Role of autophagy via PI3K/Akt signaling pathway in attenuation of doxorubicin-induced cardiomyopathy by Sevoflurane postconditioning
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摘要: 目的 评价七氟烷后处理对阿霉素大鼠心肌细胞磷脂酰肌醇-3激酶/蛋白质丝氨酸苏氨酸激酶(PI3K/Akt)通路和自噬的影响。 方法 培养H9c2心肌细胞,采用随机数字表法,将细胞分为6组:对照组(C组)、阿霉素损伤组(Dox组)、七氟烷处理组(Sev组)、LY294002抑制剂组(LY组)、溶剂对照组(DMSO组)、3-MA抑制剂组(3-MA组)。除C组外,其余5组建立阿霉素心肌细胞损伤模型。C组与Dox组不做处理,Sev组、LY组(暴露前培养基中加入LY294002)、DMSO组(加入DMSO)、3-MA组(加入3-MA)2.4%七氟烷暴露2 h后,采用ELISA法测定培养液中c Tn I和Caspase-3的浓度;Western blot法检测微管相关蛋白1轻链3-Ⅱ(LC 3-Ⅱ)、总Akt(tAkt)及磷酸化Akt(p-Akt)的表达。计量资料以x±s表示,组间比较采用单因素方差分析。 结果 与C组比较,其余5组p-Akt表达降低,LC3-Ⅱ表达、c Tn I和Caspase-3浓度升高(P<0.05);与Dox组比较,Sev组p-Akt表达升高,LC3-Ⅱ、c Tn I和Caspase-3浓度降低(P<0.05);与Sev组比较,LY组p-Akt表达降低,LC3-Ⅱ表达升高,c Tn I和Caspase-3浓度升高(P<0.05);3-MA组LC3-Ⅱ、c Tn I和Caspase-3浓度降低(P<0.05);DMSO组上述指标差异无统计学意义(P>0.05)。 结论 七氟烷后处理可减轻阿霉素性心肌细胞损伤,与激活PI3K/Akt通路抑制自噬过度激活有关。Abstract: Objective To investigate the effect of Sevoflurane postconditioning on PI3K/Akt signaling pathway and autophagy after doxorubicin-induced cardiomyopathy. Methods The H9c2 cardiomyocytes were randomly divided into 6groups (n=6):control group (group C),Doxorubicin group (group Dox),Sevoflurane group (group Sev),LY294002group (group LY),Solvent control group (group DMSO),3-MA group (group 3-MA).In addition to the group C,the other5 groups were all established doxorubicin-induced damage to cardiomyocytes model.Group C and group Dox did not receive any treatment,group Sev,group LY (LY294002 was added to the medium before expose),group DMSO (added DMSO),group 3-MA (added 3-MA) exposed to 2.4% Sevoflurane for 2 h.The concentrations of c Tn I and Caspase-3 in the culture medium was detected by ELISA;the expression of LC3-Ⅱ,total Akt (t-Akt) and phosphorylated Akt (p-Akt) was detected by Western blot.The quantitative data were presented as mean±standard deviation (x±s).The groups were compared using ANOVA. Results Compared with group C,the p-Akt expression was decreased,LC3-Ⅱ expression,apoptosis index,serum c Tn I and Caspase-3 concentration were significantly increased in the other five groups (P<0.05).Compared with group Dox,expression of p-Akt was increased,the expression of LC3-Ⅱ,apoptosis index,c Tn I and Caspase-3 were decreased in group Sev (P<0.05).Compared with group Sev,p-Akt was decreased,the expression of LC3-Ⅱ,apoptosis index,serum c Tn I and Caspase-3 concentration were increased in group LY (P<0.05);the expression of LC3-Ⅱ,c Tn I and Caspase-3 were decreased in group 3-MA (P<0.05). Conclusion Sevoflurane postconditioning on cardiomyocytes may reduce the myocardial cell injury induced by adriamycin,which may be related to the activation of PI3 K/Akt pathway to inhibit the excessive activation of autophagy.
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