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目的 探讨马兜铃酸(AA)对体外培养的人肾小管上皮细胞株(HK-2)及人肾间质成纤维细胞(hRIFs)的作用。方法 (1)用MTT比色法检测细胞增殖反应。(2)用乳酸脱氢酶(LDH)释放试验检测AA的细胞毒作用。(3)应用RT-PCR观察细胞Ⅰ型胶原(Col Ⅰ)、转化生长因子β(TGF-β)、纤溶酶原激活物抑制物1(PAI-1)、基质金属蛋白酶1(MMP-1)及金属蛋白酶组织抑制物1(TIMP-1)的mRNA表达。结果 (1)AA在10、20和40 μg/ml时对HK-2及hRIFs无刺激增殖及细胞毒效应(P>0.05);而在80和160μg/ml时却能明显杀伤上述细胞(P<0.01)。(2)40μg/ml浓度的AA孵育细胞16 h,可显著上调HK-2的TGF-β、PAI-1和TIMP-1 mRNA表达(P<0.05),也可上调hRIFs的Col Ⅰ、TGF-β、PAI-1和TIMP-1 mRNA的表达(P<0.05);而10和20μg/ml浓度的AA未观察到上述作用。结论 80和160μg/ml AA对HK-2有明显细胞毒作用,此作用可能与急性马兜铃酸肾病发病相关;而40μg/ml AA可上调HK-2及hRIFs的TGF-β、PAI-1和TIMP-1 mRNA表达,并能上调hRIFs的Col Ⅰ mRNA表达,此作用可能与慢性马兜铃酸肾病发病相关。
Objective To investigate the effect of aristolochic acid (AA) on human renal tubular epithelial cell line (HK-2) and human renal interstitial fibroblasts (hRIFs) cultured in vitro. Methods (1) The cell proliferation reaction was detected by MTT assay. (2) The cytotoxicity of AA was tested by lactate dehydrogenase (LDH) release assay. (3) The expressions of type Ⅰ collagen (Col Ⅰ), transforming growth factor β (TGF-β), plasminogen activator inhibitor 1 (PAI-1) and matrix metalloproteinase 1 ) And tissue inhibitor of metalloproteinase 1 (TIMP-1) mRNA expression. Results (1) The proliferation and cytotoxicity of HK-2 and hRIFs were not significantly inhibited by AA at 10, 20 and 40 μg / ml (P> 0.05) <0.01). (2) The expression of TGF-β, PAI-1 and TIMP-1 mRNA in HK-2 cells was significantly increased after incubation with 40μg / ml AA for 16 h (P <0.05) β, PAI-1 and TIMP-1 mRNA expression (P <0.05); while 10 and 20μg / ml concentration of AA did not observe the above effect. Conclusions 80 and 160 μg / ml AA have obvious cytotoxic effect on HK-2, which may be related to the pathogenesis of acute aristolochic acid nephropathy. However, 40 μg / ml AA can up-regulate the expressions of TGF-β and PAI-1 in HK-2 and hRIFs And TIMP-1 mRNA expression, and can up-regulate Col Ⅰ mRNA expression of hRIFs, which may be related to the pathogenesis of chronic aristolochic acid nephropathy.