Aronica S M, Mantel C, Gonin R, Marshall M S, Sarris A, Cooper S, Hague N, Zhang X F, Broxmeyer H E
Department of Medicine (Hematology/Oncology), Indiana University School of Medicine, Indianapolis 46202, USA.
J Biol Chem. 1995 Sep 15;270(37):21998-2007. doi: 10.1074/jbc.270.37.21998.
Stimulatory cytokines, including granulocyte-macrophage colony-stimulating factor (GM-CSF) and steel factor (SLF), act in a synergistic manner to stimulate the growth of hematopoietic progenitor cells, an effect also demonstrated for the growth factor-dependent human hematopoietic cell line MO7e. While little is known about the mechanisms responsible for mediating synergistic interactions of cytokines, Raf-1, a component of the MAP kinase signaling pathway, is thought to play a role in the stimulatory response evoked by several cytokines, including SLF and GM-CSF. Interferon-inducible protein-10 (IP-10) and macrophage inflammatory protein-1 alpha (MIP-1 alpha) are members of the chemokine family of suppressive cytokines. Prior exposure of hematopoietic cells to chemokines, including IP-10 and MIP-1 alpha, inhibits the synergistic action of growth factors on stimulating cell proliferation. We report that treatment of MO7e cells with the combination of GM-CSF and SLF directly stimulates statistically significant synergistic increases in the phosphorylation and activation of Raf-1 kinase, and in cellular protein synthesis levels. Pretreatment of MO7e cells with IP-10 or MIP-1 alpha blocked synergistic growth factor action, resulting in statistically significant suppression of cell proliferation, protein synthesis, and Raf-1 phosphorylation and activation. IP-10 and MIP-1 alpha treatment also evoked significant increases in intracellular cAMP levels. Pretreatment of cells with agents which serve to raise intracellular cAMP levels, or with cAMP analogs inhibited the synergistic actions of GM-CSF and SLF in a manner similar to IP-10 and MIP-1 alpha. In addition, treatment of cells with a potent inhibitor of cAMP-dependent protein kinase A blocked the suppressive action of MIP-1 alpha and IP-10 on Raf-1 kinase activity and on MO7e cell proliferation. The ability of IP-10 and MIP-1 alpha to antagonize the synergistic action of GM-CSF and SLF appears to involve inactivation of Raf-1 and the down-regulation of protein synthesis. Our findings suggest that both MIP-1 alpha and IP-10 mediate their suppressive effects in MO7e cells by stimulating increases in cellular cAMP levels and activating protein kinase A, a mechanism we believe to be unique to these chemokines and not one applied to all growth suppressive members of the chemokine superfamily (for example, interleukin 8 and platelet factor 4).
包括粒细胞-巨噬细胞集落刺激因子（GM-CSF）和干细胞因子（SLF）在内的刺激性细胞因子以协同方式作用，刺激造血祖细胞的生长，这一作用在依赖生长因子的人造血细胞系MO7e中也得到了证实。虽然对于介导细胞因子协同相互作用的机制知之甚少，但丝裂原活化蛋白激酶（MAP）信号通路的组成部分Raf-1被认为在包括SLF和GM-CSF在内的多种细胞因子所引发的刺激反应中发挥作用。干扰素诱导蛋白10（IP-10）和巨噬细胞炎性蛋白1α（MIP-1α）是抑制性细胞因子趋化因子家族的成员。造血细胞预先暴露于包括IP-10和MIP-1α在内的趋化因子会抑制生长因子对细胞增殖的协同作用。我们报告，用GM-CSF和SLF联合处理MO7e细胞可直接刺激Raf-1激酶的磷酸化和活化以及细胞蛋白质合成水平出现具有统计学意义的协同增加。用IP-10或MIP-1α对MO7e细胞进行预处理可阻断生长因子的协同作用，导致细胞增殖、蛋白质合成以及Raf-1磷酸化和活化受到具有统计学意义的抑制。IP-10和MIP-1α处理还会使细胞内cAMP水平显著升高。用能够提高细胞内cAMP水平的试剂或cAMP类似物对细胞进行预处理，以类似于IP-10和MIP-1α的方式抑制了GM-CSF和SLF的协同作用。此外，用一种有效的cAMP依赖性蛋白激酶A抑制剂处理细胞可阻断MIP-1α和IP-10对Raf-1激酶活性和MO7e细胞增殖的抑制作用。IP-10和MIP-1α拮抗GM-CSF和SLF协同作用的能力似乎涉及Raf-1的失活和蛋白质合成的下调。我们的研究结果表明，MIP-1α和IP-10均通过刺激细胞内cAMP水平升高和激活蛋白激酶A来介导它们在MO7e细胞中的抑制作用，我们认为这是这些趋化因子所特有的机制，而不是趋化因子超家族所有生长抑制成员（例如，白细胞介素8和血小板因子4）所适用的机制。
