RNA干扰维生素D受体基因表达对HK-2细胞中钠/二羧酸协同转运蛋白1表达的影响  

RNA interferes with the expression of vitamin D receptor gene and affects the expression of NaDC1 in HK-2 cells

作　　者：郭东群 李颢[1] 李康健[1] 申吉泓[1] 张白羽[1] 柯坤彬[1] 罗钰辉[1] GUO Dongqun;LI Hao;LI Kangjian;SHEN Jihong;ZHANG Baiyu;KE Kunbin;LUO Yuhui(Department of Urology,the First affiliated Hospital of Kunming Medical University,Kunming 650032,China)

机构地区：[1]昆明医科大学第一附属医院泌尿外科,昆明650032

出　　处：《实用医学杂志》2020年第4期451-455,共5页The Journal of Practical Medicine

基　　金：云南省科技计划项目[编号:2017FE468(-029)];国家自然科学基金项目(编号:81460141);云南省高层次卫生技术人才项目(编号:D-2017043)。

摘　　要：目的细胞水平上检测干扰维生素D受体(vitamin D receptor,VDR)基因表达对钠/二羧酸协同转运蛋白1(Na(+)-dicarboxylate cotransporter,NaDC)在HK-2细胞中的表达的影响,探讨VDR与NaDC1在尿枸橼酸调控中的关系。方法设计6条VDR基因的siRNA,用脂质体法转染人肾近曲小管上皮HK-2细胞株,通过qPCR和Western Blot检测HK-2细胞VDR m RNA和蛋白质的表达水平,筛选有效的siRNA,用以转染HK-2细胞株,通过qPCR和WB检测细胞株中NaDC1 m RNA和蛋白的表达水平。结果siRNA-2、siRNA-3和siRNA-4能够显著降低VDRmRNA和蛋白质表达水平(P<0.05),用以后续实验发现siRNA-2、siRNA-3和siRNA-4组NaDC1的m RNA和蛋白质表达水平显著降低(P<0.05)。结论在HK-2细胞中,在一定表达水平范围内,VDR调控NaDC1的表达,两者表达水平呈正相关。由于VDR的表达上调引起了NaDC1的表达也相应上调,从而使NaDC1的转运活性增加,尿枸橼酸在肾近曲小管的重吸收也增加,表现为低枸橼酸尿症。Objective To detect the effect of interfering VDR gene expression on the expression of NaDC1 in HK-2 cells at the cellular level,and to explore the relationship between VDR and NaDCl in the regulation of urinary citrate.Methods Six siRNAs of VDR gene"were designed and transfected into HK-2 cell line of human renal proximal convoluted tubular epithelium by liposome method.The expression levels of VDR gene and protein in HK-2 cells were detected by real-time polymerase chain reaction(qPCR)and Western Blot(WB)to screen effective siRNA.To transfect HK-2 cell line,the expression levels of NaDCl mRNA and protein in HK-2 cell line were detected by qPCR and WB.Results SiRNA-2,siRNA-3 and siRNA-4 could significantly reduce the expression of VDR and protein(P<0.05).It-was found that the expression of NaDCl in siRNA-2,siRNA-3 and siRNA-4 groups decreased significantly(P<0.05).Conclusions In HK-2 cells,within a certain range of expression levels,VDR regulated the expression of NaDCl,and the expression levels"were positively correlated.The upregulation of VDR caused the expression of NaDCl to be up-regulated accordingly,which increased the transport activity of NaDCl,and the reabsorption of urinary citrate in renal proximal tubules was also increased,manifested as hypocitrateuria.

关 键 词：低枸橼酸尿症 维生素D受体 钠/二羧酸协同转运蛋白1 RNA干扰 

分 类 号：R34[医药卫生—基础医学]