Inflammation and macromolecule oxidation are inter-related processes that are major contributors to atherogenesis. An important role of Cp in inflammation is suggested by its increased plasma level during the acute phase reaction and by its synthesis by activated macrophages in sites of inflammation. Clinical studies have shown that elevated plasma Cp is a significant risk factor for atherosclerosis, restenosis after endarterectomy, and myocardial infarction. In view of our finding that Cp copper induces oxidative modification of LDL in vitro, Cp may represent an important molecular link connecting inflammation, oxidation, and atherosclerosis. In new preliminary studies, we show that Cp binds with high affinity to LDL, and that the interaction is required for LDL oxidation by Cp. Other preliminary evidence indicates that Cp binds LDL in human plasma, and that binding to LDL may be higher in patients with cardiovascular disease. Studies with epitope-specific antibodies to Cp localized the interaction site to the second major sequence domain of Cp. These structural studies may be relevant to human inflammatory diseases, e.g., atherosclerosis, since coding-altering, single nucleotide polymorphisms (SNPs) have been identified in the human Cp gene, and two are adjacent to the Cp/LDL binding site. Preliminary studies in mice with targeted deletion of the Cp gene suggests that Cp increases oxidative modification of proteins and lipids in inflammatory sites in vivo. From previous work and our new preliminary results we propose the following hypothesis: That in sites of inflammation, macrophage-derived Cp binds to LDL and causes specific copper ion-mediated oxidative modification of the protein and lipid components. We further propose that by defining the specific Cp domains and amino acids required for pro- and anti-inflammatory activities, recombinant, monofunctional Cp can be generated that can test the function of the individual activities in vivo. Finally, we propose that human Cp containing SNPs in these functional domains alter the inflammatory properties of Cp, and also the risk for atherosclerosis in humans. We will test this hypothesis by pursuing the following Specific Aims:
Aim 1. Determine the structural requirements for pro- and anti-inflammatory Cp activities; we will analyze structural determinants of Cp required for LDL oxidase, ferroxidase, and nitrosation activities, and for Cp interaction with LDL.
Aim 2. Investigate the role of Cp in oxidation during inflammatory processes in vivo. Cp-null mice will be subjected to sepsis and peritonitis models of inflammation, and specific lipid and protein oxidation products in peritoneal lavage and plasma will be measured by LC-mass spectrometry These studies will elucidate, at the molecular, animal, and patient level, the role of Cp activities in inflammation.
Showing the most recent 10 out of 271 publications
