基于转录组测序探讨金天格胶囊调控滑膜间充质干细胞外泌体miRNA和关节软骨细胞mRNA治疗大鼠骨关节炎的作用机制  

Exploring the Mechanism of Action of Jintiange Capsules in Regulating SMSC-Exos miRNA and Articular Chondrocytes mRNA for the Treatment of Osteoarthritis in Rats Based on Transcriptome

作　　者：王智超 张雪 张小飞 史亚军 郭东艳 栾飞 翟秉涛 邹俊波 袁普卫[3] WANG Zhichao;ZHANG Xue;ZHANG Xiaofei;SHI Yajun;GUO Dongyan;LUAN Fei;ZHAI Bingtao;ZOU Junbo;YUAN Puwei(Shaanxi Province Key Laboratory of New Drugs and Chinese Medicine Foundation Research,College of Pharmacy,Shaanxi University of Chinese Medicine,Xianyang 712046,China;Affiliated Hospital of Shaanxi University of Chinese Medicine,Xianyang 712046,China;The First Clinical College,Shaanxi University of Chinese Medicine,Xianyang 712046,China)

机构地区：[1]陕西中医药大学药学院,陕西省中药基础与新药研究重点实验室,陕西咸阳712046 [2]陕西中医药大学附属医院,陕西咸阳712046 [3]陕西中医药大学第一临床医学院,陕西咸阳712046

出　　处：《中国现代应用药学》2024年第11期1464-1475,共12页Chinese Journal of Modern Applied Pharmacy

基　　金：国家中医药管理局青年岐黄学者培养项目[国中医药人教函(2022)256号];陕西省重点研发计划资助项目(2022SF-238);陕西省中医药管理局中医药重点学科建设项目(2017001);陕西省中医药管理局中医药“双链融合”中青年科研创新团队项目(2022-SLRH-LJ-001);陕西中医药大学学科创新团队项目(2019-YL11)。

摘　　要：目的对金天格胶囊治疗骨关节炎的药效进行确证,基于高通量测序技术探讨滑膜间充质干细胞外泌体(synovial mesenchymal stem cell exosomes,SMSC-Exos)与关节软骨细胞(articular chondrocytes,ACs)治疗骨关节炎的潜在作用机制。方法采用Ⅱ型胶原酶诱导大鼠骨关节炎,通过一般行为学观察、双足平衡差异试验、机械足反射阈值、MicroCT观察与番红固绿染色进行药效验证。将SMSC、ACs在适宜浓度的含药血清中培养,对照组、模型组和金天格组ACs进行mRNA测序以及SMSC-Exos进行miRNA测序。筛选差异表达mRNA与差异表达miRNA,并进行靶基因预测。通过Venn图对SMSC与ACs的差异表达基因求交集得到共同差异表达基因,运用Cytoscape软件构建miRNA-m RNA调控网络。对共同差异表达基因进行表达趋势分析以及对差异表达基因mRNA与miRNA、Micro-CT药效指标与差异表达基因mRNA进行相关性分析。结果骨关节炎病理状态下,miRNA-23a-3p、miRNA-342-3p、miRNA-146b-5p、mi RNA-501-3p、miRNA-214-3p表达下调,miRNA-222-3p、miRNA-30e-3p、miRNA-676-3p、miRNA-192-5p表达上调(P<0.05),这些miRNA的表达均在金天格胶囊含药血清干预后逆转表达,且趋势明显。Micro-CT药效指标、m RNA与miRNA之间存在一定的相关性。结论金天格胶囊治疗骨关节炎的药效作用明显,其作用机制可能与促进SMSC-Exos靶向ACs转运miRNA进而调控Serpinb10、Ntn1、Il1b、Tgm2、Megf10、Il11、Cd40、Slc15a3、Pou2f2等基因有关。OBJECTIVE To confirm the therapeutic effect of Jintiange capsules on osteoarthritis(OA)and the potential mechanism of synovial mesenchymal stem cell exosomes(SMSC-Exos)and articular chondrocytes(ACs)in the treatment of OA based on high-throughput sequencing technology.METHODS TypeⅡcollagenase-induced OA rats were used for efficacy verification through general behavioral observation,bipedal balance difference experiment,mechanical foot reflex threshold,Micro-CT observation,and Safranin O-Fast Green staining.SMSCs and ACs were cultured in suitable concentration of drug-containing serum,and mRNA sequencing was performed on ACs in the control,model,and Jintiange capsules groups,as well as miRNA sequencing on SMSC-Exos.Differential expressed mRNAs and miRNAs were screened and target genes were predicted.The common differential expressed genes between SMSC and ACs were obtained by intersecting the differential expressed genes,and a miRNA-mRNA regulatory network was constructed using Cytoscape software.The expression trend analysis of common differential expressed genes was conducted,as well as the correlation analysis between differential expressed gene mRNA and miRNA,Micro-CT efficacy indicators,and differential expressed gene mRNA.RESULTS Under the pathological state of OA,the expression of miRNA-23a-3p,miRNA-342-3p,miRNA-146b-5p,miRNA-501-3p,and miRNA-214-3p were down-regulated,while miRNA-222-3p,miRNA-30e-3p,miRNA-676-3p,and miRNA-192-5p were up-regulated(P<0.05).The expressions of these miRNAs were significantly reversed after intervention with drug-containing serum of Jintiange capsules.There was a certain correlation between Micro-CT efficacy indicators,mRNA and miRNA.CONCLUSION Jintiange capsule has obvious efficacy in the treatment of OA,and its mechanism may be related to the promotion of SMSC-Exos targeting ACs to transport miRNA and then regulate Serpinb10,Ntn1,Il1b,Tgm2,Megf10,Il11,Cd40,Slc15a3,Pou2f2 and other genes.

关 键 词：骨关节炎 转录组测序 金天格胶囊 滑膜间充质干细胞外泌体 关节软骨细胞 

分 类 号：R966[医药卫生—药理学]