Interferon-gamma constitutively expressed in the stromal microenvironment of human marrow cultures mediates potent hematopoietic inhibition
Interferon-gamma constitutively expressed in the stromal microenvironment of human marrow cultures mediates potent hematopoietic inhibition
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DOI：
10.1182/blood.v87.10.4149.bloodjournal87104149
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发表时间：
1996-05-15
期刊：
影响因子：
20.3
通讯作者：
Young, NS
中科院分区：
文献类型：
--
作者：
Selleri, C;Maciejewski, JP;Young, NS
临床和实验室研究表明，干扰素-γ参与了再生障碍性贫血的病理生理过程。再生障碍性贫血(AA)患者的T细胞在体外分泌干扰素-γ，活化的细胞毒性淋巴细胞渗入再生障碍性骨髓(BM)，并且在大多数AA患者的BM中存在正常BM中未检测到的干扰素-γmRNA。许多患者对使用抗胸腺细胞球蛋白和环孢素的免疫抑制治疗有反应。使用长期骨髓培养(LTBMC)作为造血的组织培养模型，我们发现在长期培养-启动细胞(LTC-IC)试验中，干扰素-γ是一种有效的抑制物，长期培养-起始细胞试验是人类造血干细胞的最佳体外替代试验，也是定向祖细胞(集落形成单位-粒-巨噬细胞[CFU-GM]和爆裂形成单位-红系细胞[BFU-E])的输出。在LTBMC中，需要持续加入相对较高的干扰素-γ浓度(每周1.000 U/m L或每2天200U/m L)以抑制二次克隆形成，以衡量L TC-IC数量和克隆形成能力。为了模拟局部产生的干扰素-γ，通过逆转录病毒介导的基因转移使人基质细胞表达转导的干扰素-γ基因。在LTC-IC实验中，基质细胞分泌的干扰素-γ远强于外源性的干扰素-γ。对于纯化的CD34(+)细胞，在干扰素-γ基质存在的情况下培养显著减少二次集落数量以及CFU-GM和BFU-E的产生。从这些培养上清液中只含有大约20U/ml的干扰素-γ；当将这个数量的细胞因子加入到LTBMC中时，对造血几乎没有影响。其机制可能与抑制CD34(+)细胞的细胞周期进程和诱导其凋亡有关。在细胞形态、细胞表面表型或造血生长因子基因表达方面，局部低水平的干扰素-γ产生对基质细胞功能没有明显影响。具有遗传改变的基质细胞的LTBMC提供了一种免疫抑制再生障碍性贫血的体外模型，并可能有助于测试某些治疗方法。我们从我们的数据中推断，局部产生低水平的抑制性细胞因子足以显著抑制造血，并破坏干细胞和更成熟的祖细胞。
Clinical and laboratory studies have suggested involvement of interferon-gamma (IFN-gamma) in the pathophysiology of aplastic anemia. T cells from aplastic anemia (AA) patients secrete IFN-gamma in vitro, activated cytotoxic lymphocytes infiltrate aplastic bone marrow (BM), and IFN-gamma mRNA, not detected in normal BM, is present in BM from most AA patients. Many patients respond to immunosuppressive therapy with antithymocyte globulin and cyclosporine. Using long-term BM cultures (LTBMC) as a tissue culture model of hematopoiesis, we show that IFN-gamma is a potent inhibitor in the longterm 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-gamma 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-gamma, human stromal cells were engineered by retroviral-mediated gene transfer to express a transduced IFN-gamma gene. IFN-gamma secreted by stromal cells was far more potent than exogenous IFN-gamma in its effects in the LTC-IC assay. For purified CD34(+) cells, culture in the presence of IFN-gamma 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-gamma; this quantity of cytokine, when added to LTBMC, had little effect on hematopoiesis. 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-gamma 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.