton, full-term births (at least 37 weeks gestation) to 217 referent wives (604 referent births) and 176 worker wives (513 births; 221 pre-exposure and 292 exposed). The suggestion of an increase in birth weight with high TCDD concentrations vanished when adjustment was made for motherâs education, parity, smoking during pregnancy, length of gestation, and the infantâs sex. In order to consider the possibility that the mothers might have experienced direct exposure from materials carried home by the fathers when they were engaged in their NIOSH employment during a pregnancy, another analysis was restricted to such pregnancies for the cohort members and pregnancies occurring while the NIOSH plants were in operation for the referents, or 334 referent births and 98 exposed births. In this case, the adjusted analysis found a statistically signiï¬cant increase of 31 g in birth weight with each log increase in TCDD concentration (p 0.03).
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 533 Lawson et al. (2004) also conducted an analysis of preterm births based on a total of 1,153 births: 618 referent, 238 pre-exposed, and 297 during-exposure. With or without adjustment for age of mother and for the occurrence of an ac- cident, smoking, or medication use during pregnancy, no increase in risk of pre- maturity was observed (odds ratio [OR] 0.8, 95% CI 0.6â1.1). Environmental Studies Another study of the effect of dioxins on reproductive outcomes was con- ducted in an area surrounding a municipal-waste incinerator located 10 km from Taipei that began operating in 1992 (Lin et al., 2006). The incineratorâs average emission concentration of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) for 1997 (6.47 ng TEQ/m3) was used as the input for a dispersion model (EPAâs Industrial Source Complex ModelâShort Term Model), which was used to estimate annual averages for each administrative district of Taipei. A cut point of 0.03 pg TEQ/m3, between the background concentrations for rural (0.02 pg TEQ/m3) and urban air (0.05 pg TEQ/m3), was selected. The dispersion model predicted concentrations greater than 0.03 pg TEQ/m3 for 40 districts; those were retained as the âexposedâ districts, which were further subdivided at 0.05 pg TEQ/m3. Among the districts with predicted concentrations of zero, 40 âreferenceâ districts were randomly selected; those were all in non-industrial areas at least 12 km from the incinerator. From Taiwanâs birth registry, Lin et al. (2006) assembled newborn gender, birth order, gestational age, and birth weight, plus maternal age, address, and educational level, for all singleton live births in the study districts for 1991 and 1997. After excluding those with birth weights less than 500 g, with gestational ages less than 20 weeks, or born to unwed women, 6,697 and 6,282 infants born in 1991 and 1997, respectively, were available for analysis. Analyses were adjusted for maternal age and education, sex of newborn, and birth order; information on other potential confounders (such as maternal smoking) was not available. Categorical comparisons of the predicted high and low exposed districts with the reference districts found no associations for low birth weight (under 2,500 g), PTD (less than 37 weeks of gestation), or sex ratio for 1991 (as might have been expected before exposure began) or for 1997 (after the incinerator had been in operation for 5 years). Regression on the continuous variables determined the difference between the exposed and reference districts in 1991 and in 1997 for mean birth weight or mean gestational age. In 1997, the adjusted decrease in birth weight of 5.87 g for the exposed districts was non- signiï¬cant, but the relative reduction in gestational age of 0.09 week was sig- niï¬cant (p 0.05). (For 1991, both these endpoints had non-signiï¬cantly higher values in the ânot yetâ exposed districts than in the reference districts; therefore, a comprehensive before-and-after analysis might have revealed stronger effects in the districts that ultimately were exposed.) In light of no suggestion of an
534 VETERANS AND AGENT ORANGE: UPDATE 2006 increased risk of PTD, the small reported reduction in gestational age is of ques- tionable biological signiï¬cance. The studyâs large size gave it considerable sensitivity. On the whole it used appropriate methods, but apparently an analysis was not conducted to make full use of the 1991 data as self-control information to determined the relative changes between the exposed and reference districts from 1991 to 1997. The exposure assessment is only as reliable as the estimates generated by EPAâs dis- persion model for the districts of Taipei, so exposure misclassiï¬cation could have contributed to biasing the generally negative results toward the null. In their study of birth outcomes for mothers living within about 10 km of Japanese municipal-waste incinerators emitting high concentrations of dioxin, Tango et al. (2004) found no relationship between proximity of the motherâs residence to the incinerator for low or extremely low (under 1,500 g) birth weight. Analyses for association with peak dioxin concentrations in soil (known to occur about 2 km from the incinerators) were also negative. The sex ratio at birth, evaluated in terms of deviation in the proportion of female live births from the national proportion, was also found not to be related to the distance of the maternal residence from an incinerator. Vietnam-Veteran Studies No new Vietnam veteran studies concerning exposure to the compounds of interest and birth weight or preterm delivery were published since Update 2004. Biologic Plausibility Laboratory studies of the potential male-mediated developmental toxicity of TCDD and herbicides as a result of exposure of adult male animals are too lim- ited to permit conclusions. TCDD and herbicides are known to cross the placenta leading to direct exposure of the fetus. Data from studies in experimental animals also suggest that the pre-implantation embryo and developing fetus are sensitive to the toxic effects of 2,4-D and TCDD after maternal exposure. However, the signiï¬cance of those animal effects for humans is not clear. The biologic plausibility of reproductive effects in general arising from ex- posure to the chemicals of interest is discussed at the end of this chapter. Synthesis The three studies reviewed here (two environmental and one based on an occupational cohort) did not ï¬nd an association between exposure to the com- pounds of interest and the risk of low birth weight or prematurity. The two new weakly signiï¬cant ï¬ndings may simply be spurious results arising among many
