Isolation and characterization of a complement-activating lipid extracted from human atherosclerotic lesions.

Abstract

SummaryThe major characteristics of human atherosclerotic lesions are similar to those of a chronic inflammatory reaction, namely fibrosis, mesenchymal cell proliferation, the presence of resident macrophages, and cell necrosis. Atherosclerosis exhibits in addition the feature of lipid (mainly cholesterol) accumulation . The results ofthe present report demonstrate that a specific cholesterolcontaining lipid particle present in human atherosclerotic lesions activates the complement system to completion. Thus, lipid could represent a stimulatory factor for the inflammatory reaction, whose underlying mechanistic basis may be, at least in part, complement activation . The complement-activating lipid was purified from saline extracts of aortic atherosclerotic lesions by sucrose density gradient centrifugation followed by molecular sieve chromatography on Sepharose 2B. It contained little protein other than albumin, was 100-500 nm in size, exhibited an unesterified to total cholesterol ratio of 0.58 and an unesterified cholesterol to phospholipid ratio of 1.2. The lipid, termed lesion lipid complement (LCA), activated the alternative pathway ofcomplement in a dose-dependent manner. Lesion-extracted low density lipoprotein (LDL) obtained during the purification procedure failed to activate complement. Specific generation of C3a desArg and C5b-9 by LCA indicated C3/C5 convertase formation with activation proceeding to completion. Biochemical and electron microscopic evaluations revealed that much of the C5b-9 present in atherosclerotic lesions is membraneous, rather than fluid phase SC5b-9 . The observations reported herein establish a link between lipid insudation and inflammation in atherosclerotic lesions via the mechanism of complement activation. therosclerosis shares in common with chronic inflamma-1A tion the features ofleukocyte infiltration, fibrosis, mesenchymal cell proliferation, and tissue necrosis . Complement plays a central role in many inflammatory and immune diseases, and most of the complement components have now been identified in human atherosclerotic lesions (1) . Of particular importance is the presence of C5b-9 terminal complement complexes since their assembly indicates that complement activation with C3/C5 convertase formation has taken place. The respective convertases cleave C3 and C5 generating the anaphylatoxins C3a and C5a, which in turn mediate proinflammatory leukocyte functions (2) . The C5b-9 complex, in addition to its potential cytolytic effect, can influence cell physiology in sublytic doses via activation of various calcium-dependent pathways (3) .In a previous report, we demonstrated a temporal and spatial colocalization of C5b-9 complexes and lipid in the aortic tunica intima of cholesterol-fed rabbits (4) . Lipid vesicles rich in unesterified cholesterol are an early and consistent feature of human and animal atherosclerotic lesions (5-11) . In vitro, liposomes containing a >50 mole percent of unesterified cholesterol activate complement to completion (12, 13) . We therefore su...

Enzymatically Modified Low‐Density Lipoprotein Is Present in All Stages of Aortic Valve Sclerosis: Implications for Pathogenesis of the Disease

“…The concept that we have been pursuing proposes extracellular enzymatic degradation of LDL to represent a key step leading to generation of an atherogenic lipoprotein 7, 10, 11, 30. eLDL generated by all tested proteases is endowed with the same atherogenic properties as the originally described trypsin‐generated eLDL 23, 31.…”

Immunohistochemical Demonstration of Enzymatically Modified Human LDL and Its Colocalization With the Terminal Complement Complex in the Early Atherosclerotic Lesion

“…Third, lipid moieties possessing the capacity to spontaneously activate the alternative complement pathway have been isolated from human atherosclerotic lesions. 35 Collectively designated LCA, these lipid droplets ultrastructurally resemble lipoprotein derivatives that had been isolated from atherosclerotic plaques in other laboratories. 30 -33 The lesion lipids contain relatively large amounts of free cholesterol, 30,32,33,[35][36][37] whereas the bulk of cholesterol in both LDL and ox-LDL is esterified.…”

“…35 Collectively designated LCA, these lipid droplets ultrastructurally resemble lipoprotein derivatives that had been isolated from atherosclerotic plaques in other laboratories. 30 -33 The lesion lipids contain relatively large amounts of free cholesterol, 30,32,33,[35][36][37] whereas the bulk of cholesterol in both LDL and ox-LDL is esterified. Deesterification is probably important in generating the complementactivating property of LCA, which is not shared by native LDL or ox-LDL.…”

Complement C6 Deficiency Protects Against Diet-Induced Atherosclerosis in Rabbits

“…Heterogeneously sized lipid droplets containing high amounts of free cholesterol were isolated from early lesions and were shown to be capable of spontaneously activating the alternative complement pathway. 8 The origin of this lipid, termed the lesion complement activator (LCA), was unknown, but the possibility that it represented an LDL derivative was obvious. To corroborate this assumption, attempts were undertaken to transform LDL in vitro into a complement-activating moiety.…”

“…Its complement-activating properties were possibly due to the high content of free cholesterol, as also found in LCA. 8,9 It has been known for 20 years that nonesterified cholesterol spontaneously activates the alternative complement pathway, 12,13 and that neuraminic acid restricts such activation. 14 Hence, a simple model for the creation of a complement-activating lipid via enzymatic degradation of LDL emerged.…”