Rate Of Β-oxidation Research Articles

Twenty-four hour insulin secretion and beta cell NEFA oxidation in type 2 diabetic, morbidly obese patients before and after bariatric surgery

We have previously demonstrated that type 2 diabetes resolves after bariatric surgery. To study the role of NEFA in the prompt normalisation of beta cell glucose sensitivity, insulin secretion and beta cell glucose and lipid metabolism were investigated by a model of nutrient-stimulated insulin secretion using a multiple-meal test. Hourly glucose, C-peptide and NEFA were measured in nine morbidly obese, type 2 diabetic patients before and 1 week after bariatric surgery and in six matched healthy volunteers over 24 h. A mathematical model of glucose-NEFA comodulation of insulin secretion rate (ISR) was used to compute ISR and beta-oxidation. Insulin sensitivity was measured by an OGTT minimal model. Beta cell sensitivity to glucose and NEFA was doubled after surgery, while the 24 h insulin secretion decreased from 277.1 +/- 144.4 to 198.0 +/- 107.6 nmol/m(2) (p < 0.02). Insulin sensitivity was restored. The beta-oxidation rate of beta cells was completely normalised (from 0.032 +/- 0.012 x 10(-12) to 0.103 +/- 0.031 x 10(-12) mmol/min per cell, p < 0.005). The best predictor of beta cell function improvement was the duration of diabetes. Bariatric surgery in type 2 diabetes restores beta-oxidation in beta cells, doubles glucose-NEFA sensitivity and reverses diabetes. It is likely that ISR is reduced to match insulin-sensitivity normalisation, in spite of no significant reduction in NEFA levels. We hypothesise that insulin sensitivity normalisation might appear as a consequence of nutrient exclusion from proximal intestinal transit, and that secondarily the need for insulin secretion diminishes. The insulin sensitivity increase is much higher than usually obtained by insulin-sensitising agents and is independent of weight changes.

A moderate increase in carnitine palmitoyltransferase 1a activity is sufficient to substantially reduce hepatic triglyceride levels

Nonalcoholic fatty liver disease (NAFLD), hypertriglyceridemia, and elevated free fatty acids are present in the majority of patients with metabolic syndrome and type 2 diabetes mellitus and are strongly associated with hepatic insulin resistance. In the current study, we tested the hypothesis that an increased rate of fatty acid oxidation in liver would prevent the potentially harmful effects of fatty acid elevation, including hepatic triglyceride (TG) accumulation and elevated TG secretion. Primary rat hepatocytes were transduced with adenovirus encoding carnitine palmitoyltransferase 1a (Adv-CPT-1a) or control adenoviruses encoding either beta-galactosidase (Adv-beta-gal) or carnitine palmitoyltransferase 2 (Adv-CPT-2). Overexpression of CPT-1a increased the rate of beta-oxidation and ketogenesis by approximately 70%, whereas esterification of exogenous fatty acids and de novo lipogenesis were unchanged. Importantly, CPT-1a overexpression was accompanied by a 35% reduction in TG accumulation and a 60% decrease in TG secretion by hepatocytes. There were no changes in secretion of apolipoprotein B (apoB), suggesting the synthesis of smaller, less atherogenic VLDL particles. To evaluate the effect of increasing hepatic CPT-1a activity in vivo, we injected lean or obese male rats with Adv-CPT-1a, Adv-beta-gal, or Adv-CPT-2. Hepatic CPT-1a activity was increased by approximately 46%, and the rate of fatty acid oxidation was increased by approximately 44% in lean and approximately 36% in obese CPT-1a-overexpressing animals compared with Adv-CPT-2- or Adv-beta-gal-treated rats. Similar to observations in vitro, liver TG content was reduced by approximately 37% (lean) and approximately 69% (obese) by this in vivo intervention. We conclude that a moderate stimulation of fatty acid oxidation achieved by an increase in CPT-1a activity is sufficient to substantially reduce hepatic TG accumulation both in vitro and in vivo. Therefore, interventions that increase CPT-1a activity could have potential benefits in the treatment of NAFLD.

