Isolation murine mesenchymal stem cells by positive selection
- First Online:
- 402 Downloads
Isolation and purification of mesenchymal stem cells (MSCs) from mouse via plastic adherent cultures is arduous because of the unwanted growth of hematopoietic cells and non-MSCs. In this work, homogenous populations of CD34+ MSCs from mouse bone marrow were isolated via positive selection. For this purpose, C57Bl/6 mice were killed and bone marrow cells were aspirated before incubation with magnetic bead conjugated to anti-CD34 antibody. A sample of positively selected CD34+ cells were prepared for flow cytometry to examine the expression of CD34 antigen and others were subcultured in a 25-cm2 culture flask. To investigate the mesenchymal nature, the plastic adherent cultivated cells were induced to differentiate along osteoblastic and adipogenic lineages. Furthermore, the expression of some surface markers was investigated by flow cytometry. According to the result, purified populations of fibroblast-like CD34+ cells were achieved in the first passage (1 wk after culture initiation). The cells expressed CD34, CD44, Sca-1, and Vcam-1 antigens (markers) but not CD11b and CD45. They were capable of differentiating into osteocytes and adipocytes. This study indicated that our protocol can result in the efficient isolation of homogenous populations of MSCs from C57BL/6 mouse bone marrow. We have shown that murine bone marrow-derived CD34+ cells with plastic adherent properties and capability of differentiating into skeletal lineages in vitro are MSCs.
KeywordsPositive selection Mesenchymal stem cells Murine bone marrow
- Horwitz EM, Gordon PL, Koo WK, Marx JC, Neel MD, McNall RY, Muul L, Hofmann T. Isolated allogeneic bone marrow-derived mesenchymal cells engraft and stimulate growth in children with osteogenesis imperfecta: Implications for cell therapy of bone. Proc Natl Acad Sci USA. 2002; 99:8932–8937.PubMedCrossRefGoogle Scholar
- Koc ON, Gerson SL, Cooper BW, Dyhouse SM, Haynesworth SE, Caplan AI, Lazarus HM. Rapid hematopoietic recovery after confusion of autologous-blood stem cells and culture-expanded marrow mesenchymal stem cells in advanced breast cancer patients receiving high-dose chemotherapy. J Clin Oncol. 2000; 18:307–316.PubMedGoogle Scholar