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 535 comparisons; a modest increase in average birth weight would not be construed as an adverse effect, and the small decrease in average gestation is of questionable biologic importance. Although the results overall suggest a lack of an association, they should be interpreted with caution because of some methodologic limita- tions, such as a long recall period in the cohort study and exposure misclassiï¬ca- tion in the environmental studies. Conclusions On the basis of its evaluation of the evidence reviewed here and in previous VAO reports, the committee concludes that there is inadequate or insufï¬cient evidence of an association between exposure to the compounds of interest and low birth weight or preterm delivery. BIRTH DEFECTS The March of Dimes deï¬nes a birth defect as âan abnormality of structure, function or metabolism, whether genetically determined or as the result of an en- vironmental inï¬uence during embryonic or fetal lifeâ (Bloom, 1981). Other terms, often used interchangeably, are congenital anomaly and congenital malformation. Major birth defects, which occur in 2â3 percent of live births, are abnormalities that are present at birth and are severe enough to interfere with viability or physi- cal well-being. Birth defects are detected in another 5 percent of babies through the ï¬rst year of life. The causes of most birth defects are unknown. Genetic fac- tors, exposure to some medications, exposure to environmental contaminants, oc- cupational exposures, and lifestyle factors have been implicated in the etiology of birth defects (Kalter and Warkany, 1983). Most etiologic research has focused on the effects of maternal and fetal exposures, but some work has addressed paternal exposures. Paternally mediated exposures might occur by several routes and exert effects in various ways. One way is through direct genetic damage to the male germ cell transmitted to the offspring and dominantly expressed as a birth defect. A hypothesized route is the transfer of toxic compounds through a manâs body into his seminal ï¬uid, resulting in intermittent fetal exposures throughout gesta- tion (Chia and Shi, 2002). Another, even more indirect route of paternally medi- ated exposure could be contact of family members with contamination brought into the home from the workplace, but this would not be applicable to offspring of Vietnam veterans conceived during the post-deployment period. Conclusions from VAO and Updates The committee responsible for VAO determined that there was inadequate or insufï¬cient evidence of an association between exposure to 2,4-D, 2,4,5-T or its contaminant TCDD, picloram, or cacodylic acid and birth defects among
536 VETERANS AND AGENT ORANGE: UPDATE 2006 offspring. Additional information available to the committee responsible for Update 1996 led it to conclude that there was limited or suggestive evidence of an association between at least one of the compounds of interest and spina biï¬da in the children of veterans; there was no change in the conclusions regarding other birth defects. The committee for Update 2002, which reviewed the study of female Vietnam veterans (Kang et al., 2000) reporting signiï¬cant increases in the occurrence of birth defects among their offspring, did not ï¬nd those results adequate to modify prior conclusions. Later VAO committees have not encoun- tered additional data to merit changing the conclusion that the evidence is inad- equate to support an association between exposure to the chemicals of interest and birth defects (aside from spina biï¬da) among the offspring of either male or female veterans. Summaries of the results from studies of birth defects and speciï¬cally neural- tube defects that were reviewed here and in earlier reports can be found in the Tables 7-3 and 7-4, respectively. Update of the Epidemiologic Literature Occupational Studies Lawson et al. (2004) reported on the pregnancy outcomes among wives of men from the NIOSH cohort of workers who were highly exposed to chemicals contaminated with TCDD compared to those of matched neighborhood referents. The analysis of birth defects was performed on 1,153 liveborn and 13 stillborn infants. Because only a modest number of birth defects were reported, pre- exposed and referent pregnancies were combined when they were qualitatively contrasted to the approximately 300 exposed pregnancies. Conï¬rmation from vital and medical records was attempted, but was successful for only about 50 percent of the reported birth defects. The reported categories included 6 major central nervous system defects (4 among the exposed pregnancies), plus clubfoot, hip and lower limb defects, genitourinary defects, cardiovascular defects, and cleft lip or palate. Although the numbers were too small for reliable conclusions and only descriptive data were presented, there did not appear to be any trend of increased incidence of birth defects among offspring of exposed workers. Environmental Studies The only relevant environmental study published since Update 2004 was Tango et al. (2004). The study found no associations between proximity to the incinerators and rates of fetal loss or infant deaths (before 1 week, 1 month, or 1 year) due to congenital malformation. In analyses using a âpeak-declineâ ap- proach based on soil concentrations that had been shown to peak approximately 2 km from the incinerators, a signiï¬cant outcome was found for infant death in
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 537 TABLE 7-3 Selected Epidemiologic StudiesâBirth Defects in the Offspring of Subjects Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a OCCUPATIONAL New Studies Lawson et al., Wives of workers with serum TCDD levels in 2004 NIOSH cohort 14 * (*) Studies Reviewed in Update 1998 Kristensen et al., Norwegian farmers (maternal and paternal 1997 exposure) 4,189 1.0 (1.0â1.1)b Dimich-Ward Sawmill workers et al., 1996 Cataracts 11c 5.7 (1.4â22.6) Genital organs 105c 1.3 (0.9â1.5) Garry et al., 1996 Private pesticide appliers Circulatoryârespiratory 17 1.7 (1.0â2.8) Gastrointestinal 6 1.7 (0.8â3.8) Urogenital 20 1.7 (1.1â2.6) Musculoskeletalâintegumental 30 Maternal age 30 11 0.9 (0.5â1.7) Maternal age 30 19 2.5 (1.6â4.0) Chromosomal 8 1.1 (0.5â2.1) Other 48 Maternal age 35 36 1.1 (0.8â1.6) Maternal age 35 12 3.0 (1.6â5.3) All births with anomalies 125 1.4 (1.2â1.7) Studies Reviewed in VAO Moses et al., 1984 Follow-up of 2,4,5-T male production workers 11 1.3 (0.5â3.4) Suskind and Follow-up of 2,4,5-T male production workers 18 1.1 (0.5â2.2) Hertzberg, 1984 Smith et al., 1982 Follow-up of 2,4,5-T sprayersâsprayers vs non-sprayers 13 1.2 (0.6â2.5)e Townsend et al., Follow-up of Dow Chemical plant workers 30 0.9 (0.5â1.4) 1982 ENVIRONMENTAL New Studies Cordier et al., Residents of the RhÃ´ne-Alpes region of France 2004 living near municipal solid waste incinerators (maternal and paternal exposure) Minor anomalies 518 0.9 (0.8â1.1) Chromosomal anomalies 204 1.0 (0.9â1.2) Monogenic anomalies 83 1.1 (0.8â1.4) Unknown or multifactoral etiology 964 1.1 (1.0â1.2) Tango et al., 2004 Investigated multiple pregnancy outcomes in Japanâinfant deaths from congenital defects 42 (*) NS continued
538 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-3 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Schreinemachers, Rural or farm residents of Minnesota, Montana, 2003 North and South Dakota (maternal and paternal exposure) Any birth anomaly 213 1.1 (0.9â1.3) Central nervous system anomalies 12 0.8 (0.5â1.4) Circulatory or respiratory anomalies 39 1.7 (1.1â2.6) Digestive system anomalies 24 0.9 (0.6â1.5) Urogenital anomalies 44 1.0 (0.7â1.5) Musculoskeletal or integumental anomalies 70 1.5 (1.1â2.1) Chromosomal anomalies 17 0.9 (0.6â1.6) Studies Reviewed in Update 2002 Loffredo et al., Mothers in the BaltimoreâWashington Infant 2001 Study exposed to herbicides during the ï¬rst trimester (maternal exposure) 8 2.8 (1.2â6.9) Revich et al., Residents of Chapaevsk, Russiaâcongenital 2001 malformations * (*) NS ten Tusscher Infants born in Zeeburg, Amsterdam clinics et al., 2000 1963â1965 with orofacial cleft (maternal exposure) Births in 1963 5 (*) SS Births in 1964 7 (*) SS Studies Reviewed in Update 2000 GarcÃa et al., 1998 Residents of agricultural areas in Spainâ median score on chlorophenoxy herbicides exposure duration (months) index 14 3.1(0.6â16.9) Studies Reviewed in VAO Fitzgerald et al., Persons exposed to an electrical transformer 1989 ï¬reâtotal birth defects (maternal or paternal exposure) 1 2.1 (0.05â11.85) Mastroiacovo Seveso residents (maternal, paternal, and in et al., 1988 utero exposure) Zones A and B total defects 27 1.2 (0.9â1.6)e Zones A, B, R total defects 137 1.0 (0.8â1.1)e Zones A and B mild defects 14 1.4 (0.9â2.2)e Stockbauer et al., Persons in Missouri with documented TCDD 1988 soil contamination near residence (maternal; paternal; in utero exposure) Total birth defects 17 0.8 (0.4â1.5) Major defects 15 0.8 (0.4â1.7) Midline defects 4 0.7 (0.2â2.3)
