Silencing of S100A4 gene inhibits oxygen-induced retinal neovascularization in mice
10.3760/cma.j.issn.1005-1015.2016.01.013
- VernacularTitle:钙结合蛋白S100A4基因静默对氧诱导视网膜新生血管的抑制作用及其机制
- Author:
Gumeng CHENG
;
Tao HE
;
Yiqiao XING
- Publication Type:Journal Article
- Keywords:
Retinal neovascularization/prevention & control;
Calgranulin A/antagonists & inhibitors;
Gene silencing;
Animal experimentation
- From:
Chinese Journal of Ocular Fundus Diseases
2016;32(1):52-57
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the inhibitory effects and related mechanism of S100A4 gene silencing on oxygen-induced retinal neovascularization.Methods 7-day-old C57BL/6J mice were randomly divided into 5 groups including normal group,normal-S100A4 group,oxygen induced retinopathy (OIR) group,OIR-S100A4 group,OIR-green fluorescent protein (GFP) group.To establish the OIR model,mice from all groups except normal one were exposed to (75±2) ％ oxygen for 5 days and then to room air.In the OIR-S100A4 group and OIR-GFP group,the OIR mice were given an intravitreal injection of 1 μl of 1.0 × 109 PFU/ml adenovirus of Ad-S100A4-RNAi or Ad-GFP at P12,and then returned to normoxia for the next 5 days.In the OIR group,OIR was induced in C57bl/6J mice from P7 to P17.In the normal-S100A4 group,the normal P12 mice were give an intravitreal injection of 1 μl of Ad-S100A4-RNAi adenovirus,and maintained in room air from P12 to P17.In normal group,newborn mouse litters were maintained in room air from P0 to P17 without any treatment.Mice in all five groups were euthanized at P17,and retinas were collected for biochemical assays and morphological study.Retinal neovascularization (RNV) was evaluated by counting the number of pre-retinal neovascular cells and the whole mount immunofluorescent staining of the mouse retina.Protein and mRNA expression levels of S100A4,cAMP responsive element binding protein (CREB),B cell lymphoma-2 (bcl-2),Caspase-3 were determined with western blot and real-time PCR.Results The number of pre-retinal neovascular cell nuclei in retinas from OIR-S100A4 group were obviously lower than those in the retinas from OIR group and OIR-GFP group (t=13.61,14.64;P＜0.05).In OIR-S100A4 group,the retinal neovascular tufts area and the vaso-oblitertion area were both significantly smaller than those in OIR group and OIR-GFP group (P＜0.05).Protein level of CREB and bcl-2 were significantly down-regulated in OIR-S100A4 group than those in OIR and OIR-GFP group (P＜0.05).On the contrary,protein levels of Caspase-3 were up-regulated in OIR-S100A4 group than those in OIR and OIR-GFP group (P＜0.05).Conclusion Ad-S100A4-RNAi transfer ameliorates RNV in mouse model of OIR maybe through down-regulating the expression of bcl-2 and CREB,and up-regulating the Caspase-3.
