Endocrine pancreatic cell regeneration from bone marrow

描述(申请人提供)：胰岛移植治疗1型糖尿病的疗效受到供体器官数量不足的限制。胰腺细胞的替代来源可能会绕过这个问题。我们的初步发现表明，骨髓(BM)含有多能细胞，能够植入胰岛并分泌胰岛素。我们设计了一种体内小鼠骨髓移植模型，在该模型中，表达胰岛素的供体来源细胞可以通过表达增强型绿色荧光蛋白(EGFP)来识别。供者来源的细胞可以通过荧光激活的细胞分选来分离和浓缩，并在体外进一步测试。我们假设骨髓中有一群细胞具有分化为胰腺内分泌β细胞的能力，并且这些细胞是完全分化和功能正常的。我们最初的目标是：(1)评估骨髓来源的β细胞是否表现出成熟的功能正常的β细胞的特征。胰岛素原转化为胰岛素和C-肽的过程将在这些细胞中进行评估。将研究培养的骨髓来源的EGFP阳性胰岛细胞对葡萄糖的反应性，以及对胰升糖素样肽-1的反应性。钙和三磷酸腺苷敏感钾通道的电生理特性将使用膜片钳技术进行测定。胰岛素分泌颗粒在超微结构水平的形态特征将补充这些研究。(2)评价骨髓来源细胞向胰腺内分泌细胞“转分化”的动力学和机制，并寻找增加这一“转分化”过程相对数量的策略。(3)检测骨髓移植能否成功治疗糖尿病小鼠模型。(4)确定BM内对胰腺内分泌细胞群有贡献的细胞类型。这些研究的长期目标是评估骨髓来源细胞移植作为糖尿病的潜在治疗方法。

DESCRIPTION (provided by applicant): Curative treatment of type 1 diabetes mellitus by islet transplantation is limited by insufficient numbers of donor organs. Alternative sources for pancreatic cells may circumvent this problem. Our preliminary findings suggest that bone marrow (BM) contains pluripotent cells that are capable of engrafting pancreatic islets and secreting insulin. We have devised an in vivo mouse model of bone marrow transplantation in which donor derived cells that express insulin can be identified by their expression of enhanced green fluorescent protein (EGFP). Donor-derived cells can be isolated and enriched by fluorescence activated cell sorting and further tested in vitro. We hypothesize that bone marrow contains a subpopulation of cells that has the capacity to differentiate into pancreatic endocrine beta-cells and that these are fully differentiated and functionally competent. Our initial aims are: (1) To assess whether BM derived beta-cells exhibit characteristics of mature functionally competent beta-cells. Processing of pro-insulin to insulin and C-peptide will be assessed in these cells. Studies of the responsiveness to glucose, to the incretin-hormone glucagon-like peptide-1 in cultured BM derived EGFP-positive islet cells will be conducted. Electrophysiological properties of calcium and ATP-sensitive potassium channels will be determined using the patch-clamp technique. Morphological characterization of insulin secretory granules at the ultrastructural level will complement these studies. (2) To evaluate the kinetics and underlying mechanisms of "transdifferentiation" of BM derived cells into pancreatic endocrine cells and to find strategies to increase the relative quantity of this "transdifferentiation" process. (3) To test whether diabetic mouse models can be successfully treated with bone marrow transplantation. (4) To identify the cell type within BM that contributes to the pancreatic endocrine cell population. The long-term goal of these studies is to evaluate transplantation of BM derived cells as a potential treatment for diabetes mellitus.