| 王文斌,李文高,崔文宁.微 RNA-106b-5p通过靶向 α-烯醇化酶调控类风湿关节炎滑膜成纤维细胞增殖、炎症的研究[J].安徽医药,2023,27(10):2045-2050. |
| 微 RNA-106b-5p通过靶向 α-烯醇化酶调控类风湿关节炎滑膜成纤维细胞增殖、炎症的研究 |
| Study on miR-106b-5p regulating proliferation and inflammation of synovial fibroblasts in rheumatoid arthritis through targeting α-enolase |
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| DOI：10.3969/j.issn.1009-6469.2023.10.030 |
| 中文关键词: 关节炎，类风湿 微 RNA-106b-5p α-烯醇化酶 滑膜 成纤维细胞 增殖 炎症 |
| 英文关键词: Arthritis,rheumatoid miR-106b-5p α-enolase Synovial membrane Fibroblasts Proliferation Inflammation |
| 基金项目:陕西省医学科学研究课题计划( 2018JM1255) |
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| 中文摘要: |
| 目的观察微 RNA-106b-5p、α-烯醇化酶( ENO1)对类风湿关节炎( RA)滑膜成纤维细胞( MH7A)增殖、炎症的影响，并探究二者之间的联系。方法 2018年 1月至 2021年 12月长安医院接诊手术的 8例 RA病人获得了滑膜组织， 4例意外事故引起关节损伤的无 RA、骨关节炎史健康病人作为对照。运用 RT-qPCR实验检测 RA滑膜、正常滑膜及对照组(正常 MH7A)、白细胞介素( IL)-1β组细胞( 10 μg/L IL-1β处理)中 miR-106b-5p、ENO1的表达；脂质体法将 mimic-NC组(转染 mimic)、 miR-106b-5p mimic组(转染 miR-106b-5p mimic)、 inhibitor-NC组(转染 inhibitor)、 miR-106b-5p inhibitor组(转染 miR-106b-5p inhibitor)、 pcDNA组(转染 pcDNA)、 pcDNA-ENO1组(转染 pcDNA-ENO1)、 si-NC组(转染 si-NC)、 si-ENO1组(转染 si-ENO1)、 miR-106b-5p mimic+pcDNA组(共转染 miR-106b-5p mimic+pcDNA)、 miR-106b-5p mimic+pcDNA-ENO1组(共转染 miR-106b-5p mimic+pcDNA-ENO1)、 miR-106b-5p inhibitor+si-NC组(共转染 miR-106b-5p inhibitor+si-NC)、 miR-106b-5p inhibitor+si-ENO1组(共转染 miR-106b-5p inhibitor+si-ENO1)转染至 MH7A，再 IL-1β处理。细胞计数试剂盒( CCK8)检测细胞活性；酶联免疫吸附试验(ELISA)检测细胞 IL-6、IL-8、基质金属蛋白酶 -3(MMP-3)的含量。双萤光素酶报告基因实验检测细胞荧光活性；蛋白质印迹法实验检测细胞 ENO1蛋白。结果与对照组( 0.52±0.05、147.32±12.54、0.45±0.03、1.55±0.12)相比， IL-1β组细胞活性(1.13±0.06)、 IL-6(433.21±37.84)、 IL-8(1.38±0.10)、 MMP-3(2.78±0.22)均升高( P<0.05)。与 mimic-NC组( 1.12±0.06、435.15±39.67、1.39±0.12、2.77±0.25)相比， miR-106b-5p mimic组细胞活性( 1.98±0.11)、 IL-6(164.12±15.74)、 IL-8(0.65±0.05)、 MMP-3(1.94±0.17)均降低(P<0.05)且抑制 miR-106b-5p具有相反的作用。双萤光素酶报告实验显示， miR-106b-5p靶向负调控 ENO1，且二者在 RA组织中的表达呈负相关。与 pcDNA组( 1.14±0.06、434.93±37.89、1.37±0.11、2.79±0.22)相比， pcDNA-ENO1组细胞活性(1.96±0.08)、 IL-6(867.83±71.84)、 IL-8(2.83±0.23)、 MMP-3(4.93±0.34)升高( P<0.05)；与 si-NC组相比， si-ENO1组细胞活性、 IL6、IL-8、MMP-3降低( P<0.05)。过表达 ENO1减弱 miR-106b-5p对 IL-1β处理细胞增殖、炎性因子的促进作用，敲减 ENO1减弱抑制 miR-106b-5p对 IL-1β处理细胞增殖、炎性因子的抑制作用。结论 miR-106b-5p能够抑制类风湿关节炎滑膜成纤维细胞的增殖和炎症反应，产生这种调控作用的潜在机制与靶向 ENO1有关。 |
| 英文摘要: |
| Objective To observe miR-106b-5p and α-enolase (ENO1) on the proliferation and inflammation of synovial fibroblastsin rheumatoid arthritis (RA) (MH7A), and to explore the relationship between them.Methods Synovial tissue was obtained from 8 RApatients who underwent surgery in Chang′an Hospital from January 2018 to December 2021, and 4 healthy patients with no history ofRA or osteoarthritis who had joint injuries caused by accidents were