抑制LINC00667表达对类风湿关节炎成纤维样滑膜细胞增殖、迁移、侵袭的影响

Effects of inhibition of IC06LINC00667 expression on proliferation, migration and invasion of fibroblastic synovial cells in rheumatoid arthritis

摘要:

目的 探究长链非编码RNA（LncRNA）LINC00667对类风湿关节炎成纤维样滑膜细胞（RA-FLS）增殖、迁移、侵袭的影响及其机制。方法 复制类风湿关节炎（RA）模型大鼠，SPF级Wistar大鼠分离踝关节滑膜组织，用实时荧光定量聚合酶链反应（qRT-PCR）检测RA-FLS中LINC00667和miR-19a-3p的表达。在RA-FLS中分别转染si-NC（si-NC组）、si-LINC00667（si-LINC00667组）、miR-NC（miR-NC组）、miR-19a-3p模拟物（miR-19a-3p组）、si-LINC00667与anti-miR-NC（si-LINC00667+ anti-miR-NC组）、si-LINC00667与anti-miR-19a-3p（si-LINC00667+ anti-miR-19a-3p组）。CCK-8法检测各组转染后RA-FLS的增殖抑制率。Transwell法检测RA-FLS迁移、侵袭。Western blotting检测E钙黏蛋白（E-cadherin）、N钙黏蛋白（N-cadherin）表达。双萤光素酶报告验证LINC00667和miR-19a-3p的靶向关系。结果 RA模型组滑膜组织中LINC00667表达量约为正常组的270%，miR-19a-3p表达量约为正常组的36%（P <0.05）。转染成功后，si-LINC00667组RA-FLS的增殖抑制率、E-cadherin蛋白相对表达量高于si-NC组（P <0.05），si-LINC00667组迁移、侵袭细胞数、N-cadherin蛋白相对表达量低于si-NC组（P <0.05）。LINC00667靶向miR-19a-3p，miR-19a-3p组RA-FLS的增殖抑制率、E-cadherin蛋白相对表达量高于miR-NC组（P <0.05），miR-19a-3p组迁移、侵袭细胞数、N-cadherin蛋白相对表达量低于miR-NC组（P <0.05）。si-LINC00667+ anti-miR-19a-3p组RA-FLS的增殖抑制率、E-cadherin蛋白相对表达量低于si-LINC00667+ anti-miR-NC组（P <0.05），si-LINC00667+ anti-miR-19a-3p组迁移、侵袭细胞数、N-cadherin蛋白相对表达量高于si-LINC00667+ anti-miR-NC组（P <0.05）。结论 抑制miR-19a-3p通过靶向LINC00667，抑制RA-FLS的增殖、迁移、侵袭，LINC00667或可用作诊断和治疗RA的靶点

Abstract:

Objective To explore the effects and mechanisms of long chain non-coding RNA (lncRNA) LINC00667 on the proliferation, migration and invasion of rheumatoid arthritis synovial fibroblasts (RA-FLS).Methods The model was established in SPF Wistar rats, the synovial tissue of ankle joint was isolated, and the expression of LINC00667 and miR-19a-3p in RA-FLS was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Transfect si-NC, si-LINC00667, miR-NC, miR-19a-3p simulant, si-LINC00667 and anti-miR-NC, si-LINC00667 and anti-miR-19a-3p in RA-FLS, and divide them into si-NC group, si-LINC00667 group, miR-NC group, miR-19a-3p group, si-LINC00667 + anti-miR-NC group and si-LINC00667 + anti-miR-19a-3p group. CCK-8 method was used to detect the proliferation inhibition rate of RA-FLS after different transfections, Transwell method was used to detect cell migration and invasion, and Western blotting was used to detect the expression of E-cadherin and N-cadherin. Double luciferase report verified the targeting relationship between LINC00667 and miR-19a-3p.Results The expression of LINC00667 in synovial tissue of RA model group was 270% of that of normal group, and the expression of miR-19a-3p was 36% that of normal group (P < 0.05). After successful transfection, the inhibition rate of RA-FLS and the expression of E-cadherin protein in the si LINC00667 group were higher than those in the si-NC group, and the number of migration and invasion cells and the expression of N-cadherin protein were lower than those in the si-NC group (P < 0.05). LINC00667 targeted miR-19a-3p. The inhibition rate of RA-FLS and the expression of E-cadherin protein in miR-19a-3p group were higher than those in miR-NC group, while the number of migration and invasion cells and the expression of N-cadherin protein in miR-19a-3p group were lower than those in miR-NC group (P < 0.05). The inhibition rate of RA-FLS and the expression of E-cadherin protein in si-LINC00667 + anti-miR-19a-3p group were lower than those in si-LINC00667 + anti-miR-NC group, while the number of migration and invasion cells and the expression of N-cadherin protein in si-LINC00667 + anti-miR-NC group were higher than those in si-LINC00667 + anti-miR-NC group (P < 0.05).Conclusion LINC00667 can promote the proliferation, migration and invasion of RA-FLS by targeting miR-19a-3p.