MATERIALS AND METHODS

Adult males of a hereditary HTg line, weighing 280–310 g, were used. Randombred males of the Wistar strain of a similar weight served as controls. All rats were fed a high-carbohydrate diet (70 cal% sucrose, 20 cal% protein, 10 cal% fat) ad libitum. After 3 weeks, the rats were killed by decapitation and nonfasted blood samples were collected into tubes containing Na 2 EDTA, centrifuged, aliquoted, and immediately assayed for conjugated diene concentration. Lipoproteins (LDL+ VLDL) were isolated by ultracentrifugation at 112,000 g for 20 h at 10 C with a final density of 1.055 g/mL. 10, 11 LDL+ VLDL samples were dialyzed against oxygen-free buffer at 4 C in the dark for 24 h. The susceptibility of the lipoprotein fractions to Cu++-induced oxidation in vitro was measured by continuous monitoring of the conjugated diene (CD) formation as described by Kleinveld et al., 12 with the difference that the incubation of lipoproteins was performed at 37 C, and by measuring the production of thiobarbituric-reactive substances (TBARS) as described by Thomas et al. 13 Oxidative modification of the protein moiety of the lipoproteins was measured by the emission fluorescence spectra at 430 nm (excitation, 360 nm) according to Esterbauer et al. 14 Protein concentration of the lipoproteins was determined by the method of Lowry modified to include sodium dodecyl sulfate. 15 Plasma lipid peroxide concentration was assessed by measurement of CD 16 and by fluorometric measurement of TBARS 17 with fluorometric detection (515–553 nm). Alpha-tocopherol was estimated by HPLC analysis. 18 The serum levels of triglycerides and FFA were measured by enzymatic kits (Lachema, Czech Republic, and Boehringer-Mannheim, Germany, respectively). Aortic triglycerides, cholesterol (measured enzymatically with kits), and CD concentrations were determined in lipid extracts of the aorta.