Skeletal muscle adaptation to fatty acid depends on coordinated actions of the PPARs and PGC1α: implications for metabolic disease

Dyslipidemia and intramuscular accumulation of fatty acid metabolites are increasingly recognized as core features of obesity and type 2 diabetes. Emerging evidence suggests that normal physiological adaptations to a heavy lipid load depend on the coordinated actions of broad transcriptional regulators such as the peroxisome proliferator activated receptors (PPARs) and PPAR gamma coactivator 1 alpha (PGC1 alpha). The application of transcriptomics and targeted metabolic profiling tools based on mass spectrometry has led to our finding that lipid-induced insulin resistance is a condition in which upregulation of PPAR-targeted genes and high rates of beta-oxidation are not supported by a commensurate upregulation of tricarboxylic acid (TCA) cycle activity. In contrast, exercise training enhances mitochondrial performance, favoring tighter coupling between beta-oxidation and the TCA cycle, and concomitantly restores insulin sensitivity in animals fed a chronic high-fat diet. The exercise-activated transcriptional coactivator, PGC1 alpha, plays a key role in coordinating metabolic flux through these 2 intersecting metabolic pathways, and its suppression by overfeeding may contribute to diet-induced mitochondrial dysfunction. Our emerging model predicts that muscle insulin resistance arises from a mitochondrial disconnect between beta-oxidation and TCA cycle activity. Understanding of this "disconnect" and its molecular basis may lead to new therapeutic approaches to combatting metabolic disease.

Effects of total replacement of fish oil by vegetable oils on n-3 and n-6 polyunsaturated fatty acid desaturation and elongation in sharpsnout seabream (Diplodus puntazzo) hepatocytes and enterocytes

The main aim of this work was to determine the impact of total dietary fish oil replacement by vegetable oils on the fatty acid metabolism of sharpsnout seabream hepatocytes and digestive tract enterocytes. Three isonitrogenous (48% crude protein) and isoenergetic (23 MJ/kg) experimental diets were formulated using three different lipid sources: fish oil (FO), rich in n-3 HUFAs; soybean oil (SO), rich in linoleic acid (LA), and linseed oil (LO), especially rich in linolenic acid (LNA). These diets were fed, three times a day to apparent satiation, to triplicate groups of 30 sharpsnout seabream (with an initial average weight of 14.9 g) for nine months at 23.5±1.2 °C. Inclusion of vegetable oils in sharpsnout seabream diet did not have any quantitative nutritional effects on desaturation/elongation of [1-14C] LNA and [1-14C] LA in isolated hepatocytes and total digestive tract enterocytes. Most of the radioactivity found in tissue lipid extracts (94% and 86% for hepatocytes and enterocytes, respectively) using silver nitrate thin-layer chromatography was recovered as [1-14C] C18 PUFA, clearly showing the lack of any significant desaturase/elongase activity in these cells. Only the high levels of HUFA in fish tissues pointed to the existence of some kind of regulatory mechanism, presumably based on HUFA bioaccumulation and C18 PUFA oxidation. Moreover, direct measurements of β-oxidation rates yielded very low values in all cases, the only significant difference being a higher oxidation rate in hepatocytes from fish fed LO versus FO diet.

Specificity in Beta Cell Expression of l-3-Hydroxyacyl-CoA Dehydrogenase, Short Chain, and Potential Role in Down-regulating Insulin Release

A loss-of-function mutation of the mitochondrial beta-oxidation enzyme l-3-hydroxyacyl-CoA dehydrogenase, short chain (HADHSC), has been associated with hyperinsulinemic hypoglycemia in man. It is still unclear whether loss of glucose homeostasis in these patients (partly) results from a dysregulation of beta cells. This study examines HADHSC expression in purified rat beta cells and investigates whether its selective suppression elevates insulin release. Beta cells expressed the highest levels of HADHSC mRNA and protein of all examined tissues, including those with high rates of mitochondrial beta-oxidation. On the other hand, beta cells expressed relatively low levels of other beta-oxidation enzymes (acyl-CoA dehydrogenase short, medium, and long chain and acetyl-coenzyme A acyltransferase 2). HADHSC expression was sequence-specifically silenced by RNA interference, and the effects were examined on glucose-stimulated insulin secretion following 48-72 h of suppression. In both rat beta cells and in the beta cell line INS1 832-13, HADHSC silencing resulted in elevated insulin release at low and at high glucose concentrations, which appeared not to be caused by increased rates of glucose metabolism or an inhibition in fatty acid oxidation. These data indicate that the normal beta cell phenotype is characterized by a high expression of HADHSC and a low expression of other beta-oxidation enzymes. Down-regulation of HADHSC causes an elevated secretory activity suggesting that this enzyme protects against inappropriately high insulin levels and hypoglycemia.

