2型糖尿病患者与正常人来源的iPS细胞改善动物模型脑缺血预后的比较研究

The technique that differentiates induced pluripotent stem cells from patient's own somatic cells to a variety of histiocytes, brings a new path for the treatment of many diseases. Our recent research successfully generated iPS cells from normal adult human fibroblasts by introduction of four defined transcription factors. These iPS cells had the efficiency in improving the prognosis of animal models of cerebral ischemia after transplantation. In view of the cerebral ischemia occurs mainly in type 2 diabetes, and there had been reports that fibroblasts obtained from diabetic individuals were different than those from normal individuals in characteristics. Therefore, it deserves our close attention to identify whether there is difference in the efficiency in improving cerebral ischemia between iPS cells generated from T2D patient's own fibroblasts and from normal subject. To use the iPS cells in the treatment of cerebral ischemia, there are three aspects deserve our close attention: clone formation and proliferative capacity, differentiation capacity into neuron-like cells, and the efficiency in improving the prognosis of cerebral ischemia. In this project, we will generate iPS cells from patients with different states of T2D and compare them with paired normal human iPS cells to determine whether there is difference in these three aspects and differential characteristics. In this project, cerebral ischemia common background diseases (T2D) will be included in basic research using iPS cells improve the prognosis of cerebral ischemia, to provide more reasonable evaluation for using the patient's own iPS cells in the clinical treatment of cerebral ischemia.

将病人自身体细胞诱导生成的多能干（iPS)细胞，再分化为各种组织细胞技术，为多种疾病的治疗带来全新策略。我们最近的研究将4种因子转入正常人成纤维细胞(FB)，成功制备iPS细胞，移植后有效改善动物模型脑缺血预后。鉴于脑缺血多发于2型糖尿病，而有报道称从糖尿病个体获得的FB特性不同于正常个体FB，因此用2型糖尿病人自身FB制备的iPS细胞与正常人来源的iPS细胞在治疗脑缺血的效果方面是否存在差异，值得高度重视。对用iPS细胞治疗脑缺血而言，其克隆形成和增殖能力、向神经元样细胞分化及改善脑缺血预后三个方面能力最值重视。本项目用不同病情程度2型糖尿病患者FB制备iPS细胞，并与配对的正常人iPS细胞进行上述三个方面的对比研究，明确有无差异及差异的特征。本项目将脑缺血常见的背景性疾病（2型糖尿病）纳入用iPS细胞改善脑缺血预后的基础研究，为将病人自身的iPS细胞用于脑缺血临床治疗提供更合理的评价。

将病人自身体细胞诱导生成的多能干（iPS)细胞，为多种疾病的治疗带来全新选择。我们前期发现从正常人成纤维细胞制备的iPS细胞可有效改善动物模型脑缺血预后。鉴于脑缺血多发于2型糖尿病，而从糖尿病个体获得的成体细胞特性不同于正常个体的成体细胞，因此用2型糖尿病人自身的成体细胞制备的iPS细胞与正常人的成体细胞来源的iPS细胞是否在细胞生物学特性和功能方面存在差异，是自体iPS用于治疗并发于糖尿病患者的脑卒中之前必须解决的重要问题。为了使所获得的诱导性多能干细胞携带更多的糖尿病性状，选用老年（74-89岁）重度糖尿病患者的细胞用于诱导。由于创口愈合障碍是糖尿病的重要并发症，有创性地获得成纤维细胞用于诱导iPS不被糖尿病患者接受，所以改用患者血淋巴细胞体外扩增的细胞制备患者诱导性多能干细胞。我们发现糖尿病患者的血淋巴细胞在体外生成克隆的能力、增殖能力和都显著弱于来源于正常人的血淋巴细胞，用携带4种经典诱导因子的慢病毒诱导iPS的成功率也从100%降低到20%，但改用仙台病毒后，诱导成功率可以提高到100%。将诱导的iPS培养10代后，来源于糖尿病患者和正常人的iPS在干细胞多能性、细胞活力、细胞生物学功能和对高糖的反应性等方面均无显著差异。由于异种细胞治疗受到免疫反应和免疫性炎症的影响，所以我们以同种同基因iPS为代表，观察了同种同基因小鼠iPS对小鼠缺血性脑卒中的治疗作用及其机制，发现同种同基因小鼠iPS可选择性归巢到脑损伤边缘区、减轻炎症和炎症因子表达、降低梗死面积并提高脑缺血小鼠生存率。本项目研究表明自体iPS可用于治疗并发于糖尿病患者的脑卒中。