中医正骨2025，Vol.37Issue(1)：38-44,7.
补肾强筋胶囊含药血清对膝骨关节炎软骨细胞模型自噬影响的实验研究
The effect of Bushen Qiangjin Jiaonang(补肾强筋胶囊)medicated serum on autophagy in knee osteoarthritis chondrocyte model:an experimental study
摘要
Abstract
Objective:To explore the effect of Bushen Qiangjin Jiaonang(补肾强筋胶囊,BSQJNN)medicated serum on autophagy in knee osteoarthritis(KOA)chondrocyte model based on the adenosine monophosphate activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)pathway.Methods:The human primary chondrocytes cultured in vitro were divided into normal group,model group,BSQJJN group,and AMPK inhibitor group.The chondrocytes in normal group were intervened with 10％blank serum,the ones in model group with 10％blank serum and 10 ng/mL interleukin-1 β(IL-1 β),the ones in BSQJJN group with 10％BSQJJN medicated serum and 10 ng/mL IL-1β,and the ones in AMPK inhibitor group with 10％BSQJJN medicated serum,10 ng/mL IL-1β and 10 μmol/L Dorsomor-phin(an AMPK inhibitor).After 24-hour intervention,the chondrocyte proliferation rate,chondrocyte apoptosis rate,and chondrocyte auto-phagy rate were detected by CCK-8 assay,annexin V-fluorescein isothiocyanate/propidium iodide(FITC/PI)double staining,and dansyl ca-daverine method,respectively.Furthermore,the mRNA expression levels of Beclin1,microtubule-associated protein 1 light chain 3B(LC3B),and UNC-51 like autophagy activating kinase 1(ULK1)were detected by real-time fluorescence quantitative PCR technology,and the protein expression levels of Beclin1,LC3B-Ⅰ,LC3B-Ⅱ,ULK1,phospho-AMPKα(p-AMPKα),and phospho-mTOR(p-mTOR)were de-tected by using Western blot.Results:① The chondrocyte proliferation rate.The chondrocyte proliferation rate was lower in model group compared to normal group(P=0.000),and was higher in BSQJJN group compared to model group(P=0.019),and was lower in AMPK inhibitor group compared to BSQJJN group(P=0.002).②The chondrocyte apoptosis rate.The chondrocyte apoptosis rate was higher in model group compared to normal group(P=0.000),and was lower in BSQJJN group compared to model group(P=0.000),and was higher in AMPK inhibitor group compared to BSQJJN group(P=0.000).③The chondrocyte autophagy rate.The chondrocyte autophagy rate was lower in model group compared to normal group(P=0.000),and was higher in BSQJJN group compared to model group(P=0.000),and was lower in AMPK inhibitor group compared to BSQJJN group(P=0.000).④Thie mRNA expression levels of chondrocyte autophagy-related genes.The mRNA expression levels of Beclin1,LC3B and ULK1 were lower in model group compared to normal group(P=0.002,P=0.001,P=0.003),and were higher in BSQJJN group compared to model group(P=0.008,P=0.005,P=0.012),and were lower in AMPK inhibitor group compared to BSQJJN group(P=0.000,P=0.038,P=0.004).⑤The protein expression levels of chondrocyte auto-phagy-related genes.The protein expression levels of Beclin1,ULK1,p-AMPKα and the ratio of LC3B-Ⅱ to LC3B-Ⅰ were lower,while the protein expression level of p-mTOR was higher in model group compared to normal group(P=0.028,P=0.019,P=0.007,P=0.044,P=0.025).The protein expression levels of Beclin1,ULK1,p-AMPKα and the ratio of LC3B-Ⅱ to LC3B-Ⅰ were higher,while the protein expression level of p-mTOR was lower in BSQJJN group compared to model group(P=0.004,P=0.048,P=0.040,P=0.043,P=0.031).The protein expression levels of Beclin1,ULK1,p-AMPKα and the ratio of LC3B-Ⅱ to LC3B-Ⅰ were lower,while the protein ex-pression level of p-mTOR was higher in AMPK inhibitor group compared to BSQJJN group(P=0.032,P=0.005,P=0.035,P=0.047,P=0.033).Conclusion:BSQJJN medicated serum may up-regulate the autophagy level of KOA chondrocyte model through the AMPK/mTOR pathway,thereby delaying the progression of KOA.关键词
骨关节炎,膝/补肾强筋胶囊/白细胞介素-1β/软骨细胞/自吞噬/腺苷一磷酸/蛋白激酶类/TOR丝氨酸-苏氨酸蛋白激酶Key words
osteoarthritis,knee/Bushen Qiangjin Jiaonang/interleukin-1 β/chondrocytes/autophagy/adenosine monophosphate/protein ki-nases/TOR serine-threonine kinases引用本文复制引用
胡子旋,姚楠,黄丹娥,黄雪君,甘海宁,赵自明,陈玉兴..补肾强筋胶囊含药血清对膝骨关节炎软骨细胞模型自噬影响的实验研究[J].中医正骨,2025,37(1):38-44,7.基金项目
广东省中医药局科研项目(20222011) （20222011）
广东省医学科研基金项目(B2022208) （B2022208）
广东省第二中医院科研创新基金项目(SEZYY2023A02) （SEZYY2023A02）
