Leukocyte chemoattractants have long been regarded as a group of proinflammatory factors that trigger specific cellular function such as chemotaxis, degranulation, generation of superoxide anions and activation of integrins. Results from recent studies indicate that chemoattractants can also induce the synthesis and secretion of proinflammatory cytokines including chemokines, a functions that may be crucial for the initiation and regulation of a local cytokine network and or the continued recruitment of leukocytes to site of inflammation. Preliminary results derived from our studies suggest the hypothesis that chemoattractant-induced NF-kappa B- activation is a primary mechanism for cytokine gene expression in leukocytes. It is also postulated that the signaling pathways leading to chemoattractant-induced NF(kappa)B activation differ from that utilized by other NF(kappa)B-activating agents such as TNF(alpha) and PMA. The current project aims to test these hypothesis, and contains 2 specific aims. (1) Characterization of chemoattractant-stimulated signaling events leading to transcription activation. This is the core of the application. Our primary interest is in the identification of mechanisms that unique to chemoattractant (PAF, FMLP)-stimulated NF-(kappa) B activation. A reconstitution system, based on transient co-transfection of plasmid DNA, will be employed to identify an additional myeloid differentiation-association factor required for FMLP-( and also C5a and IL-8) induced NF-(kappa)B activation. (2) Investigation of the extent and variability of chemoattractant-stimulated gene expression in monocytes and polymorphonulear neutrophils (PMN). IN this specific we will examine whether monocytes and neutrophils employ the same mechanism for gene expression in response to chemoattractant stimulation. We will also test the notion IL-8 can regulate its own gene expression through binding of the IL-8 receptors and activation of NF-(kappa) B. Achievement of these goals will extend our current understanding of the physiological functions of chemoattractants, and is an integral part of our long term research objective relating to the molecular mechanisms of leukocyte activation.
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