selected as control group. RT-qPCR test was used to detect the expression of miR-106b-5p and ENO1 in RA synovium, normal synovium, control group (normal MH7A) and IL-1β group cells (10 μg/L IL-1β). Mimic-NC group (transfected mimic), miR-106b-5p mimic group (transfected miR-106b-5p mimic), inhibitor-NC group (transfected inhibitor), miR-106b-5p inhibitor group (transfected miR-106b-5p inhibitor), pcDNA group (transfected pcDNA), pcDNA-ENO1 group (transfected pcDNA-ENO1), si-NC group (transfected si-NC), si-ENO1 group (transfected si-ENO1), miR-106b-5p mimic+pcDNA group (co-transfection miR-106b-5p mimic+pcDNA), miR-106b-5p mimic+pcDNA-ENO1 group (co-transfected miR-106b-5p mic+ pcDNA-ENO1), miR-106b-5p inhibitor+si-NC group (co-transfected miR-106b-5p inhibitor+si-NC), miR-106b-5p inhibitor+si-ENO1 group (co-transfected miR-106b-5p inhibitor+si-ENO1) were transfected into MH7A, and then IL-1β handle. Cell counting Kit (CCK8) was used to detect cell activity. The contents of IL-6, IL-8 and MMP-3 were detected by enzyme linked immunosorbent assay (ELISA).Cell fluorescence activity was detected by double luciferase reporter gene assay; ENO1 protein was detected by Western blotting.Re? sults Compared with the control group (0.52±0.05, 147.32±12.54, 0.45±0.03, 1.55±0.12), the cellular activity (1.13±0.06), IL-6 (433.21±37.84), IL-8 (1.38±0.10), and MMP-3 (2.78±0.22) in the IL-1β group were elevated (P<0.05). Compared with the mimic-NC group (1.12±0.06, 435.15±39.67, 1.39±0.12, 2.77±0.25), cell activity (1.98±0.11), IL-6 (164.12±15.74), IL-8 (0.65±0.05), MMP-3 (1.94±0.17) in the miR-106b-5p mimic group were reduced (P<0.05), and the inhibition of miR-106b-5p had the opposite effect. The double luciferase assay showed that miR-106b-5p targeted and negatively regulated ENO1, and their expression was negatively correlated in RA tissues. Compared with the pcDNA group (1.14±0.06, 434.93±37.89, 1.37±0.11, 2.79±0.22), the cell activity (1.96±0.08), IL6 (867.83±71.84), IL-8 (2.83±0.23), MMP-3 (4.93± 0.34) in pcDNA-ENO1 group were elevated (P<0.05). Compared with the si-NC group, the cell activity, IL-6, IL-8, and MMP-3 of the si-ENO1 group were decreased (P<0.05). Overexpression ENO1 attenuated the effect of miR-106b-5p on IL-1β treatment of cell proliferation and promotion of inflammatory factors, knockdown of ENO1 attenuated inhibition of miR-106b-5p on IL-1β treatment of cell proliferation and inhibition of inflammatory factors. Conclusions miR-106b-5pcan inhibit the proliferation and inflammatory response of rheumatoid arthritis synovial fibroblasts. The potential mechanism of this regulation may be related to ENO1 targeting. |
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