Fatty Acid Transport Protein 4 Is the Principal Very Long Chain Fatty Acyl-CoA Synthetase in Skin Fibroblasts

Fatty acid transport protein 4 (FATP4) is a fatty acyl-CoA synthetase that preferentially activates very long chain fatty acid substrates, such as C24:0, to their CoA derivatives. To gain better insight into the physiological functions of FATP4, we established dermal fibroblast cell lines from FATP4-deficient wrinkle-free mice and wild type (w.t.) mice. FATP4 -/- fibroblasts had no detectable FATP4 protein by Western blot. Compared with w.t. fibroblasts, cells lacking FATP4 had an 83% decrease in C24:0 activation. Peroxisomal degradation of C24:0 was reduced by 58%, and rates of C24:0 incorporation into major phospholipid species (54-64% decrease), triacylglycerol (64% decrease), and cholesterol esters (58% decrease) were significantly diminished. Because these lipid metabolic processes take place in different subcellular organelles, we used immunofluorescence and Western blotting of subcellular fractions to investigate the distribution of FATP4 protein and measured enzyme activity in fractions from w.t. and FATP4 -/- fibroblasts. FATP4 protein and acyl-CoA synthetase activity localized to multiple organelles, including mitochondria, peroxisomes, endoplasmic reticulum, and the mitochondria-associated membrane fraction. We conclude that in murine skin fibroblasts, FATP4 is the major enzyme producing very long chain fatty acid-CoA for lipid metabolic pathways. Although FATP4 deficiency primarily affected very long chain fatty acid metabolism, mutant fibroblasts also showed reduced uptake of a fluorescent long chain fatty acid and reduced levels of long chain polyunsaturated fatty acids. FATP4-deficient cells also contained abnormal neutral lipid droplets. These additional defects indicate that metabolic abnormalities in these cells are not limited to very long chain fatty acids.

β-Oxidation in hepatocyte cultures from mice with peroxisomal gene knockouts

β-Oxidation of carboxylates takes place both in mitochondria and peroxisomes and in each pathway parallel enzymes exist for each conversion step. In order to better define the substrate specificities of these enzymes and in particular the elusive role of peroxisomal MFP-1, hepatocyte cultures from mice with peroxisomal gene knockouts were used to assess the consequences on substrate degradation. Hepatocytes from mice with liver selective elimination of peroxisomes displayed severely impaired oxidation of 2-methylhexadecanoic acid, the bile acid intermediate trihydroxycholestanoic acid (THCA), and tetradecanedioic acid. In contrast, mitochondrial β-oxidation rates of palmitate were doubled, despite the severely affected inner mitochondrial membrane. As expected, β-oxidation of the branched chain compounds 2-methylhexadecanoic acid and THCA was reduced in hepatocytes from mice with inactivation of MFP-2. More surprisingly, dicarboxylic fatty acid oxidation was impaired in MFP-1 but not in MFP-2 knockout hepatocytes, indicating that MFP-1 might play more than an obsolete role in peroxisomal β-oxidation.

