ARACHIDONATE METABOLITES AND NEUTROPHIL FUNCTION

受刺激的多形核中性粒细胞（PMN）代谢其 花生四烯酸和磷脂储存成一系列生物活性物质， 产品：白三烯（LT）B4，5-羟基二十碳四烯酸（5-HETE）， 血小板活化因子（PAF）， 甘油二酯（DAG）。 我们假设这些产物相互作用 以影响如下各种组织的功能反应。 LTB 4、PAF和5-HETE通过与它们各自的受体结合来刺激细胞。 质膜受体 LTB 4和PAF的受体通过 升高胞浆Ca和激活蛋白激酶C（PKC）， 涉及GTP结合蛋白，磷脂酶C， 并将磷脂酰肌醇二磷酸裂解成肌醇 三磷酸盐和DAG。 肌醇三磷酸释放Ca 2 + 亚细胞池至胞质溶胶，而DAG通过以下方式直接激活PKC： Ca 2+增强反应。 5-HETE的受体通过 不同的机制，（其中，我们假设，是协同的， 由PAF和LTB 4使用），以升高胞质Ca 2+并促进PKC activation. 升高的Ca 2+和激活的PKC然后启动 功能 然而，PG和在一定条件下激活的PKC 抑制功能;它们可能通过下调 LTB 4、PAF和5-HETE，或通过干扰这些受体， 与GTP结合蛋白的连接。 我们的概念将在 研究中的人PMN、PMIN胞质和PMIN分离的细胞器 a）定义亚细胞分布和结合 LTB 4、PAF、5-HETE和PG的受体参数; B）检查 这些受体与Ca 2+的特异性连接和作用 流量和PKC激活：c）确定PG和 激活PKC对这些受体结合配体和功能 与GTP结合蛋白的连接;和d）评估 LTB4。 PMN对5-HETE、PG、PAF、DAG、Ca ~（2+）和PKC的反应 外源刺激（例如，趋化肽和钙 离子载体）和其它方法， 选择性消除PMN合成或对脂质的反应 产品. 我们的研究在很大程度上依赖于使用中性粒细胞， 生理条件和相关细胞的重建- 游离模型包括纯化的PMN细胞器、PKC和GTP- 结合蛋白 最终，我们的目标是确定脂质是如何 产品刺激不同的细胞类型， 在刺激-反应耦合事件中。 由于这些产品和 它们的起源细胞似乎与炎症，过敏， 过敏性和支气管痉挛反应，这项建议是 与细胞生物学、宿主防御 机制、肺生理学、药理学和临床 药

Stimulated polymorphonuclear neutrophils (PMN) metabolize their arachidonate and phospholipid stores into a series of bioactive products: leukotriene (LT)B4, 5-hydroxyicosatetraenoate (5-HETE), prostaglandins (PG), platelet-activating factor (PAF), and diacylglycerol (DAG). We postulate that these products interact to influence functional responses of various tissues as follows. LTB4, PAF, and 5-HETE excite cells by binding to their respective plasma membrane receptors. Receptors for LTB4 and PAF operate by raising cytosolic Ca and activating protein kinase C (PKC) through a linkage cascade involving GTP-binding proteins, phospholipase C, and cleavage of phosphatidylinositol diphosphate into inositol triphosphate and DAG. Inositol triphosphate releases Ca2+ from subcellular pools to cytosol whereas DAG directly activates PKC by a Ca2+-enhanced reaction. Receptors for 5-HETE operate by a different mechanism, (which, we postulate, is synergistic with that used by PAF and LTB4), to raise cytosolic Ca2+ and promote PKC activation. The elevated Ca2+ and activated PKC then initiate function. However, PG and, under certain conditions, activated PKC inhibit function; they may act by down-regulating receptors for LTB4, PAF, and 5-HETE or by interfering with these receptors' linkages to GTP-binding proteins. Our concepts will be tested on human PMN, PMIN cytoplasts, and PMIN isolated organelles in studies that: a) define the subcellular distributions and binding parameters of receptors for LTB4, PAF, 5-HETE, and PG; b) examine these receptors' specific linkages to, and effects upon, Ca2+ fluxes and PKC activation: c) determine the influences of PG and activated PKC on these receptors' binding of ligands and functional linkages to GTP-binding proteins; and d) evaluate the roles of LTB4. 5-HETE, PG, PAF, DAG, Ca2+ and PKC in PMN responses to exogenous stimuli (e.g., chemotactic peptides and calcium ionophores) using pharmacologic agents and other methods that selective abrogate PMN synthesis of or responses to the lipid products. Our studies rely heavily upon the use of PMN under physiological conditions and the reconstruction of relevant cell- free models comprised of purified PMN organelles, PKC, and GTP- binding proteins. Ultimately, we aim to define how the lipid products stimulate diverse cell types and to implicate the products in stimulus-response coupling events. Since these products and their cells of origin appear involved in inflammatory, allergic, anaphylactic, and bronchospastic reactions, this proposal is relevant to the general fields of cellular biology, host defense mechanisms, pulmonary physiology, pharmacology, and clinical medicine.