Association of liver steatosis with lipid oversecretion and hypotriglyceridaemia in C57BL/6j mice fed trans-10,cis-12-linoleic acid

Conjugated linoleic acids (CLA) have recently been recognized to reduce body fat and plasma lipids in some animals. This study demonstrated that the steatosis accompanying the fat loss induced by trans-10,cis-12-C(18:2) (CLA2) and not cis-9,trans-11-C(18:2) (CLA1) isomer in C57BL/6j mice was not due to an alteration of the liver lipoprotein production that was even increased. The 3-fold decrease in plasma triacylglycerol contents and the induction of mRNA expression of low-density lipoprotein receptors concomitantly observed in CLA2-fed mice suggested an increase in the lipoprotein clearance at the level of the liver itself. CLA1 feeding produced similar but attenuated effects on triglyceridaemia only.     

Conjugated linoleic acid isomers in mitochondria: evidence for an alteration of fatty acid oxidation

The beneficial effects exerted by low amounts of conjugated linoleic acids (CLA) suggest that CLA are maximally conserved and raise the question about their mitochondrial oxidizability. Cis-9,trans-11-C(18:2) (CLA1) and trans-10,cis-12-C(18:2) (CLA2) were compared to cis-9,cis-12-C(18:2) (linoleic acid; LA) and cis-9-C(16:1) (palmitoleic acid; PA), as substrates for total fatty acid (FA) oxidation and for the enzymatic steps required for the entry of FA into rat liver mitochondria. Oxygen consumption rate was lowest when CLA1 was used as a substrate with that on CLA2 being intermediate between it and the respiration on LA and PA. The order of the radiolabeled FA oxidation rate was PA > LA > CLA2 > CLA1. Transesterification to acylcarnitines of the octadecadienoic acids were similar, while uptake across inner membranes of CLA1 and, to a lesser extent, of CLA2 was greater than that of LA or PA. Prior oxidation of CLA1 or CLA2 made re-isolated mitochondria much less capable of oxidising PA or LA under carnitine-dependent conditions, but without altering the carnitine-independent oxidation of octanoic acid. Therefore, the CLA studied appeared to be both poorly oxidizable and capable of interfering with the oxidation of usual FA at a step close to the beginning of the beta-oxidative cycle.     

Large-scale preparation of (9Z,12E)-[1-(13)C]-octadeca-9,12-dienoic acid, (9Z,12Z,15E)-[1-(13)C]-octadeca-9,12,15-trienoic acid and their [1-(13)C] all-cis isomers

Several grams of labelled trans linoleic and linolenic acids with high chemical and isomeric purities (>97%) have been prepared for human metabolism studies. A total of 12.5 g of (9Z, 12E)-[1-(13)C]-octadeca-9,12-dienoic acid and 6.3 g of (9Z,12Z, 15E)-[1-(13)C]-octadeca-9,12,15-trienoic acid were obtained in, respectively, seven steps (7.8% overall yield) and 11 steps (7% overall yield) from 7-bromo-heptan-1-ol. The trans bromo precursors used for the labelling were synthesised by using copper-catalysed couplings. The trans fatty acids were then obtained via the nitrile derivatives. A total of 23.5 g of (9Z,12Z)-[1-(13)C]-octadeca-9, 12-dienoic acid and 10.4 g of (9Z,12Z,15Z)-[1-(13)C]-octadeca-9,12, 15-trienoic acid were prepared in five steps in, respectively, 32 and 18% overall yield. Large quantities of bromo and chloro precursors were synthesised from the commercially available acid according to Barton's procedure. In all cases, the main impurities (>0.5%) of each labelled fatty acid have been characterised.     

