Iptakalim对低氧诱导的人肺动脉平滑肌细胞增殖及TGF-β和PCNA表达的影响

Effects of Iptakalim on the proliferation and expression of TGF-β and PCNA in primary cultured human pulmonary artery smooth muscle cells induced by hypoxia

目的研究新型ATP敏感性钾通道(KATP)开放剂Iptakalim(IPT)对低氧诱导的人肺动脉平滑肌细胞(HPASMC)增殖及转化生长因子-β(TGF-β)和核增殖抗原(PCNA)表达的影响。方法原代培养HPASMC,随机分成对照组、低氧组、低氧+IPT10-7组、低氧+IPT10-6组和低氧+IPT10-5组,采用流式细胞技术检测细胞周期,用细胞免疫组织化学技术检测细胞TGF-β和PCNA表达部位分布,用Western-blot方法检测TGF-β和PCNA蛋白的表达量。结果缺氧24 h,G0/G1期细胞比例减少,S期比例增高,与对照组相比,差异具有统计学意义(P<0.05);免疫组织化学技术显示TGF-β染色主要位于细胞浆和细胞膜,PCNA染色主要位于细胞核;Western-blot结果显示低氧组TGF-β和PCNA的表达高于对照组(P<0.05)和低氧+IPT组(P<0.05),随着IPT剂量的增加,TGF-β和PCNA的表达呈下降趋势,差异具有统计学意义(P<0.05)。结论低氧能诱导原代培养的HPASMC增殖,刺激TGF-β和PCNA表达增高;IPT能浓度依赖性抑制这种作用,其机制可能与抑制TGF-β和PCNA的表达有关。

Purpose To investigate the effects of Iptakalim （IPT） , a novel ATP-sensitive potassium, on the cell proliferation and the expressions of TGF-β and PCNA of human pulmonary artery smooth muscle cells（HPASMC）induced by hypoxia. Methods The primary cultured HPASMCs were randomly divided into control group,hypoxia group, hypoxia ＋ IPT 10^-7 group, hypoxia ＋ IPT 10^-6 group and hypoxia ＋ IPT 10^-5 group. Cell cycles were detected by flow cytometry, and the protein expression location of TGF-β and PCNA were detected by immunohistochemistry, and the protein expression of TGF-β and PCNA were detected by Western-blot analysis. Results Under the condition of hypoxia after 24 hours, the rate of HPASMCs in G0/G1 period was reduced and the rate of cells in S period was increased. There was a statistical significance compared with the control group（P 〈 0.05 ）. The results of stain by immunohistochemistry indicated that TGF-β was mainly located in the cytoplasm and cytomembrane of cell and PCNA was mainly located in nucleus of cell. Western-blot analysis indicated that the expressions of TGF-β and PCNA in hypoxia group were more than those of the control group. IPT inhibited the protein expression of TGF-β and PCNA induced by hypoxia in concentration-dependent manner. Conclusion Hypoxia could induce the cell proliferation of HPASMCs, and promote the expressions of TGF-β and PCNA. Iptakalim,a novel ATP-sensitive potassium, could suppress the cell proliferation in dose-dependent man- ner, which mechanism might be related to the suppression of the expressions of TGF-β and PCNA.