有氧运动经CNPY2调控PERK-CHOP信号通路改善小鼠非酒精性脂肪肝的机制研究

Aerobic exercise improves non-alcoholic fatty liver diseases by regulating PERK-CHOP pathway via CNPY2

目的:采用冠层同源物2(CNPY2)基因敲除,结合运动干预,探讨CNPY2和有氧运动在高脂膳食诱导非酒精性脂肪肝(NAFLD)中的作用及机制。方法:(12±1)周龄SPF级C57BL/6J雄性CNPY2基因敲除(KO)小鼠及其同系同龄同窝野生型(WT)小鼠适应性喂养1周后,随机分为对照组、模型组和模型运动组。对照组予普通饲料喂养,模型组和模型运动组予高脂饲料喂养,连续17周。模型运动组从第10周开始进行连续有氧运动干预,直至第18周实验结束。HE和油红O染色观察肝组织病理形态;全自动生化仪检测小鼠血清高密度脂蛋白(HDL-c)、低密度脂蛋白(LDL-c)、血清总胆固醇(TC)、甘油三酯(TG)、天门冬氨酸氨基转移酶(AST)和丙氨酸转氨酶(ALT)水平;Western Blot法检测肝组织CNPY2、PERK、p-eIF2α、CHOP蛋白表达;qRT-PCR法检测肝组织PERK mRNA和eIF2αmRNA表达;Tunel法检测肝细胞凋亡。结果:WT小鼠模型组CNPY2表达较对照组显著升高(P<0.05),WT小鼠模型运动组CNPY2表达较模型组显著降低(P<0.05)。同时,与对照组比较,KO和WT小鼠模型组均可见明显肝细胞脂肪性变和脂滴,其血清ALT、AST、TC、TG和LDL-c水平、肝组织PERK、p-eIF2α、CHOP、PERK mRNA和eIF2αmRNA表达、肝细胞凋亡显著升高(P<0.05),血清HDL-c水平显著降低(P<0.05)。与模型组比较,KO和WT小鼠模型运动组上述指标得到显著改善(P<0.05)。与WT小鼠比较,KO小鼠肝组织脂肪性变减轻,肝组织CNPY2、PERK、p-eIF2α、CHOP、PERK mRNA和eIF2αmRNA表达显著降低(P<0.05)。结论:CNPY2基因缺失和有氧运动均可有效改善NAFLD,其机制可能与其下调CNPY2表达,抑制PERK-CHOP信号通路活性,降低PERK-CHOP信号通路相关分子表达,减少肝细胞凋亡有关。

Objective:To investigate the role of CNPY2 and aerobic exercise in nonalcoholic fatty liver disease(NAFLD)induced by high fat diet and relative mechanism through CNPY2 gene deletion combined with exercise intervention.Method:Thirty male CNPY2 knockout(KO)mice(12±1 weeks)and age-matched wild type(WT)littermates were randomly divided into control group,model group,and model with exercise group.Mice in model with exercise group were performed aerobic exercise training from the 10th week to the end of the experiment(week 18).HE and Oil Red O staining were used to observe the pathological morphology of liver tissues.Serum total cholesterol(TC),triglyceride(TG),high-density lipoprotein(HDL-c),low-density lipoprotein(LDL-c),aspartate aminotransferase(AST)and alanine aminotransferase(ALT)levels were measured by automatic biochemical analyzer.The protein expression of CNPY2,PERK,p-e IF2αand CHOP in liver were detected by Western blot.The m RNA expression of PERK and e IF2αin liver were detected by q RT-PCR,and hepatocyte apoptosis was detected by Tunel assay.Result:Among WT mice,the expression of CNPY2 in the model group was higher than that in the control group,and was lower in the exercise group than that in the model group.Meanwhile,compared with the control group,both KO and WT mice in the model group showed significant hepatic steatosis with significant inflammatory cell infiltration,and a large number of fat droplets in hepatocytes.The ALT,AST,TC,TG and LDL-c levels in serum,the expression of PERK,p-e IF2,CHOP,PERK m RNA and e IF2 m RNA in liver and the hepatocyte apoptosis were also higher.However,the HDL-c content in serum was lower.Compared with model group,aerobic exercise partially reversed the parameters in both KO and WT mice.Compared with WT mice,KO mice showed less hepatic steatosis and the expression of CNPY2,PERK,p-e IF2α,CHOP,PERK m RNA and e IF2αm RNA also decreased.Conclusion:Both CNPY2 gene deletion and aerobic exercise can effectively improve NAFLD,which may be related to the down-regulation of CNPY2 expression,thereby inhibiting the activity of PERK-CHOP pathway and decreasing the related molecule expression of PERK-CHOP pathway and reducing the hepatocyte apoptosis.