OTUD6B-AS1通过miR-21-5p对高糖诱导的ARPE-19细胞增殖、迁移的影响

目的　探讨高糖诱导的人视网膜色素上皮细胞（ARPE-19)中含有卵巢肿瘤结构域的6B反义RNA 1(OTUD6B-AS1)的功能及其作用机制。方法　取ARPE-19细胞，加入含30 mmol·L^-1葡萄糖的培养基培养，设为高糖组，另取ARPE-19细胞不做任何处理设为对照组。用Lipofectamine^TM 2000转染试剂分别将过表达空质粒（pcDNA-NC组）、OTUD6B-AS1过表达质粒（pcDNA-OTUD6B-AS1组）和OTUD6B-AS1过表达质粒＋miR-21-5p 模拟物（pcDNA-OTUD6B-AS1+miR-21-5p mimics组）转染进高糖诱导的ARPE-19细胞。实时荧光定量PCR（qRT-PCR）检测细胞中OTUD6B-AS1和miR-21-5p的表达；用CCK-8法检测各组细胞增殖能力；流式细胞术检测细胞凋亡情况；细胞划痕实验检测各组细胞迁移能力；荧光素酶报告基因实验验证OTUD6B-AS1与miR-21-5p的靶向关系。结果　与对照组比较，高糖组ARPE-19细胞中OTUD6B-AS1表达量显著降低、miR-21-5p表达量显著升高，差异均有统计学意义（均为P<0.05）。与对照组比较，高糖组细胞凋亡率显著升高，细胞增殖率和迁移率均显著降低，差异均有统计学意义(均为P<0.05）。与pcDNA-NC组比较，pcDNA-OTUD6B-AS1组中细胞增殖率和迁移率显著升高，细胞凋亡率显著降低，差异均有统计学意义(均为P<0.05）。双荧光素酶报告基因实验显示，OTUD6B-AS1可靶向miR-21-5p。与pcDNA-OTUD6B-AS1组比较，pcDNA-OTUD6B-AS1＋miR-21-5p mimics组细胞凋亡率显著升高，细胞增殖率和迁移率显著降低，差异均有统计学意义(均为P<0.05）。结论　OTUD6B-AS1可抑制高糖诱导的ARPE-19细胞凋亡，并促进细胞增殖和迁移，其作用机制与miR-21-5p有关。

Effect of ovarian tumor domain containing 6B antisense RNA1 on the proliferation and migration of high glucose-induced retinal pigment epithelial cells via miR-21-5p

Objective　To investigate the function and mechanism of ovarian tumor domain containing 6B antisense RNA1 (OTUD6B-AS1) in retinal pigment epithelial (RPE) cells induced by high glucose (HG). Methods　ARPE-19 cells cultured in medium containing 30 mmol·L^-1 glucose were set as the HG group, and ARPE-19 cells without any treatment were set as the control group. Overexpressed empty plasmids (pcDNA-NC group), OTUD6B-AS1 overexpressed plasmids (pcDNA-OTUD6B-AS1 group), and OTUD6B-AS1 overexpressed plasmids + miR-21-5p mimics (pcDNA-OTUD6B-AS1+miR-21-5p group) were transfected into HG-induced ARPE-19 cells using Lipofectamine^TM 2000 transfection reagent, respectively. The expression of OTUD6B-AS1 and miR-21-5p was detected by quantitative real-time polymerase chain reaction, the cell proliferation was detected by CCK-8 assay, the cell apoptosis was detected by flow cytometry, the cell migration was detected by wound-healing assay, and the targeting relationship between OTUD6B-AS1 and miR-21-5p was verified by luciferase reporter gene assay. Results　Compared with the control group, OTUD6B-AS1 expression was significantly decreased while miR-21-5p expression was significantly increased in the HG group (both P<0.05). Compared with the control group, the cell apoptosis was significantly increased while the cell proliferation and migration were significantly reduced in the HG group (all P<0.05). Compared with the pcDNA-NC group, the cell proliferation and migration in the pcDNA-OTUD6B-AS1 group were significantly increased while the cell apoptosis was significantly reduced (all P<0.05). Dual-luciferase reporter gene assay showed that OTUD6B-AS1 could target miR-21-5p. Compared with the pcDNA-OTUD6B-AS1 group, the cell apoptosis in the pcDNA-OTUD6B-AS1+miR-21-5p group was significantly increased while the cell proliferation and migration were significantly decreased (all P<0.05). Conclusion　OTUD6B-AS1 can inhibit the apoptosis of HG-induced ARPE-19 cells and promote their proliferation and migration, which is related to miR-21-5p.