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 539 TABLE 7-3 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Hanify et al., Residents of areas of Northland New Zealand 1981 subject to aerial 2,4,5-T sprayingd All birth malformations 164 1.7 (1.4â2.1)e All heart malformations 20 3.9 (2.1â7.4)e Hypospadias, epispadias 18 5.6 (2.7â11.7)e Talipes 52 1.7 (1.2â2.3)e Cleft lip 6 0.6 (0.3â1.3)e Isolated cleft palate 7 1.4 (0.6â3.2)e VIETNAM VETERANS Studies Reviewed in Update 2002 Kang et al., 2000 Female Vietnam-era veteransâdeployed vs non- deployed (maternal exposure) âLikelyâ birth defects * 1.7 (1.2â2.2) âModerate-to-severeâ birth defects * 1.5 (1.1â2.0) Studies Reviewed in Update 2000 AIHW, 1999 Australian Vietnam veteransâValidation Study Down syndrome 67 92 expected (73â111) Tracheo-oesophageal ï¬stula 10 23 expected (14â32) Anencephaly 13 16 expected (8â24) Cleft lip or palate 94 64 expected (48â80) Absent external body part 22 34 expected (23â45) Extra body part 74 74 expected (*) Michalek et al., Air Force Ranch Hand veterans 1998a Before service in Southeast Asia * 0.7 (*) After service in Southeast Asia * 1.5 (*) Studies Reviewed in Update 1996 Wolfe et al., 1995 High exposure Ranch Hands relative to comparisons All anomalies 57 1.0 (0.8â1.3) Nervous system 3 (*) Eye 3 1.6 (0.4â6.0) Ear, face, and neck 5 1.7 (0.6â4.7) Circulatory system and heart 4 0.9 (0.3â2.7) Respiratory system 2 (*) Digestive system 5 0.8 (0.3â2.0) Genital system 6 1.2 (0.5â3.0) Urinary system 7 2.1 (0.8â5.4) Musculoskeletal 31 0.9 (0.6â1.2) Skin 3 0.5 (0.2â1.7) Chromosomal anomalies 1 (*) continued
540 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-3 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Studies Reviewed in VAO AFHS, 1992 Air Force Operation Ranch Hand veteransâ birth defects in conceptions following service in Southeast Asia Congenital anomalies 229 1.3 (1.1â1.6) Nervous system 5 1.9 (0.5â7.2) Respiratory system 5 2.6 (0.6â10.7) Circulatory system or heart 19 1.4 (0.7â2.6) Urinary system 21 2.5 (1.3â5.0) Chromosomal 6 1.8 (0.6â6.1) Other 5 2.6 (0.6â10.7) Aschengrau and Vietnam veterans whose children were born at Monson, 1990 Boston Hospital for Women All congenital anomalies (crude OR) Vietnam veterans vs men without known military service 55 1.3 (0.9â1.9) Vietnam veterans vs non-Vietnam veterans 55 1.2 (0.8â1.9) One or more major malformations (crude OR) Vietnam veterans vs men without known military service 18 1.8 (1.0â3.1) Vietnam veterans vs non-Vietnam veterans 18 1.3 (0.7â2.4) CDC, 1989a Vietnam Experience Studyâinterview data Total anomalies 826 1.3 (1.2â1.4) Nervous system defects 33 2.3 (1.2â4.5) Ear, face, neck defects 37 1.6 (0.9â2.8) Integument 41 2.2 (1.2â4.0) Musculoskeletal defects 426 1.2 (1.1â1.5) Hydrocephalus 11 5.1 (1.1â23.1) Spina biï¬da 9 1.7 (0.6â5.0) Hypospadias 10 3.1 (0.9â11.3) Multiple defects 71 1.6 (1.1â2.5) Children of veterans reporting high exposure 46 1.7 (1.2â2.4) CDC, 1989b General Birth Defects Studyâhospital records Birth defects 130 1.0 (0.8â1.3) Major birth defects 51 1.2 (0.8â1.9) Digestive system defects 18 2.0 (0.9â4.6) Birth defectsâblack Vietnam veterans only 21 3.4 (1.5â7.6) Donovan et al., Australian Vietnam veterans 1984 Vietnam veterans vs all other men 127 1.02 (0.8â1.3) National Service veteransâVietnam service vs no Vietnam service 69 1.3 (0.9â2.0)
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 541 TABLE 7-3 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Erikson et al., Vietnam veterans identiï¬ed through the CDC 1984a Metropolitan Atlanta Congenital Defects Program Any major birth defects 428 1.0 (0.8â1.1) Multiple birth defects with reported exposure 25 1.1 (0.7â1.7) EOI-5: spina biï¬da 1 2.7 (1.2â6.2) EOI-5: cleft lip with or without cleft palate 5 2.2 (1.0â4.9) ABBREVIATIONS: AFHS, Air Force Health Study; AIHW, Australian Institute of Health and Wel- fare; CDC, Centers for Disease Control and Prevention; CI, conï¬dence interval; EOI, exposure op- portunity index; NS, not signiï¬cant; SIR, standardized incidence ratio; SS, statistically signiï¬cant. Unless otherwise indicated, studies show paternal exposure. a Given when available. b 95% conï¬dence intervals contained one for all outcomes. Anencephaly and spina biï¬da included in this calculation. c Number of workers with maximal index of exposure (upper three quartiles) for any job held up to 3 months prior to conception. d Excludes stillbirths, neonatal death, or dislocated or dislocatable hip. e 90% conï¬dence interval. * Information not provided by study authors. TABLE 7-4 Selected Epidemiologic StudiesâNeural Tube Defects in the Offspring of Subjects Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a OCCUPATIONAL Studies Reviewed in Update 1998 Blatter et al., Dutch farmers 1997 Spina biï¬daâmoderate/heavy exposure Pesticide use 8 1.7 (0.7â4.0) Herbicide use 7 1.6 (0.6â4.0)g Kristensen Norwegian farmersâspina biï¬da (maternal and paternal et al., 1997 exposure) Tractor spraying equipment 28 1.6 (0.9â2.7) Tractor spraying equipment and orchards or greenhouses 5 2.8 (1.1â7.1) Dimich- Sawmill workers Ward et al., Spina biï¬da or anencephaly 22b 2.4 (1.1â5.3) 1996 Spina biï¬da only 18b 1.8 (0.8â4.1) Garry et al., Private pesticide appliersâcentral nervous system 1996 defects 6 1.1 (0.5â2.4) continued
542 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-4 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a ENVIRONMENTALc New Studies Cordier Residents of the RhÃ´ne-Alpes region of France et al., 2004 (maternal and paternal exposure) 49 0.9 (0.6â1.2) Studies Reviewed in VAO Stockbauer Persons in Missouri with documented TCDD soil et al., 1988 contaminationâcentral nervous system defects (maternal; paternal; in utero exposure) 3 3.0 (0.3â35.9) Hanify et al., Spraying of 2,4,5-T in New Zealand (all exposures) 1981 Anencephaly 10 1.4 (0.7â2.9)d Spina biï¬da 13 1.1 (0.6â2.1)d VIETNAM VETERANS Studies Reviewed in Update 2000 AIHW, 1999 Australian Vietnam veteransâValidation Study Spina biï¬daâmaxima 50 33 expected (22â44) Anencephaly 13 16 expected (8â24) Studies Reviewed in Update 1996 Wolfe et al., Air Force Operation Ranch Hand personnelâneural 1995 tube defectse 4 (*) Studies Reviewed in VAO CDC, 1989b Vietnam Experience Study Spina biï¬da Vietnam veteransâ children 9 1.7 (0.6â5.0) Non-Vietnam veteransâ children 5 (*) Anencephaly Vietnam Veteransâ children 3 (*) Non-Vietnam veteransâ children 0 (*) Erickson Birth Defects StudyâVietnam veterans et al., Spina Biï¬da 19 1.1 (0.6â1.7) 1984a,b Anencephaly 12 0.9 (0.5â1.7) EOI-5: spina biï¬da 20f 2.7 (1.2â6.2) EOI-5: anencephaly 7f 0.7 (0.2â2.8) ADVA, 1983 Australian Vietnam veteransâneural tube defects 16 0.9 ABBREVIATIONS: 2,4,5-T, 2,4,5-trichlorophenoxyacetic acid; AIHW, Australian Institute of Health and Welfare; CDC, Centers for Disease Control and Prevention; EOI, exposure opportunity index; NR, not reported. Unless otherwise indicated, studies show paternal exposure. a Given when available. b Number of workers with maximal index of exposure (upper three quartiles) for any job held up to 3 months prior to conception. c Either or both parents potentially exposed. d 90% conï¬dence interval.
