title: Human CD34(+) bone marrow cells regulate stromal production of interleukin-6 and granulocyte colony-stimulating factor and increase the colony-stimulating activity of stroma.
abstract: Cytokines produced by stromal cells induce the proliferation and differentiation of hematopoietic cells in the marrow microenvironment. We hypothesized that cross-talk between hematopoietic cells at different stages of differentiation and stromal cells influences stromal cytokine production and is responsible for maintaining steady-state hematopoiesis and responding to stress situations. We show that coculture of primitive CD34(+) cells in contact with or separated by a transwell membrane from irradiated human bone marrow stromal layers induces a fourfold to fivefold increase in interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF) levels in the stromal supernatant (SN) during the first week. Levels of both cytokines decreased to baseline after coculture of CD34(+) cells for 3 to 5 weeks. Coculture of more mature CD15(+)/CD14(-) myeloid precursors induced only a transient 1.5- to 2-fold increase in IL-6 and G-CSF at 48 hours. Neither CD34(+) nor CD15(+)/CD14(-) cells produced IL-6, G-CSF, IL-1beta, or tumor necrosis factor alpha. When CD34(+) cells were cultured in methylcellulose medium supplemented with cytokines at concentrations found in stromal SN or supplemented with stromal SN, a fourfold to fivefold increase in colony formation was seen over cultures supplemented with erythropoietin (EPO) only. When cultures were supplemented with the increased concentrations of IL-6 and G-CSF detected in cocultures of stroma and CD34(+) cells or when CD34(+) cells were cocultured in methylcellulose medium in a transwell above a stromal layer, a further increase in the number and size of colonies was seen. The colony-forming unit-granulocyte-macrophage-stimulating activity of stromal SN was neutralized by antibodies against G-CSF or IL-6. These studies indicate that primitive CD34(+) progenitors provide a soluble positive feedback signal to induce cytokine production by stromal cells and that the observed increase in cytokine levels is biologically relevant.
authors.affiliation: Departments of Medicine and Pediatrics, University of Minnesota Medical School, Minneapolis, MN, USA.
authors.forename: P
authors.initials: P
authors.lastname: Gupta
authors.forename: B R
authors.initials: BR
authors.lastname: Blazar
authors.forename: K
authors.initials: K
authors.lastname: Gupta
authors.forename: C M
authors.initials: CM
authors.lastname: Verfaillie
mesh_terms: D018952:Antigens, CD34 / Q000032:analysis; D001854:Bone Marrow Cells / Q000378:metabolism* / Q000528:radiation effects; D002452:Cell Count; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D004921:Erythropoietin / Q000494:pharmacology; D005246:Feedback; D005786:Gene Expression Regulation / Q000187:drug effects; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis* / Q000235:genetics; D006410:Hematopoiesis / Q000502:physiology*; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D018952:Antigens, CD34; D016207:Cytokines; D015850:Interleukin-6; D004921:Erythropoietin; D016179:Granulocyte Colony-Stimulating Factor