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目的探讨体外条件下转化生长因子(TGF)β1对人近端肾小管上皮细胞系(HK-2)表达结缔组织生长因子(CTGF)的诱导情况,以及罗格列酮对其影响和作用途径。方法应用RT-PCR方法检测CTGF和α-平滑肌肌动蛋白(SMA)mRNA表达。应用Western印迹方法检测CTGF和纤连蛋白(FN)的蛋白表达。结果(1)HK-2细胞低水平表达CTGFmRNA和蛋白,TGF-β1呈剂量和时间依赖性增加HK-2细胞CTGFmRNA和蛋白表达(P<0.01)。(2)罗格列酮和15脱氧前列腺素J2(15d-PGJ2)呈剂量依赖性抑制TGF—β1(4ng/ml)诱导的HK-2细胞CTGFmRNA和蛋白的表达。(3)过氧化物酶体增殖体激活受体(PPAR)γ特异性抑制剂GW9662(1μmol/L)完全阻断罗格列酮和15d—PGJ2对CTGFmRNA和蛋白表达的抑制作用。(4)与单纯TGF-β14ng/ml刺激组相比,罗格列酮5、10μmol/L处理组HK-2细胞FN蛋白分别下调39.9%和53.4%(P<0.01),并呈剂量依赖性(P<0.01),而GW9662完全阻断该作用。结论在体外,罗格列酮可通过激活肾小管上皮细胞PPARγ抑制TGF-β1诱导的CTGF转录和表达,同时也可抑制FN的表达。
Objective To investigate the effect of transforming growth factor (TGF) β1 on the expression of connective tissue growth factor (CTGF) in human proximal tubular epithelial cell line (HK-2) in vitro and its effect and mechanism of action. Methods RT-PCR was used to detect the mRNA expression of CTGF and α-smooth muscle actin (SMA). Western blotting was used to detect the protein expression of CTGF and fibronectin (FN). Results (1) CTGF mRNA and protein were down-regulated in HK-2 cells. TGF-β1 increased CTGF mRNA and protein expression in HK-2 cells in a dose- and time-dependent manner (P <0.01). (2) Rosiglitazone and 15-deoxy prostaglandin J2 (15d-PGJ2) inhibited CTGF mRNA and protein expression in HK-2 cells induced by TGF-β1 (4ng / ml) in a dose-dependent manner. (3) GW9662 (1μmol / L), a specific inhibitor of peroxisome proliferator-activated receptor (PPAR), completely blocked the inhibitory effect of rosiglitazone and 15d-PGJ2 on CTGF mRNA and protein expression. (4) FN protein in HK-2 cells treated with rosiglitazone 5,10μmol / L was decreased by 39.9% and 53.4% (P <0.01) compared with TGF-β14ng / ml stimulation alone, (P <0.01), while GW9662 completely blocked this effect. Conclusion In vitro, rosiglitazone inhibits TGF-β1-induced CTGF transcription and expression by activating renal tubular epithelial cells PPARγ, and inhibits FN expression.