瓜子金皂苷己对异丙肾上腺素诱导新生大鼠心肌缺血损伤的保护作用及可能机制

Protective effect of PGSF on myocardial ischemia injury induced by ISO in neonatal rats and its possible mechanism

目的探讨瓜子金皂苷己(PGSF)对异丙肾上腺素(ISO)诱导新生大鼠心肌缺血(MI)损伤的保护作用及其机制。方法将50只新生SD大鼠随机分为对照组、模型组及低、中、高剂量PGSF组(5、10、50 mg/kg),每组10只。对照组大鼠予以生理盐水处理;模型组大鼠皮下注射ISO(85 mg/kg,1次/d)建立MI模型,低、中、高剂量PGSF组大鼠分别腹腔注射5、10、50 mg/kg PGSF预处理7 d后再建立MI模型。采用超声心动图评价各组大鼠心功能;HE染色观察各组大鼠心肌组织病理变化;酶联免疫吸附法(ELISA)检测各组大鼠血清中肌钙蛋白(cTnI)、肌酸激酶同工酶(CK-MB)、肌红蛋白(Mb)和乳酸脱氢酶(LDH)活性;检测心肌组织中丙二醛(MDA)和活性氧(ROS)含量;免疫组化染色检测心肌组织细胞核增殖抗原(Ki67)、天冬氨酸半胱氨酸蛋白酶-3(caspase-3)蛋白表达;蛋白免疫印迹法(Western blot)检测心肌组织中蛋白激酶B(AKT)、核因子E2相关因子2(Nrf2)蛋白表达。结果模型组大鼠心肌结构紊乱,细胞充血、肿胀,有大量炎症细胞浸润及大片坏死灶。模型组大鼠左心室壁厚度(LVWT)、左心室射血分数(LVEF)、缩短分数(FS)、心率(HR)和Ki67阳性蛋白表达低于对照组(均P<0.05);而左心室收缩末体积(LVESV),血清中cTnI、CK-MB、Mb、LDH活性,心肌组织中ROS和MDA含量以及caspase-3阳性蛋白高于对照组(均P<0.05)。与模型组比较,低、中、高剂量PGSF组新生大鼠心肌组织病变程度逐渐减轻。PGSF低、中、高剂量组新生大鼠LVWT、LVEF、FS、HR和Ki67阳性蛋白表达高于模型组(均P<0.05);而LVESV,血清中cTnI、CK-MB、Mb、LDH活性,心肌组织中ROS和MDA含量以及caspase-3阳性蛋白表达低于模型组(均P<0.05)。Western blot结果发现,模型组新生大鼠心肌组织中磷酸化AKT(p-AKT)/AKT、磷酸化Nrf2(p-Nrf2)/Nrf2蛋白相对表达低于对照组(均P<0.05);低、中、高剂量PGSF组新生大鼠心肌组织中p-AKT/AKT、p-Nrf2/Nrf2蛋白相对表达高于模型组(P<0.05)。结论PGSF对新生大鼠MI损伤有保护作用,其机制可能与抗细胞凋亡、抗氧化应激有关。

Objective To study the protective effect of polygalasaponin F(PGSF)on isoproterenol(ISO)-induced myocardial ischemia(MI)injury in neonatal rats and its possible mechanism.Methods Fifty newborn Sprague Dawley(SD)rats were randomly divided into control group,model group,low,medium and high dose PGSF groups(5,10,50 mg/kg),with 10 rats in each group.The rats in the control group were treated with normal saline;Myocardial ischemia(MI)model was established in model group by subcutaneous injection of isoproterenol(ISO,85 mg/kg,once a day);The MI model was established in rats of low,medium and high dose PGSF group after intraperitoneal injection of 5,10 and 50 mg/kg PGSF for 7 days.The cardiac function of rats in each group was evaluated by echocardiography;pathological changes of myocardial tissue of rats in each group were observed by hematoxylin and eosin(HE)staining;The serum activities of troponin I(cTnI),creatine kinase isoenzyme(CK-MB),myoglobin(Mb)and lactate dehydrogenase(LDH)of rats in each group were detected by enzyme linked immunosorbent assay(ELISA);the content of malondialdehyde(MDA)and active oxygen species(ROS)in myocardial tissue were detected;the expression of nuclear proliferation antigen(Ki67)and caspase-3 protein in myocardial tissue was detected by immunohistochemical staining;The expression of protein kinase B(AKT)and nuclear factor erythroid 2-related factor 2(Nrf2)protein in myocardium was detected by Western blot.Results In the model group,the myocardial structure was disordered,the cells were congested and swollen,and there were a lot of inflammatory cells infiltrating and large necrotic foci.The left ventricular wall thickness(LVWT),left ventricular ejection fraction(LVEF),fractional shortening(FS),heart rate(HR)and expression of Ki67 positive protein in the model group were lower than those in the control group(all P<0.05),while the left ventricular end systolic volume(LVESV),activities of cTnI,CK-MB,Mb,LDH in serum,content of ROS and MDA in myocardial tissue and caspase-3 positive protein in the model group were higher than those in the control group(all P<0.05).Compared with the model group,the degree of myocardial pathological changes in neonatal rats of the low,medium and high dose PGSF groups gradually decreased.Compared with model group,the LVWT,LVEF,FS,HR and expression of Ki67 positive protein increased in low,medium and high dose PGSF groups(all P<0.05),while the LVESV,activities of cTnI,CK-MB,Mb and LDH in serum,content of ROS and MDA in myocardial tissue and the expression of caspase-3 positive protein decreased(all P<0.05);Western blot results showed that the relative expression of phosphorylated(p)-AKT/AKT,p-Nrf2/Nrf2 protein in myocardium of model group was lower than that of control group(all P<0.05);The relative expression of p-AKT/AKT,p-Nrf2/Nrf2 protein in myocardium of low,medium and high dose PGSF groups were higher than that in the model group(all P<0.05).Conclusions PGSF has protective effect on MI injury in neonatal rats,and its mechanism may be related to anti-apoptosis and anti-oxidative stress.