Transformational platform for regenerating autologous transplantable endocrine tissue from human pancreatic matrix and pluripotent stem cells

从人胰腺基质和多能干细胞再生自体可移植内分泌组织的转化平台

• 批准号: 9169474
• 负责人: Jon S Odorico
• 金额: $ 22.31万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9169474
• 关键词: Address; Adult; Age; Anti-Inflammatory Agents; Anti-inflammatory; Autologous; Beta Cell; Biological; Biomedical Engineering; Blood Vessels; Blood flow; Cell Survival; Cell physiology; Cells; Child; Collaborations; Data; Deposition; Development; Diabetes Mellitus; Endocrine; Endocrine Glands; Endothelial Cells; Engineering; Engraftment; Exhibits; Extracellular Matrix; Family suidae; Future; Gel; Glucose; Goals; Health; Human; Hydrogels; Hypoxia; Immunosuppression; In Vitro; Inflammation; Insulin; Insulin-Dependent Diabetes Mellitus; Islets of Langerhans; Islets of Langerhans Transplantation; Kidney; Label; Life; Link; Liver; Mass Spectrum Analysis; Medical; Medicine; Mesenchymal; Methods; Modeling; Molds; Mus; Natural regeneration; Non-Insulin-Dependent Diabetes Mellitus; Nutrient; Organ; Organ Donor; Oxygen; Pancreas; Patients; Pharmacy Schools; Physiological; Pluripotent Stem Cells; Porifera; Portal vein structure; Procedures; Property; Proteins; Proteome; Proteomics; Regenerative Medicine; Replacement Therapy; Reporting; Research; Schools; Signal Transduction; Site; Staging; Stem cells; Stromal Cells; Techniques; Technology; Testing; Time; Tissue Engineering; Tissue Grafts; Tissue Sample; Tissue Transplantation; Tissues; Transplantation; allograft rejection; base; beta cell replacement; cell behavior; clinically relevant; design; diabetic; fetal; glycemic control; human embryonic stem cell; human tissue; implantation; improved; innovation; innovative technologies; insulin secretion; interest; intrahepatic; islet; minimally invasive; mouse model; new technology; novel; paracrine; prevent; regenerative; research study; scaffold; stem cell differentiation; subcutaneous

ABSTRACT Diabetes and its complications still claim the lives of millions of people despite continuing advances in insulin delivery technology primarily because insulin fails to achieve perfect glycemic control. On the other hand, beta cell replacement therapies including vascularized pancreas and isolated islet transplantation are able to fully restore normoglycemia, achieve insulin-independence and can delay end-organ complications. However, these latter therapies suffer from two key limitations, the shortage of organs and the need for life- long immunosuppression to prevent allograft rejection. Furthermore, the intrahepatic portal vein islet transplantation site used in humans is far from ideal and many islets are lost after implantation. An ideal beta cell replacement therapy strives towards both generating an abundant supply of functional beta cells and identifying a minimally invasive, well-vascularized, retrievable site for transplantation that is clinically applicable. After years of research it is now well established that human pluripotent stem cells (hPSCs) can be directed to differentiate into highly enriched physiological functional islet-like clusters (ILCs) in vitro that are capable of curing diabetes in mice. The extracellular matrix (ECM) is a critical component of the cellular niche that helps maintain cellular differentiation and provides tissue-specific signals to guide the fate and behavior of cells. Recent progress in the decellularization of organs has spurred great interest in using natural matrix for regenerative medical applications; yet, few studies have focused on the pancreas in general and the human pancreas to date has not been effectively decellularized and studied. Appreciating the importance of tissue-specific ECM, we have established effective techniques for the decellularization and delipidization of human pancreas tissue to produce several types of natural matrix constructs, including intact 3D matrix, molded sponge scaffolds and a spontaneous gelling hydrogel (hP-ECM). With the challenges of identifying a clinically applicable transplant site that provides for immediate and sufficient oxygen and nutrient delivery, we believe there is compelling rationale to take advantage of the proven proangiogenic and anti-inflammatory properties of ECs and MSCs. Thus, transplanting ILCs with hPSC-derived endothelial cells (ECs) and hPSC-derived mesenchymal stromal cells (MSCs), each providing essential properties, combined with hP-ECM into a prevascularized deviceless retrievable subcutaneous site might provide a more optimal transplant platform. Now, based on this innovative technology we aim to obtain a better understanding of the composition and function of natural hP-ECM in the context of hPSC differentiation to beta cells. The immediate objectives are to characterize human pancreatic extracellular matrix and to use this natural matrix in combination with stem cell-derived β cells, ECs and MSCs to reconstruct endocrine tissue capable of glucose- stimulated insulin-secretion after transplantation to mice. Our specific aims are to: 1) Comprehensively characterize the human pancreatic and islet ECM proteome, or matrixome, and compare the matrixome of different developmental ages using advanced quantitative mass spectrometry methods in collaboration with Dr. Linjun Li, 2) Construct a hP-ECM - cellular composite tissue graft combining hPSC-ILCs with ECs +/- MSCs and test its function in an immunodeficient murine diabetes model. Ultimately, we envision a bioengineered composite endocrine organ as a highly innovative regenerative medicine strategy for producing potentially autologous insulin-producing tissue for transplantation. These basic enabling studies are the first steps towards developing an effective, minimally invasive transplant platform that is available for all patients with diabetes.

