Mechanisms involved in the in vitro modification of low density lipoprotein by human umbilical vein endothelial cells and copper ions

In this study we evaluated the time course and mechanism of low density lipoprotein (LDL) oxidation induced by human umbilical vein endothelial cells (HUVECs), cell-free medium (CFM) and Cu2+. After incubating LDL (200 mu g/ml) with HUVECs, CFM and Cu2+ (concentration adjusted to obtain the same degree of LDL modification as with HUVECs), the extent of LDL lipid peroxidation and apoprotein B modification was monitored at different times from 0 to 24 h, This involved evaluating the time course of LDL conjugated diene, peroxide, malonyldialdehyde (MDA), fluorescence, relative electrophoretic mobility (REM), vitamin E and monounsaturated and polyunsaturated fatty acids. After incubation with HUVECs, the LDL REM was significantly higher than that obtained in CFM (p < 0.01). When balanced for the same degree of LDL modification as obtained with HUVECs, Cu2+ gave a REM similar to that obtained with HUVECs. At the different times of incubation there was no statistical difference between conjugated diene and peroxide values after incubation with HUVECs and with CFM. The values obtained with Cu2+ were significantly higher than those obtained with HUVECs and CFM (p < 0.01). MDA and LDL fluorescence were significantly higher after exposure to HUVECs than to CFM (p < 0.01), values being similar to those obtained with Cu2+. There was no statistical difference between the values of LDL oleic, linoleic, arachidonic and eicosapentaenoic acids after incubation with HUVECs and CFM. Eicosatetraynoic acid (ETYA), a lipoxygenase inhibitor, determined dose-dependent reduction of MDA formation induced by the incubation of LDL with HUVECs; it did not affect LDL conjugated diene. ETYA did not have any effect on the MDA derived from LDL after incubation with Cu2+ or CFM. The results of this study demonstrate that, unlike Cu2+, the contribution of HUVECs to LDL modification does not involve only lipid peroxidation of the lipoprotein; it also includes intracellular radical and non-radical processes.