n-6 PUFAs downregulate expression of the tricarboxylate carrier in rat liver by transcriptional and posttranscriptional mechanisms.

The tricarboxylate (citrate) carrier (TCC), a protein of the mitochondrial inner membrane, is an obligatory component of the shuttle system by which mitochondrial acetyl-CoA is transported into the cytosol, where lipogenesis occurs. The aim of this study was to investigate the molecular basis for the regulation of TCC gene expression by a high-fat, n-6 PUFA-enriched diet. Rats received for up to 4 weeks a diet enriched with 15% safflower oil (SO), which is high in linoleic acid (70.4%). We found a gradual decrease of TCC activity and a parallel decline in the abundance of TCC mRNA, the maximum effect occurring after 4 weeks of treatment. At this time, the estimated half-life of TCC mRNA was the same in the hepatocytes from rats on both diets, whereas the transcriptional rate of TCC mRNA, tested by nuclear run-on assay, was reduced by approximately 38% in the rats on the SO-enriched diet. The RNase protection assay showed that the ratio of mature to precursor RNA, measured in the nuclei, decreased with the change to the n-6 PUFA diet. These results suggest that administration of n-6 PUFAs to rats leads to changes not only in the transcriptional rate of the TCC gene but also in the processing of the nuclear precursor for TCC RNA.

Covariance of tricarboxylate carrier activity and lipogenesis in liver of polyunsaturated fatty acid (n-6) fed rats.

The mitochondrial tricarboxylate (citrate) carrier plays an important role in hepatic intermediary metabolism because, among other functions, it supplies the cytosol with acetyl units for fatty-acid synthesis. In this study, the effect of polyunsaturated fatty acids (PUFA, n-6) on the function of this mitochondrial transporter and on lipogenic enzyme activities was investigated by feeding rats for 4 weeks with a 15%-fat diet composed of high linoleic safflower oil. Citrate transport was strongly reduced in liver mitochondria isolated from PUFA-treated rats. A reduced transport activity was also observed when solubilized mitochondrial citrate carrier from PUFA-treated rats was reconstituted into liposomes. In the same animals, a decrease of cytosolic lipogenic enzyme activities was observed. These results indicate a coordinated modulation of citrate carrier and of lipogenic enzyme activities by PUFA feeding. Kinetic analysis of the carrier activity showed that only V(max) decreased, whereas K(m) was almost virtually unaffected. The PUFA-mediated effect is most likely due to the reduced mRNA level and lower content of the citrate carrier protein observed in the safflower oil-fed rats.

Oleic acid inhibits endothelial activation : A direct vascular antiatherogenic mechanism of a nutritional component in the mediterranean diet.

Because oleic acid is implicated in the antiatherogenic effects attributed to the Mediterranean diet, we investigated whether this fatty acid can modulate endothelial activation, ie, the concerted expression of gene products involved in leukocyte recruitment and early atherogenesis. We incubated sodium oleate with human umbilical vein endothelial cells for 0 to 72 hours, followed by coincubation of oleate with human recombinant tumor necrosis factor, interleukin (IL)-1alpha, IL-1beta, IL-4, Escherichia coli lipopolysaccharide (LPS), or phorbol 12-myristate 13-acetate for a further 6 to 24 hours. The endothelial expression of vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and intercellular adhesion molecule-1 was monitored by cell surface enzyme immunoassays or flow cytometry, and steady-state levels of VCAM-1 mRNA were assessed by Northern blot analysis. At 10 to 100 micromol/L for >24 hours, oleate inhibited the expression of all adhesion molecules tested. After a 72-hour incubation with oleate and a further 16-hour incubation with oleate plus 1 microg/mL LPS, VCAM-1 expression was reduced by >40% compared with control. Adhesion of monocytoid U937 cells to LPS-treated endothelial cells was reduced concomitantly. Oleate also produced a quantitatively similar reduction of VCAM-1 mRNA levels on Northern blot analysis and inhibited nuclear factor-kappaB activation on electrophoretic mobility shift assays. Incubation of endothelial cells with oleate for 72 hours decreased the relative proportions of saturated (palmitic and stearic) acids in total cell lipids and increased the proportions of oleate in total cell lipids without significantly changing the relative proportions of polyunsaturated fatty acids. Although less potent than polyunsaturated fatty acids in inhibiting endothelial activation, oleic acid may contribute to the prevention of atherogenesis through selective displacement of saturated fatty acids in cell membrane phospholipids and a consequent modulation of gene expression for molecules involved in monocyte recruitment.