REGULATION OF POLYMORPHONUCLEAR LEUKOCYTE PHAGOCYTOSIS

University Of Michigan At Ann Arbor, Ann Arbor MI

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Abstract

Polymorphonuclear leukocytes (PMNs) are recruited to sites of inflammation and tissue injury where they destroy foreign pathogens. Pathogens are engulfed by phagocytosis and contained within phagosomes where they await destruction. There are two major events that contribute to pathogen destruction: (1) the NADPH oxidase assembles on the phagosomal membrane and delivers reactive oxygen metabolites into the phagosome and (2) cytoplasmic granules fuse with the phagosomal membrane and deliver hydrolytic and bactericidal enzymes to the phagosome. Presently, the lipid signaling pathways leading to these complex events have not been clearly defined. We found that ceramide is generated at a time when the respiratory burst is terminated and that addition of cell- permeable ceramide analogues and sphingolipids block phagocytosis. Therefore, sphingolipids, including ceramide, may function as critical components of receptor-mediated signaling in PMNs. This application propose to explore the role of sphingolipid metabolism in PMN phagocytosis. Based on these key observation, we propose the following hypothesis: ceramide and sphingosine formed during receptor-mediated activation are critical to the regulation of phagocytosis in PMNs. Specific Aim 1 will demonstrate that ceramide is essential in the signaling pathways that regulate PMN phagocytosis and identify those pathways which contribute to ceramide generation. Specific Aim 2 will characterize PMN sphingomyelinase(s), define their subcellular localization and determine what factors regulate their activation. Specific AIM 3 will examine the cellular targets of ceramide and sphingosine actin during PMN phagocytosis, focusing primarily on activation of mitogen-activated kinase (MAPK), Raf kinase and how they are linked to phagosome generation.