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Pharmacological inhibition of mitochondrial fatty acid oxidation (FAO) has been clinically used to alleviate certain metabolic diseases by remodeling cellular metabolism. However, mitochondrial FAO inhibition also leads to mechanistic target of rapamycin complex 1 (mTORC1) activation-related protein synthesis and tissue hypertrophy, but the mechanism remains unclear. Here, by using a mitochondrial FAO inhibitor (mildronate or etomoxir) or knocking out carnitine palmitoyltransferase-1, we revealed that mitochondrial FAO inhibition activated the mTORC1 pathway through general control nondepressible 5-dependent Raptor acetylation. Mitochondrial FAO inhibition significantly promoted glucose catabolism and increased intracellular acetyl-CoA levels. In response to the increased intracellular acetyl-CoA, acetyltransferase general control nondepressible 5 activated mTORC1 by catalyzing Raptor acetylation through direct interaction. Further investigation also screened Raptor deacetylase histone deacetylase class II and identified histone deacetylase 7 as a potential regulator of Raptor. These results provide a possible mechanistic explanation for the mTORC1 activation after mitochondrial FAO inhibition and also bring light to reveal the roles of nutrient metabolic remodeling in regulating protein acetylation by affecting acetyl-CoA production., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: This study assessed the efficacy of INV-202, a novel peripherally restricted cannabinoid type-1 receptor (CB1R) inverse agonist, in a streptozotocin-induced type-1 diabetes nephropathy mouse model., Methods: Diabetes was induced in 8-week-old C57BL6/J male mice via intraperitoneal injection of streptozotocin (45 mg/kg/day for 5 days); nondiabetic controls received citrate buffer. Diabetic mice were randomized to 3 groups based on blood glucose, polyuria, and albuminuria, and administered daily oral doses for 28-days of INV-202 at 0.3 or 3 mg/kg or vehicle., Results: INV-202 did not affect body weight but decreased kidney weight compared with the vehicle group. While polyuria was unaffected by INV-202 treatment, urinary urea (control 30.77 ± 14.93; vehicle 189.81 ± 31.49; INV-202 (0.3 mg/kg) 127.76 ± 20; INV-202 (3 mg/kg) 93.70 ± 24.97 mg/24h) and albumin (control 3.06 ± 0.38; vehicle 850.08 ± 170.50; INV-202 (0.3 mg/kg) 290.65 ± 88.70; INV-202 (3 mg/kg) 111.29 ± 33.47 µg/24h) excretion both decreased compared with vehicle-treated diabetic mice. Compared with the vehicle group, there was a significant improvement in the urinary albumin to creatinine ratio across INV-202 groups. Regardless of the dose, INV-202 significantly reduced angiotensin II excretion in diabetic mice. The treatment also decreased Agtr1a renal expression in a dose-dependent manner. Compared with nondiabetic controls, the glomerular filtration rate was increased in the vehicle group and significantly decreased by INV-202 at 3 mg/kg. While the vehicle group showed a significant loss in the mean number of podocytes per glomerulus, INV-202 treatment limited podocyte loss in a dose-dependent manner. Moreover, in both INV-202 groups, expression of genes coding for podocyte structural proteins nephrin ( Nphs1 ), podocin ( Nphs2 ), and podocalyxin ( Pdxl ) were restored to levels similar to nondiabetic controls. INV-202 partially limited the proximal tubular epithelial cell (PTEC) hyperplasia and normalized genetic markers for PTEC lesions. INV-202 also reduced expression of genes contributing to oxidative stress ( Nox2 , Nox4 , and P47phox ) and inflammation ( Tnf ). In addition, diabetes-induced renal fibrosis was significantly reduced by INV-202., Conclusions: INV-202 reduced glomerular injury, preserved podocyte structure and function, reduced injury to PTECs, and ultimately reduced renal fibrosis in a streptozotocin-induced diabetic nephropathy mouse model. These results suggest that INV-202 may represent a new therapeutic option in the treatment of diabetic kidney disease., Competing Interests: Author GC is the Chief Medical Officer at Inversago Pharma, Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declare that this study received funding from Inversago Pharma, Inc. The funder had the following involvement with the study: study design., (Copyright © 2023 Jacquot, Pointeau, Roger-Villeboeuf, Passilly-Degrace, Belkaid, Regazzoni, Leemput, Buch, Demizieux, Vergès, Degrace, Crater and Jourdan.)

Targeting cannabinoid 1 receptors (CB1R) with peripherally restricted antagonists (or inverse agonists) shows promise to improve metabolic disorders associated with obesity. In this context, we designed and synthetized JM-00266, a new CB1R blocker with limited blood-brain barrier (BBB) permeability. Pharmacokinetics were tested with SwissADME and in vivo in rodents after oral and intraperitoneal administration of JM-00266 in comparison with Rimonabant. In silico predictions indicated JM-00266 is a non-brain penetrant compound and this was confirmed by brain/plasma ratios and brain uptake index values. JM-00266 had no impact on food intake, anxiety-related behavior and body temperature suggesting an absence of central activity. cAMP assays performed in CB1R-transfected HEK293T/17 cells showed that the drug exhibited inverse agonist activity on CB1R. In addition, JM-00266 counteracted anandamide-induced gastroparesis indicating substantial peripheral activity. Acute administration of JM-00266 also improved glucose tolerance and insulin sensitivity in wild-type mice, but not in CB1R -/- mice. Furthermore, the accumulation of JM-00266 in adipose tissue was associated with an increase in lipolysis. In conclusion, JM-00266 or derivatives can be predicted as a new candidate for modulating peripheral endocannabinoid activity and improving obesity-related metabolic disorders.

