No abstract

SummarySince tumor necrosis factor (TNF)-a, interferon (IFN)-% and transforming growth factor (TGF)-[3 have all been shown to be specific inhibitors of early human hematopoiesis, we wanted to investigate the interactions of these three cytokines on very primitive human adult bone marrow CD34 § +CD38-hematopoietic progenitor cells, using a pre-colony-forming cell (pre-CFC) assay, which detects the effects of these cytokines on the initial phases of the differentiation of these primitive progenitors, which are unresponsive to interleukin (IL) 3 alone. Surprisingly, TNF-cx was a very potent stimulator of the proliferation of CD34 + +CD38-cells and was the most potent synergistic factor for the IL-3-induced proliferation of these cells of all cytokines tested (IL-1, IL-6, granulocyte colony-stimulating factor, kit ligand). TNF-o~ was the only cytokine that, as a single added factor, induced substantial proliferation in CD34 + +CD38-cells in the presence of IL-3, except for kit ligand, which induced very limited proliferation. TNF-c~, moreover, induced a high degree of resistance to the inhibitory effects of TGF-13 in a dose-dependent way. The inhibitory effects oflFN-% however, were not affected by the presence of TNF-ot. We hypothesize that in situations of hematopoietic stress, TNF-oL may abrogate the inhibitory effect of ambient TGF-[3 in the bone marrow microenvironment to allow primitive stem cells to proliferate and differentiate in response to an increased demand for mature blood cells.

To investigate the T-lymphopoietic capacity of human adult bone marrow (ABM) hematopoietic progenitor cells, CD34+Lin−, CD34+CD38+, and CD34++CD38− cells were cultured in a severe combined immunodeficient (SCID) mouse fetal thymic organ culture (FTOC). Direct seeding of these progenitors resulted in a moderate to severe cell loss, particularly for the CD34++CD38− cell fraction, and T cells could only be generated from the CD34+Lin− fraction. Preincubation for 36 hours with interleukin-3 (IL-3) and stem cell factor (SCF) led to an improved cell survival and proliferation, although T-cell development was seen only in the CD34+Lin− fraction. Addition of tumor necrosis factor (TNF)- to IL-3 + SCF-supplemented preincubation medium resulted in optimal cell survival, cell proliferation. and T-cell generation of all 3 cell fractions. The TNF- effect resulted in an up-regulation of CD127 (ie, the IL-7 receptor -chain) in a small subset of the CD34+ cells. No evidence could be generated to support the possibility that TNF- inhibits a cell population that suppresses T-cell differentiation. A quantitatively different T-cell generation potency was still seen between the 3 subpopulations: CD34+Lin− (100% success rate) > CD34+CD38+ (66%) > CD34++CD38− (25%). These data contrast with our previous findings using fetal liver and cord blood progenitors, which readily differentiate into T-lymphocytes in FTOC, even without prestimulation with cytokines. Our results demonstrate that adult CD34++CD38− cells, known to contain hematopoietic stem cells, can differentiate into T-lymphocytes and that a significant difference exists in T-lymphopoietic activity of stem cells derived from ontogenetically different sources.

Important functional differences exist between primitive CD34 ++ CD38 − hematopoietic progenitor cells derived from human fetal liver (FL) and adult bone marrow (ABM). FL progenitors are known to have higher proliferative capacities and lower cytokine requirements than their ABM counterparts. In this study, we isolated FL and ABM CD34 ++ CD38 − cells and used a two-stage culture system to investigate the effects of transforming growth factor-␤ (TGF-␤) and blocking anti-TGF-␤ antibodies (anti-TGF-␤) on these cells. First, we demonstrate that FL progenitors are significantly less sensitive to the inhibitory effects of TGF-␤ than ABM cells. Second, whereas ABM cells are significantly stimulated by anti-TGF-␤, only very limited effects are seen on FL cells. Third, we show that the effect of anti-TGF-␤ is mainly situated at the level of the initial cell cycles of very primitive progenitor cells with a high proliferation potential. Fourth, we demonstrate that blocking the effects of endogenous TGF-␤ reduces the growth factor requirements of ABM cells in order to proliferate and differentiate. Based on these data, we hypothesize that at least part of the functional differences that exist between adult and fetal stem cells can be accounted for by a developmental different responsiveness to TGF-␤.