Embryonic Treatment with Di-(2-ethylhexyl) Phthalate Alters the Methylation Status of Sperm DNA in F3 Generation Mouse Offspring.

Plastics are a pervasive part of our everyday lives. Phthalate plasticizers, a group of two major chemicals in plastics, provide flexibility and transparency to plastics. Di-(2-ethylhexyl) phthalate (DEHP) is the most abundant phthalate with production reaching 2 million tons annually. DEHP and other phthalates are not covalently bound to plastics and thus they readily leach into the environment and are taken up daily by mammals through inhalation, digestion, and dermal contact. Evidence has linked human phthalate exposure to reduced gestational age, low birth weight, abnormal development, and abnormal reproductive and hormonal effects. More recently, we have shown that embryonic exposure of pregnant dams to DEHP resulted in disruption of testicular tubule morphology and spermatogonial stem cell proliferation in F3 generation offspring. In this study, we set out to determine if these observed transgenerational disruptions could be caused by changes in the DNA methylation pattern in sperm DNA from F3 generation offspring of DEHP-exposed dams. Time pregnant CD1 mouse dams were treated with DEHP at embryonic days E7-E14 and the subsequent F1 generation offspring were used to generate F2 and F3 generation offspring. To prepare DNA for promoter tiling array, caudal epididymal sperm from F3 generation offspring was collected for the DEHP-treated and control groups, followed by Methylated DNA Immunoprecipitation (MeDIP) enrichment and subsequent amplification. The tiling array analysis indicated that DEHP alters the DNA methylation pattern, with altered genes involved in cell fate and commitment, histone modification, meiosis, spermatogenesis and cell-cell adhesion. These data indicate that observed transgenerational disruptions in testicular morphology and spermatogonial stem cell proliferation could be due to epigenetic changes induced by embryonic DEHP exposure. (poster)