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Effects of estrogen on immature, ovariectomized mice: a multi-approach, tissue-by-tissue comparison. J.W. Burt1,3, L.D. Burgoon2,3, D. Humes1, A.S. Harney1, and T.R. Zacharewski1,3 1Department of Biochemistry & Molecular Biology, 2Department of Pharmacology & Toxicology, and 3Center for Integrative Toxicology, National Food Safety & Toxicology Center, Michigan State University, East Lansing, MI 48912 Estrogen is a key hormonal regulator of cell growth, differentiation and functions across a wide array of tissues which include the female and male reproductive tracts, and the skeletal and cardiovascular systems. Gene expression and histological experiments were run in parallel to compare and contrast changes in the uterus, mammary gland, liver, femur and kidney in response to 17a-ethynyl estradiol (EE) in immature, ovariectomized C57BL/6 mice. Mice were gavaged with 100ug/kg of EE or sesame seed oil and organs were harvested at 2, 4, 8, 12, 18, 24 hours or dosed once daily on 3 consecutive days and then organs harvested at 72 hours after the initial dose. cDNA microarrays containing 13361 mouse clones (7810 unique, annotated genes) were used to compare gene expression profiles of EE treated animals to time-matched, vehicle controls. An independent reference design of 3 biological replicates was used incorporating two independent, dye-swapped labelings per sample. Data were normalized using a semiparametic approach, and gene activity assessed using an empirical Bayes method to compare treated from vehicle responses per time-point. Active gene lists were compiled for each organ based upon filtering by P1(t) values. Comparisons of active gene lists across tissues indicate that estrogen regulates common biological processes (i.e. proliferation and xenobiotic metabolism), in addition to tissue-specific responses (i.e. lipid metabolism in the liver). Histological analysis was performed on organs from a parallel EE time course study, which allows for phenotypic anchoring of the observed gene expression changes. In the most dramatic change, the uterotropic response was seen as a 7.5 fold increase in uterine blotted weight at 72 hours, an increase in the luminal epithelial cell height and an increase in the number of BrdU stained cells at 18 and 24 hours. These physiological and morphological changes could be correlated with observed mRNA expression changes of growth factors, proto-oncogenes and cell cycle and proliferation genes. Work Supported by NIEHS ES011271 and ES07255-16 |
