H-W Liu1, Y Meng, Y-B Ren, P Sun. 1. Department of Ophthalmology, First Affiliated Hospital of Jiamusi University, Jiamusi, China. 15694548795@163.com.
Abstract
OBJECTIVE: To explore the role of microRNA-15b in diabetic retinopathy (DR) and its underlying mechanism. MATERIALS AND METHODS: Diabetic retinopathy rat model was first constructed. Retinal endothelial cells (EC) and retinal pericytes (RP) in DR rats were extracted. The mRNA expression of microRNA-15b in EC and RP cells was detected by qRT-PCR (quantitative Real Time-Polymerase Chain Reaction). Protein expression of insulin receptor substrate 1 (IRS-1) in EC and RP cells was detected by Western blot. After altering microRNA-15b expression by plasmid transfection, cell viability was detected by CCK-8 (cell counting kit-8) assay. Furthermore, the target gene of microRNA-15b was predicted by TargetScan analysis and the binding condition was verified by luciferase reporter gene assay. Finally, rescue experiments were carried out to explore the regulatory effect of microRNA-15b on IRS-1. RESULTS: MicroRNA-15b was lowly expressed, whereas IRS-1 was highly expressed in EC and RP cells. After overexpression of microRNA-15b, viabilities of EC and RP cells were decreased and β-catenin expression was inhibited. TargetScan predicted that IRS-1 was the downstream gene of microRNA-15b, which was further verified by luciferase reporter gene assay. Rescue experiments indicated that microRNA-15b was capable of regulating IRS-1 via Wnt/β-catenin signaling pathway. CONCLUSIONS: MicroRNA-15b participates in the development of diabetic retinopathy by targeting IRS-1 via Wnt/β-catenin signaling pathway.
OBJECTIVE: To explore the role of microRNA-15b in diabetic retinopathy (DR) and its underlying mechanism. MATERIALS AND METHODS:Diabetic retinopathyrat model was first constructed. Retinal endothelial cells (EC) and retinal pericytes (RP) in DR rats were extracted. The mRNA expression of microRNA-15b in EC and RP cells was detected by qRT-PCR (quantitative Real Time-Polymerase Chain Reaction). Protein expression of insulin receptor substrate 1 (IRS-1) in EC and RP cells was detected by Western blot. After altering microRNA-15b expression by plasmid transfection, cell viability was detected by CCK-8 (cell counting kit-8) assay. Furthermore, the target gene of microRNA-15b was predicted by TargetScan analysis and the binding condition was verified by luciferase reporter gene assay. Finally, rescue experiments were carried out to explore the regulatory effect of microRNA-15b on IRS-1. RESULTS:MicroRNA-15b was lowly expressed, whereas IRS-1 was highly expressed in EC and RP cells. After overexpression of microRNA-15b, viabilities of EC and RP cells were decreased and β-catenin expression was inhibited. TargetScan predicted that IRS-1 was the downstream gene of microRNA-15b, which was further verified by luciferase reporter gene assay. Rescue experiments indicated that microRNA-15b was capable of regulating IRS-1 via Wnt/β-catenin signaling pathway. CONCLUSIONS:MicroRNA-15b participates in the development of diabetic retinopathy by targeting IRS-1 via Wnt/β-catenin signaling pathway.
Authors: María Constanza Potilinski; Gustavo A Ortíz; Juan P Salica; Emiliano S López; Mariano Fernández Acquier; Eduardo Chuluyan; Juan E Gallo Journal: PLoS One Date: 2020-02-07 Impact factor: 3.240