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 543 TABLE 7-4 Continued e Of the four neural tube defects reported among Ranch Hand offspring there were two spina biï¬da (high dioxin level), one spina biï¬da (low dioxin), and one anencephaly (low dioxin). No neural tube defects were reported in the comparison cohort. 454 post-service births were studied in Ranch Hand veterans; 570 in comparison cohort. f Number of Vietnam veterans fathering a child with a neural tube defect given any exposure op- portunity index. g Calculated from data presented in the paper. * Information not provided by study authors. the ï¬rst year of life due to any congenital defect (p 0.047), but the relation- ship was slightly more signiï¬cant for all deaths in the ï¬rst year (p 0.023); no relationship was found for the intervals closer to birth. This study had several methodologic weaknesses, including the lack of individual-level information on other risk factors and of individual exposure data. No data on speciï¬c types of congenital defects were included. Vietnam-Veteran Studies No new Vietnam veteran studies concerning exposure to the compounds of interest and birth defects were published since Update 2004. Biologic Plausibility Laboratory studies have established that maternal exposure to TCDD during pregnancy is associated with a wide variety of birth defects, which depend on the timing of exposure and the species being studied. However, laboratory stud- ies of potential male-mediated developmental toxicity attributable to exposure to TCDD and herbicides, speciï¬cally with regard to birth defects, are too limited to permit conclusions. It is notable that both the aryl hydrocarbon receptor (AhR) and the AhR nuclear translocator are expressed in the human testis and sperm (Khorram et al., 2004; Shultz et al., 2003), and studies in rodents have shown that TCDD exposure results in signiï¬cant changes in gene expression in spermato- cytes and Sertoli cells; hence, cells of the testis are responsive to TCDD expo- sure (Kuroda et al., 2005; Yamano et al., 2005). Furthermore, laboratory studies have shown that TCDD inhibits spermatogenesis and reduces erectile function in animals (Moon et al., 2004; Simanainen et al., 2004). Thus, there is biologic potential for paternal exposure to contribute to TCDD-induced reproductive and developmental toxicity. The only study assessing paternal TCDD exposure in humans that was re- viewed for the present update failed to show any evidence that paternal TCDD
544 VETERANS AND AGENT ORANGE: UPDATE 2006 exposure of male chemical workers caused an increase in birth defects in their offspring. In a previous laboratory study, paternal exposure of rats to TCDD failed to result in any birth defects in the F1 generation (Chahoud et al., 1991). And an epidemiologic study of maternal serum TCDD failed to establish any link with maternal thyroid dysfunction, which might contribute to cognitive or motor impairment in offspring (Foster et al., 2005). The biologic plausibility of reproductive effects in general arising from ex- posure to the chemicals of interest is discussed at the end of this chapter. Synthesis For this update, the only new occupational study of birth defects and expo- sures to the chemicals of interest was a follow-on study of the children of men in the NIOSH cohort (Lawson et al., 2004). Because only 1,166 liveborn and stillborn infants were available for analyses, the information generated was too sparse to provide additional insights into the risks of birth defects. Birth defects were addressed indirectly by a new environmental study of residents in areas sur- rounding metropolitan waste-incineration facilities in Japan (Tango et al., 2004). On the basis of large numbers of births, the Japanese study found an association between residence in the areas with the highest soil dioxin concentrations and deaths before the ï¬rst birthday due to any congenital abnormality, but this rela- tionship did not carry over to deaths occurring in the ï¬rst month or in the ï¬rst week of life. Ngo et al. (2006) conducted a systematic review and meta-analysis of the association between Agent Orange and birth defects. Relevant studies published from 1966 to 2002 were identiï¬ed by search engines, and the authors also in- cluded unpublished conference proceedings and information from researchers on ï¬ndings from unpublished studies. They reviewed 22 studies, including 13 Vietnamese studies (11 of them unpublished) and 9 of Vietnam veterans (2 unpublished). Secondary analyses of primary data reviewed by previous VAO committees fall outside the type of ï¬ndings ordinarily factored into the evidence bases evaluated in the VAO reviews, but given the nature of the reported results, the present committee wanted to comment on the Ngo et al. review. Unpublished studies are outside VAOâs inclusion criteria, so the 13 unpublished studies con- sidered by Ngo et al. were not included in previous VAO reviews of the associa- tion between the chemicals of interest and birth defects. All the studies that Ngo et al. indicate as published were reviewed in the VAO series. The Vietnam-veteran study that provided the highest risk estimates for birth defects (Field and Kerr, 1988) was excluded from consideration by previous VAO committees because of serious methodologic issues (IOM, 1994). In limiting their review to Agent Orange, Ngo et al. also excluded the broader array of occupational and environ-
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 545 mental studies on the components of Agent Orange that previous VAO reviews have addressed. Ngo et al. (2006) reported a substantially greater strength of association between exposure to Agent Orange and birth defects in the studies of Vietnamese popula- tions (RR 3.00, 95% CI 2.19â4.12) than in those of non-Vietnamese populations (RR 1.29, 95% CI 1.04â1.59). The non-Vietnamese study populations consisted of Vietnam veterans, who were almost exclusively men, whereas the Vietnamese popu- lations had a much greater likelihood of maternal exposure. Only Kang et al. (2000) addressed birth defects among the offspring of female Vietnam War veterans, who overall constitute fewer than 10,000 of the roughly 3 million US Vietnam veterans. Ngo et al. (2006) also conducted subgroup meta-analyses based on presumed exposure intensity. They reported no signiï¬cant increased risk of birth defects among non-Ranch Hand veterans (RR 1.04, 95% CI 0.93â1.16). Combining three sequential studies of Ranch Hand veterans, who were presumed to have been more highly exposed, risk was increased (RR 1.20, 95% CI 1.08â1.34), but they were unable to control for non-independent observations among the stud- ies of Ranch Hand veterans studies. Combined risks from studies of North Viet- namese veterans and of sprayed Vietnamese civilians were successively higher. The numbers of cases reported in the studies reviewed by Ngo et al. (2006) were too small to allow meta-analysis of speciï¬c types of birth defects. In general, the relatively small number of offspring among Vietnam veterans seriously restricts the ability to detect statistically signiï¬cant increases in speciï¬c birth defects, and meta-analytic methods are the best approach to assessing the overall import of the studies of exposures to the chemicals of interest and the risk of speciï¬c birth defects. In addition, as the offspring of veterans become older, the risk of diseases stemming from congenitally transmitted defects that alter normal physiologic function, such as endocrine and reproductive function, merits increasing attention. Conclusions There were no new relevant studies on the association between parental exposure to 2,4-D, 2,4,5-T, TCDD, cacodylic acid, or picloram and spina biï¬da in offspring. The committee concludes that the evidence is still limited or sug- gestive of an association between exposure to the compounds of interest and spina biï¬da. Its evaluation of the evidence reviewed here and in previous VAO reports leads the committee to conclude that there is still inadequate or insufï¬cient evi- dence of an association between exposure to the compounds of interest and all other birth defects.
