精氨酸酶II对糖尿病视网膜小胶质细胞活化的调控机制研究

The role of Arginase II on the activation of retina microglia and the pathogenesis of diabetic retinopathy (DR) is unknown. Our previous study has demonstrated that arginase II expression level is enhanced in mice tissue macrophage upon type II diabetes and arginase II promotes M1 macrophage inflammatory responses. Meanwhile, we also found that overexpression arginase II in mouse BV-2 microglia cells enhanced expression of IL-6 mRNA level, which also increased the activation of p38 MAPK. Based on this, we hypothesize that arginase II would affect retina microglia activation through p38 MAPK signaling pathway, which further could regulate the pathogenesis of DR. So far there is no international report about this hypothesis. We would like to apply arginase II deficient mice model and cellular model to study next three aspects: 1) the connection between arginase II and retina microglia activation; 2) the relationship between arginase II and expression of inflammatory cytokines in retina microglia; 3) whether arginase II could active retina microglia through p38 MAPK signaling pathway in further to regulate the pathogenesis of DR. Our project would reveal the role of arginase II in the pathogenesis of DR, which is beneficial for providing a new target for the prevention and treatment of clinical diabetic retinopathy.

精氨酸酶II在视网膜小胶质细胞活化与糖尿病视网膜病变发生中的作用未知。申请人前期研究发现，糖尿病小鼠模型体内组织巨噬细胞精氨酸酶II表达水平呈现升高并且精氨酸酶II可促进M1型巨噬细胞炎症反应，在小鼠小胶质细胞系（BV-2细胞）中过表达精氨酸酶II后引起促炎因子白介素-6表达增加，p38MAPK活性增强。申请人据此假设：精氨酸酶II通过p38MAPK信号通路影响视网膜小胶质细胞的活化而调控糖尿病视网膜病变的发生。目前国内外尚无有关该假设的研究报道。本项目拟利用精氨酸酶II小鼠敲除模型结合细胞模型探讨，1）精氨酸酶II与视网膜小胶质细胞活化的关系；2）精氨酸酶II与小胶质细胞中炎性细胞因子表达的关系；3）精氨酸酶II是否通过p38MAPK信号活化视网膜小胶质细胞从而调控糖尿病视网膜炎性病变的发生。本项目拟揭示精氨酸酶II在糖尿病视网膜病变的作用，为临床糖尿病视网膜病变的防治提供新的靶点。

精氨酸酶II是一种可以将精氨酸转化成鸟氨酸和尿素的锰金属酶。近年来众多研究证明精氨酸酶II与多种疾病的发生发展有关，但是精氨酸酶II在视网膜小胶质细胞活化与糖尿病视网膜病变发生中的作用未知。因此，本研究通过构建精氨酸酶II调控小胶质细胞活化模型，我们发现在细胞水平试验上：①小胶质细胞中（BV-2）过表达精氨酸酶II后细胞形态由分支状变成阿米巴原虫状；WB实验检测到p-p38 mapk表达明显增强；RT-PCR实验和ELISA检测到促炎因子IL-6、MCP-1表达增强；超氧化物阴离子荧光探针法检测到小胶质细胞内线粒体活性氧自由基（Mitosox）产生明显增多，但胞质内活性氧自由基（DHE）产生无明显变化。②而在小胶质细胞中沉默精氨酸酶II后细胞形态由阿米巴原虫状变成分支状；WB实验检测到p-p38 mapk表达明显降低；RT-PCR实验和ELISA检测到促炎因子IL-6、MCP-1表达明显降低；超氧化物阴离子荧光探针法检测到小胶质细胞内线粒体活性氧自由基（Mitosox）产生被抑制，但胞质内活性氧自由基（DHE）产生无变化。这些结果表明在细胞水平方面可以验证精氨酸酶II通过p38 MAPK信号通路而引起视网膜小胶质细胞的活化，从而为深入研究精氨酸酶II在糖尿病视网膜病变的作用奠定工作基础，为临床糖尿病视网膜病变的防治提供新的靶点。