Lipoxygenase treatment render low-density lipoprotein susceptible to Cu2+-catalysed oxidation

Abstract

Oxidative modification of low-density lipoprotein (LDL) has been implicated in foam-cell formation at all stages of atherosclerosis. Since transition metals and mammalian 15-lipoxygenases are capable of oxidizing LDL to its atherogenic form, a concerted action of these two catalysts in atherogenesis has been suggested. Cu2+-catalysed LDL oxidation is characterized by a kinetic lag period in which the lipophilic antioxidants are decomposed and by a complex mixture of unspecific oxidation products. We investigated the kinetics of the 15-lipoxygenase-catalysed oxygenation of LDL and found that the enzyme is capable of oxidizing LDL in the presence of the endogenous lipophilic antioxidants. In contrast with the Cu2+-catalysed reaction, no kinetic lag phase was detected. The pattern of products formed during short-term incubations was highly specific, with cholesterol-esterified (13S)-hydroperoxy-(9Z,11E)-octadecadinoic acid being the major product. However, after long-term incubations the product pattern was less specific. Preincubation with 15-lipoxygenase rendered human LDL more susceptible to Cu2+-catalysed oxidation as indicated by a dramatic shortening of the lag period. Addition of Cu2+ to lipoxygenase-treated LDL led to a steep decline in its antioxidant content and to a greatly reduced lag period. Interestingly, if normalized to a comparable hydroperoxide content, autoxidation and addition of exogenous hydroperoxy fatty acids both failed to overcome the lag period. The local peroxide concentrations in various LDL subcompartments will be discussed as a possible reason for this unexpected behaviour.

Inhibitory Effect of Flavonoids on Low‐Density Lipoprotein Peroxidation Catalyzed by Mammalian 15‐Lipoxygenase

“…2). This result suggests poor inhibiting ability of a -tocopherol toward 15-LO-induced LDL oxidation, as previously suggested (27). The addition of 1 l M ascorbic acid before the onset of LDL oxidation substantially inhibited the lipid peroxidation while it was present in the system, that is, in the rst 3 h of incubation.…”

Oxidation of Free Fatty Acids in Low Density Lipoprotein by 15-Lipoxygenase Stimulates Nonenzymic, α-Tocopherol-mediated Peroxidation of Cholesteryl Esters

“…In support of this, addition of 18:2 to LDL previously exposed to rhLO for 60 min failed to result in further accumulation of 13-(S)(Z,E)-H(P)ODE. 3 Earlier studies noted that LDL lipid oxidation induced by various 15-LO did not display this characteristic self-inactivation (11)(12)(13)(14). It is well established that FFA peroxyl radicals are released from the active site of 15-LO during enzymic formation of 13-(S)(Z,E)-HPODE (21,38) and that this can lead to the co-oxidation of various compounds, including phenolic antioxidants, PL, cholesterol, and proteins (37,39).…”

“…Much of the literature ascribes oxidation of LDL by 15-LO to a direct reaction with esterified fatty acid substrates, including phospholipids (PL) and cholesteryl esters (CE) (11)(12)(13)(14). In LDL exposed to 15-LO, or to cells overexpressing this enzyme (15,16), the major oxidation products found are CE hydroperoxides and hydroxides (collectively referred to as CE-O(O)H).…”

“…However, the vast majority of CE are normally buried in the core of the lipoprotein particle and hence may be largely inaccessible to 15-LO. In addition, certain aspects of 15-LO catalysis, such as the rapid oxidation and characteristic suicidal inactivation of the enzyme observed with free fatty acid (FFA) substrates, are notably absent from, or distinct to, 15-LO-induced LDL oxidation (11)(12)(13)(14).…”

Kinetic study of low density lipoprotein oxidation by copper

“…In agreement with many previous studies (5, 10, 15), we found that at low Concentrations effect of 0.5 P P P P PM (A), 2.5 P P P P PM (B), 10 P P P P PM (C), 20 P P P P PM (D) and 50 P P P P PM (E) copper on maximal rates during first propagation (R 1 ) and second propagation (R 2 ) of 0.1 P P P P PM LDL Fig 9: Effect of 0.5 to 50 P P P P PM copper on the extent of 0.1 P P P P PM LDL oxidation by determine maximal conjugated diene value (CD max ) (A) and conjugated diene value at lag-time (CD lag ) (B) molar ratios of copper/LDL (at least 25), propagation of oxidation occurs during a long initial inhibited period (lag-time). The increase in conjugated diene during the propagation phase was reported to be mainly due to the formation of cholesteryl linoleate hydroperoxides (16). The rate of this type of propagation is characterized by a symmetrical profile.…”