白芍总苷经活性氧-线粒体途径改善骨关节炎软骨细胞自噬性死亡  

Total glucosides of paeony ameliorated autophagic death of osteoarthritis chondrocytes by reactive oxygen species-mitochondria pathway

作　　者：廖太阳 李晓辰 杨楠 茆军 王培民 邢润麟 LIAO Taiyang;LI Xiaochen;YANG Nan;MAO Jun;WANG Peimin;XING Runlin(Department of Orthopaedics and Traumatology,Affilliated Hospital of Nanjing University of Chinese Medicine&Jiangsu Province Hospital of Chinese Medicine,Nanjing 210029,Jiangsu,China)

机构地区：[1]南京中医药大学附属医院(江苏省中医院)骨伤科,南京210029

出　　处：《医学研究与战创伤救治》2023年第4期337-342,共6页Journal of Medical Research & Combat Trauma Care

基　　金：国家自然科学基金(81804123);南京中医药大学自然科学基金项目(XZR2020005)。

摘　　要：目的探讨白芍总苷(TGP)对白细胞介素-1β(IL-1β)诱导小鼠关节软骨细胞(MACs)的保护作用及可能机制。方法体外培养MACs细胞并通过甲苯胺蓝染色和Ⅱ型胶原免疫荧光染色鉴定;CCK-8检测TGP对MACs细胞活性的影响,根据检测结果确定后续实验的TGP剂量范围;采用10 ng/mL的IL-1β构建MACs细胞OA模型,予以不同剂量TGP(10、20及50μmol/L)处理细胞48 h,分组为:空白组,模型组,低、中、高剂量TGP干预组,DCFH-DA荧光探针检测MACs细胞中ROS荧光强度;JC-1荧光探针观察MACs细胞线粒体电位变化;流式检测MACs细胞死亡率;RT-PCR检测Beclin 1、LC3、MMP-3和MMP-13基因表达水平。结果与空白组细胞死亡率[(6.58±1.65)%]比较,模型组[(26.77±0.93)%]显著上升(P<0.05),TGP干预后,细胞死亡率显著下降;与空白组ROS荧光强度[(1.90±0.56)%]比较,模型组[(39.10±0.36)%]显著增强(P<0.05),应用TGP后ROS荧光强度显著下降(P<0.05);与空白组比较,模型组线粒体膜电位下降,而TGP组线粒体膜电位较模型组上升(P<0.05);此外,模型组Beclin 1及LC3 mRNA表达水平较空白组降低,而TGP组较模型组升高,差异具有统计学意义(P<0.05);模型组MMP-3及MMP-13 mRNA表达水平较空白组升高,而TGP组较模型组降低(P<0.05)。结论白芍总苷可通过活性氧-线粒体途径改善骨关节炎软骨细胞自噬性死亡,发挥软骨保护功效。Objective Few studies have been reported on the effect of total peony glycosides(TGP)on osteoarthritis(OA).The aim of this study was to investigate the protective effect of TGP on murine articular chondrocytes(MACs)induced by interleukin-1β(IL-1β)and its possible mechanism.Methods MACs were cultured in vitro and identified by toluidine blue staining and type II collagen immunofluorescence staining.CCK-8 was used to detect the effect of TGP on the cell viability of MACs.And the concentrations of TGP in subsequent experiments were determined according to the detection results.MACs cell OA model was established by IL-1β,and different concentrations of TGP intervention(10,20 and 50μmol/L)were administered.Cell experiments included five groups:blank group,model group,TGP low,medium and high concentration group.The content of ROS in MACs were detected by DCFH-DA.Mitochondrial potential changes in MACs observed by JC-1 fluorescent probe.Mortality of MACs was detected by flow cytometry.The mRNA expression levels of Beclin 1,LC3,MMP-3 and MMP-13 were detected by RT-PCR.Results Compared with the blank group[(6.58±1.65)%],the cell death rate in the model group[(26.77±0.93)%]was significantly increased(P<0.05).After TGP intervention,the cell death rate was significantly decreased(P<0.05).The ROS fluorescence intensity[(39.10±0.36)%]was significantly enhanced in the model group(P<0.05)and significantly decreased after the application of TGP(P<0.05).Mitochondrial membrane potential detection showed that the mitochondrial membrane potential of the model group was significantly decreased compared with the blank group.After the intervention of TGP,the mitochondrial membrane potential was significantly increased(P<0.05).The mRNA expression levels of Beclin 1 and LC3 in the model group were lower than that of the blank group,while the TGP group was higher than the model group(P<0.05).The mRNA expression levels of MMP-3 and MMP-13 were increased compared with the blank group,while the TGP group was lower than the model group(

关 键 词：白芍总苷 活性氧 线粒体 小鼠关节软骨细胞 骨关节炎 

分 类 号：R684.3[医药卫生—骨科学]