红毛五加多糖对心衰大鼠心肌细胞凋亡及相关基因表达影响的机制研究

Effect of Acanthopanax Giraldii Harms Var Hispidus Hoo polysaccharides on cardiomyocyte apoptosis and expression of related genes in rats with heart failure and its mechanism

目的探讨红毛五加多糖(AGP)对心衰大鼠心肌细胞凋亡及PI3K/AKT信号通路的影响。方法将120只2~3月龄SPF级SD大鼠(雌雄各半)随机分为假手术组、心力衰竭模型组和AGP低、中、高剂量组,每组20只。AGP低中高剂量组皮下注射AGP(10、20、30mg·kg^-1·d^-1),假手术组和模型组皮下注射等量生理盐水。预给药4d后采用左冠状动脉前降支结扎术建立大鼠心衰模型。建模42d后,通过超声检测左室短轴缩短率(FS)及左室射血分数(EF);邻苯三酚法及硫代巴比妥酸法分别测定超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量;TUNEL试剂盒测定心肌细胞凋亡率;免疫印迹检测Bcl-2、Bax、PI3K和p-ATK蛋白表达。结果与假手术组比较,模型组EF值和FS值明显降低[(31.88±2.56)%比(99.71±0.42)%、(18.02±1.52)%比(91.12±3.44)%,均P<0.05)],SOD活性明显降低[(90.21±6.12)nU/ml比(124.93±8.87)nU/ml,P<0.05)],MDA含量明显升高[(6.47±0.68)nmol/ml比(3.68±0.39)nmol/ml,P<0.05)],心肌细胞凋亡率明显升高[(22.38±1.42)%比(5.02±0.51)%,P<0.05)],Bcl-2、PI3K和p-ATK蛋白表达明显降低(0.087±0.010比0.762±0.059、0.052±0.005比0.859±0.072、0.021±0.005比0.151±0.016,均P<0.05),Bax蛋白表达明显升高(0.087±0.010比0.762±0.059,P<0.05)。与模型组比较,低、中、高剂量AGP组EF值、FS值、SOD活性均明显升高(均P<0.05),MDA含量、心肌细胞凋亡率明显降低(均P<0.05),Bcl-2、PI3K和p-ATK蛋白表达明显升高(均P<0.05),Bax蛋白表达明显降低,且有浓度依赖性(P<0.05)。结论AGP可降低心衰大鼠心肌细胞凋亡,机制可能与抑制氧化应激及激活PI3K/AKT信号通路有关。

Objective To investigate the effect of Acanthopanax Giraldii Harms Var Hispidus Hoo polysaccharides (AGP) on cardionyocyte apoptosis and PI3K/AKT signaling pathway in rats with heart failure.Methods:A total of 120 specific-pathogen-free SD rats aged 2~3 months old (female/male ratio=1) were randomly divided into the sham operation group,heart failure model group and AGP low-,medium-and high-dose groups (n=20 each).The AGP low-,medium-,and high-dose groups were injected subcutaneously with AGP (10,20 and 30 mg· kg-1· d-1),and the sham-operated group and the model group were injected subcutaneously with the same amount of normal saline.At 4d after pre-administration,the heart failure rat model was established by ligation of the left anterior descending coronary artery.At 42 d after modeling,the left ventricular short axis shortening (FS) and left ventricular ejection fraction (EF) were determined by ultrasound.The activity of superoxide dismutase (SOD) and level of malondialdehyde (MDA) were examined by pyrogallol method and thiobarbituric acid method,respectively.The apoptosis rate of cardiomyocytes was determined by TUNEL kit.The protein expression levels of Bcl-2,Bax,PI3K and p-ATK were measured by Western blotting.Results Compared with the sham operation group,the EF and FS values in the model group were significantly lower [(31.88±2.56)% vs.(99.71±0.42)%,(18.02± 1.52)% vs.(91.12±3.44)%,both P<0.05)],the SOD activity was significantly reduced [(90.21±6.12) nU/ml vs.(124.93±8.87) nU/ml,P<0.05)],the MDA level significantly increased [(6.47±0.68) nmol/ml vs.(3.68±0.39) nmol/ml,P<0.05)],the apoptosis rate of cardiomyocytes increased significantly [(22.38± 1.42)% vs.(5.02 ± 0.51)%,P<0.05)],the protein expression levels of Bcl-2,PI3K and p-ATK significantly decreased (0.087±0.010 vs.0.762±0.059,0.052±0.005 vs.0.859±0.072,0.021±0.005 vs.0.151±0.016,all P<0.05),and the protein expression level of Bax significantly increased (0.087±0.010 vs.0.762±0.059,P<0.05).Compared with the model group,the EF and FS values,the SOD activity in the low-,medium-and high-dose AGP groups significantly increased (all P<0.05),the MDA level and the apoptosis rate of cardiomyocytes significantly decreased (all P<0.05),the protein expression levels of Bcl-2,PI3K and p-ATK significantly increased (all P<0.05),and the protein expression level of Bax significantly decreased in a concentration-dependent manner (P<0.05).Conclusion AGP may reduce cardiomyocyte apoptosis in rats with heart failure.The mechanism may be related to the inhibition of oxidative stress and activation of PI3K/AKT signaling pathway.