Department of Ophthalmology, The Affiliated Wuxi People's Hospital of Nanjing Medical University, Wuxi People's Hospital, Wuxi Medical Center, Nanjing Medical University, Wuxi, 214023, Jiangsu, China.
Department of Ophthalmology, The Affiliated Wuxi People's Hospital of Nanjing Medical University, Wuxi People's Hospital, Wuxi Medical Center, Nanjing Medical University, Wuxi, 214023, Jiangsu, China.
Mol Cell Endocrinol. 2023 Oct 1;576:112042. doi: 10.1016/j.mce.2023.112042. Epub 2023 Aug 10.
The aim of this study was to investigate the mechanism underlying the role of a recently identified hsa_circ_0004805/hsa_miR-149-5p/transforming growth factor beta 2 (TGFB2) axis in the progression of diabetic retinopathy (DR). Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis suggested that hsa_circ_0004805 was highly expressed in aqueous humor samples of patients with DR, whereas hsa_miR-149-5p showed the opposite trend. Meanwhile, the results of a dual-luciferase reporter assay indicated that hsa_miR-149-5p directly interacted with both hsa_circ_0004805 and TGFB2. Using a variety of assays (Cell Counting Kit-8, EdU-labeling, Transwell, flow cytometric, wound healing, tube formation assays), we found that the overexpression of hsa_circ_0004805 significantly downregulated the level of hsa_miR-149-5p and promoted DNA synthesis, proliferation, migration, and tube formation in human retinal microvascular epithelial cells (hRECs) cultivated in a high-glucose environment. In contrast, hsa_miR-149-5p mimics inhibited DNA synthesis, proliferation, migration, and tube formation in hRECs by reducing the expression of its downstream target TGFB2 as well as the levels of phosphorylated SMAD2; however, these effects were reversed by the overexpression of hsa_circ_0004805. In a streptozotocin-induced Sprague-Dawley rat model of DR, retinal vascular leakage, capillary decellularization, loss of pericytes, fibrosis, and gliosis were evident, which could be reversed by vitreous microinjection of rat miR-149-5p mimics (rno-miR-149-5p agomir). Combined, our findings indicated that, under hyperglycemia, the hsa_circ_0004805/hsa_miR-149-5p/TGFB2 axis plays a critical role in the retinal pathophysiology associated with the development of DR, and has potential as a therapeutic target in the treatment of this condition.
本研究旨在探讨最近发现的 hsa_circ_0004805/hsa_miR-149-5p/转化生长因子β 2（TGFB2）轴在糖尿病视网膜病变（DR）进展中的作用机制。定量逆转录聚合酶链反应（qRT-PCR）分析表明，hsa_circ_0004805在 DR 患者的房水中高度表达，而 hsa_miR-149-5p 则呈现相反的趋势。同时，双荧光素酶报告基因实验的结果表明，hsa_miR-149-5p 可以直接与 hsa_circ_0004805 和 TGFB2 相互作用。通过一系列实验（细胞计数试剂盒-8、EdU 标记、Transwell、流式细胞术、划痕愈合、管形成实验），我们发现 hsa_circ_0004805 的过表达显著下调了 hsa_miR-149-5p 的水平，并促进了高糖环境中培养的人视网膜微血管上皮细胞（hRECs）的 DNA 合成、增殖、迁移和管形成。相反，hsa_miR-149-5p 模拟物通过降低其下游靶基因 TGFB2 以及磷酸化 SMAD2 的水平，抑制了 hRECs 的 DNA 合成、增殖、迁移和管形成，而 hsa_circ_0004805 的过表达则逆转了这些效应。在链脲佐菌素诱导的 DR 斯普拉格-道利大鼠模型中，视网膜血管渗漏、毛细血管去细胞化、周细胞丢失、纤维化和神经胶质增生明显，玻璃体注射大鼠 miR-149-5p 模拟物（rno-miR-149-5p agomir）可逆转这些变化。综上所述，在高血糖条件下，hsa_circ_0004805/hsa_miR-149-5p/TGFB2 轴在与 DR 发展相关的视网膜病理生理学中发挥关键作用，并且作为治疗该疾病的潜在治疗靶点具有潜力。
