阿托伐他汀激活AMPK通路抑制COPD小鼠膈肌内质网应激的作用机制

Effect and mechanism of atorvastatin on inhibiting diaphragm endoplasmic reticulum stress in COPD mice via activating AMPK pathway

目的研究阿托伐他汀激活腺苷酸活化蛋白激酶(AMPK)通路抑制慢性阻塞性肺疾病(COPD)小鼠膈肌内质网应激(ERS)的作用机制。方法野生型(WT)C57BL/6J小鼠随机分为AMPK-WT组、AMPK-WT+COPD组和AMPK-WT+COPD+阿托伐他汀组,AMPK基因敲除(KO)小鼠随机分为AMPK-KO组、AMPK-KO+COPD组和AMPK-KO+COPD+阿托伐他汀组,采用香烟烟雾吸入方法建立COPD模型,并予阿托伐他汀干预。检测0.3 s用力呼气容积与用力肺活量比(FEV0.3/FVC)、最高峰值流速(PEF)、肺组织及膈肌病理改变、膈肌细胞凋亡率及膈肌中p-AMPK、蛋白激酶R样内质网激酶(PERK)、肌醇酶1α(IRE1α)、活化转录因子6(ATF6)、磷酸化真核起始因子2α(p-eIF2α)、活化转录因子4(ATF4)、C/EBP同源蛋白(CHOP)、含半胱氨酸的天冬氨酸蛋白水解酶(Caspase)-12的表达水平。结果与AMPK-WT组比较,AMPK-WT+COPD组小鼠膈肌内肌纤维断裂,膈肌中细胞凋亡率及p-AMPK、PERK、IRE1α、ATF6、p-eIF2α、ATF4、CHOP和Caspase-12表达升高;与AMPK-WT+COPD组比较,AMPK-WT+COPD+阿托伐他汀组小鼠膈肌内肌纤维断裂明显改善,膈肌中p-AMPK表达升高,膈肌中细胞凋亡率及PERK、p-eIF2α、ATF4、CHOP和Caspase-12表达降低(P<0.05),IRE1α、ATF6表达无明显改变(P>0.05);与AMPK-WT+COPD+阿托伐他汀组比较,AMPK-KO+COPD+阿托伐他汀组小鼠膈肌中不表达p-AMPK,膈肌内肌纤维断裂加重,膈肌中细胞凋亡率及PERK、p-eIF2α、ATF4、CHOP和Caspase-12表达升高(P<0.05)。结论阿托伐他汀对COPD小鼠的膈肌具有保护作用,这与阿托伐他汀通过激活AMPK通路减轻膈肌中PERK途径介导的ERS有关。

Objective To study the effect and mechanism of atorvastatin on inhibiting diaphragm endoplasmic reticulum stress(ERS)in mice with chronic obstructive pulmonary disease(COPD)via activating adenosine monophosphate-activated protein kinase(AMPK)pathway.Methods Wild type(WT)C57BL/6J mice were randomly divided into AMPK-WT group,AMPK-WT+COPD group and AMPK-WT+COPD+atorvastatin group.AMPK gene knockout(KO)mice were randomly divided into AMPK-KO group,AMPK-KO+COPD group and AMPK-KO+COPD+atorvastatin group.The COPD model was established by cigarette smoke inhalation and intervened with atorvastatin.The ratio of 0.3 s forced expiratory volume to forced vital capacity(FEV0.3/FVC),peak flow rate(peak expiratory flow,PEF),the pathological changes of lung tissue and diaphragm,the apoptosis rate of diaphragm cells,and the expression of p-AMPK,protein kinase R-like endoplasmic reticulum kinase(PERK)and inositol-requiring enzyme1α(IRE1α),activating transcription factor 6(ATF6),p-eIF2α,ATF4,C/EBP homologous protein(CHOP)and Caspase-12 were determined.Results In AMPK-WT+COPD group,the intramuscular muscle fibers in diaphragm were broken,the apoptosis rate and the expression of p-AMPK,PERK,IRE1α,ATF6,p-eIF2α,ATF4,CHOP and Caspase-12 in diaphragm were higher than those in AMPK-WT group.Compared with AMPK-WT+COPD group,the rupture of intramuscular muscle fibers in diaphragm significantly improved,the expression of p-AMPK significantly increased,the apoptosis rate and the expression of PERK,p-eIF2α,ATF4,CHOP and Caspase-12 in diaphragm significantly decreased(P<0.05),the expression level of IRE1αand ATF6 had no significant change in AMPK-WT+COPD+atorvastatin group(P>0.05).Compared with AMPK-WT+COPD+atorvastatin group,p-AMPK was not expressed in the diaphragm of AMPK-KO+COPD+atorvastatin group,the rupture of muscle fibers in the diaphragm was aggravated,and the apoptosis rate and the expression of PERK,p-eIF2α,ATF4,CHOP and Caspase-12 in diaphragm significantly increased in AMPK-KO+COPD+atorvastatin group(P<0.05).Conclusions Atorvastatin has a protective effect on the diaphragm of COPD mice,which is related to atorvastatin alleviating PERK pathway-mediated ERS in the diaphragm via activating AMPK pathway.