546 VETERANS AND AGENT ORANGE: UPDATE 2006 CHILDHOOD CANCER The American Cancer Society estimated that 9,200 children under the age of 15 years would have a diagnosis of cancer in the United States in 2004 (ACS, 2004). Treatment and supportive care of children with cancer have improved greatly, and mortality has declined by 49 percent over the last 30 years. Despite those advances, cancer remains the leading cause of death from disease in chil- dren under the age of 15 years, and 1,510 deaths were projected for 2004 (ACS, 2004). Leukemia is the most common cancer in children. It accounts for about one-third of all childhood cancer cases; leukemia is expected to be diagnosed in nearly 2,760 children in 2004 (ACS, 2004). Of those, nearly 2,000 will have acute lymphocytic leukemia (ALL); most of the rest will have acute myelogenous leukemia (AML). AML (International Classiï¬cation of Diseases, Ninth Edition [ICD-9] 205) is commonly referred to as acute myeloid leukemia and acute nonlymphocytic leukemia. For consistency, this report uses acute myelogenous leukemia, or simply AML, regardless of usage in the source materials. ALL is most common in early childhood, peaking between the ages of 2 and 3 years, and AML is most common during the ï¬rst 2 years of life. ALL incidence is consistently higher in boys than in girls; AML incidence is similar in boys and girls (NCI, 2001). Through early adulthood, ALL rates are about twice as high in whites as in blacks; AML exhibits no consistent pattern in this respect. Chapter 6 contains additional information on leukemia as part of the discussion of adult cancer. The second-most common group of cancers in children are those of the central nervous systemâthe brain and the spinal cord. Other cancers in children include lymphomas, bone cancers, soft-tissue sarcomas, kidney cancers, eye can- cers, and adrenal gland cancers. Compared with adult cancers, relatively little is known about the etiology of most childhood cancers, especially about potential environmental risk factors and the effect of parental exposures. Conclusions from VAO and Updates The committee responsible for VAO concluded that there was inadequate or insufï¬cient evidence of an association between exposure to 2,4-D, 2,4,5-T, TCDD, picloram, or cacodylic acid and childhood cancers. Additional informa- tion available to the committees responsible for Update 1996 and Update 1998 did not change that conclusion. The committee responsible for Update 2000 reviewed the material in earlier VAO reports and newly available published literature and determined there was limited or suggestive evidence of an association between exposure to at least one of the compounds of interest and AML. After the release of Update 2000, investigators involved in one study discovered an error in their published data. The committee reconvened to evaluate the previously reviewed
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 547 and new literature regarding AML, and Acute Myelogenous Leukemia (IOM, 2002) was produced. It reclassiï¬ed AML from âlimited/suggestive evidence of an associationâ to âinadequate evidence to determine whether an association ex- ists.â Table 7-5 summarizes the results of the relevant studies. The committees responsible for Update 2000, Update 2002, and Update 2004 reviewed the mate- rial in earlier VAO reports and in newly available published literature and agreed that there remained inadequate or insufï¬cient evidence of an association between exposure and childhood cancers. Update of Epidemiologic Literature Occupational Studies Two related papers from the Childrenâs Oncology Group addressed exposures to pesticides and the risk of childhood germ-cell tumors (GCTs). One focused on residential pesticides and chemicals (Chen et al., 2006) and the other on parental occupational exposures (Chen et al., 2005), but they were based on the same overall caseâcontrol study. GCTs are very rare childhood cancers, so recruitment was done throughout the United States and included incident cases diagnosed from 1993 to 2001 in children under the age of 15 years. Controls were selected by random-digit dialing and frequency-matched on sex, year of birth, and state. Data were collected via two questionnaires (telephone and self-administered). Maternal exposure to herbicides at home from 6 months before pregnancy until termination of breastfeeding was associated with a slightly increased risk among female offspring (OR 1.4, 95% CI 1.0â2.0), but not among male offspring (OR 1.0, 95% CI 0.5â1.8). Similarly, Chen et al. (2005) reported on maternal and paternal exposures to pesticides before, during, and after pregnancy from occupational exposures. The data on analyses for exposures to herbicides, insecticides, or fungicides were not presented, but it was stated that the only positive association was for maternal exposure to herbicides at work during the postnatal period and GCTs in female offspring (OR 2.3, 95% CI 1.0â5.2); therefore, exposures with timing relevant to this review were not associated with increased risks of GCTs in the offspring. Both studies are limited by the questionable reliability of the self-reported ex- posures, reporting of the results only for the preconception through postnatal period, the small numbers of subjects, and the failure to consider residential and occupational herbicide exposures simultaneously. Environmental Studies Reynolds et al. (2005b) conducted a caseâcontrol study on childhood cancer and gestational exposure to pesticides. Compilation in Californiaâs Pesticide Use Reporting Database (CPUR) of information of amount and location of application
548 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-5 Selected Epidemiologic StudiesâChildhood Cancers Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a OCCUPATIONAL New Studies Chen et al., Parental occupational exposure to pesticide and 2005 childhood GCTs Maternal 32 1.1 (0.7â1.6) Paternal 39 0.9 (0.6â1.3) Reynolds Maternal exposure to agricultural pesticide in class of et al., 2005b âprobable human carcinogensâ (including cacodylic acid) during 9 months prior to delivery All sites 223 1.0 (0.9â1.2) Leukemias 179 1.2 (0.9â1.5) Central nervous system tumors 31 0.9 (0.5â1.4) Studies Reviewed in Update 2004 Flower Offspring of male pesticide applicators in Iowa from et al., 2004 AHS Maternal exposure to chlorophenoxy herbicides 7 0.7 (0.3â1.5) Paternal exposure to chlorophenoxy herbicides 28 1.3 (0.6â2.6) Maternal exposure to 2,4-D 7 0.7 (0.3â1.6) Paternal exposure to 2,4-D 26 1.29 (0.7â2.4) Studies Reviewed in Update 2000 Heacock Offspring of sawmill workers exposed to fungicides et al., 2000 contaminated with PCDDs and PCDFs Leukemia All workers offspringâincidence 11 1.0 (0.5â1.8) Offspring of workers with high chlorophenate exposure 5 0.8 (0.2â3.6)b Brain cancer All workers offspringâincidence 9 1.3 (0.6â2.5) Offspring of workers with high chlorophenate exposure 5 1.5 (0.4â6.9)b Buckley Childrenâs Cancer Study Groupâexposure to pesticides et al., 1989 and weed killersâAML Any paternal exposure 27 2.3 (p 0.5) Paternal exposure 1,000 days 17 2.7 (1.0â7.0) Maternal exposure 1,000 days 7 undeï¬ned
