Long-term use of oral contraceptives by women is associated with an increased risk of developing hepatic adenomas and carcinomas and thus represents an important public health problem. We and others have shown that: 1) ethinyl estradiol (EE) and mestranol are potent promoters and weak complete hepatocarcinogens in female and male rats; 2) both cause transient additive liver growth; and 3) EE and other estrogens are hepatic co-mitogens. Recently, we made a new observation that forms the basis of this renewal application. Upon chronic treatment with EE at non- hepatotoxic doses, a mitosuppressed state, characterized by reduced basal growth and decreased growth responsiveness, follows the initial transient period of growth. We hypothesize that initiated hepatocytes are differentially resistant to the growth suppressive effects of EE and that the mitosuppressed state is due to altered gene expression in the """"""""hyperplastic"""""""" livers. To explore this hypothesis we will: 1. Determine the levels of expression of TGF-beta and the mannose 6-phosphate receptor using immunohistochemical and northern blot analyses in order to compare the effects of EE with those of phenobarbital which has been shown to increase their expression; 2. Identify and clone gene products uniquely present or absent in EE mitosuppressed livers using the mRNA-PCR-based differential display technique. Clones whose differential expression is confirmed will then be characterized by sequencing and further analysis of their patterns of expression; and 3. Further characterize the mitosuppressed state in vivo with regard to its duration, its sensitivity to the extent of growth stimulation, and whether it is differentially expressed in non-initiated vs. initiated hepatocytes. Results from these studies will reveal the identity of gene products whose altered expression is associated with hepatic growth suppression and which may represent molecular biomarkers for promoters whose mechanisms of action involve the exertion of growth negative selective pressure. The results will also determine whether differential mitosuppression is involved in promotion by EE.
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