INHIBITION OF APOPTOSIS IN PANCREATIC BETA CELLS

缺乏对葡萄糖敏感的胰腺β细胞 胰岛素分泌一直是广泛发展的主要障碍， 适用于糖尿病的细胞移植疗法。 为了解决这个 问题，我们研究的最终目标是发展扩大 人β细胞或β细胞前体的群体。 这将 提供了大量的细胞，在体外生长，保留葡萄糖- 反应性胰岛素分泌或分化和获得 这个财产。 这样的细胞可以用于细胞移植 糖尿病的治疗方法 目前正在采用两种方法来扩增人类β细胞， 内分泌细胞前体细胞。 首先是培养原代细胞 在肝细胞生长的情况下对细胞外基质（ECM）的影响 因子/分散因子（HGF/SF）。 二是表达主导 细胞中的致癌基因，导致基质和生长因子 在体外独立生长。 原代细胞扩增后， 它们必须从ECM中去除并聚集成胰岛样细胞 集群（ICC）。 然而，扩增的原代细胞从 基质导致称为失巢凋亡的细胞凋亡形式。为 表达SV40 T抗原和H-ras\Val 12显性的β细胞系 癌基因，使用cre-lox重组酶去除癌基因 系统，在汇集到ICC之前，导致快速和有效的 凋亡因此，在两种不同的细胞中， 在试图扩增人类β细胞和β细胞的情况下， 文化的先驱。 在这项建议中，抑制的方法 使用一氧化氮抑制剂和腺病毒载体的细胞凋亡 将探索表达抗凋亡基因。 的影响 抗凋亡基因对移植原发性非扩增 还将研究人胰腺内分泌细胞的培养物。 这些研究对扩大人口的发展很重要 用于细胞移植的人类β细胞和β细胞前体 糖尿病的治疗方法

The shortage of pancreatic beta-cells exhibiting glucose-responsive insulin secretion has been a major obstacle to the development of widely applicable cell transplantation therapies for diabetes. To address this problem, the ultimate goal of our research is to develop expanded populations of human beta cells or beta-cell precursors. This would provide a large quantity of cells, grown in vitro, that retain glucose- responsive insulin secretion or the ability to differentiate and acquire this property. Such cells could then be used for cell transplantation therapies for diabetes. Two approaches are being pursued to expand human beta-cells and endocrine cell precursors in vitro. The first is to grow primary cells on extracellular matrix (ECM) in the presence of hepatocyte growth factor/scatter factor (HGF/SF). The second is to express dominant oncogenes in the cells, resulting in matrix and growth factor independent growth in vitro. Following the expansion of primary cells, they must be removed from the ECM and aggregated into islet-like cell clusters (ICCs). However, detachment of the expanded primary cells from the matrix results in a form of apoptosis known as anoikis. For the beta-cell line expressing the SV40 T antigen and H-ras\Val12 dominant oncogenes, removal of the oncogenes using the cre-lox recombinase system, prior to aggregation into ICCs, results in rapid and efficient apoptosis. Thus, apoptosis has been encountered in two distinct situations in attempts to expand human beta cells and beta-cell precursors in culture. In this proposal, approaches to inhibition apoptosis using inhibitors of nitric oxide and adenoviral vectors expressing anti-apoptotic genes will be explored. The effects of the anti-apoptotic genes on transplantation of primary, non-expanded cultures of human pancreatic endocrine cells will also be studied. These studies are important to the development of expanded populations of human beta cells and beta-cell precursors for cell transplantation therapies for diabetes.