gp130 and c-Kit signalings synergize for ex vivo expansion of human primitive hemopoietic progenitor cells.

Abstract

ABSTRACTgpl3O, a signal-transducing receptor component of interleukin 6 (IL-6), associates with an IL-6 and IL-6 receptor (IL-6R) complex and transduces signals. To examine the role of gpl3O signaling in the expansion of human hemopoietic progenitor cells, we tested the effects of a recombinant soluble human and/or combination with other cytokines on purified human umbilical cord blood CD34+ cells, using methylcellulose clonal assay and suspension culture in the presence or absence of serum. A combination of sIL-6R and IL-6 (sIL-6R/IL-6), but not sIL-6R or IL-6 alone, was found to dramatically stimulate expansion of hemopoietic progenitor cells as well as CD34+ cells in the presence of stem cell factor. Significant generation of multipotential hemopoietic progenitors over a period of 3 weeks in suspension culture and efficient formation of colonies, especially multilineage and blast cell colonies, in methylcellulose assay supplemented with a combination of sIL-6R/IL-6 together with stem cell factor were observed in serum-containing and serum-free culture. Addition of antigpl3O monoclonal antibodies or anti-IL-6R monoclonal antibodies to the above cultures dose-dependently inhibited the expansion of progenitor cells in suspension culture and also completely blocked the formation of multilineage colonies in methylcellulose culture. These findings demonstrated that the significant expansion of human primitive hemopoietic progenitors could be achieved with the gpl3O and c-Kit signalings initiated by the sIL-6R/IL-6 complex in the presence of stem cell factor and suggested the possible application of this method for ex vivo expansion of CD34+ cells for bone marrow transplantation.

Synergistic effects on erythropoiesis, thrombopoiesis, and stem cell competitiveness in mice deficient in thrombopoietin and steel factor receptors

“…Supplementing in vitro primary hematopoietic cell cultures with either SF or Tpo, particularly in combination with other early-acting factors, can induce HSC division and increase the recovery of totipotent cells by preventing differentiation and/or facilitating self-renewal. 25,28,44,45 Consistent with these in vitro studies, primitive cell deficiencies have been described in mice with loss-of-function alleles in Tpo or SF pathways. Tpo Ϫ/Ϫ or c-Mpl Ϫ/Ϫ mice have reduced numbers of progenitor cells of multiple lineages, including CFU-Mk, CFU-GM, CFU-E, and CFU-granulocyte-erythrocyte-megakaryocytemacrophage (CFU-GEMM).…”

“…While some single growth factors can maintain long-term repopulating cells in culture, multiple factors are required for expansion. 25,44,45 Recent findings indicate that both Tpo signaling (via JAK2) and SF signaling are required for in vitro self-renewal of multipotent hematopoietic cells. 46 However, expansion of long-term repopulating cells can be achieved in H2K-BCL-2 transgenic bone marrow cells with a single growth factor (SF), indicating that one of the in vitro requirements is inhibition of HSC apoptosis.…”

Thrombopoietin augments ex vivo expansion of human cord blood-derived hematopoietic progenitors in combination with stem cell factor and flt3 ligand

“…28 In addition, gp130 is ubiquitously expressed on CB-derived CD34 + cells. 34 As described above, most human CB-derived CD34 + cells did not express the IL- tively than IL-6 alone. In fact, Sui et al 35 have reported that a combination of sIL-6R and IL-6 dramatically stimulated the obtained in our series of experiments.…”

Cloning and expression of human B cell-specific transcription factor BACH2 mapped to chromosome 6q15

“…Mononuclear cells (MNCs) were separated by Ficoll density gradient centrifugation after depletion of phagocytes with silica (IBL) as described elsewhere (Imai et al, 1991). CB CD34+ cells were isolated from MNCs by using Dynabeads M-450 conjugated with CD34 monoclonal Ab (mAb) and DETACHaBEAD CD34 (Dynal) as described previously (Sui et al, 1995). CB CD34+107, CD34+10+ cells were puri®ed from CB CD34+ cells by cell sorting on FACS Vantage (Becton Dickinson).…”