Decreased Myocardial Free Fatty Acid Uptake in Patients With Idiopathic Dilated Cardiomyopathy: Evidence of Relationship With Insulin Resistance and Left Ventricular Dysfunction

Background Results on myocardial substrate metabolism in the failing heart have been contradictory. Insulin resistance, a common comorbidity in heart failure patients, and medical therapy may modify myocardial metabolism in complex fashions. Therefore, we characterized myocardial oxidative and free fatty acid (FFA) metabolism in patients with idiopathic dilated cardiomyopathy (IDCM) and investigated the contributions of insulin resistance and β-blocker therapy. Methods and Results Nineteen patients with IDCM (age 58 ± 8 years, ejection fraction 33 ± 8.8%) and 15 healthy controls underwent examination of myocardial blood perfusion, oxidative and FFA metabolism using positron emission tomography and [ 15O]H 2O, [ 11C]acetate and [ 11C]palmitate, respectively. Echocardiography was used to assess myocardial function, work, and efficiency of forward work. Insulin resistance was calculated using the homeostasis model assessment index (HOMA index) and the degree of β-blockade was estimated with a β-adrenoceptor occupancy test. IDCM patients were characterized by decreased cardiac efficiency (35 ± 2 versus 57 ± 12 mm Hg ·L ·g −1, P < .0001) and reduced myocardial FFA uptake (5.5 ± 2.0 versus 6.4 ± 1.2 μmol ·100 g −1 ·min −1, P < .05), but the FFA β-oxidation rate constant was not changed. In the patients, myocardial FFA uptake was inversely associated with left ventricular (LV) ejection fraction ( r = −0.63, P < .01), indicating that further depression of LV function induces an opposite switch to greater FFA uptake. The FFA β-oxidation rate constant correlated positively with the HOMA index ( r = 0.53, P < .05). In patients on β-1 selective β-blockers, β-1 adrenoceptor occupancy correlated inversely with LV work, oxidative metabolism, and FFA uptake; similar relationships were not found in patients on nonselective β-blocker. Conclusions Myocardial FFA metabolism is reduced in patients with IDCM. However, when LV function is further depressed and insulin resistance manifested, myocardial FFA uptake and oxidation are, in turn, upregulated. These findings may partly explain the discrepancies between previous studies about cardiac metabolism in heart failure.

The antiepileptic drug topiramate preserves metabolism-secretion coupling in insulin secreting cells chronically exposed to the fatty acid oleate

Topiramate (Topamax), primarily prescribed against epilepsy, was reported to reduce body weight and to ameliorate glycemic control in obese patients with diabetes. In rodent models of obesity and diabetes, topiramate treatment counteracts hyperglycemia and increases insulin levels upon glucose tolerance test. These observations suggest that topiramate might exert direct action on insulin secreting cells, in particular regarding obesity associated β-cell dysfunction. In this study, INS-1E β-cells were exposed for 3 days to the fatty acid oleate (0.4 mM) and concomitantly treated with therapeutic concentrations of topiramate before measurements of insulin secretion and metabolic parameters. In healthy cells, topiramate had no acute or chronic effects on insulin release. Exposure of INS-1E cells to oleate for 3 days increased insulin release at basal 2.5 mM glucose and blunted the response to stimulatory glucose concentration (15 mM). Such lipotoxic effects were associated with impaired mitochondrial function, as evidenced by partial loss of resting mitochondrial membrane potential and reduced hyperpolarization in response to glucose. Oil-red-O staining and triglyceride measurements revealed lipid accumulation in oleate treated cells. Topiramate treatment counteracted oleate-induced lipid load and partially protected against mitochondrial membrane dysfunction. In particular, topiramate restored glucose stimulated insulin secretion, essentially by maintaining low insulin release at basal glucose. Topiramate increased expression of the nutrient sensor PPARα and of the mitochondrial fatty acid carrier CPT-1, correlating with enhancement of β-oxidation rate. The data demonstrate that a drug originally used as mood stabilizer exerts a direct action on β-cells, protecting against lipid-induced dysfunction.

