There is strong experimental evidence that oxidized low density lipoprotein (Ox-LDL) plays an important role in atherosclerosis. However, the mechanisms by which Ox-LDL is formed in vivo are unknown. To test whether 15-lipoxygenase (15-LO) could play a role in oxidation of LDL by cells, we expressed 15-LO activity in murine fibroblasts, which do not normally have 15-LO activity, and tested their ability to modify LDL. Using a retroviral vector, we prepared fibroblasts that expressed 2- to 20-fold more 15-LO activity than control fibroblasts infected with a vector containing beta-galactosidase (lacZ). Compared with LDL incubated with lacZ cells, LDL incubated with 15-LO-containing cells were enriched with lipid hydroperoxides. When these LDL samples were subsequently subjected to oxidative stress, they were more susceptible to further oxidative modification, as judged by increased conjugated diene formation and by increased ability to compete with 125I-Ox-LDL for uptake by macrophages. These findings establish that cellular 15-LO can contribute to oxidative modification of LDL, but the quantitative significance of these findings to the in vivo oxidation of LDL remains to be established.

Apolipoprotein E3-Leiden (APOE*3-Leiden) transgenic mice have been studied to identify factors modulating chylomicron and VLDL remnant lipoprotein metabolism. Transient elevated levels of VLDL/LDL-sized lipoproteins occurred in these mice with maximal levels during the period of rapid growth (optimum at 45 d of age). After about 100 d of age, serum cholesterol and triglyceride levels stabilized to slightly elevated levels as compared to control mice. The expression of the APOE*3-Leiden transgene was not age-dependent. In young mice the in vivo hepatic production of VLDL-triglycerides was 50% increased as compared to older mice. This is sustained by in vivo VLDL-apo B turnover studies showing increased (75%) VLDL-apo B secretion rates in young mice, whereas the VLDL-apo B clearance rate appeared not to be age dependent.On a high fat/cholesterol diet, females displayed significantly higher cholesterol levels than males (10 versus 7.0 mmol/liter, respectively). Serum levels of VLDL/LDL sized lipoproteins increased upon administration of estrogens, whereas administration of testosterone gave the opposite result. As compared to male mice, in female mice the hepatic VLDL-triglyceride production rate was significantly elevated. Injection of estrogen in males also resulted in increased VLDL-triglyceride production, although not statistically significant. In vivo VLDL-apo B turnover experiments showed that the VLDL secretion rate tended to be higher in females. Although, the fractional catabolic rate of VLDLapo B is not different between males and females, administration of estrogens in males resulted in a decreased clearance rate of VLDL, whereas administration of testosterone in females resulted in an increased clearance rate of VLDL. The latter presumably due to an inhibiting effect of testosterone on the expression of the APOE*3-Leiden transgene.We conclude that hyperlipidemia in APOE*3-Leiden transgenic mice is strongly affected by age via its effect on hepatic VLDL production rate, whereas gender influences hyperlipidemia by modulating both hepatic VLDL production and clearance rate. ( J. Clin. Invest. 1996. 97:1184-1192.)

To evaluate the role of both oxidation and inflammation in atherosclerosis, we compared LDL oxidizability, in vivo lipid and cholesterol oxidation, and basal and lipopolysaccharide (LPS)-stimulated production of various cytokines in normolipidemic patients with diabetes mellitus (DM; n = 11), cigarettes smokers (n -12) and controls (// «= 14). In addition, the effects of vitamin E (600 I.U./day for 4 weeks) on these parameters were evaluated. Initial LDL oxidation characteristics before and after vitamin E were identical in the 3 groups. Plasma thiobarbituric acid reactive substances were higher in DM and smokers versus controls (0.77 ± 0.22, 0.74 ±0.14 versus 0.62 ±0.10 //mol malondialdehyde equivalents/1, respectively; P versus controls <0.05) and normalized after vitamin E supplementation. Total plasma oxysterols were higher in smokers versus controls (354 ± 104 versus 265 ± 66 nmol/1, P < 0.05) and unaffected by vitamin E. The basal and LFS-stimulated levels of interleukin-1// and tumour necrosis factor a (TNFa) and the basal level of interleukin-1-receptor antagonist (IL-1RA) were identical fot LPS-stimulated IL-1RA was higher in DM versus controls (10.7 ± 2.0 versus 8.1 ± 1.7 pmol/1, P <0.05). After in controls and smokers, and IL-1RA in smokers only. Results suggest increased in vivo t•" ess «. md inflammation in DM and smoking, which is partly overcome by vitamin E.

Dutch Diabetes Research Foundation (Diabetes Fonds).

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