**1.5.2 DDT and** *p,p* **′-DDE**

480 Toxicity and Drug Testing

compounds (i.e. vinclozolin, linuron, p,p′ DDE and phthalates etc.) and common

Fig. 3. Testicular dysgenesis syndrome. Both genetic and environmental factors affect

cell differentiation, apparent later as reduced semen quality or in the worst cases as carcinoma *in situ* (CIS) of the testis and consequent testicular cancer. (Modified from

Skakkeback et al. Human Reproduction, 2001).

**1.5.1 Polychlorinated Biphenyls (PCB)** 

testicular development and functions. Damage of the testicular cells (Leydig cells and Sertoli cells), disrupts androgen production from Leydig cells and secretion of paracrine factors from sertoli cells, leading to birth defects (hypospadias, cryptorchidism) and impaired germ

PCB make up a group of synthetic organic chemicals containing about 200 individual compounds. Some PCB bind to ERs and consequently PCB may exert their toxic effects through estrogenic activity (McKinney & Waller, 1998). Alternatively, some PCB may produce reproductive toxicity through the production of free radicals. Rats exposed to mixtures of PCB demonstrated decreased superoxide dismutase and catalase activity in the testes following exposure (Peltola et al., 1994). PCB congeners have different mechanisms of action and therefore different effects on biological systems. Longitudinal studies of children with in utero or lactational exposure to PCB and other environmental chemicals are essential to assess the long-term effects of endocrine disruption. Prenatal and lactational exposure to PCB may exert adverse effects on male reproduction during subsequent adulthood. A wellstudied population exposed to PCB/PCDF-contaminated rice oil (Yu-Cheng exposure;

environmental toxins reported to be involved in male reproductive toxicity.

The persistent pesticide, DDT, is broken down in the environment, and one of its metabolites is p,p′-DDE, which has been shown to act as an androgen receptor antagonist both in vivo and in vitro (Kelce et al., 1995). Studies in which p,p′-DDE was administered to rats during development (gestational day (GD)14–18; 100 mg/kg/day) affected androgendependent aspects of male development such that it reduced anogenital distance, caused nipple retention and, depending on the rat strain, induced hypospadias (You et al., 1998). Another DDT derivative, methoxychlor and its metabolites, have been shown to interact with both oestrogen receptors and the androgen receptor (AR). The methoxychlor metabolite, 1,1-Trichloro-2,2-bis (4 hydroxyphenyl) ethane, is an oestrogen receptor (ER)- α agonist, an ER-β antagonist and an androgen receptor (AR) antagonist (Gray et al., 2001). This illustrates that these chemicals may act by more than one mechanism to induce effects on the exposed population. Moreover, several population studies conducted to examine the effects of DDT and metabolite exposure on male reproductive health support the hypothesis that DDT exposure is related to reduced semen quality. Significantly higher seminal concentrations of *p,p*′-DDE were also reported in infertile patients compared to a fertile control group in India. Seminal fluid levels of fructose, γ-glutamyl transpeptidase, and acid phosphatase were positively correlated with *p,p*′-DDE concentrations in infertile men. The high concentration of fructose, a marker for seminal vesicle function and an important energy source for sperm, may indicate non-utilization of fructose by sperm. DDT exposure may be associated with abnormal metabolism in sperm, including decreased fuel utilization, in infertile men (Pant et al., 2004). Mexican men living in the areas where DDT was used for malaria control, but without occupational exposure to DDT, exhibited serum levels of *p,p*′- DDE approximately 350-fold greater than Canadian men exposed to background environmental levels. *p,p*′-DDE concentrations in the Mexican men were correlated with increases in SHBG concentration and negatively correlated with testosterone levels. Both semen volume and total sperm number were inversely correlated to *p,p'*-DDE levels. Thus, androgen levels and semen quality are adversely affected by high *p,p*′-DDE body burden (Ayotte et al., 2001). The studies examining DDT exposure and semen quality report consistent effects on sperm motility and sperm morphology, similar to the PCB studies. The increased SHBG concentrations associated with serum *p,p*′-DDE described by Ayotte et al.,(2001) provides a possible mechanism for observed reductions in plasma testosterone and sperm number. By inducing SHBG synthesis, *p,p*′-DDE may exert its antiandrogenic effects by reducing the amount of bioavailable testosterone, thereby impairing spermatogenesis.