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 549 TABLE 7-5 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a ENVIRONMENTAL New Studies Chen et al., Childhood GCTs and residential exposure to herbicides 2006 6 months before conception, during gestation, or through breastfeeding period Maternal exposure 47 1.3 (0.9â1.7) Daughters 36 1.4 (1.0â2.0) Sons 11 1.0 (0.5â1.8) Paternal exposure 90 1.0 (0.7â1.3) Daughters 32 1.2 (0.7â2.0) Sons 58 1.0 (0.7â1.4) Studies Reviewed in Update 2002 Daniels Neuroblastoma risk in children (caseâcontrol study) (as et al., 2001 reported by both parents) Pesticides in the home (used ever) * 1.6 (1.0â2.3) Herbicides in the garden * 1.9 (1.1â3.2) Pesticides in the garden * 2.2 (1.3â3.6) Buckley NHL diagnosed at the age of 20 years in children with et al., 2000 potential prenatal exposure to herbicides * (*)c Kerr et al., Neuroblastoma risk in children 2000 Maternal occupational exposure to insecticides 40 2.3 (1.4â3.7) Paternal exposure to dioxin 7 6.9 (1.3â68.4) Studies Reviewed in Herbicide/Dioxin Exposure and AML in the Children of Veterans Kristensen Children of agricultural workers in Norway et al., 1996 Children with AML whose parents purchased pesticides 12 1.4 (0.6â2.9) Studies Reviewed in Update 2000 Meinert Childhood cancerâpopulation-based caseâcontrol study et al., 2000 Leukemia Paternal exposure; year before pregnancy 62 1.5 (1.1â2.2) Paternal exposure; during pregnancy 57 1.6 (1.1â2.3) Maternal exposure; year before pregnancy 19 2.1 (1.1â4.2) Maternal exposure, during pregnancy 15 3.6 (1.5â8.8) Lymphomas Paternal exposure, year before pregnancy 11 1.5 (0.7â3.1) Paternal exposure, during pregnancy 10 1.6 (0.7â3.6) Maternal exposure, year before pregnancy 3 2.9 (0.7â13) Maternal exposure, during pregnancy 4 11.8 (2.2â64) Pearce and Kidney cancer in subjects (1â15 yrs) with paternal Parker, 2000 occupation in agriculture 21 0.9 (0.2â3.8) continued
550 VETERANS AND AGENT ORANGE: UPDATE 2006 TABLE 7-5 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Infante- Childhood ALL in households using herbicidesâ Rivard population-based caseâcontrol study et al., 1999 Exposure during pregnancy 118 1.8 (1.3â2.6) Exposure during childhood 178 1.4 (1.1â1.9) Studies Reviewed in Update 1996 Pesatori Seveso residents aged 0â19 yearsâ10-year follow-up, et al., 1993 morbidity, all exposure zones All cancers 17 1.2 (0.7â2.1) Ovary and uterine adnexa 2 â (0 expected) Brain 3 1.1 (0.3â4.1) Thyroid 2 4.6 (0.6â32.7) Hodgkinâs lymphoma 3 2.0 (0.5â7.6) Lymphatic leukemia 2 1.3 (0.3â6.2) Myeloid leukemia 3 2.7 (0.7â11.4) Bertazzi Seveso residents aged 0â19 yearsâ10-year follow-up, et al., 1992 mortality, all exposure zones All cancers 10 7.9 (3.8â13.6) Leukemias 5 3.9 (1.2â1.8) Lymphatic leukemia 2 1.6 (0.1â4.5) Myeloid leukemia 1 0.8 (0.0â3.1) Leukemia, others 2 1.6 (0.1â4.6) Central nervous system tumors 2 1.6 (0.1â4.6) VIETNAM VETERANS Studies Reviewed in Herbicide/Dioxin Exposure and AML in the Children of Veterans AIHW, Australian Vietnam veteransâ childrenâRevised 2001 Validation StudyâAML 12d 1.3 (0.8â4.0) Studies Reviewed in Update 2000 AIHW, 2000 Australian Vietnam veteransâ childrenâValidation StudyâAML This study, which incorrectly calculated the expected number of AML cases, is superseded by AIHW, 2001 above. Wen et al., Caseâcontrol study of childrenâs leukemia 2000 AML and ALL Father ever served in Vietnam or Cambodia 117 1.2 (0.9â1.6) 1 year in Vietnam or Cambodia 61 1.4 (0.9â2.0) 1 year in Vietnam or Cambodia 49 1.2 (0.8â1.7) AML only Father ever served in Vietnam or Cambodia 40 1.7 (1.0â2.9) 1 year in Vietnam or Cambodia 13 2.4 (1.1â5.4) 1 year in Vietnam or Cambodia 16 1.5 (0.7â3.2)
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 551 TABLE 7-5 Continued Estimated Exposed Relative Risk Reference Study Population Casesa (95% CI)a Studies Reviewed in VAO CDC, 1989b Vietnam Experience Studyâoutcomes in the offspring of veterans Cancer 25 1.5 (0.7â2.8) Leukemia 12 1.6 (0.6â4.0) Field and Cancer in children of Australian Vietnam veterans 4 (*) Kerr, 1988 Erickson CDC Birth Defects Studyâchildren of Vietnam veterans et al., 1984b âOtherâ neoplasms 87 1.8 (1.0â3.3) ABBREVIATIONS: AFHS, Air Force Health Study; AHS, Agricultural Health Study, AIHW, Aus- tralian Institute of Health and Welfare; ALL, acute lymphocytic leukemia; AML, acute myeloid leukemia; CDC, Centers for Disease Control and Prevention; CI, conï¬dence interval; EOI, exposure opportunity index; GCT, germ-cell tumor; NS, not signiï¬cant; SS, statistically signiï¬cant. a Given when available. b OR estimated using low exposure subjects as the comparison cohort. c No information on herbicides as a class, distinct from insecticides or other pesticides, was available; exposures before conception were not singled out, and no distinction between maternal and paternal exposure was made. d Of the 12 cases, 9 were observed and 3 additional cases were estimated to have occurred in the portion of the cohort whose data were not validated. * Information not provided by study authors. Unless otherwise indicated, studies show paternal exposure. by speciï¬c product began in 1990. Considering children born after September 1990 and diagnosed with cancer before the age of 5, they identiï¬ed 2,189 children entered in Californiaâs population-based cancer registry by the end of 1997. A total of 4,335 live controls were randomly selected from California birth certiï¬- cates and matched on date of birth and sex. A geographic information system approach was 99 percent successful in geocoding residence at time of birth for 2,189 case mothers and 4,388 control mothers, permitting linkage to the CPUR. Considering the amount of pesticide applied and the distance of the treated areas from the motherâs residence, cumulative exposures for the 9 months before birth were determined for several different classiï¬cations of pesticides. Californiaâs category of pesticides that are âprobable human carcinogens,â which includes cacodylic acid, is the only classiï¬cation of possible relevance for this review. Cases of all types of cancer were no more likely than controls to have experienced high exposure during gestation to âprobable human carcinogensâ (OR 1.01, 95% CI 0.85â1.20); the same was true for the subset of leukemias (OR 1.17, 95% CI 0.90â1.54) or central nervous system cancers (OR 0.86, 95% CI 0.54â1.36). No individual-level data were obtained for the study participants or
552 VETERANS AND AGENT ORANGE: UPDATE 2006 their parents regarding lifestyle or other personal characteristics to permit adjust- ment for possible confounder. The exposure assessment assumes that residential proximity is a good indicator of exposure. Distance to pesticide exposure sources was based on the childrenâs residences at birth, which would lead to at least some misclassiï¬cation due to residential mobility. Finally, although the study included many subjects, most of them lived in areas that experienced little or no agricul- tural pesticide use, so the risk estimates for the high exposure categories were based on small numbers. Vietnam-Veteran Studies No new Vietnam veteran studies concerning exposure to the compounds of interest and childhood cancer were published since Update 2004. Biologic Plausibility Paternal or maternal exposure to xenobiotics potentially could increase the susceptibility of offspring to developing cancer through multiple mechanisms. Susceptibility could be increased by inheriting a genetic predisposition, which by itself could increase the development of cancer or the likelihood of developing cancer after future exposure to a carcinogen; the mother or father would transmit either an acquired genetic defect or an epigenetic alteration that predisposed the child to cancer. Alternatively, a maternally mediated increased susceptibility to childhood cancer could result from direct exposure of a child in utero or via lactation to a xenobiotic that induces epigenetic alterations that increase cancer susceptibility or is itself a carcinogen. It has been shown that prenatal TCDD exposure of rats is associated with al- tered mammary gland differentiation and an increase in the number of mammary adenocarcinomas (Brown et al., 1998). A recent studyâs demonstration that early postnatal TCDD exposure does not increase mammary-cancer risk (Desaulniers et al., 2004) suggests that TCDD-induced changes in utero mediate the increase in cancer susceptibility. Developmental epigenetic alterations may be involved in those prenatal effects. TCDD has been shown to suppress the expression of two tumor-suppressor genes, p16Ink4a and p53, via an epigenetic mechanism that ap- pears to involve DNA methylation (Ray and Swanson, 2004). Similarly, it was re- ported that prenatal TCDD exposure increases methylation of two growth-related imprinted genes, H19 and Igf2, in the developing fetus (Wu et al., 2004). Although there is no direct evidence from animal models that TCDD in- creases the risk of childhood cancers, such as acute leukemia or GCTs, emerging research suggests that prenatal TCDD exposure can disrupt epigenetic imprinting patterns and alter organ differentiation, and this could contribute to an increased susceptibility to cancer later in life. A recent study has shown that chromosomal rearrangements associated with childhood ALL are evident in the neonatal blood
