Interferon-γ Constitutively Expressed in the Stromal Microenvironment of Human Marrow Cultures Mediates Potent Hematopoietic Inhibition

• Published in: Blood Vol. 87; no. 10; pp. 4149 - 4157
• Main Authors: Selleri, Carmine, Maciejewski, Jaroslaw P., Sato, Tadatsugu, Young, Neal S.
• Format: Journal Article
• Language: English
• Published: Washington, DC Elsevier Inc 15.05.1996; The Americain Society of Hematology
• Subjects: 3T3 Cells; Anemia, Aplastic - physiopathology; Anemias. Hemoglobinopathies; Animals; Apoptosis - drug effects; Apoptosis - physiology; Base Sequence; Biological and medical sciences; Bone Marrow - physiology; Bone Marrow Cells; Cell Cycle - drug effects; Cell Cycle - physiology; Cell Division - drug effects; Cells, Cultured; Coculture Techniques; Colony-Forming Units Assay; Connective Tissue - metabolism; Connective Tissue Cells; Diseases of red blood cells; Gene Expression; Hematologic and hematopoietic diseases; Hematopoiesis - physiology; Hematopoietic Stem Cells - cytology; Humans; Interferon-gamma - biosynthesis; Interferon-gamma - genetics; Interferon-gamma - pharmacology; Interferon-gamma - physiology; Medical sciences; Mice; Molecular Sequence Data; Polymerase Chain Reaction; Recombinant Fusion Proteins - metabolism; Transfection; Human; Cell culture; Pathophysiology; Stem cell; Cytokine; Aplastic anemia; Cell microenvironment; Hematopoietic cell; Hemopathy; Long term; Hematopoiesis; Bone marrow; Stromal cell; Inhibition; Gamma interferon
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V87.10.4149.bloodjournal87104149

Clinical and laboratory studies have suggested involvement of interferon-γ (IFN-γ) in the pathophysiology of aplastic anemia. T cells from aplastic anemia (AA) patients secrete IFN-γ in vitro, activated cytotoxic lymphocytes infiltrate aplastic bone marrow (BM), and IFN-γ mRNA, not detected in normal BM, is present in BM from most AA patients. Many patients respond to immunosuppressive therapy with anti-thymocyte globulin and cyclosporine. Using long-term BM cultures (LTBMC) as a tissue culture model of hematopoiesis, we show that IFN-γ is a potent inhibitor in the long-term culture-initiating cell (LTC-IC) assay, the best in vitro surrogate test for human hematopoietic stem cells, as well as of the output of committed progenitor cells (colony-forming unit-granulocyte-macrophage [CFU-GM] and burst-forming unit-erythroid [BFU-E]). In LTBMC, continuous addition of relatively high IFN-γ concentrations (1,000 U/mL weekly or 200 U/mL every 2 days) was required for inhibition of secondary colony formation, a measure of LTC-IC number and clonogenicity. To mimick local production of IFN-γ, human stromal cells were engineered by retroviral-mediated gene transfer to express a transduced IFN-γ gene, IFN-γ secreted by stromal cells was far more potent than exogenous IFN-γ in its effects in the LTC-IC assay. For purified CD34+ cells, culture in the presence of IFN-γ stroma dramatically reduced secondary colony numbers as well as production of CFU-GM and BFU-E. Supernatants from these cultures contained only about 20 U/mL of IFN-γ; this quantity of cytokine, when added to LTBMC, had little effect on hematopoie-sis. The mechanism of hematopoietic suppression was related to the inhibition of cell cycle progression and induction of apoptosis of CD34+ cells. There was no apparent effect of local low-level IFN-γ production on stromal cell function, as reflected in cell morphology, cell surface phenotype, or expression of hematopoietic growth factor genes. LTBMC with genetically altered stromal cells offers an in vitro model of immune suppression of hematopoiesis in AA and may be helpful in testing certain therapeutic modalities. We infer from our data that local production of low levels of inhibitory cytokine is sufficient to markedly inhibit hematopoiesis and to destroy stem cells and more mature progenitor cells. This is a US government work. There are no restrictions on its use.