Hypoxia Converts Human Macrophages Into Triglyceride-Loaded Foam Cells

Atherosclerotic lesions have regions that are hypoxic. Because the lesion contains macrophages that are loaded with lipid, we investigated whether hypoxia can influence the accumulation of lipids in these cells. Exposure of human macrophages to hypoxia for 24 hours resulted in an increased formation of cytosolic lipid droplets and an increased accumulation of triglycerides. Exposure of the macrophages to oxidized low-density lipoprotein (oxLDL) increased the accumulation of cytosolic lipid droplets because of an increase in cellular cholesterol esters. The accumulation of lipid droplets in oxLDL-treated cells was further increased after hypoxia, caused by an increased level of triglycerides. Expression analyses combined with immunoblot or RT-PCR demonstrated that hypoxia increased the expression of several genes that could promote the accumulation of lipid droplets. Hypoxia increased the mRNA and protein levels of adipocyte differentiation-related protein (ADRP). It is well known that an increased expression of ADRP increases the formation of lipid droplets. Hypoxia decreased the expression of enzymes involved in beta-oxidation (acyl-coenzyme A synthetase and acyl-coenzyme A dehydrogenase) and increased the expression of stearoyl-coenzyme A desaturase, an important enzyme in the fatty acid biosynthesis. Moreover, exposure to hypoxia decreased the rate of beta-oxidation, whereas the accumulation of triglycerides increased. The results demonstrate that exposure of human macrophages to hypoxia causes an accumulation of triglyceride-containing cytosolic lipid droplets. This indicates that the hypoxia present in atherosclerotic lesions can contribute to the formation of the lipid-loaded macrophages that characterize the lesion and to the accumulation of triglycerides in such lesions.

The role of the liver in lipid metabolism during cold acclimation in non-hibernator rodents

Cold exposure increases the demand for energy substrates. Cold acclimation of rats led to a 3-fold increase in fatty acid (FA) β-oxidation ( P < 0.01) for ex vivo livers perfused at 37 °C. This increase was preserved following perfusion at 25 °C ( P < 0.001). In vitro measurement of absolute rates of hepatic β-oxidation revealed no significant difference following cold acclimation, implying changes in fatty acid flux through β-oxidation rather than increased oxidation capacity. Total FA uptake was increased one-third following perfusion at 25 °C ( P < 0.001) and cold acclimation ( P < 0.05) and cold acclimation led to diversion of tissue FA from storage to β-oxidation ( P < 0.01). In separate experiments, in vivo hepatic lipogenesis rates for saponifiable lipids doubled ( P < 0.01) and cholesterol synthesis increased one-third ( P < 0.001). Taken together these data suggest the oxidation and synthesis of lipids occur simultaneously in hepatic tissue possibly to increase prevailing tissue FA concentrations and to generate heat through increased metabolic flux rates.

Hepatic Peroxisomal Fatty Acid β-Oxidation Is Regulated by Liver X Receptor α

Peroxisomes are the exclusive site for the beta-oxidation of very-long-chain fatty acids of more than 20 carbons in length (VLCFAs). Although the bulk of dietary long-chain fatty acids are oxidized in the mitochondria, VLCFAs cannot be catabolized in mitochondria and must be shortened first by peroxisomal beta-oxidation. The regulation of peroxisomal, mitochondrial, and microsomal fatty acid oxidation systems in liver is mediated principally by peroxisome proliferator-activated receptor alpha (PPARalpha). In this study we provide evidence that the liver X receptor (LXR) regulates the expression of the genetic program for peroxisomal beta-oxidation in liver. The genes encoding the three enzymes of the classic peroxisomal beta-oxidation cycle, acyl-coenzyme A (acyl-CoA) oxidase, enoyl-CoA hydratase/L-3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase, are activated by the LXR ligand, T0901317. Accordingly, administration of T0901317 in mice promoted a dose-dependent and greater than 2-fold increase in the rate of peroxisomal beta-oxidation in the liver. The LXR effect is independent of PPARalpha, because T0901317-induced peroxisomal beta-oxidation in the liver of PPARalpha-null mice. Interestingly, T0901317-induced peroxisomal beta-oxidation is dependent on the LXRalpha isoform, but not the LXRbeta isoform. We propose that induction of peroxisomal beta-oxidation by LXR agonists may serve as a counterregulatory mechanism for responding to the hypertriglyceridemia and liver steatosis that is promoted by potent LXR agonists in vivo; however, additional studies are warranted.

