Endre Dyrøy

Eicosapentaenoic and Docosahexaenoic Acid Affect Mitochondrial and Peroxisomal Fatty Acid Oxidation in Relation to Substrate Preference

Decreased triacylglycerol synthesis within hepatocytes due to decreased diacylglycerol acyltransferase (DGAT) activity has been suggested to be an important mechanism by which diets rich in fish oil lower plasma triacylglycerol levels. New findings suggest that eicosapentaenoic acid (EPA), and not docosahexaenoic acid (DHA), lowers plasma triacylglycerol by increased mitochondrial fatty acid oxidation and decreased availability of fatty acids for triacylglycerol synthesis. To contribute to the understanding of the triacylglycerol-lowering mechanism of fish oil, the different metabolic properties of EPA and DHA were studied in rat liver parenchymal cells and isolated rat liver organelles. EPA-CoA was a poorer substrate than DHA-CoA for DGAT in isolated rat liver microsomes, and in the presence of EPA, a markedly lower value for the triacyl[3H]glycerol/diacyl[3H]glycerol ratio was observed. The distribution of [1-14C]palmitic acid was shifted from incorporation into secreted glycerolipids toward oxidation in the presence of EPA (but not DHA) in rat liver parenchymal cells. [1-14C]EPA was oxidized to a much greater extent than [1-14C]DHA in rat liver parenchymal cells, isolated peroxisomes, and especially in purified mitochondria. As the oxidation of EPA was more effective and sensitive to the CPT-I inhibitor, etomoxir, when measured in a combination of both mitochondria and peroxisomes, we hypothesized that both are involved in EPA oxidation, whereas DHA mainly is oxidized in peroxisomes. In rats, EPA treatment lowered plasma triacylglycerol and increased hepatic mitochondrial fatty acid oxidation and carnitine palmitoyltransferase (CPT)-I activity in both the presence and absence of malonyl-CoA. Whereas only EPA treatment increased the mRNA levels of CPT-I, DHA treatment increased the mRNA levels of peroxisomal fatty acyl-CoA oxidase and fatty acid binding protein more effectively than EPA treatment. In conclusion, EPA and DHA affect cellular organelles in relation to their substrate preference. The present study strongly supports the hypothesis that EPA, and not DHA, lowers plasma triacylglycerol by increased mitochondrial fatty acid oxidation.

Antiinflammatory Effects of Tetradecylthioacetic Acid Involve Both Peroxisome Proliferator–Activated Receptor α–Dependent and –Independent Pathways

Objective—Tetradecylthioacetic acid (TTA) is a hypolipidemic antioxidant with immunomodulating properties involving activation of peroxisome proliferator–activated receptors (PPARs). Human endothelial cells express PPARs. We hypothesized that TTA could modulate endothelial cell activation at least partly through PPAR-related mechanisms. Methods and Results—We explored this hypothesis by different experimental approaches involving both in vitro studies in human endothelial cells (HUVECs) and in vivo studies in humans and PPAR-α−/− mice. Our main findings were as follows: (1) TTA suppressed the tumor necrosis factor α–induced expression of vascular cell adhesion molecule 1 (VCAM-1) and interleukin 8 (IL-8) in HUVECs. (2) No TTA-mediated attenuation of VCAM-1 and chemokine expression was seen in the liver of PPAR-α−/− mice. (3) Whereas TTA markedly enhanced PPAR-α–target genes in the liver of wild-type, but not of PPAR-α−/−, mice, no such effect on PPAR-α–target genes was seen in HUVECs. (4) The relevance of our findings to human disease was suggested by a TTA-mediated downregulation of serum levels of soluble VCAM-1 and IL-8 in psoriasis patients. Conclusion—We show that TTA has the ability to attenuate tumor necrosis factor α–mediated endothelial cell activation, further supporting antiinflammatory effects of this fatty acid, possibly involving both PPAR-α–dependent and –independent pathways.

Lipid-lowering and anti-inflammatory effects of tetradecylthioacetic acid in HIV-infected patients on highly active antiretroviral therapy.

Background  Highly active antiretroviral therapy (HAART) often leads to a dramatic improvement in clinical, viral and immunologic parameters in HIV‐infected individuals. However, the emergence of long‐term side‐effects of HAART and in particular dylipidaemia is increasingly reported. Based on the potential lipid‐lowering and immunomodulatory properties of tetradecylthioacetic acid (TTA) we examined whether TTA in combination with dietary intervention could modify lipid levels in peripheral blood in HIV‐infected patients on HAART.

Immunomodulating effects of 3-thia fatty acids in activated peripheral blood mononuclear cells

Background 3‐thia fatty acids such as tetradecylthioacetic acid (TTA) are modified fatty acids that have been suggested to change the plasma profile from atherogenic to cardio protective. Because of its interaction with peroxisome proliferator activated receptor (PPAR) we hypothesized that TTA also could have immunomodulatory properties. Based on the suggested role of inflammation in atherogenesis, any immunomodulating effects of TTA would be of particular interest for the potential use of this fatty acid in atherosclerotic disorders.

