Dynamic Culture Systems with Tissue-specific Microcarriers for Human Adipose-derived Stem Cell Expansion and Delivery

脂肪组织（脂肪）是成体干细胞的丰富来源，称为脂肪源性干细胞（ASCs），在组织工程和再生医学中具有广泛的应用前景。ASC可以被刺激增殖并分化成更专门的细胞类型，包括成熟的脂肪、骨、肌肉和软骨细胞以及其他谱系。然而，细胞培养环境显著影响ASC反应。当使用传统的细胞培养方法在组织培养聚苯乙烯上扩增时，ASC增殖速率下降，并且细胞迅速失去其多谱系分化能力。由于临床应用可能需要超过10亿个细胞的纯化干细胞群，因此非常需要新的具有成本效益的细胞培养技术，其能够从小组织活检中获得具有保留的干细胞潜力的大的、良好表征的ASC群。该研究项目的重点是开发用于人类ASC扩增和多谱系（骨，软骨，脂肪）分化的生物反应器策略。为了紧密复制ASC的天然细胞外环境的组成，将使用电喷雾建立用于制造源自人脂肪中发现的天然细胞外基质（ECM）蛋白的组织特异性微载体的方法。在这些微载体上的ASC增殖和分化的动态培养系统将被开发和优化。将系统地探索氧张力和与成熟细胞群共培养的影响，以调节细胞反应。细胞和分子生物学表征工具，包括基因和蛋白质表达的分析，将用于深入了解细胞培养微环境对ASCs的影响。该计划将为HQP在干细胞，生物材料和组织工程等多学科领域提供宝贵的培训机会，学员将发展整合工程和生物学领域的广泛技能。 总的来说，这项工作将推进基于ASC的治疗在重建、骨科、心血管和神经外科等应用中的临床转化。

Adipose tissue (fat) is a rich source of adult stem cells, termed adipose-derived stem cells (ASCs), which hold great promise for a range of applications in tissue engineering and regenerative medicine. ASCs can be stimulated to proliferate and differentiate into more specialized cell types, including mature fat, bone, muscle, and cartilage cells, amongst other lineages. However, the cell culture environment dramatically impacts the ASC response. ASC proliferation rates decline and the cells rapidly lose their multilineage differentiation capacity when expanded using traditional cell culture methods on tissue culture polystyrene. As clinical applications may require purified stem cell populations in excess of 1 billion cells, there is a significant need for new cost-effective cell culture technologies that enable the procurement of large, well-characterized ASC populations, with preserved stem cell potential, from small tissue biopsies. This research program focuses on the development of bioreactor strategies for human ASC expansion and multilineage (bone, cartilage, fat) differentiation. To closely replicate the composition of the native extracellular environment of the ASCs, methods for the fabrication of tissue-specific microcarriers derived from the natural extracellular matrix (ECM) proteins that are found in human fat will be established using electro-spraying. Dynamic culture systems for ASC proliferation and differentiation on these microcarriers will be developed and optimized. The effects of oxygen tension and co-culturing with mature cell populations will be systematically explored to tune the cell response. Cell and molecular biology characterization tools, including analyses of gene and protein expression, will be employed to develop an in-depth understanding of the impact of the cell culture microenvironment on the ASCs. This program will provide invaluable training opportunities for HQP in the multidisciplinary areas of stem cells, biomaterials, and tissue engineering, and the trainees will develop extensive skills that integrate the fields of engineering and biology. Overall, this work will advance the clinical translation of ASC-based therapies in applications including reconstructive, orthopedic, cardiovascular, and neurological surgery.