Effects of trans MUFA from dairy and industrial sources on muscle mitochondrial function and insulin sensitivity

Epidemiological studies suggest that chronic consumption of trans MUFA may alter muscle insulin sensitivity. The major sources of dietary trans MUFA (dairy fat vs. industrially hydrogenated oils) have different isomeric profiles and thus probably different metabolic consequences. These effects may involve alterations in muscle mitochondrial oxidative capacity, which may in turn promote insulin resistance if fatty acid oxidation is reduced. We report that in Wistar rats, an 8 week diet enriched (4% of energy intake) in either dairy, industrial, or control MUFA did not alter insulin and glucose responses to an intraperitoneal glucose tolerance test (1g/kg). In C2C12 myotubes, vaccenic and elaidic acids did not modify insulin sensitivity compared with oleic acid. Furthermore, the ex vivo total, mitochondrial and peroxisomal oxidation rates of [1-(14)C]oleic, vaccenic, and elaidic acids were similar in soleus and tibialis anterior rat muscle. Finally, an 8 week diet enriched in either dairy or industrial trans MUFA did not alter mitochondrial oxidative capacity in these two muscles compared with control MUFA but did induce a specific reduction in soleus mitochondrial ATP and superoxide anion production (P<0.01 vs. control). In conclusion, dietary trans MUFA of dairy or industrial origin have similar effects and do not impair muscle mitochondrial capacity and insulin sensitivity.     

Rapid effects of the intravenous injection of a medium-chain triglyceride: fish oil emulsion on the triglyceride fatty acid pattern of soleus muscle from omega3 fatty acid-depleted rats.

Second generation rats depleted in long-chain polyunsaturated omega3 fatty acids display several features of the metabolic syndrome, including visceral obesity, liver steatosis, insulin resistance, hypertension, and cardiac hypertrophy. In the framework of an extensive study on such metabolic, hormonal and functional perturbations, the phospholipid fatty acid pattern and ex vivo metabolism of D-glucose were recently investigated in the soleus muscle of these omega3-depleted rats. The present study deals with the triglyceride fatty acid content and pattern of the soleus muscle in control animals and omega3-depleted rats. Some of the latter rats were injected intravenously 60-120 minutes before sacrifice with either an omega3 fatty acid-rich medium-chain triglyceride/fish oil emulsion (omega3-FO rats) or a control medium-chain triglyceride/olive oil emulsion (omega3-OO rats). The total fatty acid content of triglycerides was comparable in control and omega3-depleted rats and, in both cases, inversely related to their C20:4omega6 relative content. At variance with the situation found in control rats, no long-chain polyunsaturated omega3 fatty acid (C18:3omega3, C20:5omega3, C22:5omega3, C22:6omega3) was detected in the omega3-depleted rats. Unexpectedly, the triglyceride content in most long-chain polyunsaturated omega6 fatty acids (C18:2omega6, C20:3omega6, C20:4omega6 and C22:4omega6) had also decreased in the latter rats. Moreover, the activity of Delta9-desaturase was apparently increased in the omega3-depleted rats, as judged from the C16:1omega7/C16:0 and C18:1omega9/C18:0 ratios. The omega3-FO rats differed from omega3-OO rats by a lower contribution of C18:2omega6 metabolites (C18:3omega6, C20:3omega6, C20:4omega6 and C22:4omega6). These findings indicate that the prior injection of the medium-chain triglyceride/fish oil emulsion, known to increase the muscle phospholipid content in long-chain polyunsaturated omega3 fatty acids, may nevertheless accentuate the depletion in long-chain polyunsaturated omega6 fatty acids otherwise found in the triglycerides of omega3-depleted rats. Such a dual effect is reminiscent of that observed, under the same experimental conditions, for selected variables in D-glucose metabolism in the soleus muscle.     

