摘要
目的:探讨miR-330-3p在颞下颌关节骨关节炎(temporomandibular joint osteoarthritis,TMJOA)软骨退变中的作用机制。方法:对SW1353软骨细胞系进行炎症及加压刺激,建立大鼠单侧前牙反(牙合)(unilateral anterior crossbite,UAC)TMJOA模型,通过RT-qPCR检测体外及体内软骨OA情况下miR-330-3p的表达变化。miR-330-3p过表达或抑制后,利用EDU荧光染色以及蛋白免疫印迹(WB)检测SW1353软骨细胞的增殖及合成分解代谢变化。生物信息学分析预测并筛选miR-330-3p的下游靶点,并在炎症及加压刺激下通过RT-qPCR和WB明确miR-330-3p对其调控作用。在miR-330-3p大鼠UAC TMJOA模型的关节腔内注射后,采用免疫组织化学方法检测靶分子的表达及软骨代谢的变化,明确miR-330-3p对TMJOA的治疗作用。采用SPSS 22.0软件包对数据进行统计学分析。结果:在体外炎症和加压刺激下,以及大鼠UAC的TMJOA模型中,miR-330-3p在软骨中的表达均下降。抑制miR-330-3p可导致软骨细胞增殖能力下降,SOX9合成降低,MMP13表达增加,而过表达的结果则相反。生物信息学分析发现CTNNB1为可能的下游靶点之一,其在体外炎症和加压刺激后表达升高。过表达CTNNB1后,软骨细胞SOX9蛋白表达下降,MMP13蛋白表达升高,抑制CTNNB1的结果则相反。miR-330-3p与CTNNB1在mRNA及蛋白表达水平上存在负相关关系。大鼠UAC TMJOA模型关节腔内注射miR-330-3p模拟物后,CTNNB1的表达减少,软骨退变减轻。结论:miR-330-3p通过抑制CTNNB1的表达,减轻TMJOA的软骨退变。
PURPOSE:To study the mechanism of miR-330-3p in cartilage degeneration in temporomandibular joint osteoarthritis(TMJOA)in rats.METHODS:Human chondrocytes SW1353 was stimulated by inflammation and stress respectively and rat TMJOA model was established through unilateral anterior crossbite(UAC).The expression of miRNA-330-3p in vitro and in vivo was detected by RT-qPCR.The proliferation and metabolism changes of SW1353 were detected by EDU fluorescence staining and immunoblotting after miRNA-330-3p overexpression or inhibition.Furthermore,bioinformatics analysis was performed to predict and screen the downstream targets of miRNA-330-3p.RT-qPCR and immunoblotting were used to clarify the regulation of miRNA-330-3p on the target and the function of target.After intraarticular injection of miRNA-330-3p in TMJOA rats,the changes in cartilage metabolism were detected by immunohistochemistry to clarify the effect of miRNA-330-3p on TMJOA.SPSS 22.0 software package was used to analyze the data.RESULTS:The expression of miRNA-330-3p decreased in chondrocytes under stress or inflammatory stimuli in vitro and in TMJOA rats.Inhibition of miRNA-330-3p decreased chondrocyte proliferation,reduced SOX9 and increased MMP13 ex pression,while overexpression of miRNA-330-3p had opposite effects.Bioinformatics analysis revealed that CTNNB1 was one of the possible downstream targets,and its expression was elevated after inflammatory/stress stimuli in vitro and in TMJOA rats.SOX9 expression decreased,and MMP13 expression increased after overexpression of CTNNB1,with opposite results for inhibition of CTNNB1.There was a negative relationship between miRNA-330-3p and CTNNB1.Intra-articular injection of miRNA-330-3p in TMJOA rats alleviated cartilage degeneration.CONCLUSIONS:miRNA-330-3p attenuates cartilage degeneration in TMJOA by inhibiting the expression of CTNNB1.
作者
王楚瑶
邹璐芗
陆川
何冬梅
WANG Chu-yao;ZOU Lu-xiang;LU Chuan;HE Dong-mei(Department of Oral Surgery,Shanghai Ninth People's Hospital,Shanghai Jiao Tong University School of Medicine,College of Stomatology,Shanghai Jiao Tong University,National Center for Stomatology,National Clinical Research Center for Oral Diseases,Shanghai Key Laboratory of Stomatology.Shanghai 200011;School of Stomatology,Qingdao University.Qingdao 266000,Shandong Province,China Received:2022-04-25 Revised:2022-06-01 Online:2022-07-20 Published:2022-08-02)
出处
《中国口腔颌面外科杂志》
CAS
2022年第4期320-327,共8页
China Journal of Oral and Maxillofacial Surgery
基金
国家重点研发计划(2017YFC1103900,2020YFC2002800)
国家自然科学基金(32071313)
上海市科委科研项目(20S31902500,20Y11903900)。