抽象的 尽管糖尿病及其并发症不断取得进展，但它仍然夺去了数百万人的生命。 胰岛素输送技术主要是因为胰岛素无法实现完美的血糖控制。另一方面 另一方面，β细胞替代疗法，包括血管化胰腺和离体胰岛移植 能够完全恢复正常血糖，实现胰岛素非依赖性，并可以延缓终末器官并发症。 然而，后一种疗法有两个关键限制，即器官短缺和生命需要—— 长期免疫抑制可防止同种异体移植排斥。此外，肝内门静脉胰岛 用于人类的移植部位远非理想，许多胰岛在植入后丢失。理想的测试版 细胞替代疗法致力于产生充足的功能性β细胞和 确定临床上微创、血管良好、可回收的移植部位 适用的。 经过多年的研究，现已证实人类多能干细胞（hPSC）可以 定向分化为高度富集的生理功能性胰岛样簇（ILC）， 能够治愈小鼠糖尿病。 细胞外基质 (ECM) 是细胞生态位的重要组成部分，有助于维持细胞 分化并提供组织特异性信号来指导细胞的命运和行为。最近的进展 器官的脱细胞化激发了人们对使用天然基质进行再生医学的极大兴趣 应用程序；然而，很少有研究关注一般胰腺，并且迄今为止人类胰腺 尚未得到有效的脱细胞和研究。认识到组织特异性 ECM 的重要性，我们有 建立了人类胰腺组织脱细胞和脱脂的有效技术 生产多种类型的天然基质结构，包括完整的 3D 基质、模制海绵支架和 自发胶凝水凝胶（hP-ECM）。 面临着确定临床适用的移植位点的挑战，该位点可以立即提供 充足的氧气和营养输送，我们相信有令人信服的理由来利用经过验证的 EC 和 MSC 的促血管生成和抗炎特性。因此，用 hPSC 衍生的 ILC 进行移植 内皮细胞 (EC) 和 hPSC 衍生的间充质基质细胞 (MSC)，各自提供必需的 特性，与 hP-ECM 结合到预血管化的无装置可回收皮下部位可能 提供更优化的移植平台。 现在，基于这项创新技术，我们的目标是更好地了解其成分和 天然 hP-ECM 在 hPSC 分化为 β 细胞的过程中的功能。近期目标是 表征人胰腺细胞外基质并结合使用这种天然基质 使用干细胞衍生的 β 细胞、EC 和 MSC 来重建能够葡萄糖- 移植到小鼠体内后刺激胰岛素分泌。 我们的具体目标是：1) 全面表征人类胰腺和胰岛 ECM 蛋白质组， 或基质组，并使用高级定量质量比较不同发育年龄的基质组 与李林军博士合作的光谱测定方法，2）构建 hP-ECM - 细胞复合组织 将 hPSC-ILC 与 EC +/- MSC 相结合的移植物并测试其在免疫缺陷小鼠糖尿病中的功能 模型。最终，我们设想生物工程复合内分泌器官作为一种高度创新的再生性器官 产生用于移植的潜在自体胰岛素产生组织的医学策略。这些 基础研究是开发有效的微创移植的第一步 适合所有糖尿病患者的平台。