CD34-NEGATIVE HEMATOPOIETIC STEM-CELLS ISOLATED FROM HUMAN PERIPHERAL-BLOOD CELLS AS ULTIMATE PRECURSORS OF HEMATOPOIETIC PROGENITORS

Background: The earliest hematopoietic stem cell is still not vet iden tified. From experiments in dogs there is evidence that the earliest h ematopoietic stem cell is a CD34(-) fibroblast-like cell which can be isolated from bone marrow stromal cultures, In this set of experiments we wanted to identify such an early CD34- fibroblast-like cell popula tion from human peripheral blood. Material and Methods: Peripheral blo od mononuclear cells (PBMNC) were isolated from healthy volunteer dono rs and incubated with interleukin-6(IL-6)-containing medium in tissue culture flasks. After establishing an almost confluent adherent layer of fibroblast-like cells which attach to the plastic surface, the cell s were stimulated with various growth factors. The emerging nonadheren t cells as well as the adherent cells were applied to a colony-forming unit (CFU) assay in semisolid agar and to long-term culture-initiatin g cell (LTC-IC) assay. Results: CD34- adherently growing cells could b e isolated from human peripheral blood by IL-6-mediated plastic adhere nce. These cells grow as fibroblast-like cells in culture and differen tiation can be induced by stem cell factor (SCF), while IL-6 induces p roliferation of these cells. While differentiating, CD34- fibroblast-l ike cells lose the ability to row adherently and start to express CD34 and eventually HLA-DR, The differentiating CD34+ cells are also capab le of producing CFU and LTC-IC. Conclusion: This set of experiments pr ovides evidence that the earliest identifiable hematopoietic stem cell is a CD34+ adherently growing fibroblast-like cell which is part of t he marrow microenvironment in vivo and can differentiate into hematopo ietic progenitors upon appropriate growth factor signaling. This kind of early stem cell can also be isolated from PBMNC.