基于线粒体自噬研究补肝散对衰老大鼠肝脏损伤的影响  

Effects of Bugan Powder on liver injury in aging rats based on mitophagy

作　　者：张月 白德高 霍冉 王杰鹏[1,2] 方芳 方朝义[1,2] 王少贤 ZHANG Yue;BAI De-gao;HUO Ran;WANG Jie-peng;FANG Fang;FANG Chao-yi;WANG Shao-xian(Hebei University of Chinese Medicine,Shijiazhuang 050200,China;Hebei Key Laboratory of Integrated Chinese and Western Medicine for Lung Disease Research,Shijiazhuang 050091,China)

机构地区：[1]河北中医学院,石家庄050200 [2]河北省中西医结合肺病研究重点实验室,石家庄050091

出　　处：《中华中医药杂志》2023年第3期1039-1045,共7页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：河北省中医药类科研计划项目(No.2020136);河北中医学院省属高校基本科研业务费专项项目(No.YJZ2019006);河北中医学院研究生创新资助项目(No.XCXZZSS2021002)。

摘　　要：目的:基于肝气虚理论和线粒体自噬探讨衰老机制,观察补肝散对衰老大鼠肝脏损伤的影响。方法:SPF级雄性SD大鼠32只,随机分为空白组、模型组、补肝散组、雷帕霉素组,每组8只。除空白组腹腔注射0.9%氯化钠溶液(10 mL/kg)外,其余各组均腹腔注射D-半乳糖溶液(D-gal,400 mg/kg),复制衰老大鼠模型。造模同时,补肝散组灌服补肝散煎液(14.06 g/kg),雷帕霉素组灌服雷帕霉素溶液(1 mg/kg),空白组和模型组灌服等量0.9%氯化钠溶液,连续56 d。测定各组大鼠体质量、抓力,血清丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性及丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、碱性磷酸酶(ALP)含量。HE染色观察各组大鼠肝组织病理学变化;RT-qPCR检测线粒体融合蛋白(Mfn)1、Mfn2、自噬关键分子酵母Atg6同系物1(Beclin1)、微管相关蛋白1轻链3(LC3)、p62 mRNA表达水平;免疫组化染色(ABC法)检测Beclin1、p62蛋白表达;Western Blot测定Mfn1、Mfn2、LC3蛋白表达。结果:与空白组比较,模型组大鼠出现肝细胞水肿,肝血窦受挤压变窄,轻度脂肪变性及少量肝细胞坏死,另可见肝细胞嗜酸性病变及凋亡的肝细胞;体质量和抓力显著降低(P<0.05);血清MDA含量升高(P<0.05),SOD活性降低(P<0.05),ALT、AST、ALP含量升高(P<0.05);肝组织中Mfn1、Mfn2、p62 mRNA及蛋白表达水平升高(P<0.05),Beclin1、LC3 mRNA及蛋白表达水平降低(P<0.05)。与模型组比较,补肝散组大鼠肝细胞水肿减轻,脂肪变性减少,嗜酸性病变及凋亡的肝细胞明显减少;体质量和抓力显著升高(P<0.05);血清MDA含量降低(P<0.05),SOD活性升高(P<0.05),ALT、AST、ALP含量降低(P<0.05);肝组织中Mfn1、Mfn2、p62 mRNA及蛋白表达水平降低(P<0.05),Beclin1、LC3 mRNA及蛋白表达水平升高(P<0.05)。结论:衰老大鼠肝脏损伤与线粒体自噬功能下降关系密切,可作为肝虚致衰的重要机制;补肝散可通过提高肝脏线粒体自噬水平,减缓衰�Objective:Based on the theory of liver-qi deficiency and mitochondrial autophagy,to explore the aging mechanism and observe the effects of Bugan Powder on liver injury in aging rats.Methods:Thirty-two SPF male SD rats were randomly divided into blank group,model group,Bugan powder group and rapamycin group,with 8 rats in each group.Except the blank group,which was intraperitoneally injected with 0.9%sodium chloride solution(10 mL/kg),all the other groups were intraperitoneally injected with D-gal solution(400 mg/kg)to replicate the aging rat model.At the same time,Bugan Powder group was given Bugansan decoction(14.06 g/kg),rapamycin group was given rapamycin solution(1 mg/kg),and the blank group and model group were given the same amount of 0.9%sodium chloride solution for 56 days.The body mass,grasping power,serum MDA content,SOD activity,ALT,AST and ALP content of rats in each group were measured.HE staining was used to observe the pathological changes of liver tissue in each group.RT-qPCR was used to detect the mrna expression levels of Mfn1,Mfn2,Beclin1,LC3 and p62.Immunohistochemical staining(ABC method)was used to detect the expression of Beclin1 and p62 proteins.The expression of Mfn1,Mfn2 and LC3 protein was determined by Western Blot.Results:Compared with the blank group,in the model group,there were edema of liver cells,narrowing of hepatic sinuses,mild steatosis and a small amount of necrosis of liver cells,as well as eosinophilic lesions and apoptotic liver cells.Body mass and grip decreased(P<0.05);Serum MDA content increased(P<0.05),SOD activity decreased(P<0.05),ALT,AST and ALP content increased(P<0.05).The mRNA and protein expression levels of Mfn1,Mfn2 and p62 in liver tissue increased(P<0.05),while the mRNA and protein expression levels of Beclin1 and LC3 decreased(P<0.05).Compared with the model group,the liver edema,fatty degeneration,eosinophilic lesions and apoptotic liver cells in the Bugan Powder group were significantly reduced.Body mass and grip increased(P<0.05);Serum MDA content decrea

关 键 词：衰老 线粒体自噬 肝脏损伤 肝气亏虚 补肝散 D-半乳糖 

分 类 号：R285.5[医药卫生—中药学]