Growth reduction in glioma cells after treatment with tetradecylthioacetic acid: changes in fatty acid metabolism and oxidative status

During aerobic metabolism, a small amount of partially reduced oxygen is produced, yielding reactive oxygen species (ROS). Peroxisomes and mitochondria are major contributors to cellular ROS production, which is normally balanced by consumption by antioxidants. The fatty acid analogue tetradecylthioacetic acid (TTA) promotes mitochondrial and peroxisomal proliferation, and may induce oxidative stress and change the growth potential of cancer cells. In the present study, we found that TTA reduced [(3)H]thymidine incorporation in the glioma cell lines BT4Cn (rat), D54Mg (human), and GaMg (human) in a dose- and time-dependent manner. The 50% inhibitory TTA doses were approximately 125 microM for BT4Cn and D54Mg cells and 40 microM for GaMg cells after 4 days. alpha-Tochopherol counteracted this inhibition in GaMg cells. TTA enhanced the oxidation of [1-(14)C]palmitic acid, which could be explained by stimulation of enzymes involved in peroxisomal (fatty acyl-CoA oxidase) and/or mitochondrial (carnitine palmitoyltransferase) fatty acid oxidation. The glutathione content and the activities of glutathione peroxidase, glutathione reductase, and glutathione S-transferase were differentially affected. Increased malondialdehyde (MDA) production was seen in TTA-treated GaMg and D54Mg cells, but not in BT4Cn cells, in vitro. In BT4Cn tumor tissue from TTA-treated rats, MDA was increased while the alpha-tocopherol content tended to decrease. TTA increased the level of cytosolic cytochrome c in BT4Cn cells, which suggests induction of apoptotic cascades. Although several mechanisms are likely to be involved in the TTA-mediated effects on growth, we propose that modulation of cellular redox conditions caused by changes in fatty acid metabolism may be of vital importance.

Tetradecylthioacetic acid reduces stenosis development after balloon angioplasty injury of rabbit iliac arteries.

BACKGROUND tetradecylthioacetic acid (TTA) is a synthetic long-chain fatty acid analogue that inhibits the oxidative modification of low-density lipoprotein particles in vitro. We examined the influence of TTA on the arterial wall response after balloon angioplasty injury in a rabbit iliac model. METHODS AND RESULTS 14 rabbits were randomized to receiving either TTA fatty acids 800 mg daily perorally (weight 3.6+/-0.1 kg) or to normal diet (weight 3.5+/-0.5 kg, P=NS). Angioplasty was performed via right carotidotomy on both iliac arteries using an oversized balloon catheter, the TTA group being pretreated for 3 weeks. After angioplasty, the lumen diameter was 2.37+/-0.18 versus 2.36+/-0.13 mm for the TTA and control groups, respectively (P=NS). At 10 weeks follow-up angiography, minimal luminal diameter was 1.64+/-0.27 versus 1.13+/-0.52 mm for the TTA and control groups respectively (P<0.05). Histomorphometry did not show significant differences in intimal hyperplasia between the two groups (maximal intimal thickness 0.22+/-0.04 versus 0.19+/-0.10 mm, P=NS and intimal area 0.32+/-0.12 versus 0.36+/-0.23 mm(2), P=NS for the TTA and the control groups, respectively). In the heart, the sum of the n-3 fatty acids was 8.9+/-2.7 in the TTA group versus 4.3+/-0.2 mol% in the control group (P<0.05). The anti-inflammatory fatty acid index, calculated as (22:5 n-3+22:6 n-3+20:3 n-6)/20:4 n-6, was 0.76+/-0.10 vs. 0.25+/-0.03 for the TTA and control groups, respectively (P<0.05). In vitro TTA (100 microM) reduced the proliferation of human smooth muscle cell by more than 50%. CONCLUSION treatment with TTA is associated with positive arterial remodeling after angioplasty injury. The significance of the in vitro inhibition of human smooth muscle cell proliferation needs to be further elucidated.

Methylated Eicosapentaenoic Acid and Tetradecylthioacetic Acid: Effects on Fatty Acid Metabolism

