Bisphenol A (BPA) has gained considerable attention as an environmental chemical with multiple adverse effects on human health and disease. The chemical composition of bisphenol A is 2,2-bis(4-hydroxyphenyl)propane (see Fig. 15.1d). BPA is produced in large quantities (greater than 6 billion pounds per year) as it is incorporated into various resins and plastics (e.g., polycarbonate). Hence, the exposure to BPA is ubiquitous in the USA, frequenting many aspects of daily living primarily through dietary consumption [8]. The Chapel Hill Bisphenol A expert panel issued a consensus statement in 2007 on the relationship between BPA and human health effects [71]. BPA appears to have endocrine disrupting effects through steroid receptor binding. BPA binds to the nuclear estrogen receptor with weak affinity. Recent evidence suggests it may act to induce physiologic responses through cell membrane estrogen receptors in low pg/mL concentrations [72]. BPA levels in humans have been measured in the parts per billion range in serum [73, 74] and urine [75, 76]. The relevance of low levels of BPA exposure in humans remains in question pertaining to significant reproductive biologic effects [77–80].

Adverse female reproductive effects of BPA exposure have been described with respect to meiotic aberrations in the oocyte. In 2000, Takai et al. published their work on the effects of BPA on early embryo development [81]. Two-cell mouse embryos were cultured with BPA at low, environmentally relevant doses in the presence or absence of Tamoxifen, a selective estrogen receptor modulator; blastocyst advancement was adversely affected. A direct link between BPA and murine aneuploidy was first reported by Hunt and colleagues, in which a dramatic increase from 1–2% to 40 % was observed in chromosomal alignment defects in the first meiotic spindle [82]. This observation was ultimately traced and attributed to the leaching of BPA from damaged polycarbonate cages and water bottles in which the female mice were housed during the final stages of oocyte maturation. The effects of in utero BPA exposure may also be transgenerational. In 2007, Susiarjo et al. reported abnormal pachytene associations and abnormalities in synaptonemal complex structures of BPA-exposed murine fetuses with higher rates of aneuploidy [83]. A similar meiotic phenotype in prophase fetal oocytes of ßERKO −/− mice with similar rates of synaptonemal aberrations (57 %), compared to those of the BPA-exposed female murine fetuses (52 %), supports the estrogen receptor ß as a potential site of interaction with BPA as a pathway for meiotic disruption.

The clinical relevance of BPA is emerging. A study from Japan was published in which 45 patients with a history of three or more first trimester miscarriages without uterine anomaly or blood karyotype abnormality (study group) and 32 healthy nonpregnant women without prior pregnancy loss (control group) underwent serum BPA testing [84]. Mean serum BPA levels were significantly higher in the study group compared to the control group (2.59 vs. 0.77 ng/mL), concluding that serum BPA is associated with recurrent miscarriage. Yet, methods employed in this study introduced several limitations, including the use of an enzyme-linked immunosorbent assay (ELISA) to assess BPA exposure [85], the cross-sectional assessment of BPA which has a half-life of only several hours, a lack of consideration for “critical” biologic windows in timing the exposure assessment (such as during the LH surge when BPA exposure is most likely to affect the meiotic transition from metaphase I to metaphase II), and the different demographics utilized for recruitment of patients and control subjects. However, additional clinical studies have recently been published with respect to BPA. Serum unconjugated and urinary conjugated BPA levels both inversely correlate with fertilization rates of exposed human oocytes [86, 87]. Additionally, BPA appears to influence estradiol production by inhibiting aromatase activity in vitro [88]. Clinical evidence supporting reduced estradiol secretion exists based on lower peak estradiol responses to gonadotropin stimulation during IVF with increasing BPA exposures [73, 89]. Another study by Hanna et al. explored the potential role of BPA in altering methylation sites in women undergoing IVF [90]. They found BPA to be associated with lower methylation of the promoter region of the TSP (testes-specific protease) 50 gene. However, no evidence has been published to date, to indicate that BPA exposure is associated with increased rates of human embryonic aneuploidy.

