高糖环境及雷帕霉素干预对肾小球足细胞Ⅳ型胶原和MMP-9表达的影响

Effects of high glucose and Rapamycin intervention on expression of type Ⅳ collagen and matrix metalloproteinase-9 in podocytes

目的探讨高糖培养环境及雷帕霉素干预对肾小球足细胞Ⅳ型胶原和基质金属蛋白酶9(MMP-9)表达的影响。方法体外培养的小鼠肾小球足细胞分为空白对照组、高渗对照组、高糖组和高糖+雷帕霉素干预组。Real-time PCR和Western blotting检测各组肾小球足细胞Ⅳ型胶原α3链(COL4A3)、α5链(COL4A5)mRNA及MMP-9mRNA和蛋白的表达;ELISA法检测细胞培养上清液中Ⅳ型胶原的含量。结果与对照组比较,高糖组足细胞COL4A3、COL4A5mRNA表达及培养上清液中Ⅳ型胶原含量均升高(P<0.05),而足细胞MMP-9mRNA和蛋白表达均降低(P<0.05)。与高糖组比较,雷帕霉素干预组足细胞COL4A3、COL4A5mRNA表达及培养上清液中Ⅳ型胶原含量均降低(P<0.05),足细胞MMP-9mRNA和蛋白表达均升高(P<0.05)。结论雷帕霉素对高糖环境中足细胞Ⅳ型胶原和MMP-9表达异常具有逆转作用。

Objective To investigate the effects of high glucose and Rapamycin intervention on the expression of type Ⅳ collagen and matrix metalloproteinase-9（MMP-9）in podocytes.Methods Mouse podocytes cultured in vitro were divided into blank control group,high permeability group,high glucose group and high glucose＋Rapamycin intervention group.The expression of type Ⅳ collagen α3 chain（COL4A3）mRNA,type Ⅳ collagen α5 chain（COL4A5）mRNA,MMP-9 mRNA and MMP-9 protein in podocytes of each group was detected by Real-time PCR and Western blotting,respectively.The contents of type Ⅳ collagen in the supernatants of culture fluid were determined by ELISA.Results Compared with control group,the expression of COL4A3 mRNA and COL4A5 mRNA in podocytes and the contents of type Ⅳ collagen in the supernatants of culture fluid in high glucose group were significantly increased（P0.05）,while the expression of MMP-9 mRNA and MMP-9 protein in podocytes was significantly decreased（P0.05）.Compared with high glucose group,the expression of COL4A3 mRNA and COL4A5 mRNA in podocytes and the contents of type Ⅳ collagen in the supernatants of culture fluid in high glucose＋Rapamycin intervention group were significantly decreased（P0.05）,while the expression of MMP-9 mRNA and MMP-9 protein in podocytes was significantly increased（P0.05）.ConclusionRapamycin could reverse the abnormal expression of type Ⅳ collagen and MMP-9 in podocytes treated with high glucose.