POMGnT1突变所致的肌肉-眼-脑病的细胞与分子机制研究

项目名称： POMGnT1突变所致的肌肉-眼-脑病的细胞与分子机制研究

项目编号： No.30870867

项目类型： 面上项目

立项/批准年度： 2009

项目学科： 金属学与金属工艺

项目作者： 杨渊

作者单位： 华中科技大学

项目金额： 35万元

中文摘要： POMGnT1等糖基转移酶突变导致肌营养不良蛋白聚糖（DG）不能有效地与细胞外基质结合，从而不能形成完整的基底膜。但至今仍然不清楚细胞与细胞外基质结合的缺陷是如何引起基底膜的破裂的。采用POMGnT1突变小鼠模型，系统地研究了POMGnT1突变小鼠的基底膜的破坏及其机制。POMGnT1突变小鼠的脑皮层及放射状胶质细胞DG糖基化受损、laminin结合力减弱或消失。突变的神经干细胞球表面的细胞外基质成份（laminin-111, collagen IV, nidogen-1和perlecan）的组装速度明显下降，laminin-111和nidogen-1在神经干细胞球表面的结合显著减少。原子力显微镜（AFM)分析发现，POMGnT1突变的ILM变薄而且拓扑结构受到破坏，POMGnT1突变基底膜（BM）全面破坏，尽管某些区域显微镜下看似正常。肌营养不良蛋白聚糖病者的基底膜主要组份减少，细胞外基质组装效率降低，是基底膜物理结构改变并最终破裂的原因，尤其在神经组织快速发育阶段。同时发现POMGnT1突变小鼠脑膜成纤维细胞可诱导脑星形胶质细胞增生，BMP7至少部分参与了其星形胶质细胞增生的过程。

中文关键词： 肌肉-眼-脑病；基底膜；POMGnT1；αG；细胞外基质

英文摘要： Neuronal ectopias in the brain and retina of dystroglycanopathies are caused by disruptions of two basement membranes (BM), the pial BM and inner limiting membrane (ILM) respectively. The underlying cause for BM disruptions is a diminished αystroglycan (αG) binding of extracellular matrix (ECM) owing to mutations in glycosyltransferases such as protein O-mannose N-acetylglucosaminyltransferase 1 (POMGnT1), is the underlying cause. It is unclear how diminished cell-ECM interactions lead to BM disruptions. We hypothesized that the rate of ECM assembly is reduced resulting in a structurally defective BM. In this study, ECM aggregation on neural stem cells was analyzed. Laminin aggregation on POMGnT1 knockout neural stem cells was reduced compared to the wildtype. Furthermore, on the surfaces of neural spheres incubated with Matrigel, the ECM contained all four major BM components, including laminin-111, collagen IV, nidogen-1, and perlecan. The rate of ECM assembly on POMGnT1 knockout spheres was significantly reduced. Immunofluorescence staining and quantitative proteomic analysis revealed that laminin-111 and nidogen-1 were reduced in POMGnT1 knockout ILM. Atomic force microscopy showed that POMGnT1 knockout ILM was thinner with an altered surface ILM topography. These results indicate that the BM in POMGnT1 knockout mice is defective even in regions that appear microscopically normal. They suggest that reduced levels of key BM components may be the cause of physical structural changes that weaken the BM in dystroglycanopathies. These changes are likely caused by reduced assembly rate in ECM during developmental expansion of the neural tissue.And our data suggested that meningeal fibroblasts are an inducer of reactive astrogliosis and that such induction is mediated at least in part by BMP7.

英文关键词： musle-eye-brain disease; basement membrane; POMGnT1; αG；extrocellular matirx