Polychlorinated biphenyls comprise a family of 209 structurally related compounds, or congeners, consisting of two carbon–hydrogen phenyl rings, with a spectrum of chlorine substituents that determine activity. The PCB congener 2,2’,4,4’,5,5′-hexachlorobiphenyl (PCB #153, see Fig. 15.1e) tends to be found most frequently in US biospecimens, although the relative contributions of various congeners to total PCB body burdens vary. PCBs were originally manufactured as chemical mixtures, variously composed to suit a wide range of industrial applications. These chemicals are ubiquitous and persistent in the environment due to their chemical stability, lipophilic character, and low water solubility [5]. While banned from production in most countries in the 1970s, the persistence of PCBs makes them relevant as environmental contaminants potentially influencing reproductive outcomes. The food chain is the primary source of human exposure, as PCBs bioaccumulate in animal fats leading to biomagnification. PCB congeners were recently classified as carcinogenic to humans by the World Health Organization’s International Agency for Research on Cancer (IARC) [91], and variously possess estrogenic and antiestrogenic properties [92]. Congeners absent a chlorine (Cl) substituent at three or four of the ortho-carbon positions can assume a “coplanar” configuration and interact with the AhR, eliciting dioxin-like effects [93], including proinflammatory properties that increase oxidative stress [94] .

The first report of PCBs causing reproductive harm came from a 1980s study based in the Netherlands. A decline in the Wassen Sea seal population was traced to reduced litters associated with PCB and DDE contamination [95]. Consumption of sport-caught fish fosters increased exposure to PCBs [96], reflected in higher levels in the serum of sport fish consumers [97, 98]. Several epidemiologic studies have reported associations between PCB body burden and outcomes related to female fertility. A recent prospective cohort study of women, exposed through the consumption of Great Lakes sport fish, reported increased time to pregnancy in association with exposure to estrogenic and antiestrogenic PCB congeners [99]. An association between estrogenic PCBs and increased menstrual cycle length was reported from the same study [100]. Another, larger prospective cohort study with preconception participant enrollment also reported associations between pregnancy delays and PCB congeners [101]. Similar results have been reported by European investigators [102, 103] . Maternal serum PCB concentrations have been documented during critical windows of development, including the periconception interval and early pregnancy [104], and PCB levels are detectable within human follicular fluid [105, 106]. More recently, follicular fluid and serum PCBs have been associated with reduced oocyte fertilization [107] and with reduced embryo implantation during IVF [108], although a systematic review concluded that the epidemiologic evidence to date is insufficient to support a causal association [109] .

A substantial number of studies in mammalian models demonstrate the effects of PCBs on ovarian function, oogenesis, and embryogenesis, corroborating positive results reported from epidemiologic investigations in women. In vitro experiments report disrupted oocyte maturation, impaired fertilization, and compromised bovine embryo growth in a dose-dependent manner [110, 111]. Earlier studies demonstrated disruption of estrus and menstrual cycles in rats and primates [112–115], and the negative effects of several PCB mixtures on the fertilizability of murine oocytes [116–118]. More recently, altered synthesis of estradiol , testosterone and progesterone by human cell cultures was reported following treatment with individual PCB congeners [119]. The “dioxin-like” PCB congener #77 was also found to decrease fertilization potential in murine oocytes when female mice were fed a contaminated diet [120]. In another study, a mix of PCBs and related compounds reduced porcine oocyte cumulus expansion in a dose-dependent manner and decreased blastocyst formation [121]. Female rabbits also experienced reduced blastocyst formation following treatment three times a week with a PCB mixture [122] .

Investigation has suggested a role for the AhR, as a mechanism to explain murine ovarian weight and cyclicity effects reported following experimental PCB exposure [123, 124]. However, contradictory evidence identified AhR −/− mice to be fertile without compromised reproduction, raising the question of whether PCB effects on reproduction are mediated via other mechanisms [125]. Further evidence against an exclusive role for AhR mediated embryo-toxic effects by PCBs is provided by post-exposure AhR independent changes in rabbit blastocyst gene expression [126]. Though at higher concentrations than typically experienced by human populations, there is clear evidence in various mammalian models that oocyte competence and early embryo development can be compromised by exposures to PCBs .

