TGF-β1对诱导肾间质成纤维细胞活化并表达MMP-9及TIMP-1的影响

Effects of TGF-β1 activating renal interstitial fibroblast cells on MMP-9 and TIMP-1 expression

  • 摘要: 目的 研究转化生长因子-β1(transforming growth factor beta-1,TGF-β1)诱导的肾间质成纤维细胞(NRK-49F)发生表型转化时基质金属降解酶MMP-9及其抑制因子TIMP-1表达的变化及对分泌纤维连接蛋白(fibronectin,FN)的影响。 方法 以不同浓度hTGF-β1(0 ng/ml、0.5 ng/ml、1 ng/ml、2 ng/ml、5 ng/ml、10 ng/ml)刺激NRK-49F细胞48 h,分别应用ELISA法检测细胞上清FN的浓度,免疫荧光检测细胞表型标记物α-SMA、Vimentin的表达,Western blot、Northern blotting方法检测MMP-9、TIMP-1的基因及蛋白质表达。 结果 0.5 ~ 10 ng/ml TGF-β1随刺激浓度增高NRK-49F细胞表达α-SMA明显增多、Vimentin明显减少,细胞上清中FN的含量显著增加(P< 0.05或P< 0.01);同时,TGF-β1浓度> 2 ng/ml时显著上调TIMP-1 mRNA及蛋白表达水平(P< 0.05或P< 0.01);以上效应均呈浓度依赖性,但对细胞MMP-9 mRNA及蛋白表达无明显影响(P> 0.05)。 结论 TGF-β1能诱导肾间质成纤维细胞活化,使FN的分泌增多,该作用可能与TGF-β1上调了TIMP-1基因及蛋白质水平,进而减少细胞外基质(extracellular matrix,ECM)的降解有关。

     

    Abstract: Objective To study the changes on MMP-9 and TMP-1 expression when renal interstitial fibroblast cells (NRK-49F)occurred phenotypic transform induced by transforming growth factor beta-1 (TGF-β1), and its effects on the secretion of fibronectin(FN). Methods Renal interstitial fi broblast cells (NRK-49F) of rats were stimulated by hTGF-β1 at different concentrations of 0 ng/ml, 0.5 ng/ml, 1 ng/ml, 2 ng/ml, 5ng/ml, 10 ng/ml. Immune fl uorescence was used to measure the expression of cell phenotype sign α-SMA and Vimentin, and ELISA was used to detect the concentration of FN in supernatant, while Western Blot and Northern Blotting technique was used to measure the expression of MMP-9 and TIMP-1. Results TGF-β1 at the concentration of 0.5 ng/ml -10 ng/ml induced the expression of α-SMA which represented myo fi broblast appearance, with increasing concentration of TGF-β1, the expression of α-SMA gradually increased and Vimentin decreased signi fi cantly, furthermore FN secretion were driven up greatly (P< 0.05 or P< 0.01), while TGF-β1 at the concentration of 2 ng/ml markly drove up the expression of TIMP-1 mRNA and protein(P< 0.05 or P< 0.01) , those effects showed concentration-dependent form. However, TGF-β1 at above concentration had no remarkable effect on MMP-9 mRNA and protein expression (P> 0.05). Conclusion TGF-β1 can induce renal interstitial fi broblast activation to be myo fi broblast, and promote fi bronectin secretion, whose effects may be related to up-regulation of TIMP-1 expression so as to restrain ECM decompounded.

     

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