Research Paper Volume 13, Issue 7 pp 10584—10602

Suppression of choroidal neovascularization by silencing of long non-coding RNA IPW

Silencing of lncRNA-IPW reduces endothelial angiogenic function in vitro. (A) RF/6A cells were transfected with scrambled (Scr) siRNA, IPW siRNA, or left untreated (Ctrl) for 24 h. qRT-PCRs were performed to detect IPW expression (n = 4; *P B and C) RF/6A cells were transfected with Scr siRNA, IPW siRNA, or left untreated for 24 h, and then exposed with CoCl2 (200 μmol/L) to mimic hypoxic stress for 24 h. The group without CoCl2 treatment was taken as the Ctrl group. Cell viability was detected by MTT assays (B; n = 4; *P n = 4; *P #P 2 + IPW siRNA versus CoCl2 + Scr siRNA; One-way ANOVA). (D–F) RF/6A cells were transfected with Scr siRNA, IPW siRNA, or left untreated (Ctrl) for 24 h. Cell proliferation was determined by EdU incorporation assay. Blue: DAPI; red: EdU. Scale bar: 20 μm (D; n = 4; *P n = 4; *P n = 4; *P

Figure 2. Silencing of lncRNA-IPW reduces endothelial angiogenic function in vitro. (A) RF/6A cells were transfected with scrambled (Scr) siRNA, IPW siRNA, or left untreated (Ctrl) for 24 h. qRT-PCRs were performed to detect IPW expression (n = 4; *P < 0.05 versus Ctrl group; One-way ANOVA). (B and C) RF/6A cells were transfected with Scr siRNA, IPW siRNA, or left untreated for 24 h, and then exposed with CoCl2 (200 μmol/L) to mimic hypoxic stress for 24 h. The group without CoCl2 treatment was taken as the Ctrl group. Cell viability was detected by MTT assays (B; n = 4; *P < 0.05 versus Ctrl group; One-way ANOVA). Apoptotic cells were detected by PI/Calcein-AM staining. Green: live cells; red: dead or dying cells. Scale bar: 50 μm (C; n = 4; *P < 0.05 versus Ctrl group; #P < 0.05 CoCl2 + IPW siRNA versus CoCl2 + Scr siRNA; One-way ANOVA). (DF) RF/6A cells were transfected with Scr siRNA, IPW siRNA, or left untreated (Ctrl) for 24 h. Cell proliferation was determined by EdU incorporation assay. Blue: DAPI; red: EdU. Scale bar: 20 μm (D; n = 4; *P < 0.05 versus Ctrl group; One-way ANOVA). The migration of RF/6A cells was measured using Transwell assays. These cells migrated through the chamber were quantified. Scale bar: 50 μm (E, n = 4; *P < 0.05 versus Ctrl group; One-way ANOVA). The tube-like structures were observed 6 h after cell seeding on the matrix. The average length of tube formation for each field was statistically analyzed. Scale bar: 200 μm (F, n = 4; *P < 0.05 versus Ctrl group; One-way ANOVA).