Ellison Michael A, Thurman Gail, Gearheart Christy M, Seewald Ryan H, Porter Christopher C, Ambruso Daniel R
Department of Pediatrics, University of Colorado School of Medicine, Aurora, CO, United States of America.
Department of Pediatrics, University of Colorado School of Medicine, Aurora, CO, United States of America; Department of Pediatrics, University of Colorado Denver, The Anschutz Medical Campus, Aurora, Colorado, United States of America; Department of Pathology, University of Colorado Denver, The Anschutz Medical Campus, Aurora, Colorado, United States of America; The Center for Cancer and Blood Disorders, Transfusion Services, Children's Hospital Colorado, Aurora, Colorado, United States of America; Hematology/Oncology and Bone Marrow Transplantation Laboratories, Aurora, Colorado, United States of America.
PLoS One. 2015 Aug 28;10(8):e0136766. doi: 10.1371/journal.pone.0136766. eCollection 2015.
The cytokine and drug interferon-γ enhances superoxide anion production by the antimicrobicidal Nox2 enzyme of neutrophils. Because mature neutrophils have a short lifespan, we hypothesized that the effects of interferon-γ on these cells might be mediated by its prolonged exposure to differentiating neutrophil precursors in the bone marrow rather than its brief exposure to mature circulating neutrophils. Effects of INF-Γ on NOX2 activity: To address this possibility we exposed the myeloid PLB-985 cell line to interferon-γ for 3 days in the presence of dimethyl sulfoxide which induces terminal differentiation of these cells. Interferon-γ was found to enhance superoxide production by Nox2 in a concentration dependent manner. In contrast, application of interferon-γ alone for 3 days failed to induce detectible Nox2 activity. Additionally, application of interferon-γ for 3 hours to pre-differentiated PLB-985 cells, which models studies using isolated neutrophils, was much less effective at enhancing superoxide anion production. Effects of INF-Γ on phox protein levels: Addition of interferon-γ during differentiation was found to upregulate the Nox2 proteins gp91phox and p47phox in concert with elevated transcription of their genes. The p22phox protein was upregulated in the absence of increased transcription presumably reflecting stabilization resulting from binding to the elevated gp91phox. Thus, increased levels of gp91phox, p47phox and p22phox likely account for the interferon-γ mediated enhancement of dimethyl sulfoxide-induced Nox2 activity. In contrast, although interferon-γ alone also increased various phox proteins and their mRNAs, the pattern was very different to that seen with interferon-γ plus dimethyl sulfoxide. In particular, p47phox was not induced thus explaining the inability of interferon -γ alone to enhance Nox2 activity. Short application of interferon-γ to already differentiated cells failed to increase any phox proteins.
Our findings indicate that interferon-γ has complex effects on phox protein expression and that these are different in cells undergoing terminal differentiation. Understanding these changes may indicate additional therapeutic uses for this cytokine in human disorders.
细胞因子及药物干扰素-γ可增强中性粒细胞的抗菌性Nox2酶产生超氧阴离子的能力。由于成熟中性粒细胞寿命较短，我们推测干扰素-γ对这些细胞的作用可能是通过其长时间作用于骨髓中正在分化的中性粒细胞前体来介导的，而非短暂作用于成熟的循环中性粒细胞。干扰素-γ对Nox2活性的影响：为探究这种可能性，我们在二甲基亚砜存在的情况下，将髓系PLB-985细胞系暴露于干扰素-γ 3天，二甲基亚砜可诱导这些细胞终末分化。结果发现，干扰素-γ以浓度依赖的方式增强Nox2产生超氧阴离子的能力。相比之下，单独应用干扰素-γ 3天未能诱导出可检测到的Nox2活性。此外，将干扰素-γ应用于预分化的PLB-985细胞3小时（这模拟了使用分离的中性粒细胞进行的研究），在增强超氧阴离子产生方面效果要差得多。干扰素-γ对吞噬氧化蛋白水平的影响：在分化过程中添加干扰素-γ可上调Nox2蛋白gp91phox和p47phox，同时其基因转录水平也升高。在转录未增加的情况下，p22phox蛋白上调，这可能反映了与升高的gp91phox结合导致的稳定性增加。因此，gp91phox、p47phox和p22phox水平升高可能是干扰素-γ介导增强二甲基亚砜诱导的Nox2活性的原因。相比之下，尽管单独的干扰素-γ也会增加各种吞噬氧化蛋白及其mRNA，但模式与干扰素-γ加二甲基亚砜时非常不同。特别是，p47phox未被诱导，这就解释了单独的干扰素-γ无法增强Nox2活性的原因。对已分化细胞短期应用干扰素-γ未能增加任何吞噬氧化蛋白。
我们的研究结果表明，干扰素-γ对吞噬氧化蛋白表达具有复杂的影响，且在经历终末分化的细胞中这些影响有所不同。了解这些变化可能为该细胞因子在人类疾病中的额外治疗用途指明方向。