Dioxins represent a class of organic chemicals that are structurally related to PCBs, typically including polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and coplanar polychlorinated biphenyls (co-PCBs). Dioxins are formed primarily as a byproduct of the incomplete combustion of chlorine-containing materials, or as inadvertent contaminants in chemical synthesis; they tend to bioaccumulate and to have very long in vivo half-lives [127]. The most potent dioxin is 2,3,7,8-tetrachlorodibenzodioxin (2,3,7,8-TCDD; see Fig. 15.1f), to which humans are generally exposed via consumption of contaminated food products and inhalation around municipal waste incinerators. TCDD has been classified as a human carcinogen by the IARC [128] and displays antiestrogenic endocrine-disrupting properties [129]. Furthermore, detectable concentrations have been reported in human follicular fluid [130]. Similar to PAHs, dioxins bind as AhR ligands with downstream gene activation and expression [131]. Though the antiestrogenic effects of dioxins are primarily attributed to an increased rate of hepatic estradiol metabolism, there is also evidence that TCDD acts as an endocrine disruptor to reduce estrogen biosynthesis within the ovarian follicle via actions on 17,20-lyase activity of the P450c17 enzyme complex [132]. TCDD may influence ovarian reserve via the AhR activation pathway, similar to PAHs [133, 134] .

The clinical evidence for dioxin exposure and impaired fertility among women comes mainly from the Seveso Women’s Health Study, which studied an Italian village population north of Milan, exposed to high levels of 2,3,7,8-TCDD from a 1976 chemical plant explosion that released ~ 30 kg into the atmosphere [135, 136]. Measurable serum levels of TCDD in Seveso residents ranged from 2.5–56,000 parts per trillion (ppt), with a background of 20 ppt among members of a nonexposed reference group [137]. The evidence that dioxins adversely affect female infertility is sparse however and further studies need to be performed to clearly define the risk at more commonly encountered levels in the environment and the potential impact on unexplained female infertility .

Polybrominated diphenyl ethers (PBDEs) comprise a family of 209 closely related lipophilic chemicals , structurally analogous to PCBs, with the exceptions of bromine substituents and an ether bond. The PBDE congener 2,2’,4,4’-tetrabromodiphenyl ether (PBDE #47, see Fig. 15.1g) tends to be found most frequently in US biospecimens, although the relative contributions of various congeners to total PBDE body burdens vary. Like PCBs, these compounds were variously manufactured as mixtures to suit particular purposes, primarily blended into commercial products as flame-retardants [138]. However, their appearance in the environment is more recent, having been introduced in the early 1970s, but now distributed to humans and biota worldwide in near ubiquitous fashion [139]. Given their environmental persistence and lipophilic nature, PBDEs tend to bioaccumulate in animal fats with long half-lives [140], raising concerns with respect to human health . Exposure is likely through inhalation and ingestion of contaminated dust, and through consumption of contaminated food products. Use of the most common PBDE mixtures are being phased out (i.e., deca-BDE) or eliminated in the USA (penta-BDE and octa-BDE), but concerns remain due to the persistence of PBDE congeners in the environment and their widespread presence in commercial products .

There have been few epidemiologic studies of human fertility and PBDE exposures published to date. An increased risk for early onset of menarche in US adolescents was recently reported from a cross-sectional investigation [141]. No association between breast milk PBDE concentrations and menstrual cycle length was reported from a small cross-sectional study conducted among pregnant women in Taiwan [142]. An increased time to pregnancy was reported in association with higher serum PBDE levels among pregnant members of a Mexican-immigrant community in California, at concentrations higher than those measured in the Taiwan study, yet no association was detected for menstrual cycle parameters [143]. In a very large French cohort study, no association was reported for time to pregnancy in association with serum concentrations of a single PBDE congener; however, the latter was infrequently detected among members of the study sample [103]. A more recent prospective study of women undergoing IVF reported an increased implantation failure rate associated with increased follicular fluid PBDE concentrations [144]. In contrast, the recent prospective study with preconception enrollment of women conceiving unassisted reported no associations with PBDEs, following adjustment for confounding variables [101].

