Oxidized LDL is detected at inflammatory loci and activates proinflammatory cytokine release from monocytes and macrophages. Extensively oxidized LDL also can induce acute inflammation in vivo. The investigators propose to define further the physiologic significance and mechanism of these observations. To do so: (1) The investigators will define the properties of oxidized LDL in inflammatory human synovial fluids (SFs). To determine the nature and extent of oxidative LDL modification in human SFs the investigators will evaluate SF LDLs for apo B degradation, the degree of apo B lysyl residue modification, enhanced recognition by macrophages, and the ability to complete in solid-phase RIAs for three epitopes present in oxidized LDL: (malondialdehyde(MDA)-lysine, 4-hydroxynonenal(HNE)-lysine, and a unique apo B epitope). Certain new apo B epitopes in oxidized LDL are highly immunogenic, e.g., autoantibodies (autoAbs) to MDA-lysine are commonly detected in vivo. Thus, the investigators will quantify such autoAbs in SFs. The effects of the isolated autoAbs on phagocyte activation and inflammation induced by oxidized LDL will then be assessed, as described below. Last, the investigators will determine the extent of modification of oxidized LDL required for it to be proinflammatory in a rat model of acute synovitis (injection into subcutaneous air pouches). (2) The investigators will define the mechanism of mononuclear phagocyte activation by oxidized LDL. The investigators will determine the extent of modification of SF LDL required to induce proinflammtory cytokines (IL-1, and the major neutrophil chemotaxin, IL-8) in freshly isolated adherent human monocytes, well-differentiated monocyte-derived macrophages (MDM), and interferon-activated MDM. The modified apo B of oxidized LDL appears to be recognized both by a scavenger receptor and a putative oxidized LDL receptor on macrophages. Thus, the investigators will define the potential ability of the following to induce these cytokines, and cytosolic calcium mobilization, or to inhibit the ability of oxidized LDL to do so: (i) Other scavenger receptor ligands (fucoidin, acetylated(Ac)-LDL, and Ac-albumin); (ii) oxidized forms of LDL that appear to bind both, or neither receptor (Ac-LDL, and reductively methylated LDL, respectively); (iii) delipidated apo B extracted from modified LDLs. Last, the investigators will assess the potential roles in activation of polar and neutral lipids extracted from oxidized LDL, of receptor-mediated endocytosis and lysosomal degradation of oxidized LDL, and of oxidized LDL-induced enhancement of arachidonate metabolism in monocyte/macrophages.
