Ex vivo expansion of bone marrow mesenchymal stem cells using microcarrier beads in a stirred bioreactor

Bone marrow mesenchymal stem cells (bmMSCs) have recently gained attention as a useful resource in the fields of regenerative medicine and tissue engineering. However, the number of bmMSCs obtained from available donors is very low. Here we developed a culture strategy for in vitro expansion of bmMSCs in a 1.5 L stirred bioreactor with microcarrier beads. First, the microcarriers (Cytodex 3) were equilibrated in culture medium containing 3% fetal bovine serum (FBS) for at least 30 min prior to cell addition. After inoculation, the FBS concentration of the medium was maintained at 3% (v/v) in the first 24 h and thereafter maintained at 1% (v/v) and a developed feeding regimen was applied over 5 days. The maximum cell density of 2.6 × 10⁶ cells/mL was achieved at day 5, corresponding to a 10.4 ± 0.8 fold increases in total cell number. Among the harvested cells, 98.95% expressed CD29 and 84.48% expressed CD90, suggesting that the majority of expanded bmMSCs still retained their differentiation potential. Therefore, the developed microcarrier-based stirred bioreactor culture system is an effective method to generate significant numbers of bmMSCs for potential applications and research studies.