柴胡加龙骨牡蛎汤含药血清调控NLRP3/Caspase-1通路介导心肌细胞焦亡的研究

Mechanism of Chaihujia Longgu Muli Decoction-containing serum regulating NLRP3/Caspase-1 pathway-mediated cardiomyocyte pyroptosis

目的探讨柴胡加龙骨牡蛎汤含药血清对缺血/缺氧诱导的心肌细胞焦亡的保护作用及对NLRP3/Caspase-1信号通路的影响。方法体外培养H9c2心肌细胞,采用CCK-8法检测不同浓度基质细胞衍生因子-1α(SDF-1α)干预下H9c2心肌细胞活性,筛选合适的SDF-1α浓度。将细胞随机分为3组:模型组心肌细胞采用缺血缺氧培养基培养,SDF-1α组心肌细胞采用缺血缺氧培养基+SDF-1α培养,含药血清组心肌细胞采用缺血缺氧培养基+SDF-1α+柴胡加龙骨牡蛎汤含药血清培养。TUNEL法检测3组H9c2心肌细胞凋亡情况;Western blot技术检测3组心肌细胞中焦亡相关蛋白NLRP3、Caspase-1、ASC蛋白表达情况。结果在100 ng/mL SDF-1α浓度干预下,H9c2心肌细胞活性最高,细胞增殖率最高。SDF-1α组和含药血清组心肌细胞凋亡率及细胞中NLRP3、Caspase-1、ASC蛋白相对表达量均明显低于模型组(P均<0.05),且含药血清组均明显低于SDF-1α组(P均<0.05)。结论柴胡加龙骨牡蛎汤能抑制由缺血/缺氧诱导的H9c2心肌细胞焦亡,其机制可能与抑制NLRP3/Caspase-1通路有关。

Objective It is to explore the protective effect of Chaihujia Longgu Muli Decoction-containing serum on ischemia/hypoxia-induced cardiomyocyte pyroptosis and its effect on NLRP3/Caspase-1 signaling pathway.Methods H9c2 cardiomyocytes were cultured in vitro,the activity of H9c2 cardiomyocytes under intervention of different concentrations of stromal cell-derived factor-1α(SDF-1α)was measured by CCK-8 method,to screen the appropriate concentration of SDF-1α.The cells were randomly divided into three groups:the cardiomyocytes in the model group were cultured with ischemic-hypoxic medium,the cardiomyocytes in the SDF-1αgroup were cultured with ischemic-hypoxic medium+SDF-1α,and the cardiomyocytes in the drug-containing serum group were cultured with ischemic-hypoxic medium+SDF-1α+Chaihujia Longgu Muli Decoction-containing serum.The apoptosis of H9c2 cardiomyocytes in the three groups was detected by TUNEL method,the expression of pyroptosis related proteins NLRP3,Caspase-1 and ASC proteins in the cardiomyocytes of the three groups was detected by Western blot technique.Results The activity and cell proliferation rate of H9c2 cardiomyocytes were the highest under the intervention of SDF-1αat concentration of 100 ng/mL.The apoptosis rate of cardiomyocytes and the relative expression of NLRP3,Caspase-1 and ASC proteins in cardiomyocytes in the SDF-1αgroup and drug-containing serum group were significantly lower than those in the model group(all P<0.05),and the drug-containing serum group was significantly lower than the SDF-1αgroup(all P<0.05).Conclusion Chaihujia Longgu Muli Decoction can inhibit the pyroptosis of H9c2 cardiomyocytes induced by ischemia/hypoxia,and the mechanism may be through inhibiting the NLRP3/Caspase-1 pathway.