Bone marrow (BM) mesenchymal stromal/stem cells (MSC) are therapeutic tools in regenerative
medicine and oncology. MSC isolation is often performed starting from a separation
step based on research-grade 1.077 g/mL density gradient media (DGM). However, MSC
clinical application should require the introduction of good manufacturing practice
(GMP) reagents. We took advantage of two novel GMP DGM with densities of 1.077 and
1.073 g/mL (Ficoll-Paque PREMIUM and Ficoll-Paque PREMIUM 1.073, respectively) to
test whether these reagents could isolate MSC efficiently while simultaneously comparing
their performance.
BM samples were processed using either 1.077 or 1.073 g/mL GMP DGM. BM mononucleated
cell (MNC) fractions were analyzed for viability, immunophenotype, clonogenic potential,
ex vivo expansion and differentiation potential.
No differences were noticed in cell recovery and viability between the groups. Fluorescence-activated
cell-sorting (FACS) analyzes on freshly isolated cells indicated that the 1.073 g/mL
GMP DGM more efficiently depleted the CD45(+) fraction in comparison with 1.077 GMP
DGM. Moreover, in the 1.073 group, fibroblastic colony-forming units (CFU-F) were
1.5 times higher and the final MSC yield 1.8 times increased after four passages.
Both reagents isolated MSC with the expected phenotype; however, 1.073-isolated MSC
showed a higher expression of CD90, CD146 and GD2. Additionally, MSC from both groups
were capable of fully differentiating into bone, adipose cells and cartilage.
Both GMP DGM enriched MSC from BM samples, suggesting that these reagents would be
suitable for clinical-grade expansions. In addition, the density of 1.073 g/mL provides
a significant advantage over 1.077 g/mL GMP DGM, impacting the quantity of MSC obtained
and reducing the ex vivo expansion time for optimized cell-based clinical applications.