Cell Graft Engineering Using Microfluids

描述（由申请人提供）：本研究的目的是证明能够从冷冻保存的细胞悬浮液中去除二甲亚砜的微流控装置的原理证明。造血细胞移植（包括干细胞和成熟细胞）已经成为广泛的血液病的治疗标准。传统的干细胞冷冻保存方法使用二甲基亚砜（DMSO）作为冷冻保护剂，移植细胞通常在患者床边解冻并直接输注。DMSO输注到人体内所造成的临床毒性是有充分记录的。使用离心或自动细胞洗涤器去除DMSO通常会导致25-30%的细胞损失。这里存在一个难题：直接输注冷冻保存的造血干细胞会导致明显的不良反应，但洗涤细胞以去除DMSO会导致严重的细胞损失，进而对移植结果产生不利影响。我们提出了一种处理解冻后细胞悬浮液的新范例：使用微流体装置。我们假设微流体装置可以用来有效地洗涤这种悬浮液，同时最大限度地减少细胞损失。液体在微尺度装置中的流动具有低速、层流的特点。该装置的横截面积足够小，长度尺度可以接近血细胞的直径（~10微米）。微流体流动的层流性质将允许基于扩散的萃取，而无需使用膜来分离流体流。因此，这些特性，加上制造大规模并行微流体系统的可能性，为评估临床使用浓度和体积的细胞处理提供了一个重要的平台。这个项目也代表了一个重要的机会，将我们对干细胞生物学和微流体的理解转化为可以直接有益于人类健康的应用。减少加工过程中的细胞损失也将使更多的人获得基于干细胞的疗法（特别是脐带血干细胞）。

DESCRIPTION (provided by applicant): The objective of this investigation is to demonstrate proof-of-principal for a microfluidic device capable of removing dimethylsulfoxide from a cryopreserved cell suspension. Hematopoietic cells grafts (both stem cells and mature cells) have become the standard of care for a wide range of hematologic diseases. Traditional methods of stem cell cryopreservation use dimethylsulfoxide (DMSO) as a cryoprotective agent and cell grafts are routinely thawed at the patient bedside and infused directly. The clinical toxicity resulting from the infusion of DMSO into humans is well documented. Removal of DMSO using centrifugation or automated cell washers typically results in a 25-30% cell loss. Therein lies the conundrum: direct infusion of cryopreserved hematopoietic stem cells results in significant adverse reactions but washing cells to remove DMSO results in significant cell losses that, in turn, adversely affect transplant outcome. We propose a new paradigm for processing of post-thaw cell suspensions: the use of microfluidic devices. We hypothesize that microfluidic devices can be used to wash such suspensions effectively while minimizing cell losses. Liquid flows through microscale devices are characterized by low velocity, laminar flow. The cross sectional areas of the devices are small enough that the length scale can approach the diameter of blood cells (~10 micron). The laminar nature of microfluidic flow will permit diffusion-based extraction without the use of a membrane to separate fluid streams. These characteristics, combined with the possibility of fabricating massively- parallel microfluidic systems, therefore, provide an important platform from which to evaluate processing of cells in concentrations and volumes used clinically. This project also represents an important opportunity to translate our understanding of both stem cell biology and microfludics into an application that can directly benefit human health. Reducing cell losses during processing will also increase the access of stem cell based therapies (in particular cord blood stem cells) to more people.