Haemopoietic colony-forming cells from peripheral blood stem cell harvests: cytokine requirements and lineage...

We have measured the in vitro growth requirements of progenitor cells released into the blood of cancer patients following administration of chemotherapy and cytokines. In order to distinguish the direct effects of cytokines on progenitors from those activating accessory cells, we have compared clonogenic growth before and after CD34-positive selection of progenitors, in serum-free conditions. CD34 selection had little effect on the cytokine requirements of erythroid colony-forming cells and single cytokines, particularly interleukin-3, could support considerable colony growth in both mononuclear and CD34+ cell suspensions. Optimal erythroid colony growth, however, usually required the addition of a combination of stem cell factor and interleukin-3, in addition to erythropoietin, which was always required. Maximal numbers of granulocyte-monocyte progenitors in mononuclear cell cultures, could be achieved with a mixture of stem cell factor, interleukin-3 and granulocyte-monocyte colony stimulating factor. However, after CD34 selection, full myeloid colony growth was only achieved when granulocyte colony stimulating factor was added to the above mixture. This presumably reflects loss of accessory cells, during CD34 selection, which produce this cytokine. When transplanted after 8 d of culture, 16/22 myeloid colonies from erythropoietin-free cultures of peripheral blood stem cell harvests, could generate secondary erythroid colonies implying that these progenitors are multipotential. However, surface marker analysis of individual erythroid colonies revealed only the occasional presence of granulocytes and monocytes. These data demonstrate that cytokine mixtures are required for optimal colony growth, particularly after CD34 selection, and that most mobilized, blood clonogenic cells are multipotential.