Plasmalogen metabolism-related enzymes in rat brain during aging: influence of n-3 fatty acid intake

Plasmalogens (Pls) are phospholipids containing a vinyl-ether bond at the sn-1 position of the glycerol backbone. They represent between 1/2 and 2/3 of the ethanolamine phospholipids in the brain. During aging, the Pls content in human brain falls down. However, the role of Pls metabolism-related enzymes in the regulation of Pls levels remains to be determined. Dihydroxyacetone phosphate acyltransferase (DHAP-AT) is the enzyme involved in the first step of Pls biosynthesis. In the brain, a phospholipase A2, which selectively acts on Pls, has been isolated (Pls-PLA2s). In this work, we aimed to evaluate the impact of DHAP-AT (a key enzyme of Pls biosynthesis) and Pls-PLA2 (a specific Pls degradation enzyme) on the evolution of Pls content in the rat brain during aging. The influence of n-3 fatty acid intake was also evaluated. Littermates from two generations of n-3 deficient rats were fed an equilibrated diet containing either alpha-LNA alone or with two doses of DHA. After weaning, 3, 9 or 21 months of diet, rats were sacrificed. Enzymatic assays were performed, Pls levels were assessed and the sn-2 position of ethanolamine Pls was analyzed. DHAP-AT activity significantly increased between weaning and 3 months with a concomitant increase of brain Pls, which reached maximal levels after 9 months. Then, Pls levels and DHAP-AT activity significantly decreased while Pls-PLA2s activity significantly increased. Dietary n-3 fatty acids had no effect on DHAP-AT activity and on Pls levels. In conclusion, the increase of brain Pls content in the first part of the life may be related to the high increase of DHAP-AT activity, probably stimulated by DHA. In aged animals, the decrease of Pls levels may mainly be caused to an increase of their degradation by Pls-PLA2. Dietary DHA may not oppose the physiologic aging.     

Pancreatic islet function in omega-3 fatty acid-depleted rats: alteration of calcium fluxes and calcium-dependent insulin release

Considering the insufficient supply of long-chain polyunsaturated omega-3 fatty acids often prevailing in Western populations, this report deals mainly with alterations of Ca(2+) fluxes and Ca(2+)-dependent insulin secretory events in isolated pancreatic islets from omega-3-depleted rats. In terms of (45)Ca(2+) handling, the islets from omega-3-depleted rats, compared with those from normal animals, displayed an unaltered responsiveness to an increase in extracellular K(+) concentration, a lower inflow rate and lower fractional outflow rate of the divalent cation, and higher (45)Ca(2+)-labeled cellular pool(s) at isotopic equilibrium. The latter anomaly was corrected 120 min after intravenous injection of a novel medium-chain triglyceride-fish oil (MCT:FO) emulsion, distinct from a control omega-3-poor MCT-olive oil (MCT:OO) emulsion. At 8.3 mM D-glucose, insulin release was higher in islets from omega-3-depleted rats vs. control animals, coinciding with a higher cytosolic Ca(2+) concentration. The relative magnitude of the increase in insulin output attributable to a rise in D-glucose as well as extracellular Ca(2+) or K(+) concentration, to the absence vs. presence of verapamil and to the presence vs. absence of extracellular Ca(2+), theophylline, phorbol 12-myristate 13-acetate, or Ba(2+), was always more pronounced in islets from omega-3-depleted rats injected with the MCT:OO compared with the MCT:FO emulsion. A comparable situation prevailed when comparing islets from noninjected omega-3-depleted and normal rats. In light of these and previous findings, we propose that an impairment of Na(+),K(+)-ATPase activity plays a major, although not an exclusive, role in the perturbation of Ca(2+) fluxes and Ca(2+)-dependent secretory events in the islets from omega-3-depleted rats.     

Rapid down-regulation of mitochondrial fat metabolism in human muscle after training cessation is dissociated from changes in insulin sensitivity

The association between impairment in mitochondrial muscle fat oxidative capacity (OX_FA) and occurrence of insulin resistance was examined in 14 healthy trained men (age, 24 ± 4 yr) submitted to 4 weeks of training cessation. Training stop induced a significant decrease in mRNA levels of proteins involved in muscle fat metabolism, particularly PPARα (−58%, P < 0.01) and PGC-1α (−30%, P < 0.05), a 21% reduction in OX_FA (P < 0.01), and reduced fat oxidation during moderately intense exercise (P < 0.05). In contrast, there was no significant alteration in insulin sensitivity. In conclusion, decline in OX_FA is a rapid metabolic event following training cessation. It is involved in the regulation of whole body fat balance but not in the deterioration of insulin sensitivity.     

