Point of Care Adult Stem Cell Isolation

描述(由申请人提供)：在美国，每年大约进行500,000例骨移植手术(Boden，2002)。自体髂骨移植仍然是黄金标准，因为它提供了骨形成的三个基本要素：骨祖细胞、骨传导基质和骨诱导分子。然而，取髂骨与相当数量的并发症有关，而且通常提供的移植物数量不足(Gupta 2001)。为了解决这些局限性，人们已经做了很多工作来改进替代移植物，包括更好的骨传导基质和外源性的骨诱导剂，如骨形态发生蛋白(BMP)(Sykaras和Opperman 2003)。相比之下，只有几种基本的护理设备来分离成骨前体细胞，成骨前体细胞是促进骨愈合的第三个也是真正必不可少的元素。一种可以在护理点从骨髓抽吸物(BMA)中分离干细胞的设备将具有显著的临床好处。在这个第一阶段的提案中，我们将为人类骨髓中被称为间充质干细胞(MSC)的多潜能细胞亚群识别特定的结合肽，这种细胞已被证明促进骨和软组织的愈合(Caplan和Bruder 2001)。界面生物材料(IFBM)将通过MSC结合序列和AffinEnergy的聚苯乙烯结合序列的化学交联组合来合成。然后将测试这些IFBM从MSC悬液、混合细胞群悬液和人骨髓抽吸物中结合和保留MSC的能力。最后，我们将开发一种结合的MSC的化学释放系统，并验证其释放后的持续活性和多潜能。这些研究在成功完成后，将验证IFBM技术在开发护理点干细胞分离设备方面的实用性。虽然超出了这项建议的范围，但我们设想增加结合成骨生长因子的IFBM，成骨生长因子也存在于骨髓中(Barnes，Kolen uik等人)。1999)，到第二代骨髓过滤设备。这种第二代设备被设想用于选择和浓缩成骨诱导分子以及成骨细胞的功能群体。最后，作为一种最有效的及时护理设备，从提取骨髓抽吸液到重新植入过滤后的成骨细胞和生长因子的整个过程应该尽可能快(理想情况下，不到1小时)。

DESCRIPTION (provided by applicant): Approximately 500,000 bone-grafting procedures are performed each year in the United States (Boden, 2002). Autologous iliac crest bone graft continues to be the gold standard because it provides the three essential elements for bone formation: osteoprogenitor cells, an osteoconductive matrix, and osteoinductive molecules. Iliac crest harvest is however associated with a significant number of complications and often provides an inadequate volume of graft (Gupta 2001). To address these limitations, much work has been done to improve alternative grafts with better osteoconductive matrices and exogenous delivery of osteoinductive agents such as the bone morphogenic proteins (BMP's) (Sykaras and Opperman 2003). In contrast, there are only a few basic point-of-care devices to isolate osteogenic progenitor cells, which are the third and truly essential element to foster bone healing. A device that could isolate stem cells from a bone marrow aspirate (BMA) at point of care would be of significant clinical benefit. In this phase I proposal, we will identify specific binding peptides for the multipotent subpopulation of cells in human bone marrow known as mesenchymal stem cells (MSC's), which have been shown to promote healing of bone and soft tissue (Caplan and Bruder 2001). Interfacial biomaterials (IFBM's) will be synthesized by chemically crosslinking combinations of MSC binding sequences with Affinergy's polystyrene binding sequences. These IFBM's will then be tested for their ability to bind and retain MSC's from suspensions of MSC's, suspensions of mixed cell populations, and human bone marrow aspirates. Finally, we will develop a chemical release system for bound MSC's and verify their continued viability and multipotentiality after release. These studies, upon successful completion, will validate the utility of IFBM technology in the development of a point of care stem cell isolation device. Although beyond the scope of this proposal, we envision the addition of IFBM's that bind osteogenic growth factors, which are also present in bone marrow (Barnes, Kostenuik et al. 1999), to a second generation bone marrow filtration device. This second generation device is envisioned to select and concentrate osteoinductive molecules as well as a functional population of osteoprogenitor cells. Finally, to be most effective as a timely point of care device, the entire process from drawing a marrow aspirate to re-implantation of the filtered osteogenic cells and growth factors should be as quick as possible (ideally, less than 1 hour).