The overall objective of this proposal is to determine the mechanisms by which phthalate esters downregulate testosterone production in the developing fetal rat testes, resulting in antiandrogenic effects on the developing male reproductive tract. Phthalate esters are a class of environmental chemicals to which humans are ubiquitously exposed and which cause antiandrogenic effects on the developing male reproductive tract in rats. The mechanisms by which phthalate esters cause their effects remain to be determined. In preliminary studies, global changes in gene expression in the developing rat testis following in utero exposure to di(n-butyl) phthalate (DBP) were examined. A significant finding was that DBP produced a reduction in key genes in pathways associated with either steroid production or the provision of substrates for this activity. It is therefore hypothesized that phthalates decrease testosterone in the developing testes because of a coordinate disruption in cholesterol transport and steroid biosynthesis. This hypothesis will be tested by investigating the following specific aims: 1) Identify the key steps involved in cholesterol transport and testosterone biosynthesis that are targets for disruption by in utero exposure to DBP in the developing fetal rat testes, 2) Determine the consequences of DBP-induced changes in target gene and protein expression identified in Specific Aim 1 on fetal testicular cell signaling pathways, cholesterol transport, and steroidogenesis, and 3) Establish the molecular mechanism by which DBP downregulates expression of genes involved in cholesterol transport and steroid biosynthesis. Quantitative RT-PCR and protein analysis will be used to identify gene targets in the developing fetal testis following in utero exposure to DBP. The promoter regions of selected gene target will be further examined in vitro to determine the mechanism by which phthalate esters act on cholesterol transport and steroidogenesis. The proposed studies will identify critical genes and pathways associated with cholesterol transport that are targets for DBP in the male rat in utero and will aid in determining potential human risks from exposure to this class of environmental chemicals. ? ?
