Department of Biomedical Engineering, University of Calgary, 2500 University Drive NW, Calgary, Alberta T2N 1N4, Canada.
Department of Chemical and Petroleum Engineering, University of Calgary, 2500 University Drive NW, Calgary, Alberta T2N 1N4, Canada.
ACS Appl Bio Mater. 2024 Sep 16;7(9):5956-5964. doi: 10.1021/acsabm.4c00513. Epub 2024 Aug 27.
Mesenchymal stromal cells (MSCs) have the potential to be used as autologous or allogenic cell therapy in several diseases due to their beneficial secretome and capacity for immunomodulation and differentiation. However, clinical trials using MSCs require a large number of cells. As an alternative to traditional culture flasks, suspension bioreactors provide a scalable platform to produce clinically relevant quantities of cells. When cultured in bioreactors, anchorage-dependent cells like MSCs require the addition of microcarriers, which provide a surface for cell attachment while in suspension. The best performing microcarriers are typically coated in animal derived proteins, which increases cellular attachment and proliferation but present issues from a regulatory perspective. To overcome this issue, a recombinant fusion protein was generated linking basic fibroblast growth factor (bFGF) to a cellulose-specific carbohydrate binding module (CBM) and used to functionalize the surface of cellulose microcarriers for the expansion of human umbilical MSCs in suspension bioreactors. The fusion protein was shown to support the growth of MSCs when used as a soluble growth factor in the absence of cellulose, readily bound to cellulose microcarriers in a dose-dependent manner, and ultimately improved the expansion of MSCs when grown in bioreactors using cellulose microcarriers. The use of CBM fusion proteins offers a simple method for the surface immobilization of growth factors to animal component-free substrates such as cellulose, which can be used alongside bioreactors to increase growth factor lifespan, decrease culture medium cost, and increase cell production in the manufacturing of therapeutic cells.
间充质基质细胞（MSCs）由于其有益的分泌组和免疫调节与分化能力，有望在多种疾病中被用作自体或同种异体细胞治疗。然而，使用 MSCs 的临床试验需要大量的细胞。作为传统培养瓶的替代方法，悬浮生物反应器为生产具有临床相关性的细胞数量提供了可扩展的平台。当在生物反应器中培养时，锚定依赖性细胞（如 MSCs）需要添加微载体，微载体为细胞附着在悬浮液中提供了表面。性能最佳的微载体通常涂有动物源性蛋白，这增加了细胞的附着和增殖，但从监管角度来看存在问题。为了解决这个问题，生成了一种将碱性成纤维细胞生长因子（bFGF）与纤维素特异性碳水化合物结合模块（CBM）连接的重组融合蛋白，并用于纤维素微载体表面的功能化，以在悬浮生物反应器中扩增人脐带 MSCs。当用作无纤维素的可溶性生长因子时，融合蛋白被证明支持 MSCs 的生长，它以剂量依赖的方式与纤维素微载体容易结合，并最终在使用纤维素微载体的生物反应器中提高了 MSCs 的扩增。CBM 融合蛋白的使用为生长因子在无动物成分的基质（如纤维素）上的表面固定提供了一种简单的方法，可与生物反应器一起使用，以延长生长因子的寿命、降低培养基成本，并增加治疗性细胞制造中的细胞产量。