Prevention of hepatic steatosis and hepatic insulin resistance in mitochondrial acyl-CoA:glycerol-sn-3-phosphate acyltransferase 1 knockout mice

In order to investigate the role of mitochondrial acyl-CoA:glycerol-sn-3-phosphate acyltransferase 1 (mtGPAT1) in the pathogenesis of hepatic steatosis and hepatic insulin resistance, we examined whole-body insulin action in awake mtGPAT1 knockout (mtGPAT1(-/-)) and wild-type (wt) mice after regular control diet or three weeks of high-fat feeding. In contrast to high-fat-fed wt mice, mtGPAT1(-/-) mice displayed markedly lower hepatic triacylglycerol and diacylglycerol concentrations and were protected from hepatic insulin resistance possibly due to a lower diacylglycerol-mediated PKC activation. Hepatic acyl-CoA has previously been implicated in the pathogenesis of insulin resistance. Surprisingly, compared to wt mice, mtGPAT1(-/-) mice exhibited increased hepatic insulin sensitivity despite an almost 2-fold elevation in hepatic acyl-CoA content. These data suggest that mtGPAT1 might serve as a novel target for treatment of hepatic steatosis and hepatic insulin resistance and that long chain acyl-CoA's do not mediate fat-induced hepatic insulin resistance in this model.

Mitochondrial dysfunction induces triglyceride accumulation in 3T3-L1 cells: role of fatty acid β-oxidation and glucose

Mitochondrial cytopathy has been associated with modifications of lipid metabolism in various situations, such as the acquisition of an abnormal adipocyte phenotype observed in multiple symmetrical lipomatosis or triglyceride (TG) accumulation in muscles associated with the myoclonic epilepsy with ragged red fibers syndrome. However, the molecular signaling leading to fat metabolism dysregulation in cells with impaired mitochondrial activity is still poorly understood. Here, we found that preadipocytes incubated with inhibitors of mitochondrial respiration such as antimycin A (AA) accumulate TG vesicles but do not acquire specific markers of adipocytes. Although the uptake of TG precursors is not stimulated in 3T3-L1 cells with impaired mitochondrial activity, we found a strong stimulation of glucose uptake in AA-treated cells mediated by calcium and phosphatidylinositol 3-kinase/Akt1/glycogen synthase kinase 3beta, a pathway known to trigger the translocation of glucose transporter 4 to the plasma membrane in response to insulin. TG accumulation in AA-treated cells is mediated by a reduced peroxisome proliferator-activated receptor gamma activity that downregulates muscle carnitine palmitoyl transferase-1 expression and fatty acid beta-oxidation, and by a direct conversion of glucose into TGs accompanied by the activation of carbohydrate-responsive element binding protein, a lipogenic transcription factor. Taken together, these results could explain how mitochondrial impairment leads to the multivesicular phenotype found in some mitochondria-originating diseases associated with a dysfunction in fat metabolism.

Effect of partial substitution of dietary fish oil by vegetable oils on desaturation and β-oxidation of [1- 14C]18:3 n−3 (LNA) and [1- 14C]20:5 n−3 (EPA) in hepatocytes and enterocytes of European sea bass ( Dicentrarchus labrax L.)

The increasing worldwide aquaculture output and concomitant decrease in the stocks of feed-grade fish used for fish oil production has made fish oil replacement in feeds a priority for the aquaculture industry. The regulation of fatty acid metabolism in fish is important in order to determine strategies for the best use of plant oils in diets for commercially important cultured fish species. We have studied the desaturation/elongation and β-oxidation of 14C-linolenic (LNA) and 14C-eicosapentaenoic (EPA) acids in hepatocytes and pyloric caecal enterocytes in European sea bass fed diets with partial substitution (60%) of fish oil (FO) with vegetable oils (rapeseed, linseed and palm oil) blended in different proportions, for 64 weeks. The rate of desaturation of 14C-LNA was very low in hepatocytes from all treatments and no significant differences were observed among treatments. The rate of desaturation of 14C-LNA in enterocytes was higher than that in hepatocytes but still low (less than 5% of total radioactivity recovered). The desaturation of 14C-EPA in enterocytes was also higher than in hepatocytes, but again was low and no significant differences were found among treatments. The rates of β-oxidation of 14C-LNA and 14C-EPA were much higher than the rates of desaturation in both hepatocytes and enterocytes; however, no significant differences were observed in either hepatocytes or enterocytes among treatments. The rates of β-oxidation of 14C-LNA were considerably higher than those of 14C-EPA in both hepatocytes and enterocytes. In conclusion, European sea bass (a carnivorous marine fish) showed very low desaturation and elongation of LNA to EPA and DHA, and EPA to DHA, higher β-oxidation of LNA than EPA, and all desaturation and oxidation activities were significantly higher in enterocytes than in hepatocytes. A second major conclusion is that no clear quantitative nutritional effects on the desaturation/elongation and β-oxidation activities in either hepatocytes or enterocytes of sea bass were observed upon the inclusion of vegetable oils in the diet.

