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目的探讨1α,25-二羟基维生素D3对高糖刺激下大鼠肾小管上皮细胞中结缔组织生长因子(connective tissue growth factor,CTGF)表达的影响及其机制。方法将大鼠肾小管上皮细胞NRK52E传代培养。将传代培养后的细胞按是否加入干预剂分为3组:正常对照(A)组(D-葡萄糖6μmol·L~(-1))、高糖(B)组(D-葡萄糖30μmol·L~(-1))和1α,25-二羟基维生素D3干预(C)组(1α,25-二羟基维生素D3 5nmol·L~(-1),预处理1h,再给予D-葡萄糖30μmol·L~(-1))。3组干预后再进行培养。将培养后的3组细胞按培养时间的不同分为3个亚组:Aa 12、24、48h3组,Bb 12、24、48h3组和Cc 12、24、48h3组。采用Real-time PCR检测各组CTGF mRNA的表达水平,采用Western blot检测各组CTGF蛋白、β-catenin蛋白的表达水平。结果与Aa 24h组比较,Bb 24h组CTGF mRNA、CTGF蛋白、β-catenin蛋白表达水平均升高(均P<0.01);与Bb 24h组比较,Cc 24h组CTGF mRNA、CTGF蛋白、β-catenin蛋白表达水平均降低(均P<0.01)。结论在高糖体外培养的大鼠肾小管上皮细胞中1α,25-二羟维生素D3能够显著降低高糖刺激的CTGF表达水平,其机制可能是阻断Wnt/β-catenin信号通路来实现的。
Objective To investigate the effect and mechanism of 1α, 25-dihydroxyvitamin D3 on the expression of connective tissue growth factor (CTGF) in rat renal tubular epithelial cells stimulated by high glucose. Methods Rat renal tubular epithelial cells NRK52E were subcultured. The subcultured cells were divided into 3 groups according to whether the interventional agent was added or not: normal control group (D-glucose 6μmol·L -1), high glucose group (D-glucose 30μmol·L -1) (-1) and 1α, 25-dihydroxyvitamin D3 intervention group C (1α, 25-dihydroxyvitamin D3 5nmol·L -1), pretreatment 1h, and then given D-glucose 30μmol·L ~ (-1)). 3 groups of intervention before further training. Three groups of cultured cells were divided into three sub-groups according to different culture time: Aa 12,24,48h3 group, Bb 12,24,48h3 group and Cc 12,24,48h3 group. The expression of CTGF mRNA in each group was detected by Real-time PCR. The expression of CTGF and β-catenin in each group was detected by Western blot. Results Compared with A24h group, the levels of CTGF mRNA, CTGF protein and β-catenin in Bb 24h group were significantly increased (all P <0.01). Compared with Bb 24h group, CTGF mRNA, CTGF protein, β-catenin The protein expression levels were decreased (all P <0.01). CONCLUSION: 1α and 25-dihydroxyvitamin D3 can significantly decrease the expression of CTGF in high glucose-induced rat renal tubular epithelial cells in vitro. The mechanism may be through blocking the Wnt / β-catenin signaling pathway.