Fatty acid composition and conjugated linoleic acid content of different tissues in rats fed individual conjugated linoleic acid isomers given as triacylglycerols small star,filled

HEAT TREATMENT OF VEGETABLE OILS GAVE RISE TO FOUR MAIN CONJUGATED LINOLEIC ACID (CLA) ISOMERS : the 9c,11t, 9t,11t, 10t,12c and 10t,12t. The diet of male Wistar rats was supplemented with 150 mg/day either 9c,11t-, 9t,11t-, 10t,12c- or 10t,12t CLA isomers for 6 days and their effects on lipid composition were investigated in liver, heart, skeletal muscle Gastrocnemius, kidneys, brain and adipose tissue. The incorporation of all isomers was low (< 1.4%) and the level was as follows : adipose tissue > Gastrocnemius > liver, kidneys > brain. The main changes in the overall lipid composition were observed in skeletal muscle (Gastrocnemius) and in heart and were associated with feeding the 10t,12c and 10t,12t isomers. The diet enriched in 10t,12t CLA decreased the total long chain polyunsaturated fatty acid proportion in Gastrocnemius (from 18.4% to 14.4%) and increased that of 20:4 n-6 in heart (from 16.9 to 19.3%). The diet enriched in 10t,12c CLA decreased the monounsaturated fatty acid proportion in Gastrocnemius (from 32.0 to 26.1%) and produced an effect similar to the 10t,12t in heart. By contrast, the 9c,11t and 9t,11t isomers did not affect fatty acid composition in all tissues and organs. We concluded that ingestion of 10t,12c and 10t,12t CLA present in oils and in CLA mixtures could change muscle lipid composition.     

In vitro desaturation or elongation of monotrans isomers of linoleic acid by rat liver microsomes

Several nutritional studies have shown the in vivo conversion of the 9c,12t-18:2 and 9t,12c-18:2 into long chain polyunsaturated fatty acids (PUFA) containing 20 carbons (geometrical isomers of eicosadienoic and eicosatetraenoic acids). In the present work, some in vitro studies were carried out in order to have precise information on the conversion of these two isomers.In a first set of experiments, studies were focused on the in vitro 6 desaturation, the first regulatory step of the biosynthesis of n-6 long chain PUFA, from 9c,12c-18:2. Rat liver microsomes were prepared and incubated under desaturation conditions with [1-¹⁴C]-9c,12c-18:2 in presence of unlabelled 9c,12t-, 9t,12c- or 9t,12t-18:2. The data show that each trans isomer induced a decrease of the 6 desaturation of the [1-¹⁴C]-9c,12c-18:2, but the 9c,12t-18:2 was the most potent inhibitor (up to 63%). Rat liver microsomes were also incubated with [1-¹⁴C]-9c,12c-18:2, [1-¹⁴C]-9c,12t-18:2 or [1-¹⁴C]-9t,12c-18:2 under desaturation conditions. The results indicated that 18:2 9c,12t is a much better substrate for desaturase than 9t,12c-18:2. Moreover, the conversion levels of [1-¹⁴C]-9c,12t-18:2 was similar to what was observed for its all cis homologue, at low substrate concentration only. In a second set of experiments, in vitro elongation studies of each mono-trans 18:2 isomers and 9c,12c-18:2 were carried out. For that purpose, rat liver microsomes were incubated with [1-¹⁴C]-9c,12c-18:2, [1-¹⁴C]-9c,12t-18:2 or [1-¹⁴C]-9t,12c-18:2 under elongation conditions. The data show that [1-¹⁴C]-9t,12c-18:2 is better elongated than 9c,12c-18:2 while the amount of product formed from [1-¹⁴C]-9c,12t-18:2 was lower than was produced from the 9c,12c-18:2.Thus, the desaturation enzymes presented a higher affinity for the 9c,12t-18:2 whereas the elongation enzyme presented a higher affinity for the 9t,12c-18:2.