EAGER: Biomanufacturing: Gene expression-based standardization of stem cells

PI：亲爱的，Eric M.提案编号：1547819基于细胞的生物制造高度依赖于其起始材料：细胞。使用定义不明确的混合细胞群体将导致不满意的结果，无论下游技术如何。提纯或浓缩高价值材料，如成人干细胞，对于现有和未来的广泛治疗策略是必要的。通过更好地控制细胞来源群体的组成，再生疗法和产品的性能被假设为更具重复性，使生物制造方法比今天更可行。目前的浓缩技术依赖于细胞表面蛋白的独特组合的存在，受到极低的细胞产量和对供体变异性的极大敏感性的限制。在这项工作中，研究人员建议开发和评估新的程序，使生产的干细胞群体具有一致的再生特性，不依赖于捐赠者。这项建议的总体目标是快速分离原代间充质干细胞，并使用创新的细胞分离设备结合基于基因表达的浓缩来丰富能够再生的亚群。该项目将由布朗大学、罗德岛医院和Endocellutions公司的研究人员合作完成。为了实现这一目标，将设计一种活细胞分子信标，以荧光方式识别能够表达脂肪基因的细胞。将使用Adi-Cellutions设备获取人类脂肪组织，并在基于基因表达的分选后检查分离株固体/液体部分中的细胞的再生特性。该建议包括两个具体目标：1)研究基于基因表达的细胞分选参数如何影响成脂细胞的高产量获取；2)建立一套能够在分离的细胞群中产生一致的再生特性的处理参数，而不依赖供体。基于基因表达的细胞浓缩是一种潜在的、范式转换的技术，极大地扩展了研究人员可以在现有的细胞表面标记方法之外进行研究的潜在靶点。更实际的是，该项目产生的方法学将为其他研究人员提供一个路线图，以获得更一致的细胞群体，用于基础科学实验和临床程序。拟议的学术和行业合作将在生物医学工程、细胞和分子生物学以及临床医学领域做出贡献。本科生、研究生和医学生将参与整个拟议项目的所有阶段，实验结果将通过布朗大学和合作医院的科学交流和教育场所分发。

PI: Darling, Eric M. Proposal Number: 1547819Cell-based biomanufacturing is highly dependent on its starting material: the cell. Using poorly defined, mixed populations of cells will result in unsatisfactory outcomes, regardless of the downstream technologies. Purification or enrichment of high-value material, like adult stem cells, is necessary for a broad range of existing and future therapeutic strategies. By better controlling the composition of the source population of cells, the performance of regenerative therapies and products is hypothesized to be more reproducible, making biomanufacturing approaches more feasible than they are today. Current enrichment techniques, which depend on the existence of unique combinations of cell surface proteins, are limited by very low cell yields and significant susceptibility to donor variability. In this work, the investigators propose to develop and evaluate new procedures that enable the production of stem cell populations with consistent regenerative characteristics, independent of donor.The overall goal of this proposal is to rapidly isolate primary, mesenchymal stem cells and enrich for subpopulations capable of regeneration using an innovative cell separation device in conjunction with gene expression-based enrichment. This project will be a collaboration among researchers at Brown University, Rhode Island Hospital, and Endocellutions, Inc. To achieve this goal, a live-cell molecular beacon will be designed that fluorescently identifies cells capable of adipogenic gene expression. Human fat tissue will be obtained using the Adi-Cellutions device, and cells within the solid/liquid fractions of the isolate will be examined for their regenerative characteristics following gene expression-based sorting. Two specific aims are included in this proposal: 1) Investigate how gene expression-based cell sorting parameters influence high-yield acquisition of adipogenic cells, and 2) Establish a set of processing parameters capable of generating consistent regenerative characteristics in isolated cell populations, independent of donor. Gene expression-based cell enrichment is a potential, paradigm-shifting technique, vastly expanding the potential targets researchers can investigate beyond current, cell surface marker approaches. More practically, the methodologies generated from this project will provide a roadmap for other investigators to obtain more consistent populations of cells for use in basic science experiments as well as clinical procedures. The proposed academic-industry collaboration will result in contributions to the fields of biomedical engineering, cell and molecular biology, and clinical medicine. Undergraduate, graduate, and medical students will participate throughout all stages of the proposed project, and experimental findings will be distributed through scientific communications and educational venues at Brown University and partnering hospitals.