LncRNA HOTAIR靶向miR-34a-5p调节类风湿关节炎滑膜细胞凋亡的机制研究  

Mechanism of LncRNA HOTAIR targeting miR-34a-5p to regulate synovial cell apoptosis in rheumatoid arthritis

作　　者：袁礼波 谭洪波 施荣茂 李安旭 邱雄 徐永清 Yuan Libo;Tan Hongbo;Shi Rongmao;Li Anxu;Qiu Xiong;Xu Yongqing(The 920 Hospital of the Joint Logistics Support Force of the Chinese people’s Liberation Army,Kunming 650000,China)

机构地区：[1]中国人民解放军联勤保障部队第九二〇医院,昆明650000

出　　处：《广西医科大学学报》2022年第2期240-245,共6页Journal of Guangxi Medical University

基　　金：国家重点研发计划资助项目(No.2017YFC1103904);云南省卫健委临床中心建设重大项目资助(No.ZX20191001)。

摘　　要：目的:研究长链非编码RNA HOX转录反义RNA(LncRNA HOTAIR)对类风湿关节炎滑膜细胞(MH7A)凋亡的调控作用及其作用机制。方法:运用脂质体将pcDNA组(转染pcDNA)、pcDNA-HOTAIR组(转染pcDNA-HOTAIR)、antagomiRNA组(转染antagomiRNA)、antagomiR-34a-5p组(转染antagomiR-34a-5p)、pcDNA-HOTAIR+agomiRNA组(共转染pcDNA-HOTAIR和agomiRNA)、pcDNA-HOTAIR+agomiR-34a-5p组(共转染pcDNA-HOTAIR和agomiR-34a-5p)转染进MH7A细胞。实时荧光定量聚合酶链式反应(RT-qPCR)检测细胞中HOTAIR、miR-34a-5p的表达;流式细胞术、蛋白免疫印迹(Western blotting)实验检测细胞凋亡率、含半胱氨酸的天冬氨酸蛋白水解酶3(Caspase-3)、B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X基因(Bax)蛋白表达;双荧光素酶报告实验检测细胞荧光活性。结果:与HFLS组相比,MH7A组细胞HOTAIR表达升高;过表达HOTAIR后,MH7A细胞的凋亡率降低,凋亡促进蛋白Caspase-3、Bax表达降低,凋亡抑制蛋白Bcl-2蛋白表达升高,差异均有统计学意义(均P<0.05);双荧光素酶报告实验显示,HOTAIR与miR-34a-5p之间存在靶向关系。抑制miR-34a-5p后,MH7A细胞具有与过表达HOTAIR相似的抑制细胞凋亡作用。过表达miR-34a-5p部分逆转过表达HOTAIR对MH7A细胞的凋亡抑制作用。结论:LncRNA HOTAIR在RA滑膜细胞中高表达,抑制RA滑膜细胞凋亡,其机制之一为靶向抑制miR-34a-5p。Objective:To observe the regulatory effect of long chain noncoding RNA Hox transcription antisense RNA(LncRNA HOTAIR) on apoptosis of rheumatoid arthritis synovial cells(MH7 A) and investigate its mechanism.Methods:MH7 A cells were transfected into pcDNA group(transfected pcDNA),pcDNA-HOTAIR group(transfected pcDNA-HOTAIR),antagomiRNA group(transfected antagomiRNA),antagomiR-34 a-5 p group(transfected antagomiR-34 a-5 p),pcDNA-HOTAIR + agomiRNA group(co-transfected pcDNA-HOTAIR and agomiRNA),and pcDNA-HOTAIR + agomiR-34 a-5 p group(co-transfected pcDNA-HOTAIR and agomiR-34 a-5 p).Real time fluorescence quantitative polymerase chain reaction(RT-qPCR) was used to detect the expression of HOTAIR and miR-34 a-5 p in cells;The apoptosis rate,the expression of caspase-3,Bax and Bcl-2 were detected by flow cytometry and western blotting,respectively.Dual luciferase assay was employed to detect the targeting relationship between HOTAIR and miR-34 a-5 p.Results:Compared with HFLS group,the expression of HOTAIR in MH7 A group was significantly increased;After overexpression of HOTAIR,the apoptosis rate of MH7 A cells markedly decreased,the expression of apoptosis promoting proteins caspase-3 and Bax substantially decreased,and the expression of apoptosis inhibiting protein Bcl-2 substantially increased.The dual luciferase report experiment showed that there was a targeting relationship between HOTAIR and miR-34 a-5 p.After inhibiting miR-34 a-5 p,MH7 A cells had the similar inhibitory effect on apoptosis as that in overexpression of HOTAIR.Overexpression of miR-34 a-5 p partially reversed the inhibitory effect of overexpression of HOTAIR on apoptosis of MH7 A cells.Conclusion:LncRNA HOTAIR is highly expressed in RA synovial cells and inhibits apoptosis of RA synovial cell.One of its mechanisms is related to targeted-inhibition of miR-34 a-5 p.

关 键 词：类风湿关节炎滑膜细胞 长链非编码RNA HOTAIR miR-34a-5p 凋亡 

分 类 号：R332[医药卫生—人体生理学]