A versatile technological platform for the preparation of functionalized porous hydrogels detined to cell culture applications (INNOV Phase IA)

大多数用于疾病研究、药物发现/功效和毒理学分析、干细胞/器官工程的商业细胞培养材料都是二维功能化表面。然而，已知它们会促进细胞变平，导致与体内环境相比显著的细胞行为差异。为了克服这个问题，用于细胞培养的3D多孔功能化水凝胶材料已经开始出现在市场上。例如，这些材料将为阿尔茨海默病或心脏疾病的药物筛选分析提供更多生理学相关的工具。商业上可行的工艺应该产生生物相容性材料，精确控制其三维微观结构-孔径分布，体积分数，孔隙互联性，以方便细胞播种和增强质量运输（营养物质，生长因子，O2，废物等），并具有功能性的孔表面，以促进细胞粘附，存活和分化。这个过程应该是可扩展的，允许灭菌和长期储存。满足所有这些要求是一个挑战。据我们所知，市场上没有这种材料。这个i2i1a项目的主要目标是评估我们实验室开发的一项新技术的商业潜力，该技术用于设计和制备用于细胞培养应用的3D微结构、功能化多孔藻酸盐凝胶，在这里展示了将分化为神经元（用于阿尔茨海默病研究）或心肌细胞（用于心脏组织损伤愈合研究）的间充质干细胞。该工艺使用商业加工聚合物设备来制备共连续聚合物混合物，用于制备海藻酸盐水凝胶成型的多孔聚合物。霉菌的提取产生多孔海藻酸盐凝胶。我们的工艺允许控制凝胶孔隙度：(i) 40-60%孔隙分数；（ii）平均孔径为10 ~ 500微米；（iii）孔隙完全互连。具体目标为：(1)优化多孔藻酸盐凝胶的制备；(2)优化凝胶功能化；(3)评价其在细胞播种、粘附、存活和分化方面的性能；(4)制定灭菌/贮存程序；(5)评估工艺和材料的商业潜力。**********************

Most commercial cell culture materials for disease research, drug discovery/efficacy and toxicology assays,**stem cell/organ engineering, are 2D functionalized surfaces. However, they are known to promote cell flattening, resulting in significant cell behavior differences as compared to their in vivo environment. To overcome this problem, 3D porous, functionalized hydrogel materials for cell culture have started to appear on the market. Such materials will provide, for examples, more physiologically relevant tools for drug screening assays for Alzheimer disease, or cardiac pathologies. A commercially viable process should yield biocompatible materials, with precise control over their 3D microstructure - pore size distribution, volume fraction, pore interconnectivity to ease cell seeding and to enhance mass transport (nutrients, growth factors, O2, wastes, etc.), and with functionalized pore surface to promote cell adhesion, survival and differentiation. The process should be scalable, allow sterilization, and long-term storage. Meeting all of these requirements is a challenge. To the best of our knowledge, there is no such material on the market. The main objective of this I2I 1A project is to assess the commercial potential of a new technology developed in our laboratory for the design and preparation of 3D microstructured, functionalized porous alginate gels for cell culture applications, demonstrated herein with mesenchymal stem cells that will differentiate into neurons (for Alzheimer research) or cardiomyocytes (for research on healing of cardiac tissues injuries). The process uses commercial processing polymer equipment to prepare cocontinuous polymer blends, used to prepare porous polymers into which alginate hydrogels are molded. Extraction of the molds yields porous alginate gels. Our process allows control over gel porosity: (i) 40-60% pore fraction; (ii) 10 to 500 microns average pore size; (iii) fully interconnected pores. The specific objectives are (1) to optimize the preparation of porous alginate gels; (2) to optimize gel functionalization; (3) to evaluate their performance for cell seeding, adhesion, survival and differentiation; (4) to develop sterilization/storage procedures; (5) to assess the commercial potential of the process and materials.**********************