Targeting Müller cells for therapy of diabetic retinopathy

靶向穆勒细胞治疗糖尿病视网膜病变

• 批准号: 289242253
• 负责人: Professorin Dr. Antje Grosche
• 金额: --
• 依托单位: Institut für Physiologische Genomik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2016
• 资助国家: 德国
• 起止时间: 2015-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/289242253?language=en
• 关键词: Targeting; ller; cells; therapy; diabetic

Diabetic retinopathy (DR) is a retinal disease of highest relevance in societies of developed countries being the most prevalent reason of vision loss in the working population. Müller cells, the principal macroglia of the retina, undergo major changes in gene expression associated with partial loss of their cell functions in DR. Strikingly little is known about the modulation and mechanistic relevance of this Müller cell activation. The goal of our study is to significantly improve our understanding of the role of this glial reaction and to promote neuronal survival in DR by manipulating Müller cell gene expression. Mice carrying a mutant leptin receptor (db/db mice) will be used to model type II diabetes and associated retinal changes. To generate a constantly high neuronprotective microenvironment in the diabetic retina, we aim to overexpress the neuroprotective factor Norrin in Müller cells of db/db mice by specifically transducing them with ShH10, an adeno-associated virus serotype with Müller cell tropism. Using tools for expression analysis (qRT-PCR, Western blot) and to test for Müller cell and retinal functions (patch-clamp, electroretinogram, fluorescence angiography), we will validate the therapeutic potential of this approach. Next, we seek to overexpress the potassium channel Kir4.1 in Müller cells as it is the essential potassium channel for Müller cell homeostatic functions that gets lost in the diabetic retina. By Kir4.1 re-expression, we will test the hypothesis that a stabilized neuron-supportive Müller cell phenotype in the diabetic retina has the potential to slow down or even halt disease progression. In parallel to these proof of principle experiments, comprehensive data on the gene expression (RNA and protein) from Müller glia isolated from healthy and diabetic retinae in comparison with expression data collected from microglia, endothelial cells and retinal neurons will be generated to identify novel candidate genes specifically involved in Müller cell activation. This costly and laborious approach will give intriguing insights into the interplay of these respective cell types in the context of DR. Integration of these data with data from genome-wide association studies for DR may identify Müller cell genes potentially serving as targets for novel DR therapies in humans. After functional characterization of most promising candidate genes, we aim at implementing ShH10-driven modulation of one selected candidate to preserve key Müller cell functions fostering neuronal survival in the diabetic retina. In sum, we are confident that data collected in the present project will considerably contribute to the development of novel long-term efficient treatment options for DR that to date cannot be satisfactorily addressed by conventional therapeutic approaches owing to its complex disease etiology.

糖尿病视网膜病变（DR）是在发达国家的社会中最相关的视网膜疾病，是工作人群中视力丧失的最普遍原因。米勒细胞，视网膜的主要大胶质细胞，经历重大的基因表达变化与部分丧失其细胞功能的DR。引人注目的是，很少有人知道这个米勒细胞激活的调制和机械相关性。我们研究的目的是通过操纵Müller细胞基因表达来显著提高我们对这种胶质反应的作用的理解，并促进DR中神经元的存活。携带突变体瘦素受体的小鼠（db/db小鼠）将用于模拟II型糖尿病和相关的视网膜变化。为了在糖尿病视网膜中产生恒定的高神经元保护微环境，我们的目标是通过用ShH 10特异性转导db/db小鼠的Müller细胞中的神经保护因子Norrin，ShH 10是一种具有Müller细胞嗜性的腺相关病毒血清型。使用表达分析工具（qRT-PCR，Western blot）和测试Müller细胞和视网膜功能（膜片钳，视网膜电图，荧光血管造影），我们将验证这种方法的治疗潜力。接下来，我们试图在Müller细胞中过表达钾通道Kir4.1，因为它是Müller细胞稳态功能的必需钾通道，在糖尿病视网膜中丢失。通过Kir4.1的重新表达，我们将检验糖尿病视网膜中稳定的神经元支持Müller细胞表型有可能减缓甚至停止疾病进展的假设。与这些原理性实验的证明平行，将生成关于从健康和糖尿病视网膜分离的Müller胶质细胞的基因表达（RNA和蛋白质）的综合数据，与从小胶质细胞、内皮细胞和视网膜神经元收集的表达数据进行比较，以鉴定特异性参与Müller细胞活化的新候选基因。这种昂贵而费力的方法将为DR背景下这些相应细胞类型的相互作用提供有趣的见解。将这些数据与DR全基因组关联研究的数据整合，可以确定可能作为人类新型DR治疗靶点的Müller细胞基因。在对最有希望的候选基因进行功能表征后，我们的目标是对一个选定的候选基因进行ShH 10驱动的调制，以保留关键的Müller细胞功能，促进糖尿病视网膜中的神经元存活。总之，我们相信，本项目中收集的数据将大大有助于开发新的长期有效的DR治疗方案，迄今为止，由于其复杂的疾病病因，传统的治疗方法不能令人满意地解决。