环状 RNA 分子 MAP3K4 通过靶向 miR-193a-3p/PLCδ3 轴保护人晶状体上皮细胞免受 HO 诱导的年龄相关性白内障功能障碍。
CircMAP3K4 protects human lens epithelial cells from HO-induced dysfunction by targeting miR-193a-3p/PLCD3 axis in age-related cataract.
机构信息
Department of Ophthalmology, Zhengzhou University, Affiliated Hospital 5, Zhengzhou, Henan, China.
College of Medical Technology and Engineering, Zhengzhou Railway Vocational Technology College, Zhengzhou, China.
出版信息
Cell Cycle. 2023 Feb;22(3):303-315. doi: 10.1080/15384101.2022.2114587. Epub 2022 Sep 7.
Circular RNAs (circRNAs) have shown pivotal regulatory roles in multiple human ocular diseases, including age-related cataract (ARC). Here, we explored the role of circRNA mitogen-activated protein kinase kinase kinase 4 (circMAP3K4, hsa_circ_0078619) in ARC pathology and its associated mechanism. The expression of RNAs and proteins was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot assay. Cell viability, senescence, proliferation, and apoptosis were analyzed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, senescence-associated-β-galactosidase (SA-β-Gal) staining, 5-ethynyl-20-deoxyuridine (EdU) assay, and flow cytometry. The oxidative stress status of SRA01/04 cells was analyzed using the commercial kits. The interaction between microRNA-193a-3p (miR-193a-3p) and circMAP3K4 or phospholipase C delta 3 (PLCD3) was verified by dual-luciferase reporter assay, RNA immunoprecipitation (RIP) assay, and RNA-pull down assay. CircMAP3K4 was significantly down-regulated in ARC patients and HO-induced SRA01/04 cells. HO treatment restrained the viability and proliferation and promoted the senescence, apoptosis, and oxidative stress of SRA01/04 cells, and circMAP3K4 overexpression protected SRA01/04 cells from HO-induced dysfunction. MiR-193a-3p was a direct target of circMAP3K4, and circMAP3K4 overexpression-mediated protective effects in HO-induced SRA01/04 cells were largely reversed by the accumulation of miR-193a-3p. MiR-193a-3p interacted with the 3' untranslated region (3'UTR) of PLCD3, and PLCD3 knockdown largely overturned miR-193a-3p silencing-induced protective effects in HO-induced SRA01/04 cells. CircMAP3K4 up-regulated the expression of PLCD3 via sponging miR-193a-3p in SRA01/04 cells. In conclusion, circMAP3K4 protected SRA01/04 cells from HO-induced dysfunction in ARC through mediating miR-193a-3p/PLCD3 axis.
环状 RNA（circRNA）在多种人类眼部疾病中表现出关键的调节作用，包括年龄相关性白内障（ARC）。在这里，我们探讨了环状 RNA 丝裂原活化蛋白激酶激酶激酶 4（circMAP3K4，hsa_circ_0078619）在 ARC 病理中的作用及其相关机制。通过逆转录定量聚合酶链反应（RT-qPCR）和 Western blot 分析检测 RNA 和蛋白质的表达。通过 3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四唑溴盐（MTT）测定、衰老相关-β-半乳糖苷酶（SA-β-Gal）染色、5-乙炔基-20-脱氧尿苷（EdU）测定和流式细胞术分析细胞活力、衰老、增殖和凋亡。使用商业试剂盒分析 SRA01/04 细胞的氧化应激状态。通过双荧光素酶报告基因检测、RNA 免疫沉淀（RIP）检测和 RNA 下拉检测验证 microRNA-193a-3p（miR-193a-3p）与 circMAP3K4 或磷脂酶 C 三角洲 3（PLCD3）之间的相互作用。在 ARC 患者和 HO 诱导的 SRA01/04 细胞中，circMAP3K4 显著下调。HO 处理抑制 SRA01/04 细胞的活力和增殖，促进衰老、凋亡和氧化应激，而过表达 circMAP3K4 可保护 SRA01/04 细胞免受 HO 诱导的功能障碍。miR-193a-3p 是 circMAP3K4 的直接靶标，而过表达 circMAP3K4 介导的 HO 诱导的 SRA01/04 细胞中的保护作用在 miR-193a-3p 积累时被大大逆转。miR-193a-3p 与 PLCδ3 的 3'非翻译区（3'UTR）相互作用，而 PLCδ3 的敲低在 HO 诱导的 SRA01/04 细胞中大大逆转了 miR-193a-3p 沉默诱导的保护作用。circMAP3K4 通过海绵吸附 miR-193a-3p 在 SRA01/04 细胞中上调 PLCD3 的表达。总之，circMAP3K4 通过介导 miR-193a-3p/PLCD3 轴保护 SRA01/04 细胞免受 HO 诱导的 ARC 功能障碍。
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