Endocrine pancreatic cell regeneration from bone marrow

描述（由申请人提供）：胰岛移植治疗1型糖尿病由于供体器官数量不足而受到限制。胰腺细胞的替代来源可以避免这个问题。我们的初步研究结果表明，骨髓（BM）含有能够移植胰岛和分泌胰岛素的多能细胞。我们设计了一种体内小鼠骨髓移植模型，在这种模型中，可以通过表达增强的绿色荧光蛋白（EGFP）来鉴定供体来源的表达胰岛素的细胞。供体来源的细胞可以通过荧光激活细胞分选分离和富集，并进一步在体外测试。我们假设骨髓中含有一个细胞亚群，这些细胞有能力分化为胰腺内分泌β细胞，并且这些细胞是完全分化的，具有功能能力。我们最初的目的是：(1)评估BM来源的β细胞是否表现出成熟功能胜任的β细胞的特征。将在这些细胞中评估前胰岛素转化为胰岛素和c肽的过程。在培养的BM衍生的egfp阳性胰岛细胞中，将进行对葡萄糖的反应性研究，对胰高血糖素样肽-1的反应性。钙和atp敏感钾通道的电生理特性将使用膜片钳技术来确定。胰岛素分泌颗粒在超微结构水平上的形态表征将补充这些研究。(2)评估骨髓源性细胞向胰腺内分泌细胞“转分化”的动力学和潜在机制，并寻找增加这种“转分化”过程相对数量的策略。(3)验证骨髓移植能否成功治疗糖尿病小鼠模型。(4)确定乳腺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BM来源的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.