Development of cell-enhanced chitosan scaffolds to overcome long gaps after peripheral nerve injury

Abstract

of well-defined serum-free and xeno-free media formulations which represent important milestones towards the production of MSC for cellular therapies. In this work, a microcarrier-based suspension culture was explored for the scale-up of MSC expansion in xeno-free medium using synthetic peptide acrylate surface beads. Cells were maintained on Corning Synthemax II polystyrene and CELLstart -coated Solohill plastic microcarriers (dos Santos et al, 2011) for 14 days in xeno-free medium. Bone marrow (BM) derivedand adipose tissue (AT) derived-MSC were seeded at 50,000 cells/mL with 4.5 cm2/mL of microcarriers in spinner flasks (80 mL volume). To maximize cell seeding, the adhesion step was performed in 50% final volume during the first 24 hours. The efficiency of initial cell adhesion to microcarriers was similar for both cell types; slightly better cell adhesion was observed on Synthemax II compared to CELLstart-coated microcarriers: 48% and 43% for AT MSC and 42% to 37% for BM MSC. The homogenous cell distribution on the first days of culture resulted in a higher expansion rate with exponential growth from day 4 to day 6 for AT MSC and day 4 to day 9 for BM MSC. The longer growth phase observed for BM MSC resulted in higher cell densities of 350,000 cells/mL compared to 250,000 cell/mL for AT MSC cultures. Expanded cells maintained their characteristic immunophenotype and multilineage differentiation potential. Ongoing work includes the translation of this culture system to a fully controlled stirred-tank bioreactor (1 L) and the study of the impact of different culture parameters namely feeding regime and dissolved oxygen concentration on cell productivity. This scalable xeno-free, microcarrierbased culture platform is anticipated to enable cost effective and robust production of MSC meeting the needs of the allogeneic “off-the-shelf” MSC therapy sector.