HO-1介导黄芪甲苷抗原代心肌细胞缺氧/复氧损伤作用研究

Study of the protective effect of Astragaloside Ⅳ mediated by HO-1 against hypoxia/reoxygenation induced cell injury in primary cardiomyocytes

关键词:  黄芪甲苷  血红素氧化酶1  心肌细胞  缺氧/复氧  CoPP  ZnPP

Key words:  Astragaloside Ⅳ  heme oxygenase-1  cardiomyocytes  hypoxia/reoxygenation  CoPP  ZnPP

摘要:

[目的]研究黄芪甲苷（AS-Ⅳ）对乳鼠原代心肌细胞缺氧/复氧（H/R）损伤的保护作用及机制。[方法]培养乳鼠原代心肌细胞，以缺氧4 h，复氧4 h建立心肌H/R损伤模型。四甲基噻唑蓝（MTT）法检测细胞活力。检测细胞培养液中心肌肌钙蛋白（cTnT）、乳酸脱氢酶（LDH）含量，炎症细胞因子超敏C反应蛋白（hs-CRP）、肿瘤坏死因子-α（TNF-α）浓度。分别以聚合酶链式反应逆转录（RT-PCR）、蛋白免疫印迹（Western blot）法检测心肌细胞血红素氧化酶1（HO-1）mRNA及蛋白表达水平。[结果]与H/R组比较，AS-Ⅳ、HO-1激动剂原卟啉氯化钴（CoPP）均可显著降低细胞上清中cTnT、LDH含量（P<0.01），降低炎症因子hs-CRP、TNF-α水平（P<0.01），而HO-1拮抗剂原卟啉Ⅸ锌（Ⅱ）络合物（ZnPP）作用趋势则相反。与H/R组比较，CoPP组及AS-Ⅳ组HO-1 mRNA、蛋白表达水平显著升高（P<0.01），ZnPP组则显著下降（P<0.01）。[结论]AS-Ⅳ对心肌细胞缺氧/复氧损伤具有显著的保护作用，其机制与诱导具有保护作用的HO-1表达有关。

Abstract:

[Objective] To study the protective effect and mechanism of Astragaloside Ⅳ(AS-Ⅳ) against hypoxia/reoxygenation (H/R) induced cell injury in primary cardiomyocytes.[Methods] Neonatal rat primary cardiomyocytes were cultured, then H/R injury model was established by hypoxia followed by reoxygenation for 4 h respectively. Cell viability was measured by MTT method. Besides, different treatment factors impacts on cTnT, LDH and inflammatory cytokine hs-CRP and TNF-α level in cell culture medium was examined. Protein level of HO-1 was detected by Western blot analysis, and mRNA level was detected by RT-PCR.[Results] Compared with H/R group, both AS-Ⅳ and CoPP, HO-1 inducer, could significantly protect cells from damage induced by H/R, and inhibit inflammatory cytokine hs-CRP and TNF-α level(P<0.01). While ZnPP, HO-1 inhibitor, showed a remarkable effect of cell damage based on H/R injury. AS-Ⅳ showed a similar effect to CoPP, which could further increased HO-1 mRNA and protein expression(P<0.01).[Conclusion] AS-Ⅳ could inhibit cell damage caused by H/R by induction of HO-1 expression.