Cell therapy of diabetes using broad spectrum multipotent stem cells

使用广谱多能干细胞治疗糖尿病

• 批准号: 7293549
• 负责人: SHAY SOKER
• 金额: $ 50万
• 依托单位: PLUREON CORPORATION
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7293549
• 关键词: Address; Adenovirus Vector; Amniotic Fluid; Animal Disease Models; Animal Model; Animals; Beta Cell; Biological Assay; Blood Circulation; Blood Glucose; Brain Death; C-Peptide; Cell Aging; Cell Therapy; Cells; Clinical; Clonal Expansion; Condition; Development; Diabetes Mellitus; Diabetic mouse; Disease; Doctor of Philosophy; Electroporation; Embryo; Face; Generations; Genetic screening method; Germ Layers; Glucose; Goals; Heart; Home environment; Human; Immune; In Vitro; Injection of therapeutic agent; Insulin; Insulin-Dependent Diabetes Mellitus; Islets of Langerhans; Kidney; Kidney Failure; Multipotent Stem Cells; Mus; Natural regeneration; Neuropathy; Non-Insulin-Dependent Diabetes Mellitus; Numbers; Organ; Pancreas; Patients; Phenotype; Plasmid Cloning Vector; Population; Principal Investigator; Procedures; Production; Protocols documentation; Recombinants; Regulation; Regulator Genes; Research; Retinal Diseases; Source; Staging System; Stem cells; Streptozocin; Structure; Structure of beta Cell of islet; Teratoma; Testing; Therapeutic; Toxin; Translations; Transplantation; Treatment Protocols; amniotic fluid derived stem cell; beta cell replacement; blood glucose regulation; capsule; cell type; desire; diabetes mellitus therapy; diabetic; embryonic stem cell; expression vector; glucose production; human stem cells; implantation; in vivo; islet; novel; prenatal; prevent; progenitor; programs; research study; restoration; stem; transcription factor; tumor; vector

DESCRIPTION (provided by applicant): Transplantation of insulin-producing cells in many diabetes patients potentially would restore normal glucose homeostasis and prevent the severe long-term complications of the disease. However, the scarcity of donated pancreata currently limits transplantation of the whole organ or of isolated pancreatic islets to a tiny fraction of those patients who might benefit from such treatment. Directed differentiation of stem cells offers a possible means to generate abundant insulin-producing cells. A promising new source of stem cells has been identified, namely, amniotic fluid collected for prenatal genetic testing. Amniotic fluid-derived stem (AFS) cells can be expanded extensively in culture, do not form teratoma tumors, and have the capacity to yield a variety of specialized cell types. Preliminary experiments with mouse AFS cells showed that they can give rise to insulin-producing cells resembling beta-cells of the pancreas. Differentiation along this lineage was promoted by transient expression of the pancreatic transcription factor Pdx-1. The proposed project will determine whether human AFS cells, driven by Pdx-1 provided from an expression vector, can similarly yield insulin-producing cells of the pancreatic lineage. Two (2) complementary strategies will be explored to develop a stem cell-based therapy for diabetes. 1 approach will be to generate clusters of insulin-producing cells in culture that resemble pancreatic islets ("neo-islets") and could be utilized in a transplantation procedure that has proven successful with isolated islets, the Edmonton Protocol. Human AFS cells will be induced to express human Pdx-1 by introduction of a plasmid vector via nucleofection, a high efficiency form of electroporation. The cells will be cultured in a 2-stage system shown previously to support the production from mouse AFS cells, transduced with a Pdx-1 vector, of insulin-producing cells in neo-islet structures. The resulting differentiated human cells will be assayed for multiple markers of pancreatic beta-cells and for the capacity to synthesize insulin and to secrete it in a glucose-responsive manner. The potential therapeutic value of neo-islets produced from human AFS cells will be tested by transplantation under the kidney capsule in immune-deficient mice rendered diabetic by streptozotocin, a toxin that destroys endogenous pancreatic beta-cells. The second approach will be to introduce the Pdx-1 expression vector into the human stem cells and then inject them directly into the circulation of diabetic mice. The treated animals will be tested for restoration of normal regulation of blood glucose, the production of human-specific insulin and C-peptide, and the regeneration of pancreatic islets by the human cells. Preliminary studies with mouse AFS cells suggest that this approach can provide long-term reversal of diabetes.

描述（由申请人提供）：许多糖尿病患者中产生胰岛素细胞的移植可能会恢复正常的葡萄糖稳态，并防止严重的长期并发症。但是，捐赠的胰腺的稀缺性目前限制了整个器官或孤立的胰岛移植到可能从这种治疗中受益的那些患者中的一小部分。干细胞的定向分化提供了产生丰富的胰岛素产生细胞的可能手段。已经确定了一种有希望的新的干细胞来源，即收集用于产前基因检测的羊水。羊水衍生的茎（AFS）细胞可以在培养中广泛扩展，不形成畸胎瘤肿瘤，并且具有产生各种专用细胞类型的能力。小鼠AFS细胞的初步实验表明，它们可以产生类似于胰腺β细胞的胰岛素产生细胞。通过胰腺转录因子PDX-1的瞬时表达促进了沿该谱系的分化。拟议的项目将确定由表达载体提供的PDX-1驱动的人类AFS细胞是否可以同样产生胰腺谱系的胰岛素产生细胞。将探讨两（2）个互补策略，以开发用于糖尿病的干细胞疗法。 1方法是在类似于胰岛（“新伊斯兰”）的培养物中生成产生胰岛素的细胞簇，并且可以在移植程序中使用，该程序已证明已通过孤立的胰岛（Edmonton方案）证明成功。人AFS细胞将通过通过核反射（一种高效的电穿孔形式）引入质粒载体来诱导人PDX-1。这些细胞将在先前显示的2阶段系统中培养，以支持新isleT结构中产生胰岛素的细胞的小鼠AFS细胞的产生，该细胞的产生，该细胞的产生。将对胰腺β细胞的多个标记物以及合成胰岛素合成并以葡萄糖反应方式分泌的能力分析所得分化的人类细胞。由人AFS细胞产生的新志尿菌的潜在治疗价值将通过在免疫缺陷型小鼠中通过肾脏胶囊的肾囊下的移植来测试，这是链霉菌素，链蛋白酶，抗蛋白酶，一种破坏内源性胰腺betaβ细胞的毒素。第二种方法是将PDX-1表达载体引入人类干细胞，然后将其直接注入糖尿病小鼠的循环中。治疗的动物将进行测试，以恢复血糖的正常调节，人类特异性胰岛素和C肽的产生以及人类细胞的再生胰岛的再生。使用小鼠AFS细胞的初步研究表明，这种方法可以长期逆转糖尿病。