Murine mesenchymal stem cells isolated by low density primary culture system

Murine mesenchymal stem cells (mMSC) and the difficult task of isolation and purification of them have been the subject of rather extensive investigation. The present study sought to isolate these cells from two different mouse strains, one outbred and the other inbred, primarily through a relatively simple but novel approach, the most important feature of which was the low density primary culture of bone marrow cells. For this purpose, mononuclear cells from either NMRI or BALB/c bone marrow were plated at about 500 cells per well of 24-well plates and incubated for 7 days. At this point, the fibroblastic clones that had emerged were pooled together and expanded through several subcultures. To investigate the mesenchymal nature, we differentiated the cells into the osteoblastic, chondrocytic and adipocytic lineages in different subcultures up to passage 10. According to the results, 1 week after culture initiation, several clones each comprising several fibroblastic cells appeared in each plate. The cells from different passages were capable of differentiating into corresponding skeletal tissues. In the present investigation, the best culture condition for maximum proliferation and also the expression of certain surface marker on isolated cells were examined. In this term the two murine strains showed some differences.