We introduced methyl or ethyl groups to the 2- or 3-position of the eicosapentaenoic acid (EPA) molecule to investigate whether the branching of EPA could influence its hypolipidemic effect in rats. The most effective branching involved two methyl groups in the 2-position and one methyl group in the 3-position. These EPA derivatives increased hepatic mitochondrial and peroxisomal beta-oxidation and decreased plasma lipids concomitant with suppressed acetyl-coenzyme A (CoA) carboxylase (EC 6.4.1.2) and fatty acid synthase (EC 2.3.1.85) activities. This was followed by elevated activities of camitine O-palmitoyltransferase (EC 2.3.1.21) and possibly 2,4-dienoyl-CoA reductase (EC 1.3.1.34), as well as induced mRNA levels of these enzymes and fatty acyl-CoA oxidase. The fatty acid composition in liver changed, with an increased 18:1 n-9 content, whereas the expression of delta9-desaturase remained unchanged. We investigated the flux of fatty acids in cultured hepatocytes, and found that oxidation of [1-14C]-labeled palmitic acid increased but the secretion of palmitic acid-labeled triglycerides decreased after addition of 2-methyl-EPA. The fatty acyl-CoA oxidase (EC 1.3.3.6) activity in these cells remained unchanged. A significant negative correlation was obtained between palmitic acid oxidation and palmitic acid-labeled synthesized triglycerides. To investigate whether the hypolipidemic effect occurred independently of induced peroxisomal beta-oxidation, we fed rats 2-methyl-tetradecylthioacetic acid. This compound increased the peroxisomal but not the mitochondrial beta-oxidation, and the plasma lipid levels were unchanged. In conclusion, EPA methylated in the 2- or 3-position renders it more potent as a hypolipidemic agent. Furthermore, this study supports the hypothesis that the mitochondrion is the primary site for the hypolipidemic effect.

Tetradecylselenoacetic Acid, a PPAR Ligand With Antioxidant, Antiinflammatory, and Hypolipidemic Properties

Objective—Antioxidants protect against oxidative stress and inflammation, which, in combination with hyperlipidemia, are important mediators of atherogenesis. Here we present a selenium-substituted fatty acid, tetradecylselenoacetic acid (TSA), which is hypothesized to have antioxidant, antiinflammatory, and hypolipidemic properties. Methods and Results—We show that TSA exerts antioxidant properties by delaying the onset of oxidation of human low density lipoprotein (LDL), by reducing the uptake of oxidized LDL in murine macrophages, and by increasing the mRNA level of superoxide dismutase in rat liver. TSA also showed antiinflammatory effects by suppressing the release of interleukin (IL)-2 and -4, and by increasing the release of IL-10 in human blood leukocytes. In addition, TSA decreased the plasma triacylglycerol level and increased the mitochondrial fatty acid &bgr;-oxidation in rat liver. In pigs, TSA seemed to reduce coronary artery intimal thickening after percutaneous coronary intervention. In HepG2 cells TSA activated all peroxisome proliferator-activated receptors (PPARs) in a dose-dependent manner. Conclusions—Our data suggest that TSA exert potent antioxidant, antiinflammatory, and hypolipidemic properties, potentially involving PPAR-related mechanisms. Based on these effects, it is tempting to hypothesize that TSA could be an interesting antiatherogenic approach to atherosclerotic disorders.

Thia fatty acids with the sulfur atom in even or odd positions have opposite effects on fatty acid catabolism.

As tools for mechanistic studies on lipid metabolism, with the long-term goal of developing a drug for the treatment of lipid disorders, thia FA with the sulfur atom inserted at positions 3–9 from the carboxyl group were fed to male Wistar rats for 1 wk to determine their impact on key parameters in lipid metabolism and hepatic levels of thia FA metabolites. Thia FA with the sulfur atom in even positions decreased hepatic and cardiac mitochondrial β-oxidation and profoundly increased hepatic and cardiac TAG levels. The plasma TAG level was unchanged and the hepatic acyl-CoA oxidase activity increased. In contrast, thia FA with the sulfur atom in odd positions, especially 3-thia FA, tended to increase hepatic and cardiac FA oxidation and acyl-CoA oxidase and carnitine palmitoyltransferase-II activities, and decreased the plasma TAG levels. The effects seem to be related to differences in the catabolic rate of the thia FA. Differences between the two groups of acids were also observed with respect to the regulation of genes involved in FA transport and catabolism. Feeding experiments with 3- and 4-thia FA in combination indicated that the 4-thia FA partly attenuated the effects of the 3-thia FA on mitochondrial FA oxidation and the hepatic TAG level. In summary, the position of the sulfur atom in the alkyl chain, especially whether it is placed in the even or odd position, is crucial for the biological effect of the thia FA.

Synthesis and Analysis of Novel Glycerolipids for the Treatment of Metabolic Syndrome

Tetradecylthioacetic acid (TTA) 1 is a peroxisome proliferator-activated receptor (PPAR) agonist found to improve insulin sensitivity, lower blood lipid levels, enhance fatty acid oxidation, and promote anti-inflammation in vivo. In an attempt to enhance these properties, two key thioether fatty acid (Thefa) lipids, ditetradecylthioacetyl phosphatidylcholine 2 and tritetradecylthioacetyl glycerol 3, are synthesized and administered po to male Wistar rats at two different doses to study and compare metabolic outcomes relative to the administration of 1 alone after 6 days. Liposomal formulations of 1 and 2 are also prepared to evaluate acute metabolic responses (at 3 h) post i.v. injection. Across all metrics measured, 1-induced responses post po administration are in line with previous data. Responses induced from 3 are mostly equivalent to 1-induced responses. By contrast, 2-induced responses almost always outperform those of 1 and 3. Therefore, 2 may represent a new lead for the treatment of metabolic syndrome.