Microfluidic-based high-efficiency cell fusion for studying nuclear reprogramming

基于微流控的高效细胞融合研究核重编程

基本信息

批准号：
7449496
负责人：
Joel Voldman
金额：
$ 22.06万
依托单位：
MASSACHUSETTS INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-09-01 至 2010-08-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): We propose to develop a new microfluidic technology to efficiently create 1,000's of fusion events between embryonic stem cells and somatic cells in order to study early events in nuclear reprogramming. Our device uses a three-step loading procedure and a special geometry to efficiently arrange cells side-by-side to create a large fraction of properly paired hybrid cells. We are developing this technology to study nuclear reprogramming, which is the process whereby a somatic cell's nuclear program is rebooted to bring that cell back into a more primitive, pluripotent state. Factors within pluripotent or totipotent cells can reprogram somatic cells, and the current approaches center on methods of transferring these factors from one cell to the other. The standard approach to reprogramming nuclear transfer, in which the somatic cell nucleus is transplanted into an unfertilized egg is tedious and limited to small numbers of cells. Cell fusion, whereby a somatic cell is fused to an embryonic stem cell, merging the cellular contents, can also be used to reprogram the somatic cell's nucleus. Cell fusion has the potential to be easier and more scalable than nuclear transfer. One significant technical limitation that prevents the widespread use of this approach for studying reprogramming is that current approaches to cell fusion use randomly paired cells. This results in a small number of desired hybrid fusions (stem cell + somatic cell) in a large background of unwanted fusions and unfused cells. In order to purify the desired fusions from this background, researchers undertake antibiotic selection steps that add days to the procedure and prevent researchers from studying early events in nuclear reprogramming. By developing a device to efficiently pair cells, we are addressing the dominant technological challenge in the creation of hybrids, and believe that our device will have significant impact for studying fusion-induced reprogramming.

描述（由申请人提供）：我们建议开发一种新的微流体技术，以有效地在胚胎干细胞和体细胞之间创建1,000次融合事件，以研究核重编程中的早期事件。我们的设备使用三步加载过程和特殊的几何形状并排排列单元格，以创建大量合适的配对杂种细胞。我们正在开发这项技术来研究核重编程，这是重新启动体细胞核计划以使该细胞重新恢复到更原始的多能状态的过程。多能或全能细胞内的因素可以重新编程体细胞，并且当前方法集中在将这些因子从一个细胞转移到另一个细胞的方法。重编程核转移的标准方法，其中体细胞核被移植到未施用的卵中是乏味的，并且仅限于少量细胞。细胞融合将体细胞融合到胚胎干细胞并融合细胞含量的融合，也可以用于重新编程体细胞的核。细胞融合具有比核转移更容易和更可扩展的潜力。一种重要的技术限制，阻止这种方法在研究重编程中广泛使用这种方法，即当前的细胞融合使用方法随机配对细胞。这会导致在大的不良融合和未散发细胞的巨大背景下进行少量所需的杂交融合（干细胞 +体细胞）。为了净化此背景中所需的融合，研究人员采取了抗生素选择步骤，从而为程序增加了天数，并阻止研究人员研究核重编程中的早期事件。通过开发有效配对细胞的设备，我们正在解决杂种创建的主要技术挑战，并认为我们的设备将对研究融合引起的重编程产生重大影响。