ULTRAVIOLET-TREATED LIPOPROTEINS AS A MODEL SYSTEM FOR THE STUDY OF THE BIOLOGICAL EFFECTS OF LIPID PEROXIDES ON CULTURED-CELLS .3. THE PROTECTIVE EFFECT OF ANTIOXIDANTS (PROBUCOL, CATECHIN, VITAMIN-E) AGAINST THE CYTOTOXICITY OF OXIDIZED LDL OCCURS IN 2 DIFFERENT WAYS

Comparison of the protective effect of three antioxidants (from three different chemical classes) against cell injury due to LDL oxidation, allowed us to clearly discriminate between two different lines of defence. The ultraviolet-induced lipid peroxidation of LDL was strongly inhibited by 10-mu-mol/l catechin and 25-mu-mol/l probucol, but only poorly by 100-mu-mol/l vitamin E. The ultraviolet-treated LDL protected by catechin or probucol (i.e. LDL irradiated by ultraviolet in the presence of effective concentrations of antioxidants inhibiting the lipid peroxidation) were much less 'cytotoxic' than unprotected ultraviolet-treated LDL. In contrast, LDL treated by ultraviolet in the presence of 100-mu-mol/l vitamin E were 'cytotoxic' similarly to unprotected LDL. The level of 'cytotoxicity' of LDL treated by ultraviolet in the presence of antioxidants (protected ultraviolet-treated LDL) was well correlated with their content in lipid peroxidation markers. Therefore these markers can be useful for predicting the 'cytotoxicity' of oxidized LDL and subsequently the protective effect of the tested antioxidants. The 'cytotoxicity' of unprotected ultraviolet-treated LDL (i.e. LDL irradiated by ultraviolet in the absence of exogenous antioxidant) can be effectively blocked by preincubation of the cells with antioxidants. Catechin (10-mu-mol/l) and vitamin E (100-mu-mol/l) are very effective cytoprotective agents, whereas probucol (up to 50-mu-mol/l) was completely ineffective under these experimental conditions. The cytoprotective effect of vitamin E was associated to a complete inhibition of the cellular TBARS formation induced by ultraviolet-treated LDL, whereas the cytoprotective effect of catechin was relatively independent on the TBARS inhibition. All these results showed that: (1) probucol (25-mu-mol/l) is very effective to prevent lipid peroxidation of LDL and their subsequent 'cytotoxicity', but it cannot protect cells against the 'cytotoxicity' of previously oxidized LDL; (2) vitamin E (100-mu-mol/l) prevents poorly the ultraviolet-induced lipid peroxidation of LDL, but is able to block simultaneously the cellular oxidative stress and the 'cytotoxicity' induced by previously oxidized LDL; and (3) catechin (10-mu-mol/l) exhibited two types of protective effects: it inhibits the lipid peroxidation of LDL (and their subsequent 'cytotoxicity') and very effectively protects the cells against 'toxicity' of previously oxidized LDL (with only little inhibition of the cellular oxidative stress). These results demonstrate that protective agents can constitute two lines of defence (at least) in the prevention of cell injury induced by LDL oxidation: the first line consists of the prevention of LDL oxidation (inhibition of cytotoxic compounds formation and prevention of the subsequent 'cytotoxicity'); the second one consists of direct protection of cells against the 'cytotoxic' effect of lipid peroxidation products contained in the ultraviolet-treated LDL. The use of drugs protecting LDL and cells against the oxidative stress, is discussed in view of obtaining better prevention of the 'cytotoxicity' of oxidized LDL and to slow down the atherosclerotic process.