A perfluorocarbon-based culture device for beta cell biology applications (Phase

用于 β 细胞生物学应用的基于全氟化碳的培养装置（Phase

基本信息

批准号：
8314435
负责人：
Juan Dominguez-Bendala
金额：
$ 59.74万
依托单位：
OPHYSIO, INC.
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-10 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8314435
关键词：
Academia Address Administrator Adult Air Award Basic Science Beta Cell Biological Testing Biology Blood flow California Cell Culture Techniques Cell Death Cell Differentiation process Cell Survival Cell Therapy Cell physiology Cells Cellular biology Cessation of life Clinical Custom Development Devices Diabetes Mellitus Diffusion Digestive System Disorders Discipline Employee Strikes Ensure Equipment Evolution Fluorocarbons Funding Future Generations Goals Growth Human Hyperoxia Hypoxia Immunodeficient Mouse In Vitro Industry Institutes Insulin-Dependent Diabetes Mellitus Islet Cell Islets of Langerhans Islets of Langerhans Transplantation Kidney Diseases Knowledge Laboratories Legal patent Link Marketing Medical Membrane Methods Mission Modeling Organ Outcome Oxygen Oxygen measurement, partial pressure, arterial Pancreas Pattern Phase Physicians Physiological Production Public Health Regenerative Medicine Replacement Therapy Reproducibility Research Research Institute Silicones Speed Stem Cell Research Stem cells Sterilization Structure of beta Cell of islet System Technology Temperature Testing Tissues Translational Research Translations Transplantation Universities Washington Work Xenograft procedure base beta cell replacement cell type commercialization design diabetes mellitus therapy diabetic embryonic stem cell experience fetal flasks improved interest islet islet stem cells manufacturing process novel phase 1 study pre-clinical prevent prospective scale up stem cell differentiation stem cell therapy success technology development tissue culture tissue/cell culture tool trend

项目摘要

ABSTRACT Conventional culture vessels are not designed for physiological oxygen delivery. Both hyperoxia and hypoxia - commonly observed when culturing cells and tissues in regular plasticware- have been linked to reduced cellular function and death. An adequate means to provide oxygenation is also critical for stem cell applications in which the differentiation outcome is dependent on oxygen tension levels. We have addressed this problem by devising a novel culture device, the "oxygen sandwich". This simple system is designed to deliver oxygen in a quasi-physiological fashion by means of a basal air-permeable perfluorocarbon-silicone (PFC/Si) membrane. Our long-term goal is to establish this system as a new standard for tissue culture, both for applications in which oxygen diffusion rates become limiting (such as 3D culture) and for those that require precise adjustments of oxygenation to steer stem cell differentiation in the desired direction. We will focus our proof-of- concept work on islet/beta cell biology. This is a rapidly expanding market that includes clinical uses (islet transplantation), active in vitro research on adult/fetal islets of several species, and pre-clinicl stem cell research with the potential to revolutionize the treatment of diabetes within the next 5-10 years. The pertinence of our model choice is highlighted by two well-documented observations: [1] Pancreatic beta cells (the current end product used in clinical therapies for diabetes) are highly sensitive to sub- and super-physiological oxygen concentrations; and [2] Stem cell differentiation into beta cells (the subject of worldwide research to replace cadaveric islets for future clinical uses) is exquisitely dependent on evolving oxygen tensions. Our Phase I studies aimed at demonstrating that the enhanced in vitro survival and function observed in PFC/Si-cultured islets also resulted in better pre-clinical transplantation outcomes using a marginal mass xenotransplantation model (human islets into diabetic nu/nu immunodeficient mice). After the successful completion of these studies, our Phase II proposal is based on the following specific aims: (1) Scaling-up and definition of manufacturing process (QA & QC, sterilization) for mass production of PFC/Si culture devices; and (2) biological testing and reproducibility studies in human islets and embryonic stem cells (hESc). This application benefits from the assembly of first-rate teams with highly complementary expertise. Our Phase I results are strongly supportive of the feasibility of this proposal. Success in our research would fill a widely acknowledged gap in our ability to preserve islet cell function and survival in vitro confirming this system as a potential new standard for beta cell biology and differentiation studies. As such positive outcome might ultimately speed up the applicability of new-generation beta cell replacement therapies, this project is greatly relevant to the mission of the National Institutes of Diabetes and Digestive and Kidney Diseases (NIDDK). Success in these studies will also provide proof of principle of the superiority of our oxygen enhancing technology for many other cell culture applications.

抽象的传统的培养容器不是为生理氧气输送而设计的。高氧和缺氧（在普通塑料器皿中培养细胞和组织时常见）都与细胞功能下降和死亡有关。提供氧合的适当方法对于干细胞应用也至关重要，其中分化结果取决于氧张力水平。我们通过设计一种新型培养装置“氧气三明治”解决了这个问题。这个简单的系统旨在通过基础透气的全氟碳硅 (PFC/Si) 膜以准生理方式输送氧气。我们的长期目标是将该系统建立为组织培养的新标准，既适用于氧扩散速率受到限制的应用（例如 3D 培养），也适用于需要精确调整氧合以引导干细胞分化的应用。想要的方向。我们将把概念验证工作的重点放在胰岛/β 细胞生物学上。这是一个快速扩张的市场，包括临床应用（胰岛移植）、对多个物种的成人/胎儿胰岛的活跃体外研究以及临床前干细胞研究，有可能在未来 5-10 年内彻底改变糖尿病的治疗年。两个有据可查的观察结果强调了我们模型选择的相关性：[1] 胰腺β细胞（目前用于糖尿病临床治疗的最终产品）对亚生理和超生理氧浓度高度敏感； [2] 干细胞分化为β细胞（全球研究的主题，以取代尸体胰岛用于未来的临床用途）完全依赖于不断变化的氧张力。我们的第一阶段研究旨在证明，使用边缘质量异种移植模型（将人类胰岛移植到糖尿病 nu/nu 免疫缺陷小鼠中），在 PFC/Si 培养的胰岛中观察到的体外存活和功能增强也导致了更好的临床前移植结果。成功完成这些研究后，我们的第二阶段建议基于以下具体目标：（1）扩大和定义PFC/Si培养装置大规模生产的制造工艺（QA和QC、灭菌）； (2) 人类胰岛和胚胎干细胞 (hESc) 的生物测试和再现性研究。该应用程序受益于具有高度互补专业知识的一流团队的集合。我们的第一阶段结果强烈支持该提案的可行性。我们研究的成功将填补我们在体外保存胰岛细胞功能和存活的能力方面广泛承认的空白，从而证实该系统作为β细胞生物学和分化研究的潜在新标准。由于这种积极的结果可能最终会加速新一代 β 细胞替代疗法的应用，因此该项目与美国国立糖尿病、消化和肾脏疾病研究所 (NIDDK) 的使命密切相关。这些研究的成功还将为我们的增氧技术在许多其他细胞培养应用中的优越性提供原理证明。