附子多糖保护缺氧/复氧乳鼠心肌细胞及其抗内质网应激的机制研究

目的: 观察附子多糖对缺氧/复氧后心肌细胞的保护,并探讨附子多糖的保护机制是否与其抑制内质网应激反应有关。方法: 建立乳鼠心肌细胞缺氧/复氧模型,将乳鼠心肌细胞分为正常对照组、缺氧/复氧组(缺氧3 h后复氧6 h)和不同浓度附子多糖(0.1 g/L、1 g/L、10 g/L、20 g/L)+缺氧/复氧组。MTT法测定心肌细胞存活率,流式细胞术检测心肌细胞凋亡率,Western blotting分析葡萄糖调节蛋白78(GRP78)、CCAAT增强子结合蛋白同源蛋白(CHOP)及caspase-12的表达,荧光定量PCR进一步检测CHOP及caspase-12 mRNA表达。结果: 缺氧复氧后,心肌细胞中内质网应激反应标志蛋白GRP78表达增加,内质网应激凋亡相关蛋白CHOP及caspase-12蛋白和mRNA表达亦明显升高。与缺氧/复氧组相比较,附子多糖预处理24 h后可以有效抑制缺氧/复氧引起的GRP78、 CHOP和caspase-12的表达上调,增加心肌细胞的存活率,抑制心肌细胞凋亡的发生。附子多糖的保护效应呈剂量依赖形式,10 g/L剂量时达到峰值。结论: 附子多糖保护缺氧/复氧后心肌细胞的可能机制与其抑制内质网应激所介导的细胞凋亡途径相关。

Fuzi polysaccharide protects neonatal rat cardiomyocytes with hypoxia-reoxygenation by inhibiting endoplasmic reticulum stress

AIM:To investigate whether the protection mechanism of Fuzi polysaccharide(FPS) is related to inhibition of endoplasmic reticulum stress in cultured neonatal rat cardiomyocytes with hypoxia/reoxygenation(H/R). METHODS:Cultured rat myocardial cells were divided into control group,H/R group(hypoxia for 3 h and reoxygenation for 6 h) and different concentrations of FPS(0.1 g/L,1 g/L,10 g/L or 20 g/L) + H/R groups.The cell survival was detected by MTT assay and cell apoptosis of cardiomyocytes was measured by flow cytometry using Annexin V-FITC staining. The expression of glucose-regulated protein 78(GRP78),CCAAT/enhancer-binding protein homologous protein (CHOP) and caspase-12 were determined by Western blotting.The mRNA expression of CHOP and caspase-12 was detected by quantitative PCR.RESULTS;After reoxygenation,the expression of GRP78,CHOP and caspase-12 in cardiomyocytes was increased.Compared with H/R group,the expression of GRP78,CHOP and caspase-12 in FPS + H/R groups was significantly inhibited,the survival rate of cardiomyocytes was increased and the apoptosis of cardiomyocytes was inhibited.This protective effect of FPS was in a dose-dependent manner and reached its peak at 10 g/L.CONCLUSION: Fuzi polysaccharide protects cardiomyocytes from H/R injury.The mechanism is related to inhibiting endoplasmic reticulum stress.