花旗松素调控内质网应激PERK-ATF4通路减轻高血压大鼠心肌肥厚的机制研究

Mechanism of taxifolin regulating endoplasmic reticulum stress via the PERK-ATF4 pathway to reduce myocardial hypertrophy in hypertensive rats

目的探讨花旗松素(TAX)对自发性高血压大鼠(SHR)心肌肥厚的影响及分子机制。方法24只SHR分为SHR对照组(SHR组)、TAX组(20 mg/kg)、TAX+PERK激活剂CCT020312(CCT)组(20 mg/kg TAX+2 mg/kg CCT),每组8只;另选8只正常血压Wistar-Kyoto(WKY)大鼠作为正常对照组(WKY组),给予相应的药物持续干预8周。实验过程中观察大鼠血压变化,并于干预结束后超声心动图检测大鼠舒张期室间隔厚度(IVSd)、收缩期室间隔厚度(IVSs)、左心室射血分数(LVEF)判断心肌肥厚程度和心脏功能,计算心脏指数、左心室指数,苏木精-伊红(HE)染色、小麦胚芽凝集素(WGA)染色和Masson染色评估心肌组织病理学变化,实时荧光定量PCR(qRT-PCR)检测心肌组织中心房钠尿肽(ANP)、B型利钠肽(BNP)、I型胶原蛋白α1链(COL1A1)和Ⅲ型胶原蛋白α1链(COL3A1)mRNA表达,Western blot检测心肌组织蛋白激酶R样内质网激酶(PERK)-转录激活因子4(ATF4)通路相关蛋白表达。结果干预结束后,与WKY组相比,SHR组收缩压(SBP)、舒张压(DBP)、IVSd、IVSs、心脏指数、左心室指数、心肌细胞横截面积、胶原容积分数(CVF)、心肌组织ANP、BNP、COL1A1和COL3A1 mRNA表达、葡萄糖调节蛋白78(GRP78)、ATF4、C/EBP同源蛋白(CHOP)蛋白水平和p-PERK/PERK比值升高(均P<0.05),LVEF降低(P<0.05);与SHR组相比,TAX组SBP、DBP、IVSd、IVSs、心脏指数、左心室指数、心肌细胞横截面积、CVF、心肌组织ANP、BNP、COL1A1和COL3A1 mRNA表达、GRP78、ATF4、CHOP蛋白水平和p-PERK/PERK比值降低(均P<0.05),LVEF升高(P<0.05);CCT020312可部分逆转TAX对心脏功能和心肌肥厚的保护作用。结论TAX可通过抑制内质网应激(ERS),改善高血压心肌肥厚,其作用机制可能与抑制PERK-ATF4通路有关。

Objective To investigate the effect and molecular mechanism of taxifolin(TAX)on myocardial hypertrophy in spontaneously hypertensive rats(SHRs).Methods Twenty-four SHRs were divided into an SHR control group(SHR group),TAX group(20 mg/kg),and TAX+PERK activator CCT020312(CCT)group(20 mg/kg TAX+2 mg/kg CCT)with eight SHRs per group.Eight Wistar-Kyoto(WKY)rats with normal blood pressure were used as the normal control group(WKY group).All animals were administered corresponding drugs for 8 weeks of continuous treatment.During the experiment,changes in blood pressure were observed.After the treatments,the thickness of the diastolic ventricular septum(IVSd),the thickness of the systolic ventricular septum(IVSs),and the left ventricular ejection fraction(LVEF)were measured by echocardiography to determine the degree of myocardial hypertrophy and cardiac functions.The cardiac index and left ventricular index were calculated.Hematoxylin-eosin(HE),wheat germ agglutinin(WGA),and Masson staining were performed to evaluate pathological changes of myocardial tissue.Real-time quantitative PCR(qRT-PCR)was performed to measure the mRNA expression of atrial natriuretic peptide(ANP),B-type natriuretic peptide(BNP),type I collagenα1 chain(COL1A1),and typeⅢcollagenα1 chain(COL3A1)in myocardial tissues.Western blot was performed to detect expression of protein kinase R-like endoplasmic reticulum kinase(PERK)-activator of transcription 4(ATF4)pathway-related proteins in cardiac muscle.Results After the treatments,compared with the WKY group,systolic blood pressure(SBP),diastolic blood pressure(DBP),IVSd,IVSs,cardiac index,left ventricular index,myocardial cell cross-sectional area,collagen volume fraction(CVF),myocardial tissue ANP,BNP,COL1A1 and COL3A1 mRNA expression,glucose-regulated protein 78(GRP78),ATF4,and C/EBP homologous protein(CHOP)protein levels and the p-PERK/PERK ratio were increased in the SHR group(all P<0.05),and LVEF was decreased(P<0.05).Compared with the SHR group,SBP,DBP,IVSd,IVSs,cardiac index,left ventricular index,myocardial cell cross-sectional area,CVF,myocardial tissue ANP,BNP,COL1A1,and COL3A1 mRNA expression,GRP78,ATF4,CHOP protein levels,and the p-PERK/PERK ratio were decreased in the TAX group(all P<0.05),and LVEF was increased(P<0.05).CCT020312 partially reversed the protective effects of TAX on cardiac functions and hypertrophy.Conclusions TAX improves hypertensive myocardial hypertrophy by inhibiting endoplasmic reticulum stress,and its mechanism may be related to inhibiting the PERK-ATF4 pathway.