Insulin Producing Cells from Amniotic Stem Cells for Diabetes Therapy

简介（由申请人提供）：糖尿病是一个日益严重的世界性问题。在美国，它影响了1800多万人，每年的医疗费用超过1300亿美元。1型糖尿病和晚期2型糖尿病患者的胰岛素治疗不能预防严重的长期并发症，包括神经病变、血管疾病、视网膜病变和肾功能衰竭。胰岛移植恢复胰岛素的产生提供了重要的前景。然而，供体胰腺的供应远远不能满足医疗需求。为了充分发挥细胞治疗糖尿病的潜力，需要新的胰岛素生成细胞来源。我们提出通过从羊水中分离的干细胞体外分化产生胰腺β谱系细胞。这些“AFS细胞”能够广泛扩展和分化为所有三个胚胎胚层的衍生物。我们的初步研究表明，在胰腺转录因子PDX-1的表达促进下，小鼠AFS细胞可以产生胰岛素产生细胞和胰岛样细胞团（“新胰岛”）。我们现在建议从人类和非人灵长类动物（NHP）的AFS细胞中产生新胰岛。为了有效地生成胰岛素生成细胞，我们将优化表达PDX-1的质粒载体的递送，并将系统地测试先前显示的促进胰腺细胞分化的生长因子和底物。由此产生的新胰岛将与真正的人类和NHP胰岛进行比较，使用用于临床移植的测试。为了评估其恢复葡萄糖代谢控制以及胰岛素和c肽产生的能力，将新胰岛植入经链脲佐菌素（STZ）治疗的糖尿病免疫缺陷小鼠。stz治疗的NHP移植将在生理学上更类似于人类的模型中评估新胰岛的功能。将在孕妇羊膜穿刺术后获得NHP AFS细胞系。这些干细胞将用于产生新的胰岛，用于自体移植到相应的后代中。同样的供体细胞将在异体受体中使用临床相关的免疫抑制方案进行比较。成功开发可移植的胰岛素生成细胞的丰富来源可能对治疗一个重大的公共卫生问题产生深远的影响。

DESCRIPTION (provided by applicant): Diabetes mellitus is a growing problem worldwide. In the US it affects over 18 million people and results in annual health care costs exceeding $130 billion. Insulin therapy of Type 1 diabetes, and in advanced cases of Type 2 diabetes, does not prevent serious long-term complications including neuropathy, vascular disease, retinopathy and renal failure. Transplantation of pancreatic islets to restore insulin production offers significant promise. However, the supply of donor pancreata falls far short of meeting the medical need. New sources of insulin producing cells will be required to realize the full potential of cell therapy for diabetes. We propose to generate pancreatic beta lineage cells by in vitro differentiation of stem cells isolated from amniotic fluid. These "AFS cells" are capable of both extensive expansion and differentiation into derivatives of all three embryonic germ layers. Our Preliminary Studies showed that mouse AFS cells can yield insulin producing cells and islet-like cell clusters ("neo-islets"), promoted by expression of the pancreatic transcription factor PDX-1. We now propose to produce neo-islets from human and non-human primate (NHP) AFS cells. To efficiently generate insulin producing cells, we will optimize delivery of a plasmid vector to express PDX-1, and will systematically test growth factors and substrates shown previously to promote pancreatic beta cell differentiation. The resulting neo-islets will be compared with authentic human and NHP pancreatic islets using tests developed for clinical transplantation. To assess their ability to restore control of glucose metabolism and production of insulin and C-peptide, neo-islets will be implanted in immunodeficient mice made diabetic with streptozotocin (STZ). Transplantation in STZ-treated NHP will assess the function of neo-islets in a model physiologically more similar to humans. NHP AFS cell lines will be derived after amniocentesis of pregnant mothers. These stem cells will be used to generate neo-islets for autologous transplantation into the corresponding offspring. The same donor cells will be compared in allogeneic recipients using clinically relevant immunosuppression regimens. Successful development of an abundant source of transplantable insulin producing cells potentially would have a profound impact on the treatment of a major public health problem.