Abstracts: ISEE 22nd Annual Conference, Seoul, Korea, 28 August-1 September 2010: Reproductive Health and Environment
Epigenetic changes disrupting the regulation of gene expression are a common process in the development of cancer. Testicular germ cell tumor (TGCT) is the most common malignancy in young males, and its incidence has been rising in recent years. Epigenetic mechanisms have been known to be associated with gene silencing in TGCT. DNA hypermethylation can cause gene silencing, including the inactivation of tumor suppressor genes. Many tumor suppressor genes have been found to be silenced in TGCT following DNA hypermethylation. Phthalates are endocrine disruptors and are abundantly used as industrial plasticizers. Di-2-(ethylhexyl) phthalate (DEHP) is the most common phthalate found in consumer products and has been reported to be associated with hypermethylation. After exposure, Di-2-(ethylhexyl) phthalate is rapidly hydrolyzed into its active form, mono-(2-ethylhexyl) phthalate (MEHP), a testicular toxicant. The purpose of this study is to examine gene promoter-region methylation in testicular cells and assess the relationship between exposure to phthalates and expression and methylation status of tumor suppressor genes: Testisin and RASSF1A. Alteration of gene expression of xenobiotic metabolizing enzymes is also known to be associated with TGCT, thus genes, such as GSTP1 and CYP1A2, will also be studied.
Promoter hypermethylation and gene expression was examined for each genes in testicular germ cells exposed to MEHP in a dose- and time-dependent manner at concentrations of 10 nM, 1 μM, 10 μM, 100 μM, and 1 mM at 24-, 48,- 72-, and 96-hour points, respectively. The methylation status of the tumor suppressor genes Testisin, RASSF1A, and xenbiotic metabolizing enzyme genes: GSTP1 and CYP1A2 were assessed by methylation specific (MSP)-PCR and gene expression by qRT-PCR.
Exposures inversely increased cell proliferation, cell viability and methylation, and decreased gene expression. The results indicative of an association between DNA methylation and gene expression following exposure to phthalate will be presented and discussed.
The findings of this study will allow for a better understanding of the role phthalates play in DNA methylation and the subsequent expression of methylated genes in testicular germ cells.