罗格列酮对肺成纤维细胞TGF-β1信号转导途径的干预作用

The Effect of Rosiglitazone on Signal Pathways of Transforming Growth Factor-β1 in Human Lung Fibroblast

目的研究转化生长因子-β1(TGF-β1)对人肺成纤维细胞结缔组织生长因子(CTGF)、CTGF mRNA、核因子-κB(NF-κB)及激活子蛋白-1(AP-1)表达的影响及其罗格列酮对肺成纤维细胞TGF-β1信号转导途径的影响及可能机制。方法以人肺成纤维细胞系(HLF-02)为研究对象,用Western blot法检测TGF-β1、姜黄素、吡咯烷二硫基甲酸盐(PDTC)、罗格列酮作用后的CTGF、NF-κB及AP-1蛋白的表达,免疫组织化学方法检测CTGF蛋白、RT-PCR技术测CTGF mRNA的表达。结果①与对照组比较,1ng/mLTGF-β1作用15min后,CTGF表达水平即增强(P<0.01),并随着作用时间延长而逐渐增强。CTGF mRNA的表达亦随着时间的延长逐渐增加,作用24h表达最多。②1ng/mLTGF-β1作用30min后,NF-κB及AP-1的表达水平增强(P<0.01),以后表达趋于稳定,抑制剂(PDTC100μmol/L、姜黄素50μmol/L)抑制这两条通路后CTGF的表达明显下降。③低(5μmol/L)、中(10μmol/L)、高(15μmol/L)浓度罗格列酮预处理细胞30min+TGF-β1作用24h组较单纯TGF-β1处理24h组CTGF、NF-κB及AP-1蛋白表达明显降低(P<0.01),并且可以明显抑制1ng/mLTGF-β1作用24h引起的CTGF mRNA的表达(P<0.01)。结论在体外,罗格列酮可通过NF-κB及AP-1信号转导途径启动人肺成纤维细胞过氧化物酶体增殖物激活受体γ(PPAR-γ)抑制TGF-β1诱导CTGF转录和表达。

Objective To investigate the expression of CTGF, CTGF mRNA, NF-κB and AP-1 induced by TGF-β1 in human lung fibroblast （HLF-02）, and study the effect and possible mechanism of rosiglitazone on signal pathways of TGF-β1 in HLF-02. Methods The effects of TGF-β1, eurcumin, PDTC and rosiglitazone on the expression of CTGF, NF-κB and AP-1 were evaluated with Western blot. The expression of CTGF was also detected with immunohistochemistry assay. The level of CTGF mRNA was detected with RT-PCR. Results （1） The CTGF protein levels of HLF-02 cells were significantly up-regulated after incubated with 1 ng/mL TGF-β1 for 15 rain （P〈0.01 vs Control）. The CTGF mRNA was also up-regulated. The levels of CTGF protein and CTGF mRNA expression was up-regulated by TGF-β1 in a time-dependent manner. （2） The expression of NF-κB and AP-1 increased after HLF-02 cells were incubated with 1 ng/mL TGF-β1 for 30 min （P〈0. 01 vs Control）. The CTGF protein levels were inhibited obviously after HLF-02 cells were incubated with PDTC or curcumin. （3） The expression of CTGF, NF-κB and AP-1 decreased after pre-incubation with different doses of rosiglitazone （P〈0. 01 vs TGF-β1 group）. The CTGF mRNA were also markedly inhibited （P〈0.01 vs TGF-β1 group）. Conclusion It is supposed that rosiglitazone inhibits CTGF expression induced by TGF-β1 in HLF-02 cells by activating PPARγ through NF-κB and AP-1 signal transduction pathways.