健肝消脂方调控PINK1/Parkin通路介导的线粒体自噬治疗非酒精性脂肪肝  

Jian'gan Xiaozhi Decoction Regulates PINK1/Parkin Pathway Mediated Mitochondrial Autophagy to Treat Non-alcoholic Fatty Liver Disease

作　　者：廖加抱 宋云 王斯 王宁 LIAO Jiabao;SONG Yun;WANG Si(Jiaxing Hospital of Traditional Chinese Medicine,Jiaxing,314000,China;Yunnan University of Chinese Medicine;The First People's Hospital of Anning City)

机构地区：[1]嘉兴市中医医院,浙江嘉兴314000 [2]云南中医药大学 [3]安宁市第一人民医院

出　　处：《浙江中医药大学学报》2024年第8期905-914,共10页Journal of Zhejiang Chinese Medical University

基　　金：国家自然科学基金青年项目(82104820);浙江省自然科学基金项目(LTGY23H270009);嘉兴市科技局应用性基础研究项目(2023AY11035)。

摘　　要：[目的]探究健肝消脂方(Jian’gan Xiaozhi Decoction,JGXZ)对非酒精性脂肪肝(non-alcoholic fatty liver disease,NAFLD)模型小鼠的药效作用,并从人第10号染色体缺失的磷酸酶及张力蛋白同源基因诱导的激酶1(PTEN induced putative kinase 1,PINK1)/E3泛素-蛋白连接酶(Parkin)信号通路介导的线粒体自噬角度探究其作用机制。[方法]60只C57BL/6J小鼠适应性喂养1周后,随机分为6组,包括正常组(Control),模型组(NAFLD),多烯磷脂酰胆碱胶囊组(polyene phosphatidyl choline,PPC)和JGXZ低、中、高剂量组。除正常组外,另外5组均给予高脂饮食。治疗过程持续8周,每周统计小鼠体质量,8周后采集小鼠血液,分离血清,测定丙氨酸氨基转移酶(alanine aminotransferase,ALT)和天冬氨酸氨基转移酶(aspartate aminotransferase,AST)水平;小鼠肝脏称重后固定,取同一部位进行苏木精-伊红(hematoxylin-eosin,HE)与油红O染色,观察肝组织病理学变化;采用试剂盒检测肝组织中的甘油三酯(triglyceride,TG)、总胆固醇(total cholesterol,TC)、白细胞介素-1β(interleukin-1β,IL-1β)、白细胞介素-6(interleukin 6,IL-6)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、丙二醛(malondialdehyde,MDA)水平,以及谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)、超氧化物歧化酶(superoxide dismutase,SOD)活性,以评估JGXZ对NAFLD小鼠炎症反应及氧化应激的影响;免疫印迹法检测电压依赖性阴离子通道蛋白1(voltage dependent anion channel 1,VDAC1)、线粒体外膜转位酶20(translocase of outer mitochondrial membrane 20,TOM20)、细胞色素C氧化酶Ⅳ亚型(cytochrome c oxidase subunitⅣ,COXⅣ)、PINK1、Parkin、苄氯素1(Beclin1)、微管相关蛋白轻链3(light chain 3,LC3)以及选择性自噬接头蛋白P62(prostacyclin 62,P62)蛋白表达,以评估JGXZ对NAFLD模型小鼠线粒体自噬的影响。[结果]与模型组比较,JGXZ干预明显提升NAFLD模型小鼠体质量,降低肝指数,缓解NAFLD模型小�[Objective]To investigate the therapeutic effect of Jian'gan Xiaozhi Decoction(JGXZ)on non-alcoholic fatty liver disease(NAFLD)model mice and explore its mechanism from the perspective of mitochondrial autophagy mediated by the PTEN induced putative kinase 1(PINK1)/E3 uniquitin-protein ligase(Parkin)signaling pathway.[Methods]Sixty C57BL/6J mice were randomly and equally divided into 6 groups:control,model(NAFLD),polyene phosphatidyl choline(PPC)group 180 mg/kg intragastric administration,and JGXZ low,medium and high dose groups(8,16,32 g/kg)intragastric administration.Except for control group,the other 5 groups were given a high fat diet.The treatment lasted for 8 weeks,and the body weight of the mice was recorded weekly.After 8 weeks,blood samples were collected,the levels of serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)were measured,liver tissue was weighted and fixed,and histological changes in liver tissue were observed by hematoxylin-eosin(HE)and oil red O staining.The levels of total cholesterol(TC),triglyceride(TG),interleukin-1β(IL-1β),IL-6,tumor necrosis factor-α(TNF-α),malondialdehyde(MDA)andactivities of glutathione peroxidase(GSH-Px),superoxide dismutase(SOD)in liver tissue were measured to evaluate the effects of JGXZ oninflammation and oxidative stress in NAFLD mice.Western blot was used to detect the protein expression levels of voltage-dependentanion channel 1(VDAC1),translocase of outer mitochondrial membrane 20(TOM20),cytochrome c oxidase subunitⅣ(COXⅣ),phosphatase and PINK1,Parkin,Beclin1,microtubule-associated protein light chain 3(LC3),and P62 to evaluate the effects of JGXZ onmitochondrial autophagy in NAFLD mice.[Result Result]Compared with model group,JGXZ intervention significantly improved the body weightof NAFLD model mice,reduced liver index,alleviated liver tissue lesions in NAFLD model mice,reduced TC,TG,ALT,AST levels,decreased IL-1β,IL-6,TNF-αand MDA levels,increased GSH-Px and SOD activity,down-regulated VDAC1,TOM20,COXⅣandP62 protein expressi

关 键 词：健肝消脂方 非酒精性脂肪肝 PINK1/Parkin信号通路 线粒体自噬 氧化应激 炎症 

分 类 号：R331[医药卫生—人体生理学]