ACTIVATION OF THE HUMAN NEUTROPHIL

描述：(改编自调查人员摘要)炎症性 中介物肿瘤坏死因子-α诱导中性粒细胞分泌 细胞外氧自由基的释放和颗粒内容物有助于 组织损伤。研究人员假设激活不同的 脂类介体介导的蛋白激酶C亚型既有阳性也有 在这些事件中扮演了负面的监管角色。肿瘤坏死因子-α触发血管内皮细胞活化 效应酶PLC-γ和PI3激酶，导致产生这样的 脂质介质如二甘油三酯和PIP3以及PKC亚型的激活。 产生超氧化物的积极信号是通过 依赖DG的PKC亚型对p47-Phox的磷酸化作用。负性 例如，由PKC发出的信号导致肿瘤坏死因子-α结合的下调 依赖于DG和PIP3的p60肿瘤坏死因子受体的磷酸化 Zeta PKC。研究人员进一步提出，基于PKC的信令 需要1)胞浆PLC-γ和PI3激酶进入细胞膜 相关脂质，2)胞浆PKC亚型靶向PLC和PI3 3)PKC的靶向 与它们的底物有相同的类型。该模型将通过实验进行测试。 重点关注四个具体目标：1)确定Zeta PKC是否催化 P60TNFR在死亡区域的磷酸化，从而下调 感受器。2)使用依赖DG的PKC亚型的选择性缺失来 定义阿尔法、贝塔和德尔塔PKC在超氧化物生成中的作用。3) 肿瘤坏死因子-α诱导磷脂酶C和磷脂酶PI3的激活 PIP3和DG对PKC亚型的伽马和差异性激活。4)定义 Rack(活化C激酶受体)在PLC共定位中的作用 依赖DG的PKC亚型与PKC和超氧化物歧化 一代。长期目标是定义积极和消极 肿瘤坏死因子-α激活超氧化物歧化的信号转导途径 中性粒细胞，从而潜在地导致靶向分子策略 用于控制炎症和组织损伤。

DESCRIPTION: (Adapted from investigator's abstract) The inflammatory mediator TNF-alpha elicits secretion by adherent neutrophils and the extracellular release of oxygen radicals and granule contents contributes to tissue damage. The investigator hypothesizes that activation of different protein kinase C isotypes by lipid mediators plays both positive and negative regulatory roles in these events. TNF-alpha triggers activation of effector enzymes PLC-gamma and PI3 kinase, leading to production of such lipid mediators as diglyceride and PIP3 and to activation of PKC isotypes. Positive signalling for superoxide generation is mediated through phosphorylation of p47-phox by DG-dependent PKC isotypes. Negative signalling by PKC elicits down regulation of TNF-alpha binding, for example by phosphorylation of the p60 TNF receptor by DG-independent, PIP3-dependent zeta PKC. The investigator further proposes that PKC-based signalling requires 1) access of cytosolic PLC-gamma and PI3 kinase to membrane associated lipids, 2) targeting of cytosolic PKC isotypes to PLC and PI3 kinase and thus to lipid mediators DG and PIP3, and 3) targeting of PKC isotypes to their substrates. This model will be tested through experiments focusing on four specific aims: 1) Determine if zeta PKC catalyzes phosphorylation of p60 TNFR in the death domain and thereby down regulates the receptor. 2) Use selective deletion of DG-dependent PKC isotypes to define roles for alpha, beta, and delta PKC in superoxide generation. 3) Establish TNF-alpha elicited activation of PI3 kinase and phospholipase C gamma and differential activation of PKC isotypes by PIP3 and DG. 4) Define a role for RACK (receptor for activated C kinase) in co-localization of PLC and DG-dependent PKC isotypes and activation of PKC and superoxide generation. The long term objective is to define positive and negative signalling for TNF-alpha induced activation of superoxide generation by neutrophils, thereby potentially leading to targeted molecular strategies for control of inflammation and tissue damage.