大肠杆菌K5多糖对糖尿病视网膜病变小鼠视网膜新生血管形成作用的研究

目的 探究大肠杆菌K5多糖对糖尿病视网膜病变小鼠视网膜新生血管形成的影响.方法 发酵培养大肠杆菌K5菌株,对培养液进行分离纯化制备K5多糖,并分析K5多糖的纯度;雄性ICR小鼠被随机分为正常纽、模型组和干预组,其中正常组是健康小鼠,而模型组和干预组均使用链脲佐菌素建立糖尿病小鼠模型,干预组于造模后使用K5多糖干预4周,造模后8周各组取小鼠眼球组织,行HE染色及视网膜铺片ADPase染色;采用Real-time PCR和Western blotting检测各组小鼠眼球组织血管内皮生长因子(vascular endothelial gTowth factor,VEGF)及基质金属蛋白酶-2(matrix metalloproteinase,MMP-2)的表达情况.结果 HPLC结果显示制备的K5多糖纯度在95％以上:HE染色及ADPase染色结果显示正常组小鼠视网膜只见少量血管,模型组小鼠视网膜血管数目显著高于正常组,而干预组小鼠视网膜血管数目较模型组显著下降(P＜ 0.05);Real-time PCR和Western blot结果显示,模型组小鼠眼球组织VEGF和MMP-2的mRNA及蛋白表达水平显著高于正常组(均为P＜0.05),而干预组VEGF和MMP-2的mRNA及蛋白表达水平显著低于模型组(均为P＜0.05).结论 大肠杆菌K5多糖能够抑制糖尿病视网膜病变小鼠视网膜新生血管形成,其作用机制可能与抑制VEGF及MMP-2的表达有关.

Effect of E.coli K5 polysaccharide on retinal neovascularization in mice with diabetic retinopathy

Objective To explore the influence of E.coli K5 polysaccharide on retinal neovascularization in mice of diabetic retinopathy.Methods E.coli K5 strain was cultivated by fermentation and the supernatant was used to prepare K5 polysaccharide for analyze its purity by high performance liquid chromatography (HPLC).Moreover,male ICR mice were divided into normal group,model group and K5 polysaccharide treatment group (treatment group for short).The mice in the normal group were healthy and left untreated,while the mice in the model group and treatment group were administrated with streptozotocin to establish diabetic model,and the treatment group received K5 polysaccharide 4 weeks after the successful modeling.Then the eyeball tissues in the three groups were collected 8 weeks after modeling for HE staining and ADPase staining,and real-time PCR and Western blot were used to detect the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-2 (MMP-2) in eyeball tissues.Results HPLC resuits showed that the purity of K5 polysaccharide exceeded 95％.HE staining and ADPase staining suggested that a few of new vessels were observed,and the number of vessels in the model group was significantly higher than those in the normal group,while the treatment group was significantly lower than the model group (P ＜ 0.05).Real-time and Western blot showed that the expression of VEGF and MMP-2 mRNA and protein in the model group were significantly higher than those in the normal group,while the treatment group was significantly lower than the model group (P ＜ 0.05).Conclusion E.coli K5 polysaccharide can inhibit the retinal neovascularization generation in mice with diabetic retinopathy,and the mechanism of its action may be associated with the suppressed expression of VEGF and MMP-2.