Environmental Toxicants Induced

Male Reproductive Disorders: Identification and Mechanism of Action 483

compounds and only a few induce male reproductive tract abnormalities. Gray et al., *(*2000) compared the ability of six phthalate esters (diethylhexyl phthalate, DEHP; benzylbutyl phthalate, BBP; diisononyl phthalate, DINP; dimethyl phthalate, DMP; diethyl phthalate, DEP; dioctyl terephthalate, DOTP; all administered at 750 mg/kg body weight from GD14 to postnatal day (PND) 3) to induce malformations of the reproductive tract. This study assessed changes in many androgenic endpoints and found that only DEHP, BBP and to a lesser degree DINP induced alterations in all aspects of androgen-regulated male reproductive endpoints. Exposure to diethyl hexyl and dibutyl phthalates is associated with adverse effects on sperm motility (Fredricsson et al., 1993). Animal studies consistently demonstrated that phthalate esters are male reproductive toxicants (Park et al., 2002), with exposure associated with testicular atrophy, spermatogenetic cell loss, and damage to the Sertoli cell population. Phthalate monoesters target Sertoli cell functions in supporting the spermatogenesis process. This may be due to the effect of phthalates in reducing the ability of Sertoli cells to respond to FSH (Hauser et al., 2005). Initial reports on the effects of phthalates on male reproductive tract development focussed on the gross changes such as reduced anogenital distance, hypospadias, malformed epididymis and, in later studies, nipple retention (Mylchreest et al., 2000). Only a few studies give a more detailed account of the histological changes observed in the testis after in utero phthalate exposure and demonstrate that the fetal testis is directly affected by phthalates during fetal and neonatal testis differentiation (Parks et al., 2000; Fisher et al., 2003). Some of these alterations are permanent and affect the function of the testis in adult life and are similar to the histological changes which are now being shown in patients with testicur dysgenesis syndrome (TDS; Skakkebaek et al., 2003). The production of testosterone is critical for the normal masculinization of the male reproductive tract, as already discussed. It has been shown that DBP and DEHP are both capable of inhibiting the production of testosterone by the fetal testis (Parks et al., 2000; Fisher et al., 2003). Testosterone synthesis by the fetal testis is first detectable by GD15, reaches a peak at around GD18/19, and remains high until birth. However, phthalate treatment induces a 60–85% reduction in testosterone synthesis during this critical developmental window, reducing testosterone levels to a similar level to those found in females (Parks et al., 2000). This reduction in testosterone is a factor in the occurrence of hypospadias and cryptorchidism observed after phthalate treatment. This is not to suggest that phthalate exposure causes TDS in humans, merely that the administration of very high doses of DBP to pregnant rats induces a TDS-like syndrome in the male offspring that shows many analogous features to human TDS. It is plausible, given how highly conserved the pathways of fetal development are, that phthalate administration may disrupt some common mechanistic pathways which if altered in humans could be helpful in determining the pathogenesis of human TDS. In both the human syndrome and the rodent model, abnormal testicular development or dysgenesis is evident by the abnormal organisation of these tissues. In humans, histological evidence of testicular dysgenesis (immature seminiferous tubules with undifferentiated Sertoli cells, microcalcifications and Sertoli cell only (SCO) tubules, Leydig cell hyperplasia, morphologically distorted tubules and the presence of carcinoma *in situ* (CIS) cells) have been found in biopsies of the contralateral testes of testicular germ cell cancer patients and in biopsies from patients with infertility, hypospadias and cryptorchidism (Skakkebaek et al., 2003). These studies support the hypothesis that all of these disorders (low sperm counts,

Further, more extensive correlation studies on the serum levels of DDT and metabolites in mothers of infertile men are required. These studies would provide insight into the effects of in utero exposure to DDT on semen quality of adult males.
