Humans and other species have suffered adverse health effects thought to result from exposure to environmental chemicals that interact with the endocrine system. A significant portion of these effects have been manifested by decreased reproductive potential - often through their effects in utero leading to abnormal development of the reproductive tract. In humans and rodents these effects have been clearly demonstrated with diethylstilbestrol (DES). HOX genes are highly evolutionarily conserved and impart segmental tissue identity during the development of undifferentiated body axes. We have demonstrated that HOX genes play an essential role in reproductive tract development and have shown that expression of HOX genes in the reproductive tract is modulated by sex steroids. We have also recently shown that HOX gene expression is altered after DES exposure in utero. We hypothesize that HOX gene expression may be altered by estrogen- line endocrine disruptors in a manner analogous to the way in which they are altered by DES. The objective of this proposal is to test the hypothesis that an important mechanism by which endocrine disruptors effect the development of the reproductive tract is by altering the expression of HOX genes. We will approach this by assessing their effects on HOX gene expression both in vitro and in vivo. Both a mouse model and human cell culture model will be used; HOX gene expression in these systems has been well characterized in our laboratory. First we will identify alterations of HOX gene expression in human uterine and cervical cell culture following exposure to endocrine disruptors. Second, we will correlate these finding with altered Hox gene expression patterns in the developing and adult mouse reproductive tract following in utero exposure to these chemicals. Finally, we will determine if the molecular mechanism by which endocrine disruptors alter HOX gene expression is through interference with the estrogen receptor binding to the estrogen response elements which we have identified in the 5' flanking regions of HOX genes. The functional importance of these interactions will be verified using reporter genes linked to each HOX regulatory element. Localization of the site of endocrine disruption to the estrogen response element will be confirmed by mutational and deletional analysis. We have previously shown that Hox genes are necessary for reproductive tract development in the embryo and for reproductive function in the adult; altered Hox expression leads to developmental or functional alterations. Here we expect to demonstrate that an important mechanism of endocrine disruptor action on the reproductive tract is via the alteration of HOX gene expression. No good model exists to explain the mechanism of, and to rapidly detect the trans-generational effects of endocrine disruptors following exposure in utero. An understanding of the molecular mechanisms by which endocrine disruptors effect reproductive potential will provide a powerful tool to assess the risk of, diagnose, and perhaps lead to the treatment of he effects of endocrine disruptors.
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