Silencing of S100A4, a metastasis-associated protein, inhibits retinal neovascularization via the downregulation of BDNF in oxygen-induced ischaemic retinopathy
Silencing of S100A4, a metastasis-associated protein, inhibits retinal neovascularization via the downregulation of BDNF in oxygen-induced ischaemic retinopathy
复制标题
在氧诱导的缺血性视网膜病中，转移相关蛋白 S100A4 的沉默通过下调 BDNF 抑制视网膜新生血管形成
作者：
Cheng, G.;He, T.;Xing, Y.
目的探讨S100 A4基因在氧诱导视网膜病变（OIR）小鼠模型视网膜新生血管（RNV）形成中的作用及其机制。方法在出生后12天（P12）玻璃体内注射腺病毒S100 A4-RNAi或腺病毒绿色荧光蛋白（GFP）。在P17，使用荧光显微镜和蛋白质印迹分析评估腺病毒基因转移的功效。RNV通过小鼠视网膜的整体包埋免疫荧光染色和通过计数视网膜前新生血管细胞的数量来评估。采用Western blot和real-time PCR检测视网膜S100A4、脑源性生长因子（BDNF）和血管内皮生长因子（VEGF）的蛋白和mRNA表达水平。在OIR-S100A4-RNAi组中，与OIR组相比，视网膜中S100A4的蛋白和mRNA表达水平在P17时均显著降低。在Ad-S100A4-RNAi转移后5天，在视网膜的许多层中通过GFP荧光清楚地证明了Ad-S100A4-RNAi转移。视网膜的整体免疫荧光染色和视网膜前新生血管细胞的定量表明RNV被显著抑制。OIR-S100A4-RNAi组BDNF、VEGF和HIF-1 α的转录和翻译水平均明显降低。相关机制可能涉及沉默S100A4以减少BDNF的激活，BDNF通过HIF-1 α下调VEGF表达。这一发现为眼部新生血管疾病的治疗提供了新的靶点。
Background To investigate the underlying mechanism of S100A4 function and whether it has a role in retinal neovascularization (RNV) in a mouse model of oxygen-induced retinopathy (OIR).Methods Retinas from a mouse model of OIR were treated with and without an intravitreous injection of adenoviral-S100A4-RNAi or adenoviral green fluorescence protein (GFP) at postnatal day 12 (P12). At P17, the efficacy of adenoviral gene transfer was assessed using fluorescence microscopy and western blot analysis. RNV was evaluated by whole-mount immunofluorescence staining of the mouse retina and by counting the number of pre-retinal neovascular cells. Protein and mRNA expression levels of S100A4, brain-derived growth factor (BDNF), and vascular endothelial growth factor (VEGF) were measured using western blot analysis and real-time PCR.Results Retinal S100A4 levels were positively correlated with the progression of RNV. In the OIR-S100A4-RNAi group, both protein and mRNA expression levels of S100A4 in the retina significantly decreased at P17 compared with those in the OIR group. Ad-S100A4-RNAi transfer was clearly demonstrated by GFP fluorescence in many layers of the retina 5 days after the Ad-S100A4-RNAi transfer. Whole-mount immunofluorescence staining of the retina and quantification of the pre-retinal neovascular cells demonstrated that RNV was significantly inhibited. Meanwhile, the levels of the transcription and translation of BDNF, VEGF, and hypoxia-inducible factor-1 alpha (HIF-1 alpha) significantly decreased in the OIR-S100A4-RNAi group.Conclusions Ad-S100A4-RNAi transfer ameliorates RNV. The related mechanism may involve silencing S100A4 to decrease the activation of BDNF, which downregulates VEGF expression via HIF-1 alpha. This finding could provide a new therapeutic target for the treatment of ocular neovascularization diseases.