Neutrophils and other circulating phagocytes generate high levels of reactive oxygen species (ROS) in response to a variety of infectious or inflammatory stimuli in a process known as the respiratory burst. This response is attributed to the activity of NADPH oxidase, which produces superoxide, a precursor of ROS that are important microbicidal agents and mediators of inflammation. Patients with chronic granulomatous disease (CGD) have NADPH oxidase deficiencies and suffer from enhanced susceptibility to microbial infections and aberrant inflammatory responses. This project explores the cellular mechanisms regulating the respiratory burst in phagocytes and is characterizing oxidative responses of related enzymes expressed in a variety of non-immune cells. In work aimed at defining signal transduction pathways triggering activation of the phagocyte oxidase (phox), we have engaged two gene transfection approaches: 1) reconstitution of receptor-mediated activation of the respiratory burst in undifferentiated (K562) cells and 2) expression of modified signaling molecules in differentiated myeloid (PLB-985) cells. The reconstituted chemotactic peptide receptor responses have been delineated into both calcium-dependent and independent pathways, while the latter approach has provided new evidence for involvement of the small GTPase, ADP-ribosylation factor-6 (ARF-6), and phospholipase D in the respiratory burst. Information on signaling intermediates affecting the respiratory burst in phagocytes will provide a basis (pharmacological targets) for therapeutic strategies designed to inhibit or enhance oxidative responses of phagocytes. In other studies we are characterizing sources of reactive oxygen species in other tissues (colon, kidney, brain, and vascular tissue). In these sites, the oxidative changes can serve as redox """"""""second messengers"""""""" promoting inflammatory signals, oxygen sensing, and changes in gene expression patterns (proliferation responses to growth factors, differentiation, cellular senescence, apoptosis or programmed cell death). Studies in the p47phox-deficient mouse model of CGD indicate an essential role for p47phox in the release of reactive oxidants by microgial cells in response to neutrophil agonists or beta-amyloid, which may relate to mechanisms of development of neurodegenerative processes associated with Alzheimer's disease. The normal oxidative responses of aortic smooth muscle cells following stimulation by PDGF or angiotensis II are also absent in p47phox-deficient mice, indicating involvement of a phagocyte-like oxidase in vascular tissue. We have also identified a distinct gp91phox homologue in the kidney (renal oxidase or Renox) that is expressed in proximal convoluted tubules and proposed to serve as an oxygen sensor regulating erythropoietin synthesis. When expressed in transfected fibroblasts, Renox causes superoxide release and induces cellular senescence. Finally, a colon oxidase homologue has been characterized (Mox-1), which is expressed on epithelial surfaces and induced by lipopolysaccharide, interferon-gamma or terminal differentiation, and may function in host defense or inflammatory responses in the gut.
Showing the most recent 10 out of 41 publications
