Ex vivo expansion of mature neutrophils from hematopoietic progenitor cells and cytokine-mediated signal transduction

在CD 34阳性造血祖细胞中，干细胞因子（SCF）、粒细胞集落刺激因子（G-CSF）、血小板生成素（TPO）、粒细胞-巨噬细胞集落刺激因子（GM-CSF）、白细胞介素（IL）-3和肿瘤坏死因子（TNF）-α介导细胞外信号调节激酶（ERK）的磷酸化，IL-1和TNF-α介导p38 MAP激酶的磷酸化，TNF-α介导的c-Jun N-末端激酶（JNK）磷酸化。p38 MAP激酶抑制剂可抑制IL-1b诱导的CD 11b和CD 15表达。IL-6/sIL-6 R、G-CSF、TPO、GM-CSF、IL-3、IFN-α和IFN-γ介导的信号转导子和转录激活子（STAT）3-Y的磷酸化，SCF、IL-6/sIL-6 R、G-CSF、TPO、GM-CSF、IL-3和TNF-α介导的STST 3-S的磷酸化，G-CSF、TPO、GM-CSF、IL-3和IFN-α介导的STST 5a的磷酸化，G-CSF、GM-CSF、IL-3、IFN-α和IFN-γ介导的STAT 5 b的磷酸化，IFN-α和IFN-γ介导的STAT 1磷酸化。体外扩增具有正常功能（趋化性、吞噬作用和超氧化物释放）的成熟人中性粒细胞。将CD 34阳性造血祖细胞与SCF、IL-3、GM-CSF和G-CSF一起培养7天，然后将非贴壁细胞单独与G-CSF一起进一步培养7天。TPO在分化的早期阶段刺激成熟中性粒细胞的离体扩增。人中性粒细胞表达细胞凋亡抑制因子（cIAP 1、cIAP 2和X连锁cIAP）家族成员。G-CSF选择性上调c-IAP 2表达。JAK 2特异性抑制剂可抑制G-CSF诱导的cIAP 2上调、STAT 3磷酸化和抗凋亡作用。在单核细胞中，G-CSF抑制LPS诱导的TNF-α产生并激活STST 3。这些作用被JAK 2的抑制剂阻止。

In CD34 positive hematopoietic progenitor cells, stem cell factor(SCF), granulocyte colony-stimulating factor(G-CSF), thrombopoietin(TPO), granulocyte-macrophage colony-stimulating factor(GM-CSF), interleukin(IL)-3, and tumor necrosis factor(TNF)-α mediated phosphorylation of extracellular-signal regulated kinase(ERK), IL-1 and TNF-α mediated phosphorylation of p38 MAP kinase, TNF-α mediated phosphorylation of c-Jun N-terminal kinase (JNK). Inhibitors of p38 MAP kinase inhibited expression of CD11b and CD15 induced by IL-1b. IL-6/sIL-6R,G-CSF,TPO,GM-CSF,IL-3,IFN-α, and IFN-γ mediated phosphorylation of signal transducer and activator of transcription(STAT)3-Y,SCF,IL-6/sIL-6R,G-CSF,TPO,GM-CSF,IL-3, and TNF-α mediated phosphorylation of STST3-S,G-CSF,TPO,GM-CSF, IL-3, and IFN-a mediated phosphorylation of STST5a,G-CSF,GM-CSF,IL-3,IFN-α, and IFN-γ mediated phosphorylation of STAT5b, and IFN-α and IFN-γ mediated phosphorylation of STAT 1.Mature human neutrophils with normal functions (chemotaxis, phagocytosis and superoxide release) were ex vivo expanded from haematopoietic progenitor cells as the following procedure. CD34 positive haematopoietic progenitor cells were cultivated with SCF,IL-3,GM-CSF, and G-CSF for 7 days, and thereafter non-adherent cells were further cultivated for 7 days with G-CSF alone. TPO stimulated ex vivo expansion of mature neutrophils in the early stages of differentiation. Human neutrophils were expressed members of the inhibitor of apoptosis (LAP) family (cIAP1,cIAP2, and X-linked LAP). G-CSF selectively up-regulated c-IAP2 expression. A specific inhibitor of Janus kinase 2(JAK2) inhibited G-CSF-induced up-regulation of cIAP2, phosphorylation of STAT3, and the antiapoptotic effects. In monocyte, G-CSF inhibited LPS-induced TNF-α production and activated STST3. These effects were prevented by an inhibitor of JAK2.