Department of Ophthalmology, People's Hospital of Zhaoyuan City, Zhaoyuan, Shandong, China.
Department of Ophthalmology, Shanghai Changzheng Hospital, China Naval Medical University, Shanghai, China.
Braz J Med Biol Res. 2020;53(5):e9608. doi: 10.1590/1414-431x20209608. Epub 2020 Apr 27.
Cataract, an eye disease that threatens the health of millions of people, brings about severe economic burden for patients and society. MicroRNA (miR)-378a-5p and miR-630 were recognized as essential regulators in multiple cancers. However, the exact functions of miR-378a-5p and miR-630 in cataract are still unclear. The expression of miR-378a-5p, miR-630, and E2F transcription factor 3 (E2F3) in tissues and cells was measured by quantitative real-time polymerase chain reaction. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay was used to evaluate cell viability. Flow cytometry was conducted to analyze cell apoptosis. The interaction between E2F3 and miR-378a-5p or miR-630 was confirmed by dual-luciferase reporter assay. The expression of proteins E2F3, B cell lymphoma (Bcl-2), Bcl-2 associated X (Bax), and cleaved caspase 3 was detected by western blot assay. The expression of miR-378a-5p and miR-630 was up-regulated whereas E2F3 was down-regulated in human cataract lens tissues compared with normal lens tissues. Depletion of miR-378a-5p or miR-630 enhanced proliferation and reduced apoptosis of human lens epithelial cells. Interestingly, up-regulation of E2F3 exhibited the same trend. Next, dual-luciferase reporter assay validated the interaction between E2F3 and miR-378a-5p or miR-630. The rescue experiments further revealed that E2F3 knockdown could recover miR-378a-5p, and miR-630 inhibitor induced promotion of cell proliferation and inhibition of apoptosis in cataract. miR-378a-5p and miR-630 repressed proliferation and induced apoptosis of lens epithelial cells by targeting E2F3 in cataract, representing a prospective alternative therapy for cataract.
白内障是一种威胁数百万人健康的眼部疾病,给患者和社会带来了沉重的经济负担。microRNA (miR)-378a-5p 和 miR-630 被认为是多种癌症中的重要调节因子。然而,miR-378a-5p 和 miR-630 在白内障中的具体功能尚不清楚。通过实时定量聚合酶链反应测量组织和细胞中 miR-378a-5p、miR-630 和 E2F 转录因子 3 (E2F3) 的表达。3-(4,5-二甲基-2-噻唑基)-2,5-二苯基-2-H-四唑溴盐法用于评估细胞活力。流式细胞术用于分析细胞凋亡。双荧光素酶报告基因检测证实了 E2F3 与 miR-378a-5p 或 miR-630 之间的相互作用。Western blot 检测 E2F3、B 细胞淋巴瘤 (Bcl-2)、Bcl-2 相关 X (Bax) 和 cleaved caspase 3 蛋白的表达。与正常晶状体组织相比,人白内障晶状体组织中 miR-378a-5p 和 miR-630 的表达上调,而 E2F3 的表达下调。miR-378a-5p 或 miR-630 的耗竭增强了人晶状体上皮细胞的增殖并减少了细胞凋亡。有趣的是,E2F3 的上调也呈现出相同的趋势。接下来,双荧光素酶报告基因检测验证了 E2F3 与 miR-378a-5p 或 miR-630 之间的相互作用。挽救实验进一步表明,E2F3 敲低可以恢复 miR-378a-5p 和 miR-630 抑制剂诱导的白内障中细胞增殖的促进和凋亡的抑制。miR-378a-5p 和 miR-630 通过靶向 E2F3 抑制白内障中晶状体上皮细胞的增殖并诱导其凋亡,为白内障提供了一种有前景的替代治疗方法。