Serum-free culture of enriched murine haemopoietic stem cells. II: Effects of growth factors and haemin on development.

Abstract

A serum-free culture system was used to determine the effects of growth factors on the clonogenic development of a population of cells highly enriched for multipotential day 12 spleen colony forming cells (CFU-S) (FACS-BM). Under these conditions, interleukin-3 (IL-3) was found to be primarily a proliferative stimulus, the progenitor cells developing in the clonal assay systems produced colonies of morphologically undifferentiated cells for up to 20 days. No such induction of proliferation without maturation was observed with other growth factors (eg. granulocyte-macrophage colony stimulating factor (GM-CSF)). However, combinations of IL-3 plus secondary growth factors such as GM-CSF, macrophage colony stimulating factor (M-CSF), granulocyte colony-stimulating factor (G-CSF) or interleukin-1 (IL-1) led to the formation of colonies containing mature haemopoietic cells of the granulocytic, megakaryocytic or monocytic lineages. In contrast, erythroid development did not occur unless the protoporphyrin, haemin, was added to the cultures. Under these conditions mature erythroid cells were produced in cultures containing either IL-3 or GM-CSF (with or without erythropoietin (epo)). In replating experiments it was determined that the FACS-BM cells were able to generate large numbers of clonogenic cells for up to 30-40 free cultures. Such cultures, therefore, may be useful for investigating the biological and basis of the generation of clonogenic cells and of haemopoietic cell differentiation and development in response to growth factors.