Isolation of adipose and bone marrow mesenchymal stem cells using CD29 and CD90 modifies their capacity for osteogenic and adipogenic differentiation

Mesenchymal stem cells isolated from rats are frequently used for tissue engineering research. However, considerabledifferences have been identified between rat mesenchymal stem cells and those derived from humans, and no definedpanel of markers currently exists for the isolation of these cells. The aim of this study was to examine the effects of cellsorting for CD29+/CD90+ cells from rat adipose and bone marrow tissues on their differentiation and expression of stemcell–associated genes. Flow cytometry showed 66% and 78% CD29+/CD90+ positivity within passage 1 of adipose and bonemarrow cultures, respectively. CD29+/CD90+ cells showed a reduction in both osteogenic and adipogenic differentiationwhen compared with unsorted cells, as determined by alizarin red and Oil Red-O staining, respectively. These findings could not entirely be explained by fluorescence-activated cell sorting–induced cell injury as sort recovery was only modestlyaffected in adipose-derived cells. Maintaining cells in fluorescence-activated cell sorting buffer did not affect adipose-derivedcell viability, but a significant (p < 0.05) reduction was found in bone marrow–derived cell viability. Additionally, CD29+/ CD90+ selection was associated with a significant decrease in the expression of Lin28, Sox2, Nanog and CD73 in adiposederived cell cultures, whereas differences in stem cell–associated gene expression were not observed in sorted bone marrow–derived cell cultures. In summary, this study demonstrated that fluorescence-activated cell sorting had differential effects on adipose-derived cells and bone marrow–derived cells, and both CD29+/CD90+ cells displayed a significantly reduced capacity for osteogenic/adipogenic differentiation. In conclusion, we identify