EPO通过激活PI3K/Akt途径上调HSP70的表达对低温下心衰大鼠起保护作用

摘要

目的: 评价红细胞生成素(erythropoietin,EPO)对低温环境下心衰大鼠的影响,探讨其可能的作用机制.方法: 80只SD大鼠随机分5组,分别为对照组(CON组)、心衰低温组(HFLT组)、心衰常温组(HFNT组)、心衰低温EPO组(HFLT+EPO组)和心衰低温LY294002组(HFLT+EPO+LY组).然后将所有大鼠放置于气候箱中(CON、HFLT、HFLT+EPO和HFLT+EPO+LY组为低温环境;HFNT组为常温).超声心动图检测各组大鼠心功能,然后处死大鼠并留取心脏标本,TUNEL法测心肌细胞凋亡,荧光定量PCR测心肌组织中Fas和PI3K mRNA的表达,Western blot测大鼠心肌组织中热休克蛋白70(HSP70)、Akt和p-Akt的水平.结果: 低温下心衰大鼠心功能明显低于常温下心衰大鼠,HFLT+EPO组大鼠心功能较HFLT组明显改善,给予LY294002干预后HFLT+EPO+LY组大鼠心功能明显低于HFLT+EPO组;HFLT组及HFNT组凋亡指数均高于CON组(P<0.01),HFLT组凋亡指数又明显高于HFNT组(P<0.05),HFLT+EPO组凋亡指数明显低于HFLT组(P<0.01).Fas mRNA在HFLT组心肌组织中的表达明显高于HFNT组,PI3K mRNA在HFLT组和HFNT组心肌组织中的表达差异无统计学显著性,HFLT+EPO组心肌组织中Fas的mRNA表达明显低于HFLT组(P<0.01)和HFLT+EPO+LY组(P<0.05),而PI3K的mRNA表达则明显高于HFLT组和HFLT+EPO+LY组(P<0.05).HFLT+EPO组大鼠心肌组织中p-Akt和HSP70的水平明显高于HFLT组和HFLT+EPO+LY组(P<0.05),CON、HFLT和HFNT组大鼠心肌组织中p-Akt和HSP70的蛋白水平差异无统计学显著性.所有大鼠心肌组织中Akt的水平差异无统计学显著性.结论: EPO活化PI3K/Akt后,上调内源性保护途径中HSP70的表达并抑制细胞凋亡,从而对低温下的心衰大鼠心脏起保护作用.%AIM: To investigate the effect oferythropoietin (EPO) on the rats with heart failure (HF) in hypothermia and to explore its underlying mechanism.METHODS: The Sprague-Dawley rats (n=80) were randomly divided into five groups: control group (CON group), HF in low-temperature group (HFLT group), HF in normal temperature group (HFNT group), HF with EPO in low temperature group (HFLT+EPO group), and HF with EPO and LY2940002 in low temperature group (HFLT+EPO+LY group).All rats were housed in artifitial climate chamber.The animals in CON, HFLT, HFLT+EPO and HFLT+EPO+LY groups were under the low-temperature environment, while those in HFNT group were under normal temperature.The heart function was evaluated by echocardiography.The rats were then executed and the hearts were harvested.The apoptosis of myocytes was assessed by TUNEL method.The mRNA expression of Fas and PI3K was detected by fluorescence quantitative PCR (qPCR) and the protein levels of HSP70, Akt and p-Akt in the myocardial tissues were determined by Western blot.RESULTS: The rat cardiac functions in HFLT group were significantly deteriorated compared with HFNT group.The cardiac functions in HFLT+EPO group were improved compared with HFLT group.The cardiac functions in HFLT+EPO+LY group were significantly pejorated compared with HFLT+EPO group.The apoptotic index of the myocardium in HFLT group and HFNT group was significantly higher than that in CON group (P<0.01).The apoptotic index of the myocardium in HFLT group was significantly higher than that in HFNT group (P<0.05).The apoptotic index of the myocardium in HFLT+EPO group was significantly lower than that in HFLT group (P<0.01).The mRNA expression of Fas in HFLT group was significantly higher than that in HFNT group, and no obvious difference of the mRNA expression level of PI3K between HFLT group and HFNT group was observed.The mRNA expression of PI3K in HFLT+EPO group was significantly lower than that in HFLT group and HFLT+EPO+LY group (P<0.05), and that in HFLT+EPO group was significantly higher than that in HFLT group and HFLT+EPO+LY group (P<0.05).The protein levels of p-Akt and HSP70 in HFLT+EPO group was also higher than those in HFLT group and HFLT+EPO+LY group (P<0.05), and no obvious difference of the protein levels of p-Akt and HSP70 in CON, HFLT and HFNT groups was found.The protein level of Akt had no significant difference in each group.CONCLUSION: The pathway of PI3K/Akt may be one of the cardioprotective ways of EPO.EPO activates the PI3K/Akt pathway, upregulates the experssion of HSP70 (an endogenous protective factor) and inhibits the apoptosis, thus protecting the cardiac functions in the rats with HF in hypothermia.