Growth Factor and Antiapoptotic Gene Delivery to Human Islets

描述(申请人提供)：人胰岛移植作为治疗胰岛素依赖型糖尿病的有效手段具有很大的潜力。原发无功能是胰岛移植失败的主要原因，需要多个供体的移植。我们将验证这样的假设，即通过表达生长因子基因(如促进胰岛血运重建的人血管内皮生长因子(HVEGF))和抗凋亡基因(如人白介素1受体拮抗剂(hIL-IRA))可以同时增强胰岛移植，以防止移植胰岛在宿主体内的凋亡。 在初步研究中，携带hVEGF和绿色荧光蛋白的双顺反子腺病毒载体(ACV-GFP-hVEGF)的人胰岛分泌了较高水平的hVEGF，而AdV-GFP转染组和模拟转染组的hVEGF分泌水平很低。转基因胰岛的胰岛素释放与假转基因胰岛相当。促凋亡细胞因子干扰素-γ和肿瘤坏死因子-α不能诱导AdV-GFP-hVEGF诱导胰岛细胞凋亡。 我们的具体目的是确定i)编码hVEGF和hIL-IRA的腺病毒载体(AdvhVEGF-hLL-LRA)是否能够有效地转染胰岛并改善其功能；ii)体外转染腺病毒Adv-hVEGF-hLL-IRA是否可以防止原发胰岛无功能，并减少恢复正常血糖所需的胰岛质量。这项研究的意义在于，所提出的体外基因治疗将促进胰岛血运重建，防止细胞凋亡，并减少达到正常血糖所需的胰岛数量。这些数据将为人类胰岛移植的成功提供有益的参考。

DESCRIPTION (provided by applicant): Transplantation of human islets has great potential as an effective means of treating insulin dependent diabetes mellitus. Primary non-function is the main cause of islet graft failure and it results in the need for multi-donor transplants. We will test the hypothesis that islet engraftment can he enhanced simultaneously by expressing growth factor gene like human Vascular Endothelial Growth Factor (hVEGF) that promotes islet revascularization, and antiapoptotic gene like human interleukin-1 receptor antagonist (hIL-IRa) to prevent apoptosis of transplanted islets in the host. In preliminary studies, higher levels of hVEGF were secreted from human islets transfected with bicistronic adenoviral vector encoding hVEGF and Green Fluorescent Protein (Acv-GFP-hVEGF), while hVEGF secretion from Adv-GFP transfected and mock-transfected islets was very low. Insulin release from transfected islets was comparable to mock-transfected islets. Proapoptotic cytokines IFN-gamma and TNF-alpha did not induce apoptosis when islets were transfected with Adv-GFP-hVEGF. Our specific aims are to determine whether i) adenoviral vectors encoding hVEGF and hIL-IRa (AdvhVEGF- hlL-lRa) will efficiently transfect islets and improve their function; and ii) ex vivo transfection with Adv-hVEGF-hlL-IRa will prevent primary islet nonfunction and reduce the islet mass needed for restoring normoglycemia. The significance of this research is that the proposed ex vivo gene therapy will promote islet revascularization, prevent apoptosis and decrease the number of islets required for achieving normoglycemia. The data will be beneficial to successful human islet transplantation.