虎金方调节SIRT1/PGC1α信号通路改善代谢相关脂肪性肝病小鼠的脂质代谢研究

Hujin Prescription Improves Lipid Metabolism in Mice with Metabolic Associated Fatty Liver Disease by Regulating SIRT1/PGC1αSignaling Pathway

目的基于沉默信息调节因子1(SIRT1)/过氧化物酶体增殖物激活受体γ共激活因子1α(PGC1α)通路探讨虎金方对代谢相关脂肪性肝病(MAFLD)小鼠的作用及机制。方法将C57BL/6小鼠随机分为6组:正常组、模型组、水飞蓟宾组(48 mg·kg^(-1))及虎金方高(21.24 g·kg^(-1))、中(10.62 g·kg^(-1))、低(5.31 g·kg^(-1))剂量组。通过给予高脂饲料连续饲喂10周复制MAFLD小鼠模型。从第11周开始,正常组和模型组给予蒸馏水灌胃,其余各组按上述剂量灌胃给药(10 mL·kg^(-1)),每日1次,连续给药4周。采用HE及油红O染色法观察肝脏组织病理变化;生化试剂盒检测血清总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)、天门冬氨酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT)水平;RT-PCR及Western Blot法检测肝脏组织SIRT1、PGC1α、核呼吸因子1(NRF1)、线粒体转录因子A(TFAM)mRNA及蛋白表达。结果与正常组比较,模型组小鼠的体质量、肝质量及肝脏系数显著升高(P<0.01);NAS病理评分、脂滴面积占比显著升高(P<0.01);血清TC、TG、LDL-C、ALT、AST水平显著升高(P<0.01),HDL-C水平显著下降(P<0.01);肝脏组织中SIRT1、PGC1α、NRF1、TFAM mRNA及蛋白表达水平显著降低(P<0.01)。与模型组比较,各给药组小鼠的体质量、肝质量均显著降低(P<0.01),水飞蓟宾组和虎金方中、高剂量组小鼠的肝脏系数显著降低(P<0.05,P<0.01);各给药组小鼠的NAS病理评分、脂滴面积占比及血清TG、LDL-C、ALT、AST水平显著降低(P<0.05,P<0.01),水飞蓟宾组和虎金方低、高剂量组血清HDL-C水平显著升高(P<0.01),水飞蓟宾组和虎金方高剂量组血清TC水平显著降低(P<0.05,P<0.01);各给药组小鼠肝脏组织中SIRT1、TFAM mRNA表达水平显著升高(P<0.01),水飞蓟宾组和虎金方中、高剂量组小鼠肝脏组织中PGC1α、NRF1 mRNA表达水平显著升高(P<0.05,P<0.01);水飞蓟宾组和虎金方高剂量组小鼠肝脏组织中SIRT1蛋白表达水平显著升高(P<0.05,P<0.01),水飞蓟宾组和虎金方中、高剂量组小鼠肝脏组织中PGC1α、NRF1、TFAM蛋白表达水平显著升高(P<0.05,P<0.01)。结论虎金方可能通过调控SIRT1/PGC1α通路来影响肝脏线粒体的生物发生,减少线粒体氧化应激,促进肝脏脂质代谢,减轻脂质沉积,从而发挥缓解MAFLD的作用。

Objective To investigate the therapeutic effect and mechanism of Hujin Prescription on mice with metabolic associated fatty liver disease(MAFLD)based on silent information regulatory I(SIRT1)/peroxisome proliferatoractivated receptorγco-activator 1α(PGC1α)signaling pathway.Methods C57BL/6 mice were randomly divided into six groups:the normal group,the model group,silibinin group(4.8 mg·kg^(-1)),Hujin Prescription high-(21.24 g·kg^(-1)),medium-(10.62 g·kg^(-1)),and low-(5.31 g·kg^(-1))dose groups.The mice in the normal group were given the basal diet,while the other groups were given high fat diet(HFD)for 10 weeks to replicate the MAFLD animal model.At the beginning of the 11th week,the normal group and the model group were given distilled water by gavage,and the other groups were given corresponding drugs(10 mL·kg^(-1)),once a day for four weeks.The pathological changes of liver tissues were observed using HE and oil red O staining.Biochemical reagent kits were used to detect serum levels of total cholesterol(TC),triglycerides(TG),low-density lipoprotein cholesterol(LDL-C),high-density lipoprotein cholesterol(HDL-C),aspartate aminotransferase(AST),and alanine aminotransferase(ALT).The mRNA and protein expression levels of SIRT1,PGC1α,nuclear respiratory factor 1(NRF1),and mitochondrial transcription factor A(TFAM)in liver tissues were detected using RT-PCR and Western Blot.Results Compared with the normal group,the body mass,liver mass and liver index of the model group were significantly increased(P<0.01).NAS pathology score and the proportion of lipid droplets area were significantly increased(P<0.01).The serum levels of TC,TG,LDL-C,ALT and AST were significantly increased(P<0.01),but the level of HDL-C was significantly decreased(P<0.01).The mRNA and protein expression levels of SIRT1,PGC1α,NRF1,and TFAM in the liver tissues were significantly decreased(P<0.01).Compared with the model group,the body mass and liver mass of mice in each treatment group were significantly decreased(P<0.01).Moreover,liver index of mice in silibinin group,Hujin Prescription high-and medium-dose groups were significantly decreased(P<0.05,P<0.01).NAS pathology score and the proportion of lipid droplets area were significantly decreased(P<0.05,P<0.01).The serum levels of TG,LDL,ALT and AST of mice in each treatment group were significantly decreased(P<0.05,P<0.01).The serum level of HDL-C of mice in silibinin group,Hujin Prescription low-and high-dose groups was significantly increased(P<0.01),while the serum level of TC of mice in silibinin group and Hujin Prescription highdose group were significantly decreased(P<0.05,P<0.01).The mRNA expression levels of SIRT1 and TFAM in liver tissues of mice in each treatment group were significantly increased(P<0.01),but the mRNA expression levels of PGC1αand NRF1 in liver tissues of mice in silibinin group,Hujin Prescription medium-and high-dose groups were obviously increased(P<0.05,P<0.01).The protein expression levels of SIRT1 in liver tissues of mice in silibinin group and Hujin Prescription high-dose group were obviously increased(P<0.05,P<0.01).The protein expression levels of PGC1α,NRF1,and TFAM in liver tissues of mice in silibinin group,Hujin Prescription medium-and highdose groups were obviously increased(P<0.05,P<0.01).Conclusion Hujin Prescription may regulate the SIRT1/PGC1αpathway,affect the biogenesis of liver mitochondria,reduce mitochondrial oxidative stress,promote liver lipid metabolism,and reduce lipid deposition,thus exerting the effect of alleviating MAFLD.