Wen-Zhi Yang1, Jin Yang2, Li-Ping Xue1, Li-Bo Xiao1, Yan Li3. 1. Department of Ophthalmology, Fourth Affiliated Hospital of Kunming Medical University, Kunming, Yunnan 650021, China. 2. Department of Osteology, First Affiliated Hospital of Kunming Medical University, Kunming, Yunnan 650032, China. 3. Department of Ophthalmology, First Affiliated Hospital of Kunming Medical University, Kunming, Yunnan 650032, China. Electronic address: liyanr@hotmail.com.
Abstract
AIM: The aims of this study are to investigate the relative regulation between miR-126 and VEGF/PI3K/AKT signaling pathway in retinal vascular endothelial cells. METHODS: Rhesus macaque choroid-retinal endothelial cell line (RF/6A) cells were cultured in high glucose to imitate the conditions occurring in DR. First, we detected the expression of miR-126, VEGFA and PIK3R2 in RF/6A cells on the condition of high glucose by q-PCR and western blot. Then, after addition of miR-126 mimics and miR-126 inhibitor, we investigated the function of miR-126 in RF/6A cells by scratch wound, Transwell migration and tube formation assays, and the effect of miR-126 on the expression of VEGFA, PIK3R2 and AKT. Moreover, bioinformatics analysis and luciferase array were used to confirm the direct or specific regulation of miR-126 to VEGFA or PIK3R2. RESULTS: Here, first, we found that high glucose could induce the decrease of miR-126 and the increase of VEGFA and PIK3R2 in RF/6A. Then, by scratch wound, Transwell migration and tube formation assays, we found that miR-126 overexpression could inhibit the migration and sprouting of RF/6A cells induced by high glucose, while knockdown of miR-126 led to the opposite results. Moreover, overexpression of miR-126 inhibited the increased expression of VEGFA, PIK3R2, SDF-1α, VCAM-1, and SPRED1, and the activation of AKT1 induced by high glucose and miR-126 inhibitor caused the opposite results which were determined by q-PCR and western blot. In addition, by luciferase assay, we found that miR-126 could directly negatively regulate VEGFA and PIK3R2. CONCLUSION: Our results suggest that miR-126 overexpression inhibits the migration and sprouting of RF/6A cells induced by high glucose which might possibly be by blocking VEGFA and PIK3R2 in the VEGF/PI3K/AKT signaling pathway.
AIM: The aims of this study are to investigate the relative regulation between miR-126 and VEGF/PI3K/AKT signaling pathway in retinal vascular endothelial cells. METHODS:Rhesus macaque choroid-retinal endothelial cell line (RF/6A) cells were cultured in high glucose to imitate the conditions occurring in DR. First, we detected the expression of miR-126, VEGFA and PIK3R2 in RF/6A cells on the condition of high glucose by q-PCR and western blot. Then, after addition of miR-126 mimics and miR-126 inhibitor, we investigated the function of miR-126 in RF/6A cells by scratch wound, Transwell migration and tube formation assays, and the effect of miR-126 on the expression of VEGFA, PIK3R2 and AKT. Moreover, bioinformatics analysis and luciferase array were used to confirm the direct or specific regulation of miR-126 to VEGFA or PIK3R2. RESULTS: Here, first, we found that high glucose could induce the decrease of miR-126 and the increase of VEGFA and PIK3R2 in RF/6A. Then, by scratch wound, Transwell migration and tube formation assays, we found that miR-126 overexpression could inhibit the migration and sprouting of RF/6A cells induced by high glucose, while knockdown of miR-126 led to the opposite results. Moreover, overexpression of miR-126 inhibited the increased expression of VEGFA, PIK3R2, SDF-1α, VCAM-1, and SPRED1, and the activation of AKT1 induced by high glucose and miR-126 inhibitor caused the opposite results which were determined by q-PCR and western blot. In addition, by luciferase assay, we found that miR-126 could directly negatively regulate VEGFA and PIK3R2. CONCLUSION: Our results suggest that miR-126 overexpression inhibits the migration and sprouting of RF/6A cells induced by high glucose which might possibly be by blocking VEGFA and PIK3R2 in the VEGF/PI3K/AKT signaling pathway.