Activation of the human neurtrophil

描述（由申请人提供）： 嗜中性粒细胞是杀微生物活性所必需的，但也可引起 在疾病如败血症、类风湿性关节炎、 关节炎哮喘配体如肿瘤坏死因子（TNFa）、免疫调节剂 复合物和趋化性肽如fMet-Leu-Phe，引起 氧自由基和颗粒含量以及对表面的粘附。我们假设 分离的蛋白激酶C（PKC）同种型在蛋白质合成中既有正性作用， 在促炎反应信号中的作用。促炎信号可以 在许多水平上受PKC同种型调节，包括：a）受体，B） i）第二信使Ca 2+，和ii）脂质辅因子的产生和调节， 和c）组装复合物如用于O2-产生的NADPH氧化酶。使用 反义策略消除分化的HL 60中的特异性PKC同种型 （dHL 60），我们证明了a-PKC和b-PKC在 氧气生成相反，b-PKC下调配体启动的Ca 2+摄取， 5-PKC负性调节p60 TNF受体（p60 TNFR）的功能。 反义策略将用于选择性地耗尽特定的PKC同种型， 确定脂质和脂质依赖性PKC同种型在信号转导中的作用， 促炎反应的转导。我们将： 钙依赖性a-PKC和b-PKC亚型在NADPH活化中的作用 氧化酶和p47 phox的磷酸化。二、确定选择性角色 钙离子依赖性a-PKC和b-PKC亚型在粘附激活和细胞凋亡中的作用 MARCKS的脱颗粒和磷酸化。III确定PKC同种型 负责调节配体启动的细胞内 Ca 2+和钙库操纵的Ca 2+通道。四.审查选择性作用， 钙离子依赖性a-PKC和b-PKC亚型在配体启动的细胞凋亡调控中的作用 涉及效应酶PLCb/PLCg和PLD 1的信号传导，以及涉及 PIP 2的再生。V确定d-PKC在下调细胞凋亡中的作用。 p60 TNFR引发的信号通路。确定d-PKC是否作用于 在质膜区域或死亡结构域中的丝氨酸残基。

DESCRIPTION (provided by applicant): Secretion of oxygen radicals by neutrophils is essential for microbicidal activity, but can also give rise to the tissue damage of inflammation in diseases such as sepsis, rheumatoid arthritis, asthma. Ligands such as tumor necrosis factor (TNFa), immune complexes and chemotactic peptides such as fMet-Leu-Phe, elicit secretion of oxygen radicals and granule contents and adherence to a surface. We hypothesize that discrete protein kinase C (PKC) isotypes play both positive and negative roles in signaling for proinflammatory responses. Proinflammatory signaling can be regulated by PKC isotypes at numerous levels including: a) the receptor, b) generation and regulation of i) second messenger Ca2+, and ii) lipid cofactors, and c) assembly of complexes such as the NADPH oxidase for O2-generation. Using antisense strategies to deplete specific PKC isotypes in differentiated HL60 (dHL60), we demonstrated positive signaling roles for both a-PKC and b-PKC for O2- generation. Conversely, b-PKC downregulates ligand-initiated Ca2+ uptake, and 5-PKC negatively regulates the p60TNF receptor (p60TNFR) function. Antisense strategies will be used to selectively deplete specific PKC isotypes, to establish roles for lipids and lipid-dependent PKC isotypes in signal transduction for proinflammatory responses. We will: I Establish selective roles for Ca2+-dependent a-PKC and b-PKC isotypes in activation of the NADPH oxidase, and phosphorylation of p47phox. II Determine selective roles for Ca2+-dependent a-PKC and b-PKC isotypes in activation of adherence and degranulation and phosphorylationof MARCKS. III Determine the PKC isotypes responsible for regulation of ligand-initiated mobilization of intracellular Ca2+ and store operated Ca2+ channels. IV Examine selective roles for Ca2+-dependent a-PKC and b-PKC isotypes in the regulation of ligand initiated signalling involving effector enzymes PLCb/PLCg, and PLD1, and enzymes involved in the regeneration of PIP2. V Establish the role of d-PKC in downregulation of signalling pathways triggered by the p60TNFR. Determine if d-PKC acts on a serine residue(s) in the juxtamembrane region or the death domain.