桃仁膝康丸通过PI3K/AKT/mTOR信号通路激活软骨细胞自噬抗膝骨性关节炎的机制研究  

Taoren Xikang Pill Inhibits Knee Osteoarthritis by Activating Chondrocyte Autophagy Through PI3K/AKT/mTOR Signaling Pathway

作　　者：秦娜[1] 魏立伟[1] 张虹[1] Qin Na;Wei Liwei;Zhang Hong(Luoyang Orthopedic Traumatology Hospital of Henan Province·Henan Provincial Orthopedic Hospital,Luoyang 471000)

机构地区：[1]河南省洛阳正骨医院·河南省骨科医院,洛阳471000

出　　处：《中药药理与临床》2021年第4期165-169,共5页Pharmacology and Clinics of Chinese Materia Medica

基　　金：河南省中医药科学研究专项课题(编号:2016ZY2085)。

摘　　要：目的:探讨桃仁膝康是否通过抑制磷脂酰肌醇3-激酶(PI3K)/丝氨酸/苏氨酸激酶(AKT)/哺乳动物雷帕霉素靶蛋白(mTOR)信号通路诱导大鼠骨性关节炎(OA)软骨细胞自噬。方法:采用改良的Hulth法制备大鼠膝关节OA模型,造模成功后随机分为:模型对照组、桃仁膝康丸1.25 g/kg组和桃仁膝康丸1.25 g/kg^(+)3-甲基腺嘌呤(3-MA)1.5 mg/kg组,另设假手术对照组。术后2 w开始灌胃,给予桃仁膝康丸或生理盐水,腹腔注射给予3-甲基腺嘌呤,连续用药12 w。HE和番红O-固绿染色观察软骨组织形态学变化并进行Mankin评分,Western blot检测软骨组织中ULK1、Beclin-1、微管相关蛋白1轻链3(LC3)-II/I、磷酸化PI3K (p-PI3K)、磷酸化AKT (p-AKT)、磷酸化mTOR(p-mTOR)蛋白的表达,Real-time RT PCR检测膝关节软骨组织中Pi3k、Akt和Mtor mRNA的表达。结果:与假手术对照组比较,模型对照组Mankin评分显著升高(P<0.05),软骨组织中ULK1、Beclin-1和LC3-II/I蛋白质表达明显下调(P<0.05),Pi3k、Akt和Mtor mRNA的表达明显上调和p-PI3K、p-AKT和p-mTOR蛋白质与相应总蛋白的比值增加(P<0.05);与模型对照组比较,桃仁膝康丸能明显降低Mankin评分(P<0.05),上调软骨组织中ULK1、Beclin-1蛋白和增加LC3-II/I比值(P<0.05),同时下调Pi3k、Akt、Mtor mRNA的表达和降低p-PI3K、p-AKT和p-mTOR蛋白质与相应总蛋白的比值(P<0.05)。结论:桃仁膝康可以通过抑制PI3K/AKT/mTOR信号通路诱导软骨细胞自噬,从而延缓大鼠膝关节OA的软骨退变。Objective:To study whether Taoren Xikang Pill induces chondrocyte autophagy in rats with osteoarthritis(OA) by inhibiting the phosphoinositide 3-kinase(PI3 K)/serine/threonine kinase AKT/mammalian target of rapamycin(mTOR) signaling pathway. Methods: The rat OA model was established using the improved Hulth’s method. After successful modeling, the rats were randomly divided into the normal control group, 1.25 g/kg Taoren Xikang Pill group, 1.25 g/kg Taoren Xikang Pill ^(+) 1.5 mg/kg autophagy inhibitor 3-methyladenine(3-MA) group, and the sham operation group. Two weeks after operation, the rats were orally administrated with Taoren Xikang Pill, normal saline, or intraperitoneally injected with 3-MA for 12 successive weeks. The morphological changes in cartilage tissue were observed by HE and safranin O staining, followed by Mankin scoring. The protein expression levels of ULK1, Beclin-1, microtubule-associated protein 1 light chain 3(LC3-II/I), phosphorylated PI3 K(p-PI3 K), phosphorylated AKT(p-AKT) and phosphorylated mTOR(p-mTOR) were assayed by Western blotting, and the mRNA expression levels of PI3 K, AKT, and mTOR were detected by Real-time PCR. Results: Compared with the sham operation group, the model group exhibited significantly increased Mankin score(P<0.05), down-regulated ULK1, Beclin-1, and LC3-II/I protein expression in cartilage tissue(P<0.05), up-regulated PI3 K, AKT, and mTOR mRNA expression, and elevated p-PI3 K/total protein, p-AKT/total protein, and p-mTOR/total protein ratios(P<0.05). Compared with the model group, Taoren Xikang Pill significantly reduced Mankin score(P<0.05), up-regulated ULK1, Beclin-1, and LC3-II/I protein expression in cartilage(P<0.05), down-regulated Pi3k,Akt, and Mtor mRNA expression, and decreased p-PI3 K/total protein, p-AKT/total protein, and p-mTOR/total protein ratios(P<0.05). Conclusion: Taoren Xikang Pill induces chondrocyte autophagy by inhibiting the PI3 K/AKT/mTOR signaling pathway, thus delaying knee cartilage degeneration of OA rats.

关 键 词：桃仁膝康丸 磷脂酰肌醇3-激酶/丝氨酸/苏氨酸激酶/哺乳动物雷帕霉素靶蛋白信号通路 自噬 骨性关节炎 

分 类 号：R285.5[医药卫生—中药学]