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 553 spots; this suggests that childhood leukemias begin before birth and that mater- nal and perinatal exposures to xenobiotics may contribute to genetic mutations (Smith et al., 2005). Synthesis The studies reviewed for this update did not ï¬nd signiï¬cant associations between the relevant exposures and childhood cancers. As with other outcomes in the offspring of Vietnam veterans, the small number of these rare childhood cancers expected among the circumscribed number of Vietnam veterans would seriously hinder detection of any actual increases; meta-analytic methods may be the best approach for assessing the overall signiï¬cance of the association between exposures to the chemicals of interest and the risk of speciï¬c childhood cancers. Conclusions On the basis of its evaluation of the evidence reviewed here and in previous VAO reports, the committee concludes that there is inadequate or insufï¬cient evidence of an association between exposure to the compounds of interest and childhood cancers. SUMMARY Overall Biologic Plausibility of Reproductive and Developmental Effects This section summarizes the general biologic plausibility of a connection be- tween exposure to the compounds of interest and reproductive and developmental effects on the basis of data from animal and cellular studies. Chapter 3 presents details of the committeeâs evaluation of data from the recent studies. TCDD induces a variety of adverse reproductive and developmental effects in laboratory animals. The administration of TCDD to male animals elicits re- productive toxicity by affecting testicular and seminal vesicle weight and func- tion and by decreasing the rate of sperm production. The mechanisms of those effects are not known, but a primary hypothesis is that they are mediated through dysregulation of testicular steroidogenesis. Exposure to TCDD is associated with increased estradiol secretion and decreased testosterone secretion; both hormones regulate sperm production. However, high doses of TCDD were required to elicit those effects, and TCDD-exposed male rats were able to sire viable fetuses. Thus, there is little evidence from laboratory studies that TCDD exposure of men con- tributes to reduced fertility. Many studies also have examined the effects of TCDD on the female re- productive system. Two primary mechanisms that probably contribute to abnor-
554 VETERANS AND AGENT ORANGE: UPDATE 2006 mal follicle development and decreased numbers of ova after TCDD exposure are crosstalk of the AhR with the estrogen receptor and dysregulation of the hypothalamic-pituitary-gonadal axis. Interaction of the activated AhR leads to inhibition of estradiol-induced gene expression and to enhancement of estrogen- receptor protein degradation; both activities may contribute to TCDDâs anties- trogenic effects. TCDD also dysregulates the secretion pattern of preovulatory gonadotropin hormones, and this leads to abnormal and reduced follicle devel- opment. In addition, oocytes are directly responsive to TCDD. Thus, TCDDâs effects on hormone concentrations, hormone-receptor signaling, and ovarian responsiveness to hormones all probably contribute to TCDD-induced female reproductive toxicity. On the basis of animal data, male and female reproductive effects of TCDD in humans are biologically plausible. Furthermore, a recent study has shown that dioxins and dibenzofurans signiï¬cantly correlate with dysregulation of estrogen metabolism in pregnant women (Wang et al., 2006)â evidence of a biologically plausible mechanism of TCDDâs adverse effects on human reproduction. In animal studies, TCDD crosses the placenta and is transferred via breast milk. In humans, TCDD has been measured in circulating maternal blood, cord blood, placenta, and breast milk (Suzuki et al., 2005), and it is estimated that an infant breast fed for 1 year accumulates a dose of TCDD that is 6 times higher than an infant not breast fed (Lorber and Phillips., 2002). Thus, the exposure of human infants in utero and via lactation has been demonstrated. In animal studies, TCDD exposure during pregnancy can reduce body weight at birth, but this typically occurs at high doses. Thus, there is little evidence that TCDD exposure is associated with reduction in birth weight. TCDD is a potent teratogen in all laboratory species that have been studied, although the pattern of induced birth defects is often species-speciï¬c. For exam- ple, ï¬sh, mice, and avian embryos exhibit substantial alterations in craniofacial developmentâshortened jaw in piscine species, cleft palate in mice, and beak malformations in birds. The developing cardiovascular system is also a com- mon target for TCDD-induced teratogenicity, and it has been shown that cardiac myocytes and the endothelial lining of the heart and blood vessels are primary target sites of TCDDâs effects. Cytochrome P450 1A1 induction or alterations in pathways controlled by vascular endothelial growth factor might mediate the early lesions that result in TCDD-related vascular derangements. That an- tioxidant treatment provides protection against TCDD-induced embryotoxicity in some systems suggests that reactive oxygen species might be involved in the teratogenic effects of exposure to TCDD. Several reports of studies in exposed animals and humans suggest that low perinatal exposure to TCDD and 2,4-D could impair brain development. Outcomes can be subtle, ranging from altered learning and memory to modiï¬ed sex-related behavior. The mechanisms of those effects are unclear. Studies in rodents show that a single maternal dose of TCDD produces
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 555 malformations of the external genitalia and functional reproductive alterations in female progeny, including decreased fertility rate, reduced fecundity, cystic en- dometrial hyperplasia, and disrupted estrous cycles. Those effects depend on the timing of exposure. Similarly, male progeny exhibit alterations in reproductive- organ development and function. Maternal exposure to TCDD impairs prostate growth and seminal vesicle weight and branching and decreases sperm production and caudal epididymal sperm number in offspring. Little research has been conducted on the offspring of male animals exposed to herbicides. Feeding of simulated Agent Orange mixtures to male mice pro- duced no adverse effects in offspring; a statistically signiï¬cant excess of fused sternebrae in the offspring of the two most highly exposed groups was attributed to an anomalously low rate of this defect in the controls (Lamb et al., 1981). Altered sex ratio might reï¬ect the effects of exposure to the chemicals of interest on reproductive capability. It has been hypothesized that concentrations of parental hormones at conception or induction of lethal mutations before birth could affect sex ratio. There has been no work with experimental animals that speciï¬cally examined the effects of TCDD on sex ratios of offspring, nor have any alterations in sex ratio been reported in animal studies that examined devel- opmental effects of TCDD on offspring. The mechanisms