益肾健骨丸对膝骨关节炎大鼠软骨组织AMPK/Cyclin Y/CDK16通路表达及软骨细胞自噬、凋亡的影响  

Effects of Yishen Jiangu Pills on AMPK/Cyclin Y/CDK16 Pathway Expressions and Chondrocyte Autophagy and Apoptosis in Cartilage Tissue of Knee Osteoarthritis Rats

作　　者：柴喜平 李盛华 周明旺 陈威 宋学文 柴英英 CHAI Xiping;LI Shenghua;ZHOU Mingwang;CHEN Wei;SONG Xuewen;CHAI Yingying(The First Affiliated Hospital of Gansu University of Chinese Medicine,Lanzhou 730050,China)

机构地区：[1]甘肃中医药大学第一附属医院,甘肃兰州730050

出　　处：《中国中医药信息杂志》2024年第10期89-95,共7页Chinese Journal of Information on Traditional Chinese Medicine

基　　金：甘肃省自然科学基金(22JR5RA634);国家中医药管理局高水平重点学科(中医骨伤科学)建设项目(203)。

摘　　要：目的观察益肾健骨丸对膝骨关节炎(KOA)大鼠软骨组织AMPK/Cyclin Y/CDK16通路及软骨细胞自噬、凋亡相关蛋白表达的影响,探讨其治疗KOA的作用机制。方法将60只2月龄SD大鼠随机分为假手术组、模型组、激动剂组和中药低、中、高剂量组,每组10只。除假手术组外,其余各组行右后膝内侧半月板切除及交叉韧带切开术建立KOA大鼠模型,并分别予相应干预14 d。采用Lequesne MG评分进行大鼠行为学评估,HE及番红O-固绿染色观察软骨组织形态,并进行Mankin评分,Western blot检测软骨组织AMPK/Cyclin Y/CDK16通路蛋白,自噬蛋白UNC-51样激酶1(ULK1)、Beclin-1、LC3B、p62,凋亡蛋白Bcl-2、Bax、胱天蛋白酶(Caspase)-3、Caspase-9表达,透射电镜观察自噬小体。结果与假手术组比较,模型组大鼠Lequesne MG评分明显升高(P<0.01),软骨表层缺损明显,软骨层变薄,软骨细胞排列杂乱、形态不规整,Mankin评分明显升高(P<0.01),软骨组织腺苷酸活化蛋白激酶(AMPK)、细胞周期素Y(Cyclin Y)、周期蛋白依赖性激酶16(CDK16)、ULK1、Beclin-1、LC3BⅡ/Ⅰ、Bcl-2蛋白表达降低,p62、Bax、Caspase-3、Caspase-9蛋白表达升高(P<0.01);与模型组比较,激动剂组和中药高剂量组大鼠Lequesne MG评分明显降低(P<0.01),中药高剂量组大鼠软骨表面较光滑,软骨细胞排列较规则,软骨层结构基本完整,软骨组织增生减弱,Mankin评分明显降低(P<0.01),激动剂组和中药中、高剂量组大鼠软骨组织AMPK、Cyclin Y、CDK16、ULK1、Beclin-1、LC3BⅡ/Ⅰ、Bcl-2蛋白表达明显升高(P<0.05,P<0.01),p62、Bax、Caspase-3、Caspase-9蛋白表达明显降低(P<0.05,P<0.01)。结论益肾健骨丸可能通过激活KOA模型大鼠软骨组织AMPK/Cyclin Y/CDK16通路促进软骨细胞自噬并抑制细胞凋亡,从而减少软骨破坏。Objective To investigate the effects of Yishen Jiangu Pills on the expressions of AMPK/Cyclin Y/CDK16 pathway and autophagy and apoptosis-related proteins in cartilage tissue of rats with knee osteoarthritis(KOA);To discuss its mechanism for the treatment of KOA.Methods Totally 602-month-old SD rats were selected and randomly divided into sham-operation group,model group,agonist group,and TCM low-,medium-and high-dosage groups,with 10 rats in each group.Except for the sham-operation group,each group underwent medial meniscectomy and cruciate ligamentotomy to establish the KOA rat model,and the corresponding interventions were given for 14 d.Lequesne MG score was used to evaluate rat behavior,morphology of cartilage tissue was observed by HE and Safranin O-fixed green staining,and Mankin score was performed,Western blot was performed to detect expression of AMPK/Cyclin Y/CDK16 pathway proteins and autophagy proteins such as ULK-1,Beclin-1,LC3B,p62 and apoptotic proteins such as Bcl-2,Bax,Caspase-3,Caspase-9 in cartilage tissue,and autophagosome were observed using transmission electron microscopy.Results Compared with the sham-operation group,Lequesne MG score significantly increased in the model group(P<0.01),there was a significant defect in the surface layer of cartilage,thinning of the cartilage layer,disordered arrangement and irregular morphology of chondrocytes,and a significant increase in Mankin score(P<0.01),the expressions of AMPK,Cyclin Y,CDK16,ULK1,Beclin-1,LC3BⅡ/Ⅰand Bcl-2 protein in cartilage tissue decreased,while the expressions of p62,Bax,Caspase-3 and Caspase-9 protein increased(P<0.01).Compared with the model group,the Lequesne MG scores of rats in the agonist group and TCM high-dosage group were significantly decreased(P<0.01),the TCM high-dosage group showed smoother cartilage surface,more regular arrangement of chondrocytes,basic integrity of cartilage layer structure,weakened cartilage tissue proliferation,and significantly decreased Mankin score(P<0.01),the expressions of AMPK,Cyclin Y

关 键 词：膝骨关节炎 益肾健骨丸 软骨细胞 自噬 AMPK通路 大鼠 

分 类 号：R285.5[医药卫生—中药学]