Isolation of BM mesenchymal stem cells by plastic adhesion or negative selection: phenotype, proliferation kinetics and differentiation potential

Abstract

Background: BM mesenchymal stem cells (MSC) have the capacity for renewal and the potential to differentiate into multiple tissues. In this study, we compared different enrichment methods to obtain MSC from BM.

Methods: Three different methods were compared with a view to obtaining MSC more rapidly from BM: negative selection (RosetteSep and MACS) and plastic adhesion. The three cell fractions were grown in complete alpha-minimum essential medium in order to evaluate their proliferative capacity, their phenotype during culture and their potential to differentiate into adipocytes, osteocytes and chondrocytes. Identification of MSC was performed by immunofluorescence with putative mesenchymal markers SH2 and SH3 but also with hematopoietic markers.

Results: After negative selection, only 1+/-0.2% and 2.9+/-0.8% of cells were recovered from BM with the RosetteSep and MACS methods, respectively. However, negative depletion permitted a homogeneous population of MSC, with more than 90% SH2+ and SH3+ cells, to be obtained rapidly and in large quantities after 10 days of culture. Similar homogeneity was observed after three passages if the plastic adhesion was used as selection method and after an average of 25-30 days of culture. Different levels of MSC maturity were also suggested by the variable level expression of Stro-1.

Discussion: Depleting selection by RosetteSep may represent an easy method of obtaining MSC rapidly from BM with the aim of potential therapeutic use.