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Analysis of gene expression during uterine induction and regression in immature, ovariectomized rats following treatment with ethynyl estradiol. JC Kwekel1, 2, HA Dalgleish1 LD Burgoon2, 3,JR Harkema2,4 TR Zacharewski1, 2, 3. 1Dept of Biochemistry & Molecular Biology, 2Center for Integrative Toxicology, National Food Safety & Toxicology Center, 3Dept of Pharmacology & Toxicology, 4Dept of Pathobiology & Diagnostic Investigation, Michigan State University, East Lansing, MI, USA. Changes in uterine physiology, cell type morphology and gene expression were evaluated in a comprehensive time course study after oral administration of 100 µg/kg bw ethynyl estradiol (EE) to immature, ovariectomized SD rats. Animals were dosed once or once daily on 3 consecutive days and uteri were harvested 2, 4, 8, 12, 18, 24, 72, 84, 96, 120, 144, or168 hrs after treatment. Animals in the histological study received an i.p. injection of 5-bromo-2-deoxyuridine (BrdU, 50 mg/kg bw) 2 hours prior to sacrifice. Uterine wet weight, a classical marker of estrogen exposure, was maximally induced ~6-fold at 72 hrs. Global changes in gene expression were examined using custom rat cDNA microarrays consisting of 8,567 clones representing 5,818 unique genes. 1,755 genes exhibited a significant change at one or more time points as determined using a model-based t-statistic/Empirical Bayes approach. In order to phenotypically link these changes in gene expression to the physiological alterations observed in the uterotrophic response, complementary histological and morphometric assessment of uterine mid-horn sections stained with eosin and hematoxylin were performed. Additional histological slides were stained with anti-BrdU for assessments of proliferation. Pathological findings included indications of edema, eosinophil infiltration, hypertrophy, hyperplasia, angiogenesis and apoptosis consistent with the temporal changes in gene expression at time points preceding or overlapping these phenotypic markers. These assessments provide baseline data characterizing estrogen elicited responses in the rodent uterus, as a comparator for future estrogenic endocrine disruptor studies. Funding provided by NIH grant ES011271 and NIEHS Training Grant: ES07255-16 |
