Netrin-1修饰的ASC对失神经支配后糖尿病小鼠血管新生的影响及分子机制

糖尿病周围神经血管病变(DPNV)发病率高、病程长、防治困难，血管合并神经病变是其难治愈的主要原因。脂肪干细胞(ASC)促进DPNV血管新生展示较好应用前景，但我们研究发现ASC在靶血管损伤部位存活及分化能力减低，是影响疗效的关键。作为重要的生长因子，Netrin-1可促进神经及血管损伤的修复，且对内皮细胞、干细胞具有保护作用。由此我们提出假设：通过使ASC过表达Netrin-1，可能会更好的促进失神经支配后糖尿病血管的新生。本项目拟在前期基础上，通过核转染使ASC过表达Netrin-1，体外检测其对高糖刺激下ASC存活、迁移、凋亡及分化的影响；体内移植治疗失神经支配糖尿病小鼠，检测新生血管血流、管径、密度变化，应用Netrin-1相关受体阻断剂及siRNA技术探讨相关信号通路的变化，阐明Netrin-1修饰的ASC促进失神经支配后血管新生的分子机制，为DPNV的防治提供新思路。

Diabetic peripheral nerve and vascular disease (DPNV) has high incidence, long course, and the early prevention and treatment are difficult. Adipose derived stem cells (ASC) play a pivotal role in promoting angiogenesis in diabetes, which shows a wide prospect of clinical application. However, our previous studies showed that ASC weakened in the target vessel parts of the survival, and differentiation capacity, which is the key to affect our active effects. As an important neurotrophic guiding factor, Netrin-1 not only takes part in the development of peripheral neuropathy and angiogenesis but also promotes endothelial cells and stem cells survival, migration, and blood vessel formation, however, its expression is lower in ASC. Thus we propose a hypothesis that overexpression of Netrin-1 by transfection in ASC might be better to promote its survival, migration and improve angiogenesis ability in diabetic mice after denervation. This study will based on our previous results, nuclear transfection will be adopted to make ASC overexpressing Netrin-1 and explore the influence of Netrin-1 on ASC survival, migration, and the differentiation capacity into endothelial cells. A unique diabetic mouse model of denervation will be established, and GFP gene will be used for tracing the roles of Netrin-1-ASC in promoting angiogenesis after cell transplantation. The numbers, diameters, and density of new vessels will also be analyzed at different transplantation times. Netrin-1 receptor DCC blocking agents and RNA Interference Silencing will be used to explore the possible effects of Netrin-1 transfected ASC in promoting angiogenesis in diabetic mouse model of denervation. The associated signal pathway will be discussed and the molecular mechanisms will be elucidated, which will bring new insights and theoretical basis into the prevention and treatment of diabetic peripheral nerve and vascular disease.