Endocrine disruptive effects have been reported for PBDEs and their OH-PBDE metabolites using experimental models [145] . Estrogen and progesterone receptor binding with agonistic or antagonistic effects have been demonstrated using in vitro systems and in vivo using rodents systems [146, 147], as has binding to the androgen receptor, also with antagonistic effects in vitro and in vivo [146, 148]. Furthermore, OH-PBDE metabolites inhibit aromatase activity in vitro [149]. Gestational exposure at environmentally relevant doses elicited structural changes in ovaries, and altered folliculogenesis at higher concentrations [150, 151] . Changes to uterine estrogen-mediated gene expression and the distribution of uterine progesterone receptors was also reported in association with nontoxic PBDE exposures [152]. While many experimental effects have been reported at doses exceeding those usually experienced by humans, differential sensitivities between women and model organisms, and long-term human exposures complicate extrapolation of these data. Moreover, additive or synergistic effects between PBDEs and related compounds such as PCBs may potentiate the impact of low doses [153], and in doing so play an important role in unexplained female infertility .

Fluorotelomer and sulfonamide alcohols are widely employed for industrial and commercial applications, including use as surfactants and chemical intermediates, and integrated into food packaging, stain resistant and “non-stick” coatings, and “breathable” waterproof fabrics [154]. Though first used in the 1950s, only recently did the widespread distribution of their breakdown products, the most prevalent of which are perfluoroctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) (see Fig. 15.1h–i), become widely appreciated by the scientific community [155]. Due to a highly stable carbon-fluorine backbone, environmental persistence is essentially indefinite and their tendency to bind serum proteins and to undergo enterohepatic circulation results in human bioaccumulation. People are generally thought to be exposed through drinking water contamination and use of relevant products. Concerns have led to a voluntary reduction and phase out of PFOS and PFOA by leading US manufacturers .

Similar to PBDEs, human investigation into potential effects on female fertility has been sparse to date. A large prospective epidemiologic study recently reported an increased time to pregnancy in association with higher maternal PFOS and PFOA measured early in gestation [156]. Yet, questions with respect to the temporal nature of the association in that study have been raised by a more recent retrospective investigation [157, 158], complicating the interpretation of these data. Two additional prospective studies recently reported no associations for PFOS or PFOA with conception [101, 159]. Experimental studies in mice have reported altered estrous cyclicity following PFOS treatment [160], and dose-dependent increases in total litter resorption following PFOA treatment [161], albeit at levels exceeding those typically experienced by women. Additional studies suggest PFOS and PFOA possess estrogenic and antiestrogenic activities [162, 163]. No adverse reproductive effects were detected at nontoxic PFOS doses in an earlier two-generation rat study [164] although these results were limited by rapid metabolism and elimination of perfluorinated compounds by the female rat, whereas the elimination half-life in humans is measured in years. Given the widespread distribution and persistence of these compounds, additional investigation is necessary to more conclusively evaluate the potential for reproductive toxicity and possible contributions to unexplained female infertility .

Another area of emerging interest is the potential effect of toxic elements on female reproductive potential, including arsenic (As), cadmium (Cd), lead (Pb), and mercury (Hg). Though nonessential, toxic elements are distributed in ubiquitous fashion, with detectable levels reported for the vast majority of the US population [8], and investigators have reported accumulation of these agents in female reproductive tissues [165]. There are several environmental sources of exposure, including inhalation of polluted air and consumption of contaminated water and food. Seafood is an important exposure source due to bioaccumulation, and in some cases biomagnification, in the aquatic food chain [166, 167]. Consumption of shellfish, especially bivalves, has been shown to increase exposure to Cd [168]. As mentioned previously, though cigarette smoke is a potent source of Cd exposure, it also contributes to Pb and As exposure. Consumption of predatory fish species has been shown to contribute to the bodily accumulation of the organic species of Hg in particular (methyl-Hg), exposure to which is considered a modifiable health risk . The US National Research Council (NRC) recommends dietary exposure to no more than 0.1 µg/kg per day methyl-Hg, approximately equivalent to a blood Hg level of < 5.8 µg/L [169]. Recently published data from the US National Health and Nutrition Examination Survey (NHANES) reveal that Asian women have significantly higher blood Hg than other races surveyed [170], indicating that they are potentially at greater risk for reproductive toxicity from methyl-Hg. For many, consumption of As contaminated drinking water is an important source of exposure to the highly toxic inorganic As species as well as to Pb [171]. Whereas seafood exposes consumers mostly to the relatively innocuous organic species of As [172]. In addition, the widespread use of herbal health and complementary therapies in the USA [173] and elsewhere may also place some groups at an increased risk for reproductive toxicity [174, 175]. A recent case of Pb intoxication resulting from the use of a contaminated herbal infertility treatment underscores the potential significance of this source [176] .

