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目的以细胞凋亡的线粒体信号通路为着眼点,对附子多糖对缺氧/复氧后心肌细胞的保护机制进行探讨。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、缺氧后适应组和附子多糖组。缺氧/复氧组给予心肌细胞缺氧3 h后复氧6 h;缺氧后适应组在细胞缺氧3 h后,复氧前即给予3个循环的5 min复氧/5 min缺氧,随后复氧6 h;附子多糖组在缺氧3 h后,将心肌细胞换入含附子多糖浓度为10 g.L?1的培养液中常规培养6 h。流式细胞仪测定心肌细胞凋亡率和线粒体膜电位,进行凋亡诱导因子(AIF)的Western blotting分析,检测心肌细胞内SOD活性和MDA含量。结果与缺氧/复氧组相比较,附子多糖后处理可以保护心肌细胞SOD活性,减少MDA的生成,阻止线粒体膜电位的下降,抑制AIF自线粒体向胞浆的释放,减少心肌细胞凋亡率。结论附子多糖后处理对缺氧/复氧后心肌细胞的保护机制与其抗氧化损伤,抑制细胞凋亡的线粒体信号途径有关。
Objective To investigate the mitochondrial signaling pathway of apoptosis and to explore the protective mechanism of monkshood polysaccharide on cardiomyocytes after hypoxia / reoxygenation. Methods The neonatal rat cardiomyocytes hypoxia / reoxygenation model was established. The cardiomyocytes were divided into normal control group, hypoxia / reoxygenation group, hypoxia adaptation group and monkshood polysaccharide group. Hypoxia / reoxygenation group was given cardiomyocytes 3h after reoxygenation 6h; After hypoxia adaptation group 3h after cell hypoxia, reoxygenation before giving 3 cycles of 5min reoxygenation / 5min hypoxia , Followed by reoxygenation for 6 h. After treated with hypoxia for 3 h, the mononuclear polysaccharide group was used to culture cardiomyocytes for 6 h in medium containing monkshood polysaccharide (10 gL? 1). The apoptotic rate of cardiomyocytes and mitochondrial membrane potential were measured by flow cytometry. Western blotting analysis of apoptosis inducing factor (AIF) was performed to detect the activity of SOD and the content of MDA in myocardial cells. Results Compared with the hypoxia / reoxygenation group, the treatment of aconite polysaccharide could protect the activity of SOD in cardiomyocytes, decrease the production of MDA, decrease the mitochondrial membrane potential, inhibit the release of AIF from the mitochondria to the cytoplasm, and decrease the apoptosis rate of cardiomyocytes . Conclusions The protective mechanism of aconite polysaccharide postconditioning on cardiomyocytes after hypoxia / reoxygenation is related to its antioxidant activity and mitochondrial signal pathway.