Involvement of testicular growth factors in fetal Leydig cell aggregation after exposure to phthalate in utero

Abstract

Exposures to di-(2-ethylhexyl) phthalate (DEHP) have been shown to be associated with decreased adult testosterone (T) levels and increased Leydig cell numbers. As yet, little is known about DEHP effects in utero on fetal Leydig cells (FLC). The present study investigated effects of DEHP on FLC function. Pregnant Long-Evans female rats received vehicle (corn oil) or DEHP at 10, 100, or 750 mg/kg by oral gavage from gestational day (GD)2-20. At GD21, T production, FLC numbers and distribution, and testicular gene expression were examined. The percentage of FLC clusters containing 6 -30 cells increased in all treatment groups, with 29 ؎ 2% in control vs. 37 ؎ 3, 35 ؎ 3, and 56 ؎ 4% in rats receiving 10, 100, and 750 mg/kg DEHP, respectively. In contrast, FLC numbers were 33% and 39% lower than control after exposures to 100 and 750 mg/kg DEHP, respectively. At these doses, mRNA levels of leukemia inhibitory factor (LIF) increased. LIF was found to induce cell aggregation in FLCs in vitro, consistent with the hypothesis that DEHP induced FLC aggregation. Testicular T levels were doubled by the 10 mg/kg dose and halved at 750 mg/kg. The mRNA levels of IGF-1 and c-Kit ligand (KITL) were induced by 10 mg/kg DEHP. These results, taken together, indicate that fetal exposures to DEHP have effects on FLC number, distribution, and most importantly, steroidogenic capacity and suggest that abnormal expressions of IGF1, KITL, and LIF genes may contribute to the reproductive toxicity of phthalates.di-(2-ethylhexyl) phthalate ͉ testosterone ͉ reproduction ͉ endocrine disruptor ͉ steroidogenesis P hthalates, widely used as plasticizers and solvents, are commonly found in a variety of consumer products including cosmetics, toys, medical tubing, and catheters and in the environment as an industrial waste product. Increasing public concern over lack of regulation on their use in the United States, in contrast to the European Union and 14 other countries (1), has arisen in response to reports that exposures to phthalates may be linked to abnormal reproductive development in the human male (2, 3). Epidemiological studies show statistical correlations between serum concentrations of phthalate monoesters, the primary metabolites of phthalates, and the incidence of anomalies such as cryptorchidism and shortened anogenital distance (AGD) (4, 5). Di-(2-ethylhexyl) phthalate (DEHP), the most abundant phthalate in the environment, has been shown to have adverse effects on androgen synthesis in the rodent (6).The Agency for Toxic Substances and Disease Registry reported that, although exposure to DEHP is generally low, the exposures of preterm infants can be as high as 10-20 mg per day (7). Controversy exists over whether DEHP, at the levels found in the environment, is harmful to humans, because most studies have been conducted in rodents administered high doses. In previous studies, we showed that the administration of low-dose (10 mg/kg body weight) DEHP for 28 days during pubertal development caused elevations in testosterone (T) (8, 9)....

Novel molecular targets associated with testicular dysgenesis induced by gestational exposure to diethylhexyl phthalate in the rat: a role for estradiol

“…We observed no decrease across the DEHP exposures; a slight decrease at 100 mg was suggested. This is consistent with previous results reporting no decrease in expression of P450scc protein (Lehmann et al 2004) or its activity (Lin et al 2008) until at least a 500 mg exposure was reached. Obviously alterations in gene expression occur at lower exposures than those resulting in proteomic changes.…”

“…Previous work suggested that clustering of fetal rat Leydig cells occurs following in utero exposure to 10 mg DEHP/kg from GD2-GD20 (Lin et al 2008). Testosterone content in the fetal testis was not altered until a 750 mg/kg exposure was reached.…”

“…!100 mg/kg) were used. Lin et al (2008) observed Leydig cell clusters at 10 and 100 mg and the Leydig cells were decreased in both size and number. As pre-term infants can be exposed to 10-20 mg/day (Agency for Toxic Substances and Disease Registry 2006), we wanted to more carefully evaluate the dose response for the 10-100 mg/kg range.…”

“…Except for one study in which pregnant rats were exposed to doses 10, 100, and 750 mg DEHP/kg (Lin et al 2008), there have been few in vivo studies of phthalates where low doses (i.e. !100 mg/kg) were used.…”

The DLK1-MEG3 locus in malignant cells of proposed primordial germ cell origins.

“…for the testes, leydig cells also secrete other hormones, including IGF1 and testosterone [148][149][150]. This indicates that, like many other cancers, exogenous growth factors in serum are likely a major source of growth stimulation for TGCTs such as EC.…”