NSF/FDA Scholar In Residence: 3D Cell Adhesion Assay for Cellularized Scaffold Characterization and Enhancement

摘要：Fisher，JohnProposal编号：16410873D打印(3DP)通过快速制造定制支架来增加组织工程(TE)的应用，其中精确的架构可以模仿天然组织。涉及多能干细胞、3DP结构和细胞外基质(ECM)材料的TE应用已变得越来越普遍。为了将这些技术转化为治疗学，有必要了解细胞在3D环境中的行为。FDA与马里兰大学(UMD)正在进行的一项合作导致了一种细胞黏附离心法(CAC)的开发和表征，以加强从骨髓中分离和分化间充质干细胞(MSCs)。这一建议将增强已建立的用于3D构造的检测的实用性。它还将提供对细胞-支架相互作用的三个方面的有用洞察，这将为标准化标准提供信息。这项工作将与马里兰大学和FDA的研究人员、教职员工和学生合作。CAC分析的开发适应3D支架设计，通过提供一种机制来表征和纯化3D结构中的细胞，有可能将TE疗法的平移距离降至最低。该细胞捕获设备将稳定3D支架，并允许基于整合素-配体结合来表征非黏附细胞。本研究的主要目的是：1)通过调整CAC方法，研究种植在3D支架上的骨髓间充质干细胞的行为和纯化，其中将评估离心力、孵育时间和3D构建物结构的影响：2)通过检测ECM添加剂对3DP支架黏附和分化能力的多向分化潜能，研究MSCs与3D ECM-聚合物支架的相互作用；以及3)利用CAC分析方法研究收集的非黏附细胞的捕获和鉴定。这将使FDA能够进一步开发CAC试验，并评估其作为一种表征基于细胞成分3D结构的疗法的方法的实用性。

ABSTRACTPI: Fisher, JohnProposal Number: 16410873D Printing (3DP) is increasing tissue engineering (TE) applications through rapid manufacture of custom scaffolds wherein precision architectures can mimic native tissue. TE applications involving multipotent stem cells, 3DP constructs, and extracellular matrix (ECM) materials have become increasingly prevalent. In order to translate these technologies as therapeutics it is necessary to understand cell behavior within the 3D environment. An ongoing FDA-University of Maryland (UMD) collaboration has led to the development and characterization of a cell adhesion centrifugation (CAC) assay to enhance isolation and differentiation of mesenchymal stem cells (MSCs) from bone marrow. This proposal will enhance the utility of the established assay for use with 3D constructs. It will also provide useful insight into cell-scaffold interaction in three dimensions that will inform standardization criteria. The work will partner researchers, faculty and students at the University of Maryland and the FDA.The development of the CAC assay to accommodate 3D scaffold designs has the potential to minimize the translational distance for TE therapies by providing a mechanism to characterize and purify cells within 3D constructs. The cellular capture device will stabilize 3D scaffolds and allow characterization of non-adherent cells based on integrin-ligand binding. The key aims are: 1) investigate MSC behavior and purification from bone marrow seeded onto 3D constructs through adaptation of the CAC assay wherein the impact of centrifugation force, incubation time, and 3D construct architecture will be evaluated: 2) investigate the interaction of MSCs with 3D ECM-polymer scaffolds through examination of multi-lineage differentiation potential enabled by ECM additives to 3DP scaffold adhesion and differentiation capacity: and 3)investigate the capture and characterization of non-adhered cells collected using the CAC assays. This will allow the FDA to further develop the CAC assay and to evaluate its utility as a method to characterize the cellular component 3D construct based therapies.