生理性缺血训练改善心衰大鼠心脏结构和功能的最佳干预强度及生物学标志物研究

Optimum intervention intensity and effect biomarkers of physiological ischemic training to improve cardiac structure and function in rats with heart failure

目的:探索生理性缺血训练(PIT)改善心衰大鼠心脏结构、功能的最佳干预强度及生物学标志物。方法:30只SD大鼠随机分为假手术组(SO组),单纯心衰组(HF组),生理性缺血训练1min组(PIT-S组)、5min组(PIT-M组)和10min组(PIT-L组),每组6只。SO组按心衰模型建立步骤,穿线过左冠状动脉前降支不结扎,不训练;HF组建立心衰大鼠模型;PIT组在HF组基础上接受训练,单次缺血时长分别为1min、5min、10min,再灌注均为5min,5次/天,5天/周,持续8周。训练结束后采用经胸壁超声心动图检测心功能,Masson染色检测心肌纤维化程度,qRT-PCR检测Bcl-2、Bax、MMP9、TIMP1 mRNA的相对表达水平,ELISA法检测血清中NT-proBNP、Bradykinin、TNF-α、IL-6、IL-10等生物学标志物含量。结果:与SO组相比,各组Bax、心肌纤维化程度、MMP9/TIMP1比值显著升高,左室射血分数(LVEF)、Bcl-2、Bcl-2/Bax比值、TIMP1显著降低(P<0.05);与HF组相比,3组PIT组LVEF显著升高,Bax、心肌纤维化程度、MMP9、MMP9/TIMP1比值显著降低(P<0.05);3组PIT大鼠中,PIT-M组LVEF、Bcl-2/Bax比值显著高于另外2组,而Bax、MMP9/TIMP1比值显著低于另外2组(P<0.05)。除PIT-S组Bradykinin含量外,各组血清各标志物含量均明显高于SO组;PIT-M组各标志物含量均显著低于HF组,其中IL-6、Bradykinin、TNF-α显著低于PIT-S组,IL-10、Bradykinin、TNF-α显著低于PIT-L组(P<0.05)。各标志物血清含量与LVEF均呈负相关(P<0.01),其中NT-proBNP相关性最为密切(r=﹣0.936,P<0.0001)。结论:相比于单次1min缺血/5min灌注、10min缺血/5min灌注,5min缺血/5min灌注的PIT方案对心衰大鼠心脏的远隔保护作用更显著,NT-proBNP可作为反映其保护作用的最佳生物学标志物。

Objective:To explore the optimum intervention intensity and effect biomarkers of physiological ischemic training(PIT) on improving cardiac structure and function in rats with heart failure.Method:Thirty rats were randomized into sham operation(SO),heart failure(HF),physiological ischemic training for 1 min(PIT-S),5 min(PIT-M) and 10 min(PIT-L),with 6 rats in each group. In the SO group,according to the heart failure model establishment,the line was threaded through the anterior descending branch of the left coronary artery without ligation or training. Heart failure model was established in HF group. PIT group received training on the basis of HF group, accompanied with ischemia for 1 min, 5 min, 10 min respectively and reperfusion for 5 min each time,5 cycles/day,5 days/week,lasting for 8 weeks. After the training,cardiac function was detected by transthoracic echocardiography,the degree of myocardial fibrosis was detected by Masson staining, the relative expression level of Bcl-2, Bax, MMP9,TIMP1 mRNA was detected by q RTPCR,and the content of NT-proBNP,Bradykinin,TNF-α,IL-6,IL-10 in serum was detected by ELISA.Result:Compared with SO group,Bax,MMP9/TIMP1 ratio and the degree of myocardial fibrosis in other groups were significantly increased, while the left ventricular ejection fraction(LVEF), Bcl-2, Bcl-2/Bax ratio and TIMP1 were significantly decreased(P<0.05). Compared with HF group,LVEF in 3 PIT groups was significantly increased,while Bax,the degree of myocardial fibrosis,MMP9 and MMP9/TIMP1 ratio were significantly decreased(P<0.05). The LVEF and Bcl-2/Bax ratio in PIT-M group were significantly higher while the Bax and MMP9/TIMP1 ratio were significantly lower than those in the other two PIT groups(P<0.05). Except the Bradykinin in PIT-S group, the content of other biomarkers in each group was significantly higher than those in SO group. The content of each biomarker in PIT-M group was significantly lower than that in HF group,among which IL-6,Bradykinin,TNF-α were significantly lower than that in PIT-S group,and IL-6,Bradykinin, TNF-α were significantly lower than that in PIT-L group(P<0.05). The serum content of the above biomarkers were all negatively correlated with LVEF(P<0.01), and NT-proBNP was the most closely correlated(r=﹣0.936,P<0.0001).Conclusion:Compared with ischemia for 1 min or 10 min,the PIT program of 5 min ischemia/5 min reperfusion has a more significant cardiac protection effect on heart failure rats,and NT-proBNP can be used as the best effect biomarker to reflect the protection effect.