by which TCDD induces birth defects have not been es- tablished and are probably species- and organ-speciï¬c. Nonetheless, studies have consistently demonstrated that TCDD-induced developmental toxicity required the AhR. That has been deï¬nitively established in mice that lack AhR expression. When pregnant AhR-null mice are exposed to TCDD, the fetuses fail to exhibit any of the typical developmental malformations associated with TCDD exposure. The activated AhR mediates changes in gene transcription, so the inappropriate and sustained activation of AhR by TCDD during development appears to be a key ï¬rst step in mediating TCDDâs developmental toxicity. Although structural differences in the AhR have been identiï¬ed among species, it functions similarly in animals and humans. Therefore, a common mechanism probably underlies the reproductive and developmental toxicity of TCDD in humans and animals, and data on animals support a biologic basis of TCDDâs toxic effects. Little information is available on the reproductive and developmental effects of exposure to the herbicides discussed in this report. Studies indicate that 2,4-D does not affect male or female fertility and does not produce fetal abnormalities. Offspring of pregnant rodents exposed to 4-(2,4-dichlorophenoxy)butyric acid (2,4- DB) exhibit a reduced growth rate and increased mortality (Charles et al., 1999), but only after very high doses. Exposure to 2,4-D also alters the concentration and function of reproductive hormones and prostaglandins. One study reported an increased incidence of malformed offspring of male mice exposed to a mixture of 2,4-D and picloram in drinking water; however, paternal toxicity was observed in the high-dose group, and there was no clear doseâresponse relationship (Blakley et al., 1989). Picloram alone produced fetal abnormalities in rabbits at doses that
556 VETERANS AND AGENT ORANGE: UPDATE 2006 are also toxic to the pregnant animals (John-Greene et al., 1985), but that effect has not been seen in many studies. 2,4,5-T was toxic to fetuses when administered to pregnant rats, mice, and hamsters; its ability to interfere with calcium homeostasis in vitro has been documented and linked to its teratogenic effects on the early de- velopment of sea urchin eggs. Cacodylic acid is toxic to rat, mouse, and hamster fetuses at high doses that are also toxic to the pregnant mother. There is growing evidence from laboratory animal and human studies that exposures during fetal or postnatal development can lead to adverse effects later in life that are not immediately apparent as structural malformations or functional deï¬cits. For example, exposure of humans and rats to TCDD in early postnatal life induces dental aberrations and reduces enamel maturation of teeth (Alaluusua et al., 2004; Gao et al., 2004). A study of human exposure to background con- centrations of dioxins, furans, and PCBs during prenatal development (Nakajima et al., 2006) suggests possibly more relationship with reduced motor development in 6-month-old infants than with their mental development; however, the few sig- niï¬cant correlations found among dozens of comparisons made were for speciï¬c congeners with low relative potency (TEFs), so the study is essentially negative for developmental effects arising from prenatal exposure to TCDD. The foregoing suggests that a connection between TCDD exposure and hu- man reproductive and developmental effects is, in general, biologically plausible. However, more deï¬nitive conclusions about the potential for such TCDD toxic- ity in humans are complicated by differences in sensitivity and susceptibility among individual animals, strains, and species; by the lack of strong evidence of organ-speciï¬c effects among species; by differences in route, dose, duration, and timing of exposure; and by substantial differences in the toxicokinetics of TCDD between laboratory animals and humans. Experiments with 2,4-D and 2,4,5-T indicate that they have subcellular effects that could constitute a biologi- cally plausible mechanism for reproductive and developmental effects. Evidence from animals, however, indicates that they do not have reproductive effects and that they have developmental effects only at very high doses. There is insufï¬cient information on picloram and cacodylic acid to assess the biologic plausibility of those compoundsâ reproductive or developmental effects. Considerable uncertainty remains about how to apply toxicologic informa- tion to the evaluation of potential health effects of herbicide or TCDD exposure on the offspring of Vietnam veterans. Scientists disagree over the extent to which information derived from animal and cellular studies can be used to predict hu- man health outcomes and over the extent to which the health effects of high-dose exposure can be extrapolated to low-dose exposure. The biologic mechanisms that underlie TCDDâs toxic effects continue to be a subject of active research, and future VAO updates are likely to have more and better information on which to base conclusions, at least for TCDD.
REPRODUCTIVE AND DEVELOPMENTAL EFFECTS 557 Synthesis The studies reviewed for this update did not ï¬nd any signiï¬cant associa- tions between the relevant exposures and reproductive outcomes. The scientiï¬c evidence supports the biologic plausibility of a connection between exposure to the chemicals of interest and reproductive effects, but the epidemiologic studies of occupational cohorts, exposed communities, and Vietnam veterans have not provided conclusive evidence of any additional associations between exposures and an array of reproductive outcomes and conditions among the offspring of exposed parents. The mechanisms by which the chemicals exert their biologic effects are still subjects of scientiï¬c investigation. With the aging of the Vietnam- veteran population, additional studies on fertility, spontaneous abortion, and sex ratio cannot be expected, although there may be additional studies of reproductive outcomes in other populations with exposure to the chemicals of interest. The possibility of structural or functional abnormalities in the offspring of exposed people will continue to be of interest. Conclusions There is inadequate or insufï¬cient evidence of an association between expo- sure to 2,4-D, 2,4,5-T, TCDD, picloram, or cacodylic acid and altered hormone concentrations; semen quality; infertility; spontaneous abortion; late fetal, neo- natal, or infant death; low birth weight or preterm delivery; birth defects other than spina biï¬da; and childhood cancers. There is limited or suggestive evidence of an association between exposure to the compounds of interest and spina biï¬da. There is limited or suggestive evidence that the speciï¬c combination of pa- ternal exposure to TCDD is not associated with risk of spontaneous abortion. REFERENCES1 Abell A, Juul S, Bonde JP. 2000. Time to pregnancy among female greenhouse workers. Scandinavian Journal of Work, Environment and Health 26(2):131â136. ACS. 2004. Cancer Facts and Figures 2004. http://www.cancer.org/downloads/STT/CAFF_ï¬nalP- WSecured.pdf (Accessed September 13). ADVA (Australia Department of Veterans Affairs). 1983. CaseâControl Study of Congenital Anoma- lies and Vietnam Service. Canberra. AFHS (Air Force Health Study). 1992. An Epidemiologic Investigation of Health Effects in Air Force Personnel Following Exposure to Herbicides. Reproductive Outcomes. Brooks AFB, TX: USAF School of Aerospace Medicine. AL-TR-1992-0090. 602 pp. 1Throughout the report the same alphabetic indicator following year of publication is used con- sistently for the same article when there were multiple citations by the same ï¬rst author in a given year. The convention of assigning the alphabetic indicator in order of citation in a given chapter is not followed.
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