Effect of KO on hepatic fatty acid oxidation.

<p>(A) CPT I activity was measured in liver mitochondria freshly isolated from rats at the times indicated. The values are expressed as nanomoles of DTNB reduced min⁻¹<b>·</b>mg protein⁻¹ and were calculated as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038797#s4" target="_blank">Methods</a> section. (B) Liver carnitine levels were also determined at the times indicated. Data are means ± SD (<i>n</i> = 4). *<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet.</p

Effect of KO on plasma triglycerides.

<p>The levels of plasma triglycerides were determined at the times indicated, using commercial kits. The values reported in the figure represent the means ± SD (<i>n</i> = 4). **<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet.</p

Effect of KO on lipogenic enzyme activities.

<p>The activities of ACC (A) and FAS (B) were measured in the cytosol of rat hepatocytes at the times indicated. The values are expressed as nanomoles of NADH (ACC) or NADPH (FAS) oxidized min⁻¹ mg protein⁻¹ and represent the means ± SD (<i>n</i> = 4). *<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet.</p

Effect of KO on body weight.

<p>Body weights of rats fed control (filled circle), HF (open circle) and HF+KO (filled square) diets are indicated for the treatment periods in weeks. Each point represents the mean ± SD for 10 animals. *<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet.</p

Composition of diets (%).

<p>The control group of animals received a standard diet (Global Diet 2018S from Harlan Teklad). The HF group received a diet with 35% fat (Diet TD.03584 from Harlan Teklad) and the KO group was fed with the above reported HF diet supplemented with 2.5% KO. Fatty acids were extracted from the three diets and analyzed by gas-liquid chromatography.</p

Mitochondrial respiratory efficiency.

<p>Respiratory control ratio (RCR) was calculated as the ratios of the rate of oxygen uptake in the presence of added ADP (V₃) to the rate observed when added ADP had been completely phosphorylated to ATP (V₄).</p>*<p><i>P</i><0.05 <i>vs.</i> rats fed control diet;</p>#<p><i>P</i><0.05 <i>vs.</i> rats fed HF diet; <i>n</i> = 3.</p

Km and Vmax of citrate transport in a reconstituted system.

<p>Km and Vmax values were measured in a reconstituted system at the times indicated. Proteoliposomes were reconstituted with the CIC as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0038797#s4" target="_blank">Methods</a> section. [^l4C] Citrate, 0.04–0.40 mM, was added to proteoliposomes containing 10 mM citrate. The citrate/citrate exchange was stopped 1 min after the addition of the radiolabeled substrate by 20 mM 1,2,3-BTA. Km and Vmax values were calculated by linear regression.</p

Liver metabolic pathways influenced by the addition of KO to a HF diet.

<p>A red X symbolizes inhibition; a green triangle symbolizes stimulation. OAA, oxaloacetate.</p

Effect of KO on liver lipids.

<p>(A) Cresyl violet staining of liver histological sections from rats fed for 12 weeks with control, HF or HF+KO diet. The levels of liver triglycerides (B) and cholesterol (C) were determined at the times indicated. Each point represents the mean ± SD for 4 liver samples. *<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet.</p

Effect of KO on protein levels of mitochondrial CIC.

<p>Liver mitochondrial proteins from control, HF or HF+KO-fed rats were separated by SDS-PAGE, transferred to nitrocellulose and then immunodecorated with antisera against either the rat CIC or the mammalian porin. The values reported in the graph represent the means ± SD (<i>n</i> = 4; *<i>P</i><0.05 <i>vs.</i> rats fed control diet; ^#<i>P</i><0.05 <i>vs.</i> rats fed HF diet). The amount of CIC revealed by immunodecoration at the beginning of dietary treatment was set to 100%.</p