Transcriptional inhibition of intestinal NHE8 expression by glucocorticoids involves Pax5.
Transcriptional inhibition of intestinal NHE8 expression by glucocorticoids involves Pax5.
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DOI：
10.1152/ajpgi.00227.2010
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发表时间：
2010-07
期刊：
影响因子：
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通讯作者：
Hua Xu;Bo Zhang;Jing Li;Huacong Chen;Chunhui Wang;F. Ghishan
中科院分区：
文献类型：
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作者：
Hua Xu;Bo Zhang;Jing Li;Huacong Chen;Chunhui Wang;F. Ghishan
钠/氢交换器(NHEs)是一类蛋白质，通过将质子移出细胞，将钠离子输送到细胞内。它们在钠吸收、细胞体积调节和细胞内pH调节中起着重要作用。在已鉴定的九种NHEs中，有三种(NHE2、NHE3和NHE8)表达在肠上皮细胞的顶膜上。糖皮质激素已被发现在肠道中调节NHE3的功能，但它们是否对NHE8的表达具有类似的功能尚不清楚。有趣的是，肠道中高糖皮质激素水平与NHE8高表达到NHE3高表达的时间变化相一致。研究旨在探讨糖皮质激素在肠道成熟过程中对NHE8表达的影响。分离肠上皮细胞刷状缘膜泡，用免疫印迹法检测甲基强的松龙对雄性乳鼠NHE8蛋白表达的影响。实时定量聚合酶链式反应(Real-Time-PCR)定量检测大鼠和Caco-2细胞中NHE8mRNA的表达。以Caco-2细胞为靶细胞，检测人NHE8启动子的活性。凝胶迁移率改变分析(GMSA)用于鉴定参与糖皮质激素调节的启动子序列和转录因子。结果表明，糖皮质激素处理的大鼠和人肠上皮细胞(Caco-2)NHE8mRNA和蛋白的表达均降低。糖皮质激素处理后，人NHE8基因启动子在Caco-2细胞中的活性也降低。GMSA表明，糖皮质激素存在时启动子活性的降低是由于转录因子Pax5与NHE8近端启动子区域结合增强所致。综上所述，本研究表明糖皮质激素通过增加Pax5与NHE8基因启动子的结合来抑制NHE8基因的表达，提示Pax5在肠道成熟过程中起重要作用。
Sodium/hydrogen exchangers (NHEs) are a family of proteins that transport sodium ions into the cells by moving protons out of the cells. They play a major role in sodium absorption, cell volume regulation, and intracellular pH regulation. Three out of nine identified NHEs (NHE2, NHE3, and NHE8) are expressed on the apical membrane of intestinal epithelial cells. Glucocorticoids have been found to regulate NHE3 function in the intestine, but it is unknown if they have a similar function on NHE8 expression. Interestingly, high glucocorticoid levels in the intestine coincide chronologically with the change from high expression of NHE8 to high expression of NHE3. Studies were performed to explore the role of glucocorticoids on NHE8 expression during intestinal maturation. Brush-border membrane vesicles were isolated from intestinal epithelia, and Western blotting was performed to determine NHE8 protein expression of suckling male rats treated with methylpredisolone. Real-time PCR was used to quantitate NHE8 mRNA expression in rats and Caco-2 cells. Human NHE8 promoter activity was characterized through transfection of Caco-2 cells. Gel mobility shift assays (GMSAs) were used to identify the promoter sequences and the transcription factors involved in glucocorticoid-mediated regulation. Our results showed that the expression of NHE8 mRNA and protein was decreased in glucocorticoid-treated rats and human intestinal epithelial cells (Caco-2). The activity of the human NHE8 gene promoter transfected in Caco-2 cells was also reduced by glucocorticoid treatment. GMSAs suggested that the reduction in promoter activity in the presence of glucocorticoids was due to enhanced transcription factor Pax5 binding on the NHE8 proximal promoter region. In conclusion, this study showed that glucocorticoids inhibit NHE8 gene expression by increasing Pax5 binding on NHE8 gene promoter, suggesting an important role for Pax5 during intestinal maturation.