Mildly Oxidized LDL Induces Platelet Aggregation Through Activation of Phospholipase A ₂

Abstract

Native LDL and LDL oxidized under various conditions were compared in terms of their ability to activate platelets. Native LDL did not induce platelet shape change or aggregation, even at high concentrations (2 mg protein/mL). LDL was mildly oxidized with either CuSO4 (mox-LDL) or 3-(N-morpholino)sydnonimine (SIN-1-LDL). Analysis of mox-LDL and SIN-1-LDL showed a small increase of dienes (E234nm from 0.28 +/- 0.04 to 0.55 +/- 0.09, mean +/- SD) and thiobarbituric acid-reactive substance (from 0 to 10.6 +/- 1.5 nmol/mg, mean +/- SEM), no change in apo B electrophoretic mobility, and a minor (12% to 30%) decrease in polyunsaturated fatty acid content. Interestingly, this small oxidative modification of LDL dramatically changed its effect on platelets. Irreversible aggregation and secretion were induced by a threshold concentration of 0.4 mg protein/mL. In contrast, LDL thoroughly oxidized with CuSO4 (ox-LDL) did not aggregate platelets. Although mox-LDL was depleted in antioxidants (alpha- and gamma-tocopherol, alpha- and beta-carotene, and other carotenoids), incubation of mox-LDL with exogenous alpha-tocopherol did not reverse its ability to induce platelet aggregation and secretion. Preincubation of platelets with the cyclooxygenase inhibitor aspirin or the phospholipase A2 inhibitors trifluoperazine, quinacrine, 4-bromophenacyl bromide, and propranolol completely prevented platelet aggregation and secretion caused by mox-LDL or SIN-1-LDL. These results indicate that mildly oxidized LDL activates platelets through a phospholipase A2/cyclooxygenase-dependent pathway. The complete inhibition of mox-LDL-induced platelet aggregation by aspirin could contribute to its beneficial effect in cardiovascular disease.

Two-photon microscopy of aorta fibers shows proteolysis induced by LDL hydroperoxides

“…In our short-term incubation of fresh aorta rings, we used LDL with a limited amount of lipid hydroperoxides, Յ 30 nmol/mg of cholesterol, a concentration 10 -20 times lower than in copper-oxidized LDL, usually referred to as ox-LDL [32], and in the "minimally oxidized LDL" obtained by storage for 2-4 months [26]. Notably, the level of lipid hydroperoxides in our minimally oxidized LDL is below the detection limit of usual analytical tests, so that unless a chemiluminescence method is used, they would be considered as native "nonoxidized" LDL.…”

Low-Density Lipoprotein Enhances Platelet Secretion Via Integrin-α IIb β 3 –Mediated Signaling

“…Even after 30 minutes of LDL-platelet contact, the release of 14 C-serotonin was the same as in LDL-free suspensions (Ͻ5% of maximal). Also, Weidtmann et al 49 reported that native LDL failed to induce dense-granule secretion. However, Kaplan et al 50 reported that ␣-granule secretion precedes dense-granule secretion at low concentrations of thrombin, collagen, and Ca 2ϩ -ionophore, resulting in release of minor amounts of granular fibrinogen.…”

HDL abnormalities in nephrotic syndrome and chronic kidney disease

“…Furthermore, increased production of nitric oxide by HDL inhibits platelet activation and adhesion. Finally, activation of phospholipase A2 by oxidized LDL promotes platelet aggregation and adhesion 30 ; therefore, reversal and/or prevention of LDL oxidation by normal HDL prevents oxidized-LDL-mediated platelet activation 31 (FIG. 2).…”