桃叶珊瑚苷对膝骨关节炎小鼠的作用及机制实验研究  

Effects and mechanism of aucubin on knee osteoarthritis in mice

作　　者：夏红丽 田俊斌[2] 李治琴 XIA Hongli;TIAN Junbin;LI Zhiqin(Xi’an No.3 Hospital,Xi’an 710021,China)

机构地区：[1]西安市第三医院,陕西西安710021 [2]西安交通大学第二附属医院,陕西西安710004

出　　处：《陕西医学杂志》2023年第9期1150-1154,1167,共6页Shaanxi Medical Journal

基　　金：陕西省自然科学基础研究计划项目(S2021-JC-YB-0939)。

摘　　要：目的:探讨桃叶珊瑚苷对膝骨关节炎(KOA)小鼠的作用及对细胞自噬和磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(PI3K/AKT/mTOR)信号通路的影响。方法:采用改良的Hulth法制备膝关节KOA小鼠模型,随机分为模型组、桃叶珊瑚苷组、激动剂组、桃叶珊瑚苷+激动剂组,另设正常对照组,每组各10只。桃叶珊瑚苷组小鼠给予80 mg/kg桃叶珊瑚苷灌胃;激动剂组小鼠腹腔注射0.5μmol/kg 740Y-P;桃叶珊瑚苷+激动剂组小鼠给予80 mg/kg桃叶珊瑚苷灌胃,同时0.5μmol/kg 740Y-P腹腔注射;正常对照组和模型组小鼠分别灌胃和腹腔注射等体积0.9%氯化钠溶液,1次/d,连续12周。采用HE染色和番红O-固绿染色观察软骨组织病理形态学变化,Mankin评分评估KOA严重程度。ELISA法检测软骨组织中白细胞介素-6(IL-6)、IL-1β和肿瘤坏死因子-α(TNF-α)水平。实时荧光定量PCR(RT-qPCR)法检测自噬相关基因(Atg5、Atg12、Baclin-1、LC3)、PI3K、AKT和mTOR mRNA水平。蛋白印迹法检测自噬相关蛋白、LC3Ⅱ/Ⅰ、p-PI3K、p-AKT、p-mTOR蛋白水平。结果:与正常对照组比较,模型组关节软骨表面粗糙,细胞结构层次混乱、数量减少,潮线排列破坏严重;激动剂组软骨组织病变程度更重;桃叶珊瑚苷+激动剂组小鼠关节软骨表面稍光滑,细胞排列相对整齐、数量稍减少,潮线受损程度减轻;桃叶珊瑚苷组软骨组织病变程度明显改善。与模型组比较,桃叶珊瑚苷组Mankin评分、IL-6、IL-1β和TNF-α水平以及PI3K、AKT和mTOR mRNA水平降低,Atg5、Atg12和Baclin-1 mRNA和蛋白表达升高,LC3 mRNA和LC3Ⅱ/Ⅰ升高,p-PI3K、p-AKT和p-mTOR蛋白水平降低(均P<0.05)。与桃叶珊瑚苷组比较,桃叶珊瑚苷+激动剂组Mankin评分、IL-6、IL-1β和TNF-α水平以及PI3K、AKT和mTOR mRNA水平升高,Atg5、Atg12和Baclin-1 mRNA和蛋白表达降低,LC3 mRNA和LC3Ⅱ/Ⅰ降低,p-PI3K、p-AKT和p-mTOR蛋白水平升高(均P<0.05)。与激动剂组比Objective:To investigate the effects of aucubin on autophagy and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/AKT/mTOR)signaling pathway in knee osteoarthritis(KOA)mice.Methods:A knee KOA mouse model was established by modified Hulth method and randomly divided into model group,aucubin group,agonist group,and aucubin+agonist group,and normal control group was set up,with 10 mice in each group.The mice in the aucubin group were given 80 mg/kg aucubin by gavage,the mice in agonist group were intraperitoneally injected with 0.5μmol/kg 740Y-P,the mice in the aucubin+agonist group were given 80 mg/kg aucubin by gavage and 0.5μmol/kg 740Y-P by intraperitoneal injection,the mice in the normal control group and the model group were given intragastric administration and intraperitoneal injection of the same volume of 0.9%sodium chloride solution,once a day for 12 weeks,respectively.The pathological changes of cartilage tissue were observed by HE staining and saffron O-solid green staining,and the severity of KOA was evaluated by Mankin score.The levels of IL-6,IL-1βand TNF-αin cartilage tissue were detected by ELISA.The mRNA levels of autophagy-related genes(Atg5,Atg12,Baclin-1,LC3),PI3K,AKT and mTOR were detected by RT-qPCR.The protein levels of autophagy-related proteins,LC3Ⅱ/Ⅰ,p-PI3K,p-AKT,and p-mTOR were detected by Western blot.Results:Compared with the normal control group,the articular cartilage surface of the model group was rough,the cell structure was disorderly,the number of cells was reduced,and the tide line arrangement was seriously damaged.The pathological changes of cartilage tissue in agonist group were more severe.The articular cartilage surface of mice in the aucubin+agonist group was slightly smooth,the cells were relatively orderly arranged,the number of cells was slightly reduced,and the damage degree of the tide line was alleviated.The pathological changes of cartilage tissue in the aucubin group were significantly improved.Compared with the model group,t

关 键 词：膝骨关节炎 桃叶珊瑚苷 磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白信号通路 自噬 小鼠 

分 类 号：R285.5[医药卫生—中药学]