1-磷酸鞘氨醇通过PI3K/Akt信号通路对冠心病大鼠心肌细胞凋亡的调控作用研究

Study on the Regulatory Effect of Sphingosine-1-phosphate on Myocardial Cell Apoptosis in Rats with Coronary Heart Disease Through PI3K/Akt Signaling Pathway

目的:观察1-磷酸鞘氨醇通过磷脂酰肌醇3-激酶(Phosphatidylinositol-3-OH kinase,PI3K)/蛋白激酶B (Protein kinase B,Akt)信号通路对冠心病(Coronary heart disease,CHD)大鼠心肌细胞凋亡的调控作用。方法:将45只清洁级Wistar雄性大鼠,随机分为假手术组、模型组和1-磷酸鞘氨醇(Sphingosine-1-phosphate,S1P)组,每组各15只。模型组和S1P组诱导CHD模型。模型诱导成功1周后,进行尾静脉注射给药。S1P组按1.0 mg/kg剂量给予S1P,假手术组和模型组分别给予等体积的生理盐水,每周1次,连续给药4周。采用TTC染色法测定各组大鼠的心肌梗死面积,使用TUNEL染色方法观察计算各组大鼠心肌细胞的凋亡指数,通过Western blot检测PI3K/Akt信号通路中相关蛋白PI3K、Akt及凋亡因子含半胱氨酸的天冬氨酸蛋白水解酶3 (Cysteinyl aspartate specific proteinase 3,Caspase-3)的表达情况。结果:TTC染色检测结果显示,与假手术组比较,模型组大鼠均出现大面积的心肌梗死(P<0.01);与模型组比较,S1P组大鼠心肌梗死面积显著减小(P<0.01)。TUNEL染色结果显示,与假手术组比较,模型组大鼠心肌细胞凋亡数量明显增加,其凋亡率显著升高(P<0.01);与模型组比较,S1P组大鼠心肌细胞的凋亡指数显著降低(P<0.05)。Western blot检测结果显示,各组间PI3K、Akt蛋白表达量无显著性差异(P>0.05);与假手术组比较,模型组心肌细胞内p-PI3K、p-Akt的表达量显著下降(P<0.01),Caspase-3的表达水升高(P<0.05);与模型组比较,S1P组p-PI3K、p-Akt的表达水平显著增加(P<0.01)。Caspase-3的表达水平降低(P<0.05)。结论:S1P可能是通过调节PI3K/Akt信号通路中相关蛋白的表达来有效抑制CHD大鼠心肌细胞凋亡的。

Objective:To observe the regulatory effect of sphingosine-1-phosphate on myocardial cell apoptosis in rats with coronary heart disease(CHD) through phosphodylinositol-3-OH kinase(PI3 K)/protein kinase B(Akt) signaling pathway.Methods: Divided 45 Wistar male rats of clean grade randomly into the sham operation group, the model group and the sphingosine-1-phosphate(S1 P) group,with 15 rats in each group. The model group and the S1 P group induced the CHD model. One week after successfully inducing the model,gave the drug by tail intravenous injection. The S1 P group was given1.0 mg/kg dose of S1 P,while the sham operation group and the model group were respectively given the same volume of normal saline once a week continuously for 4 weeks. Determined the myocardial infarction area of rats in each group by TTC staining,observed and calculated the apoptosis index of myocardial cells of rats in each group by TUNEL staining method and detected the expressions of associated protein PI3 K and Akt in PI3 K/Akt signaling pathway and cysteinyl aspartate specific proteinase 3(Caspase-3) by Western blot. Results: TTC staining results showed that, when compared with the sham operation group, a large area of myocardial infarction was found in the model group(P<0.01);compared with the model group,the myocardial infarction area in S1 P group was evidently decreased(P<0.01). TUNEL staining results showed that,when compared with the sham operation group,the amount of myocardial cell apoptosis in the model group was obviously increased and the apoptosis rate was significantly increased(P<0.01);compared with the model group,the apoptosis index of myocardial cells of rats in the S1 P group was evidently decreased(P<0.05). The detection results of Western blot showed that there was no significant difference being found in the expressions of PI3 K and Akt among the groups(P>0.05);compared with the sham operation group,the expressions of p-PI3 K and p-Akt in the myocardial cells in the model group were evidently decreased(P<0.01) and the expression levels of Caspase-3 were increased(P<0.05);compared with the model group, the expression levels of p-PI3 K and p-Akt in the S1 P group were significantly increased(P<0.01). The expression levels of Caspase-3 were decreased(P<0.05). Conclusion: S1 P may effectively inhibit the myocardial cell apoptosis in rats with CHD by regulating the expression of associated protein in PI3 K/Akt signaling pathway.