Enhancement of epithelial sodium channel expression in renal cortical collecting ducts cells by advanced glycation end products
Enhancement of epithelial sodium channel expression in renal cortical collecting ducts cells by advanced glycation end products
复制标题
DOI：
10.1093/ndt/gfl668
复制
发表时间：
2007-03-01
影响因子：
6.1
通讯作者：
Yang, Chih-Wei
中科院分区：
文献类型：
--
作者：
Chang, Chiz-Tzung;Wu, Mai-Szu;Yang, Chih-Wei
背景资料。上皮性钠通道(ENaC)是一种复合体，ENaCα亚基在醛固酮诱导的远端肾单位钠摄取中起着至关重要的作用。尽管实验性糖尿病动物模型已证实肾皮质集合管细胞αENaC表达上调，但其分子机制尚不清楚。晚期糖基化终产物(AGEs)是长期高血糖的副产物，是糖尿病肾病的重要致病因素。我们假设AGEs在调节αENaC基因表达中起作用。将小鼠CD细胞(mpkCCDCL(4))与AGE共同培养，观察AGE对α-ENaC表达和钠摄取的影响。实时定量聚合酶链式反应检测ENaC基因表达，荧光染料钠指示剂(SBFI-AM)测定钠摄取。本研究通过Western blotting分析了丝裂原活化蛋白激酶的信号通路。将携带荧光素酶报告基因的α-ENaC启动子与表达野生型或突变型血清和糖皮质激素诱导的激酶1(SGK1)基因的质粒共转染细胞，用AGE刺激细胞，以确定信号转导途径。AGEs以时间和剂量依赖的方式刺激mpkCCDCL(4)细胞，促进αENaC mRNA表达和钠摄取。AGEs还以时间和剂量依赖的方式显著刺激SGK1mRNA和SGK1活性。共转染突变型SGK1的mpkCCDCL(4)细胞可明显抑制AGEs激活的α-ENaC启动子驱动的荧光素酶活性。实验结果表明，AGEs可诱导肾小管上皮细胞表达α-ENaC，增加细胞对钠的摄取。AGEs激活αENaC表达的机制可能是通过激活mpkCCDCL(4)细胞中的SGK1。
Background. The epithelial sodium channel (ENaC) is a complex, and the alpha ENaC subunit has a crucial role in sodium uptake induced by aldosterone in the distal nephron. Although experimental animal models of diabetes have demonstrated up-regulation of alpha ENaC expression in renal cortical collecting duct (CCD) cells, the molecular mechanism remains unclear. Advanced glycation end products (AGEs) are by-products of long-term hyperglycaemia and comprise a significant pathogenic factor in diabetic nephropathy. We hypothesize that AGEs play a role in regulating alpha ENaC gene expression.Methods. Mouse CCD cells (mpkCCDcl(4)) were cultured with AGE to determine the effects of AGE on alpha ENaC expression and sodium uptake. Gene expressions of ENaC were measured by real-time PCR and sodium uptake was measured with fluorescent dye as a sodium indicator (SBFI-AM). This study analysed mitogen-activated protein kinases signalling pathways by western blotting. Cells co-transfected with plasmids of the alpha ENaC promoter carrying a luciferase reporter and plasmids expressing wild-type or mutant serum- and glucocorticoid-induced kinase 1 (Sgk1) mRNA were stimulated with AGE to identify the signalling pathway.Results. The AGEs, stimulated in a time- and dose-dependent manner, enhanced alpha ENaC mRNA expression and sodium uptake in mpkCCDcl(4) cells. The AGEs also significantly stimulated Sgk1 mRNA and Sgk1 activity in a time- and dose-dependent manner. Co-transfected with plasmid expressing mutant Sgk1 significantly limited stimulated alpha ENaC promoter-driven luciferase activity by AGEs in mpkCCDcl(4) cells.Conclusion. Experimental results indicate that AGEs induced alpha ENaC expression and increased sodium uptake in renal CCD cells. The mechanism through which AGEs activate alpha ENaC expression may be via activation of Sgk1 in mpkCCDcl(4) cells.