In vitro initial expansion of mesenchymal stem cells is influenced by the culture parameters used in the isolation process

Abstract

In the last years, there were many studies based on the use of human bone marrow mesenchymal stem cells (hMSCs) in cell therapy and tissue engineering. Although hMSCs can be easily obtained and expanded in culture, a large number of cells are often needed. The expansion of hMSCs depends on the culture conditions, such as media, cell density or culture flasks. Moreover, growth factors are often added to improve cell proliferation. In this study, we compared the effect of two culture media (DMEM and α-MEM), two culture flasks (75 or 25 cm²) and two different mononuclear cell seeding densities (1×10⁴ or 5×10⁴ MNC/cm²) on the isolation of hMSCs from bone marrow samples and analyzed if the isolation conditions affected the expansion of these cells in the first two passages. Experiments were performed without the addition of exogenous growth factors. Our results showed that α-MEM is the optimal culture medium for both, isolation and expansion of mesenchymal stem cells. Moreover, the cell seeding density of 50,000 MNC/cm² in 25 cm² culture flasks seems to be the best condition for the isolation step.