Neuronal, mesenchymal and hematopoietic cell derived from CD34− lin− cell from adult bone marrow

Recent studies have demonstrated that fibroblast like CD34− cells derived from the hematopoietic system have the potential to give rise to cells of diverse tissues types. To investigate the plasticity of the bone marrow derived cells, we performed experiments examining the effects of series of different, targeted culture conditions. CD34−/Lin− cells isolated from human adult bone marrow were used in cultures specific for hematopoietic stem cell (HSC), mesenchymal stem cell (MSC) and neural stem cell (NSC) differentiation. After 3 to 4 weeks, the cultures were harvested and evaluated by flow cytometry. MSC cultures, which consisted of MSC growth medium and growth factors, resulted in the development CD45−Cytokeratin+ (epithelial) and CD45−Vimentin+ (mesenchymal) cells. When vascular endothelial growth factor was added to the culture medium, cells expressing von Willebrands Factor were detected. Non-adherent cells expressing hematopoietic phenotype CD45+CD34+CD38+ were isolated from hematopoietic growth culture medium supplemented with hematopoietic growth factors. Cells cultured in neuronal growth medium supplemented with neuronal growth factors were observed to express neurofilament. The morphology of the cells in these three growth specific culture media was markedly different. To extend this work and assess the in vivo attributes of in vitro-derived cells, bone marrow CD34−Lin− cells cultured for 3 weeks under conditions for neuronal cell differentiation were injected via the tail vein into irradiated NOD/SCID mice. After 18 days, mice were sacrificed and cells from the brain, muscle, thymus, spleen, and bone marrow were collected and stained with antibodies specific to human CD45 and MHC class-1. Human cells (CD45+ or MHC class 1+) were only identified in the brain of transplanted mice. To confirm the FACS observation, PCR specific for human globin was performed and has demonstrated the presence of human cells only in the brain of the transplanted mice. These data indicate that HSC, MSC and NSC may originate from a common precursor or pool of precursors in the CD34−/Lin− fraction of human bone marrow. These precursor cells may have strong requirements for defined growth environments to enable their differentiation into particular lineages. The data also suggest that the homing of the injected cells may be organ specific. Immunohistochemistry should further reveal the nature of these cells.