Amassing evidence in vitro and in vivo demonstrates adverse reproductive outcomes associated with environmental exposures to toxic elements, including early pregnancy loss and decreased fecundity associated with Hg, Cd, Pb, and As exposures in women [3, 177]. Adverse female reproductive effects have been demonstrated rather clearly at high levels of exposure, such as those encountered in the workplace [178]. For example, Rowland et al. found reduced fertility outcomes among female dental assistants exposed to Hg vapor [179]. However, the effects of the lower, or “trace” doses frequently encountered in the environment are more controversial. Epidemiologic studies of populations lacking occupational exposure reported altered sex-steroid hormone economy [180], and have been associated with clinical female infertility [181–183], increased time to pregnancy [184, 185], and altered oocyte maturation [186, 187], oocyte fertilization [188], embryo development [189], and embryo implantation [190, 191] during IVF. In contrast, other studies do not support an association between pregnancy and female exposures to environmental levels of toxic elements [192–195]. These studies are preliminary though intriguing and warrant further investigation to better understand the potential harm on female reproductive potential caused by exposures to toxic elements widespread throughout the environment .

Using experimental systems in vitro and in vivo, investigators have identified several potential biologic mechanisms by which toxic elements might contribute to unexplained female infertility. Elements including As, Cd, Pb, and Hg enter eukaryotic cells [196] and in doing so can disrupt cytoskeletal function [197, 198], and increase oxidative damage by depletion of protective antioxidant molecules [199] and generation of reactive oxygen species [200]. Investigators also report estrogen-receptor interaction for each As, Cd, Pb, and Hg at concentrations similar to those to which humans are exposed through background sources and with physiologic effects downstream [201–204]. In addition, altered progesterone synthesis has been reported in association with Cd exposure by several studies employing in vitro granulosa cell culture models or in vivo murine models [205]. Disrupted progesterone synthesis has also been demonstrated in association with Hg treatment in vitro and in vivo, using fish oocytes [206]. Potentially inheritable genetic modifications involving gene regulation, rather than nucleic acid substitutions, or “epigenetic” changes have also been linked to toxic element exposure in experimental studies [207, 208] as well as in human observational studies [90]. Alterations in the expression of genes involved in cell replication, or the synthesis of factors required by an early embryo for navigating the early stages of uterine invasion and placentation could feasibly contribute to unexplained female infertility . While many potential biologic mechanisms exist, harmonization of the experimental and epidemiologic literature at environmentally relevant doses will require additional investigation so that the risk to female unexplained infertility can be more definitively assessed .

There is ever increasing evidence that environmental contaminant exposures may adversely affect human reproduction through effects on female reproductive potential. While cigarette smoking and its component chemicals are known toxic agents affecting the quality and quantity of oocytes, other toxic substances ubiquitous in our environment such as BPA, PCBs, dioxins , PBDEs, perfluorinated compounds , and toxic elements appear to have reproductive toxic effects at environmentally relevant levels as well. That said further studies are clearly needed to fully understand the impact of these various toxic environmental exposures on female as well as male infertility. Large, prospective epidemiologic-based studies are needed to address potentially subtle effects of various environmental contaminants on reproductive health. Within the framework of such studies, we can realize and appropriately counsel the infertile female patient on best practices regarding environmental and lifestyle exposures to toxic substances.