Quantitative proteomics reveals the role of ERbeta in late pregnant rat uteri

• Main Authors: Guan-yuan Chen, 陳冠元
• Other Authors: Mei-ling Tsai
• Format: Others
• Language: en_US
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/68842913064067559136

碩士 === 國立成功大學 === 生理學研究所 === 96 === To accommodate the growing embryo, uterine remodeling occurs. During late-gestation, uterine enlargement is associated with wall thinning and uterine quiescence in late pregnancy. Correspondingly, the estrogen level continuously rises to the maximal level in late-gestation. Since the expression of two estrogen receptors (ERα and ERβ) is gestation-dependent, the purpose of this study was to explore the possible role of ERβ in pregnant uteri. Our first objective was to establish a database of uterine proteome by subcellular fractionation coupled proteomic analysis of rat uteri on gestation day 18 (G18). The list of uterine proteins from late-gestation uteri allowed us to hypothesize that alteration in the abundance of proteins related to cytokskeleton, extracellular matrix (ECM), and PKA-mediated pathways contributed to uterine remodeling. Western blotting analysis showed that expression of SMMHC, α actin, β actin, nucleolin, α2-macroglobulin, and Bcl-xL were higher in G18 uteri than in G7. The expression of fibronectin, collagen VI, TH, PKAIIRα and prohibitin was lower in G18 uteri than in G7. The expression of integrinβ1, VDAC-1, Ras, and Src was indifferent between G18 and G7 uteri. Contraction analysis was to further examine the influence of cAMP on uterine contractions. These data suggested the presence of cellular hypertrophy and wall thinning with ECM degradation and ROS-induced apoptosis. Sympathetic denervation in late pregnant uteri may increase the sensitivity of late-gestation uteri to cAMP, which contributes to uterine quiescence. The second objective was to quantify the proteins modulated by genistein, an ER�� activator. Higher expression of ERβ in G18 than in G7 suggests that the estrogenic action in the late pregnancy was mainly determined by ERβ. Repeated quantitative proteomic approaches had identified 159 proteins in 14 bands. The list of the proteins modulated by genistein allowed us to hypothesize that genistein caused uterine remodeling by disturbing the balance between proteases and protease inhibitors. The imbalance may contribute to the reduction of ECM and cytoskeletal proteins. Western blotting showed the reduction of protease inhibitors, such as ��2-macroglobulin. Correspondingly, the expression of collagen VI, fibronectin, Bcl-xL, α actin and α tubulin were decreased by genistein at 10-9M. Even cell survival and uterine contraction were suppressed by genistein. Both Western blotting and functional assays supported our hypothesis. In conclusion, although ERβ is highly expressed in pregnant uteri, its role in pregnant uteri is largely unknown. This study provides an insight that ERβ plays an important role in morphological and functional remodeling of pregnant uteri by shifting the balance between proteases and protease inhibitors. The novel role of ER�� may serve as an alternative remedy for preterm labor.