A multidisciplinary approach towards cell therapy for Diabetes

描述（由申请人提供）：越来越多的人认识到成人胰腺可能是胰岛祖细胞的真正来源，因此该组织代表了糖尿病细胞治疗的可行选择。然而，由于缺乏对胚胎后阶段器官中祖细胞/干细胞生态位的了解，阻碍了为此目的的实验操作。我们已经获得了成年胰腺中两个特定群体的证据，其满足以下标准：1）胰腺祖细胞上皮群体，其表现出多谱系潜能，和2）胰腺间充质细胞群体，其当与多能上皮细胞一起培养时，允许离体扩增。这两个群体的特点是视黄酸的生产。我们在这里成立了一个多研究者团队，以解决与这种产生视黄酸的上皮细胞的存在、功能和谱系效力以及受控分化相关的具体问题。这将涵盖成人胰腺组织的研究，采用基因工程，慢病毒载体，旨在表达自我裂解多聚蛋白模仿最近成功的工作，在体细胞重编程的其他人。我们证明了一个多基因/单载体的方法，允许在非胰腺细胞中的胰岛素基因激活的成功开发。我们的具体目标如下：Leach组：表征来自小鼠胰腺的两个aldefluor阳性细胞群的向前分化和连续生长。Oberholzer组：鉴定和表征成人内分泌胰腺中与鼠细胞相比的Aldefluor阳性细胞。詹森组：补充Leach和Oberholzer小组在Aldefluor人群研究中的工作，使用生物信息学，并在功能上测试和开发改良的多基因/单载体递送载体。鲑鱼组：为其他成员提供一系列编码单个蛋白质或可裂解多聚蛋白的慢病毒载体，用于成年鼠和人胰腺祖细胞的编程测试。 相关性：该项目与糖尿病细胞治疗的进展相关，该项目整合了对胰腺维甲酸产生祖细胞的分离和培养的理解，并寻求将其分化为功能齐全的胰腺胰岛素产生细胞。此外，还采用了先进的基因传递系统，其中包含能够进行内分泌细胞编程的可自切割多聚蛋白。

DESCRIPTION (provided by applicant): There is a growing understanding that adult pancreas may be a true source for islet progenitor cells, and that this tissue therefore represents a viable option for cell therapy in diabetes. However, a lacking understanding of progenitor/stem cell niches in the organ at post-embryonic stages, hinders experimental manipulation towards this end. We have obtained evidence for two specific populations in adult pancreas, that fulfill criteria of 1) a pancreatic progenitor epithelial population, exhibiting multi-lineage potency, and 2) a pancreatic mesenchymal cell population, that when cultured together with the multipotent epithelial cells, allows for ex-vivo expansion. Both populations are characterized by retinoic acid production. We have here formed a multi-investigator team to address specific issues related to the existence, the function and lineage potency, as well as the controlled differentiation of such retinoic acid producing epithelial cells. This will cover studies of adult human pancreatic tissue employing genetic engineering using lentiviral vectors designed to express self-cleavable polyproteins modeled over recent successful work in somatic cell reprogramming by others. We demonstrate the successful development of a multiple-gene/single vector method that allows insulin gene activation in non-pancreatic cells. Our specific goals are as follows: Leach group: to characterize the two aldefluor-positive cell populations from the murine pancreas with respect to forward differentiation and continuous growth. Oberholzer group: to identify and characterize Aldefluor-positive cells in the adult human endocrine pancreas comparatively to the murine cells. Jensen group: Complement the Leach and Oberholzer groups in studies of the Aldefluor populations, using bioinformatics and to functionally test and develop modified multiple gene/single-vector delivery vectors. Salmon group: to provide to other members a series of lentiviral vectors encoding individual proteins, or cleavable polyproteins, for testing in programming of adult murine, and human, pancreatic progenitors. RELEVANCE: This project is relevant for the advance of cell-based therapy in Diabetes, the project integrates efforts to understand the isolation and culture of pancreatic retinoic-acid producing progenitor cells, and seeks to differentiate such into fully function pancreatic insulin producing cells. The furthermore employs advanced gene delivery systems incorporating self-cleavable polyproteins capable of endocrine cellular programming.