PERIPHERAL-BLOOD SELL HARVESTS YIELD PRIMITIVE MULTILINEAGE PROGENITOR CELLS IN THE CD34+/33- FRACTION

The presence of primitive hematopoietic progenitor cells or stem cells in peripheral blood (PBSC's) harvests was investigated in a single cell culturing assay and compared with the results obtained in aspirates of normal bone marrow. Based on the presence of CD33, rather differentiated progenitor cells (CD34+/33+) were distinguished from more primitive cells (CD34+/33-). The growth potential of CD34+/33+ and CD34+/33- cells have been studied. Single cell sorting was performed from peripheral blood harvests, obtained from three patients with multiple myeloma during hematopoietic recovery after treatment with high dose cyclophosphamide and rhu-GM-CSF. To test the effect of ''stem cell recruiting factors'' the cells were sorted in 96-well plates, prefilled with liquid medium both in the presence of IL-3 + G-CSF + GM-CSF + Epo and the same growth factors supplemented with SCF + IL-6. Addition of SCF and IL-6 to the culturing medium enhanced the plating efficiency of CD34+/33- cells considerably more than that of CD34+/33+ cells. This was observed in harvests of peripheral blood as well as in aspirates of normal bone marrow. The differences between CD34+/33+ and CD34+/33- were even more pronounced when only the large colonies (>500 cells/well) were taken into consideration. Assuming that IL-6 and SCF are ''stem cell recuiting factors'' the CD34+/33- fraction contains more clonogenic cells than the CD34+/33+ fraction. In all three patients the first CD34+ cells appearing in the peripheral blood (PB) after cytoreductive treatment were predominantly CD34+/33- (>80%). At later stages when the leukocyte counts had reached higher values the CD34+/33+ cells predominated. We conclude that peripheral blood stem cell harvests contain primitive multipotential clonogenic progenitor cells (of the CD34+/33- phenotype).