Toxicology

The p53 Protein Influences the Sensitivity of Testicular Germ Cells to Mono-(2-Ethylhexyl) Phthalate-Induced Apoptosis by Increasing the Membrane Level of Fas and DR5 and Decreasing the Intracellular Amount of c-FLIP

Abstract
The consequence of mono-(2-ethylhexyl) phthalate (MEHP)-induced injury of testicular Sertoli cells is the Fas-dependent apoptotic elimination of germ cells. In addition to p53's well-known ability to regulate the transcription of various apoptotis-associated proteins, p53 has also been implicated in mediating the localization of Fas to the plasma membrane of various cell types in a transcription-independent manner. To resolve p53's role in MEHP-mediated testicular toxicity, we utilized wild-type (p53+/+) and p53 knockout (p53-/-) mice. A significantly lower incidence of TUNEL-positive germ cells was observed in p53-/- mice as compared to p53+/+ mice at 1h, 1.5h, and 24h after MEHP exposure. In these same mice, an induction of Fas and death receptor 5 (DR5) in testicular membrane preparations was observed only in p53+/+ mice. Analyses of mRNA levels in testes of p53+/+ and p53-/- mice by RT-PCR revealed that increases in membrane levels of Fas occurred in the absence of their transcriptional up-regulation. Processing of procaspase-8 was only observed in MEHP-treated p53+/+ mice and this correlated with the observed incidence of germ cell apoptosis. Interestingly, the p53 status of mice also influenced the stability of c-FLIP (L), a caspase-8 inhibitory protein, that was measured at ~2-5 fold higher levels in p53-/- mice after MEHP-exposure as compared to p53+/+ mice. Taken together, these data suggest that MEHP-induced germ cell apoptosis is dependent, in part, on the p53 protein and on its ability to increase the localization of Fas and DR5 on the germ cell membrane as well to decrease the cellular levels of c-FLIP (L).

Key words: Testis • Toxicology • Apoptosis • Sertoli cells • Signal transduction

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