Fatty acid metabolism in Atlantic salmon ( Salmo salar L.) hepatocytes and influence of dietary vegetable oil

Isolated hepatocytes from Atlantic salmon ( Salmo salar), fed diets containing either 100% fish oil or a vegetable oil blend replacing 75% of the fish oil, were incubated with a range of seven 14C-labelled fatty acids. The fatty acids were [1- 14C]16:0, [1- 14C]18:1n-9, 91- 14C]18:2n-6, [1- 14C]18:3n-3, [1- 14C]20:4n-6, [1- 14C]20:5n-3, and [1- 14C]22:6n-3. After 2 h of incubation, the hepatocytes and medium were analysed for acid soluble products, incorporation into lipid classes, and hepatocytes for desaturation and elongation. Uptake into hepatocytes was highest with [1- 14C]18:2n-6 and [1- 14C]20:5n-3 and lowest with [1- 14C]16:0. The highest recovery of radioactivity in the cells was found in triacylglycerols. Of the phospholipids, the highest recovery was found in phosphatidylcholine, with [1- 14C]16:0 and [1- 14C]22:6n-3 being the most prominent fatty acids. The rates of β-oxidation were as follows: 20:4n-6 > 18:2n-6 = 16:0 > 18:1n-9 > 22:6n-3 = 18:3n-3 = 20:5n-3. Of the fatty acids taken up by the hepatocytes, [1- 14C]16:0 and [1- 14C]18:1n-9 were subsequently exported the most, with the majority of radioactivity recovered in phospholipids and triacylglycerols, respectively. The major products from desaturation and elongation were generally one cycle of elongation of the fatty acids. Diet had a clear effect on the overall lipid metabolism, with replacing 75% of the fish oil with vegetable oil resulting in decreased uptake of all fatty acids and reduced incorporation of fatty acids into cellular lipids, but increased β-oxidation activity and higher recovery in products of desaturation and elongation of [1- 14C]18:2n-6 and [1- 14C]18:3n-3.

Hepatic β-oxidation and carnitine palmitoyltransferase I in neonatal pigs after dietary treatments of clofibric acid, isoproterenol, and medium-chain triglycerides

A suckling piglet model was used to study nutritional and pharmacologic means of stimulating hepatic fatty acid beta-oxidation. Newborn pigs were fed milk diets containing either long- or medium-chain triglycerides (LCT or MCT). The long-chain control diet was supplemented further with clofibric acid (0.5%) or isoproterenol (40 ppm), and growth was monitored for 10-12 days. Clofibrate increased rates of hepatic peroxisomal and mitochondrial beta-oxidation of [1-(14)C]-palmitate by 60 and 186%, respectively. Furthermore, malonyl-CoA sensitive carnitine palmitoyltransferase (CPT I) activity increased 64% (P < 0.05) in pigs receiving clofibrate. Increased CPT I activity was not congruent with changes in message, as elevated abundance of CPT I mRNA was not detected (P = 0.16) when assessed by qRT-PCR. Neither rates of beta-oxidation nor CPT activities were affected by dietary MCT or by isoproterenol treatment (P > 0.1). Collectively, these findings indicate that clofibrate effectively induced hepatic CPT activity concomitant with increased fatty acid beta-oxidation.

