Microfluidic Cell Separation for Tissue Engineering and Regenerative Medicine

描述(申请人提供)：本提案的主要目标是创建微流控细胞分离系统，以分离或丰富组织工程和再生医学的关键细胞类型。在传统的组织工程中，在种植到支架上之前，必须丰富功能细胞类型。在以细胞为基础的组织修复和再生方法中，驻留在不同组织类型中的干细胞和祖细胞必须在使用之前进行分离和鉴定。用于这些应用的微流控细胞分离系统的设计和制造是基于以下观察结果。首先，在功能细胞浓缩方面，与预镀和使用细胞过滤器等最先进的方法相比，微流控技术更加系统化。其次，微流控系统可以处理少量(微升量级)的样本，使细胞能够有效地从少量捐赠者组织中分离出来。这些系统可以与体外细胞培养设备结合在一起，而且它们成本低，在临床环境中易于现场操作。第三，PI实验室最近的工作证明了微流控设备能够根据大小和亲和力分离细胞亚群。这项提案将侧重于创造微流控细胞分离技术，用于四个领域的组织工程应用：心脏组织、皮肤、胃肠组织和血管组织。拟议的工作将与组织工程专家合作进行：Milica Radisic博士(心脏)、Rebecca Carrier博士(肠道)、Virna Sales&John Mayer(血管)和Yaakov Nahmias&Martin Yarmush(皮肤/烧伤)。在为期3年的项目期间，将沿着独立的轨道追求以下目标：(1)设计和制造基于尺寸和黏附的微流控分离设备，以分离心脏和肠道组织中的细胞群体；(2)设计基于黏附的微流体分离方法，通过正选择分离内皮祖细胞和皮肤干细胞用于再生应用；以及(3)设计基于黏附的微流控分离方法，通过负选择分离心脏前体细胞和肠道干细胞。 与公共健康相关：近年来，组织工程学从一小部分捐献组织中培养出特定组织类型的切片的能力得到了极大的发展。最近的研究还展示了驻留在成人组织中的干细胞和祖细胞如何在修复组织中发挥作用。这两种方法的一个关键组成部分是，在将某些类型的细胞用于修复和再生应用之前，需要分离或收获这些类型的细胞，本提案旨在为此目的创造新的工具。

DESCRIPTION (provided by applicant): The major goal of this proposal is to create microfluidic cell separation systems to isolate or enrich key cell types for tissue engineering and regenerative medicine. In conventional tissue engineering, functional cell types must be enriched prior to seeding onto scaffolds. In cell-based approaches to tissue repair and regeneration, stem and progenitor cells resident in different tissue types must be isolated and characterized prior to their use. The design and fabrication of microfluidic cell separation systems for these applications is motivated by the following observations. First, in functional cell enrichment, microfluidic techniques are more systematic compared to state of the art methods such as pre-plating and using cell strainers. Second, microfluidic systems can handle small (microliter-order) sample volumes, enabling effective cell separation from small quantities of donor tissue. These systems can be incorporated with in-vitro cell culture equipment and furthermore, they are low-cost and easily operated on-site in clinical settings. Third, recent work in the PI's laboratory has demonstrated the ability of microfluidic devices to separate cell subpopulations based on size and affinity. This proposal will focus on the creation of microfluidic cell separation technologies for tissue engineering applications in four areas: cardiac tissue, skin, gastrointestinal tissue, and vascular tissue. The proposed work will be carried out in collaboration with experts in tissue engineering: Drs. Milica Radisic (cardiac), Rebecca Carrier (intestinal), Virna Sales & John Mayer (vascular), and Yaakov Nahmias & Martin Yarmush (skin/burns). During the 3-year project period, the following aims will be pursued along independent tracks: (1) design and fabricate size- and adhesion-based microfluidic separation devices to separate cell populations in cardiac and intestinal tissue; (2) design an adhesion-based microfluidic separation approach to isolate endothelial progenitor cells and skin stem cells by positive selection for regenerative applications; and (3) design an adhesion-based microfluidic separation approach to isolate cardiac progenitor cells and intestinal stem cells by negative selection. PUBLIC HEALTH RELEVANCE: The ability to grow sections of certain tissue types from a small sample of donor tissue, tissue engineering has grown tremendously in recent years. Recent work has also demonstrated how stem and progenitor cells resident in adult tissue can play a role in repairing tissue. A key component of either of these approaches is the requirement to isolate or harvest certain cell types prior to utilizing them for repair and regenerative applications and this proposal aims to create novel tools for this purpose.