Kinetics of LCFA Inhibition on Acetoclastic Methanogenesis, Propionate Degradation and β-Oxidation

Kinetics of long-chain fatty acids (LCFAs) inhibition on acetoclastic methanogenesis, propionate degradation and β-oxidation were studied with granular sludge under mesophilic batch conditions. Mathematical expressions used for reaction rates were as shown below: The simulated results revealed that the methane production rates from acetate decreased with an increase in both concentration and the number of double bonds of LCFAs. The concentrations of oleate (C18:1), linoleate (C18:2), palmitate (C16:0), and stearate (C18:0) were 0.54 mM, 0.11 mM 1.62 mM, and 2.58 mM, respectively, at which the methane production rates from acetate dropped 10%, and 3.10 mM, 0.72 mM, 5.71 mM, and 5.37 mM, respectively, at which the rates dropped 50%. The inhibitory effects of LCFAs on propionate degradation showed a similar tendency with acetoclastic methanogenesis; however, were less severe. The concentrations of oleate, linoleate, palmitate, and stearate were 1.02 mM, 0.18 mM, 2.34 mM, and 1.92 mM, respectively, at which the propionate degradation rates dropped 10%, and 4.38 mM, 1.17 mM, 5.88 mM, and 5.18 mM, respectively, at which the rates dropped 50%. The observed maximum β-oxidation rates of oleate, linoleate, palmitate, and stearate were 0.21 mmol (g VSS)−1 d−1, 0.09 mmol (g VSS)−1 d−1, 0.12 mmol (g VSS)−1 d−1, and 0.08 mmol (g VSS)−1 d−1, respectively. The lag-phase times in β-oxidation were also dependent on LCFA concentrations. The concentrations of oleate, linoleate, palmitate, and stearate, at which the lag-phase times became 5 days, were 5.93 mM, 2.24 mM, 4.02 mM, and 2.81 mM, respectively.

Stearoyl-CoA desaturase-1 deficiency reduces ceramide synthesis by downregulating serine palmitoyltransferase and increasing β-oxidation in skeletal muscle

Stearoyl-CoA desaturase (SCD) has recently been shown to be a critical control point of lipid partitioning and body weight regulation. Lack of SCD1 function significantly increases insulin sensitivity in skeletal muscles and corrects the hypometabolic phenotype of leptin-deficient ob/ob mice, indicating the direct antilipotoxic action of SCD1 deficiency. The mechanism underlying the metabolic effects of SCD1 mutation is currently unknown. Here we show that SCD1 deficiency reduced the total ceramide content in oxidative skeletal muscles (soleus and red gastrocnemius) by approximately 40%. The mRNA levels and activity of serine palmitoyltransferase (SPT), a key enzyme in ceramide synthesis, as well as the incorporation of [14C]palmitate into ceramide were decreased by approximately 50% in red muscles of SCD1-/- mice. The content of fatty acyl-CoAs, which contribute to de novo ceramide synthesis, was also reduced. The activity and mRNA levels of carnitine palmitoyltransferase I (CPT I) and the rate of beta-oxidation were increased in oxidative muscles of SCD1-/- mice. Furthermore, SCD1 deficiency increased phosphorylation of AMP-activated protein kinase (AMPK), suggesting that AMPK activation may be partially responsible for the increased fatty acid oxidation and decreased ceramide synthesis in red muscles of SCD1-/- mice. SCD1 deficiency also reduced SPT activity and ceramide content and increased AMPK phosphorylation and CPT I activity in muscles of ob/ob mice. Taken together, these results indicate that SCD1 deficiency reduces ceramide synthesis by decreasing SPT expression and increasing the rate of beta-oxidation in oxidative muscles.

Comparative study on biodegradability of polyethylene wax by bacteria and fungi

Biodegradability of polyethylene wax (PEwax) was studied comparatively between degraders in two different classes: bacteria and fungi. Gel permeation chromatography patterns were obtained before and after cultivation of a bacterial consortium KH-12 and a fungus, Aspergillus sp. AK-3 in culture media containing PEwax as their sole carbon sources for three weeks, and they were introduced into a mathematical model based on two primary factors: the gradual weight loss of large molecules due to β-oxidation processes and the direct consumption of small molecules by cells. The total consumption rates and the β-oxidation rates for both of these degraders were determined numerically, and their degradability was compared.