White adipose tissue (WAT) possesses the endocannabinoid system (ECS) machinery and produces the two major endocannabinoids (ECs), arachidonoylethanolamide (AEA) and 2-arachidonoylglycerol (2-AG). Accumulating evidence indicates that WAT cannabinoid 1 receptors (CB1R) are involved in the regulation of fat storage, tissue remodeling and secretory functions but their role in controlling lipid mobilization is unclear. In the present study, we used different strategies to acutely increase ECS activity in WAT and tested the consequences on glycerol production as a marker of lipolysis. Treating lean mice or rat WAT explants with JLZ195, which inhibits ECs degrading enzymes, induced an increase in 2-AG tissue contents that was associated with a CB1R-dependent decrease in lipolysis. Direct treatment of rat WAT explants with AEA also inhibited glycerol production while mechanistic studies revealed it could result from the stimulation of Akt-signaling pathway. Interestingly, AEA treatment decreased lipolysis both in visceral and subcutaneous WAT collected on lean subjects suggesting that ECS also reduces fat store mobilization in Human. In obese mice, WAT content and secretion rate of ECs were higher than in control while glycerol production was reduced suggesting that over-produced ECs may inhibit lipolysis activating local CB1R. Strikingly, our data also reveal that acute CB1R blockade with Rimonabant did not modify lipolysis in vitro in obese mice and human explants nor in vivo in obese mice. Taken together, these data provide physiological evidence that activation of ECS in WAT, by limiting fat mobilization, may participate in the progressive tissue remodeling that could finally lead to organ dysfunction. The present findings also indicate that acute CB1R blockade is inefficient in regulating lipolysis in obese WAT and raise the possibility of an alteration of CB1R signaling in conditions of obesity., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Buch, Muller, Leemput, Passilly-Degrace, Ortega-Deballon, Pais de Barros, Vergès, Jourdan, Demizieux and Degrace.)

Objective: Dyslipidemia observed in type 2 diabetes (T2D) is atherogenic. Important features of diabetic dyslipidemia are increased levels of triglyceride-rich lipoproteins and small dense LDL particles, which all have apolipoprotein B100 (apoB100) as a major apolipoprotein. This prompted us to study the effect of the GLP-1 agonist liraglutide on the metabolism of apoB100-containing lipoproteins., Research Design and Methods: We performed an in vivo kinetic study with stable isotopes (L-[1- 13 C]leucine) in 10 patients with T2D before and after 6 months of treatment with liraglutide (1.2 mg/day). We also evaluated in mice the effect of liraglutide on the expression of genes involved in apoB100-containing lipoprotein clearance., Results: In patients with T2D, liraglutide treatment significantly reduced plasma apoB100 (0.93 ± 0.13 vs. 1.09 ± 0.11 g/L, P = 0.011) and fasting triglycerides (1.76 ± 0.37 vs. 2.48 ± 0.69 mmol/L, P = 0.005). The kinetic study showed a significant increase in indirect catabolism of VLDL 1 -apoB100 (4.11 ± 1.91 vs. 2.96 ± 1.61 pools/day, P = 0.005), VLDL 2 -apoB100 (5.17 ± 2.53 vs. 2.84 ± 1.65 pools/day, P = 0.008), and IDL-apoB100 (5.27 ± 2.77 vs. 3.74 ± 1.85 pools/day, P = 0.017) and in catabolism of LDL-apoB100 (0.72 ± 0.22 vs. 0.56 ± 0.22 pools/day, P = 0.005). In mice, liraglutide increased lipoprotein lipase (LPL) gene expression and reduced proprotein convertase subtilisin/kexin type 9 (PCSK9), retinol-binding protein 4 (RBP4), and tumor necrosis factor-α (TNF-α) gene expression in adipose tissue and decreased PCSK9 mRNA and increased LDL receptor protein expression in liver. In vitro, liraglutide directly reduced the expression of PCSK9 in the liver., Conclusions: Treatment with liraglutide induces a significant acceleration of the catabolism of triglyceride-rich lipoproteins (VLDL 1 , VLDL 2 , IDL) and LDL. Liraglutide modifies the expression of genes involved in apoB100-containing lipoprotein catabolism. These positive effects on lipoprotein metabolism may reduce cardiovascular risk in T2D., (© 2021 by the American Diabetes Association.)

Diabetic dyslipidemia, characterized by increased plasma triglycerides and decreased HDL cholesterol levels, is a major factor contributing to nonalcoholic steatohepatitis and cardiovascular risk in type 2 diabetes. Activation of the cannabinoid-1 receptor (CB1R) and activation of inducible nitric oxide synthase (iNOS) are associated with nonalcoholic steatohepatitis progression. Here, we tested whether dual-targeting inhibition of hepatic CB1R and iNOS improves diabetic dyslipidemia in mice with diet-induced obesity (DIO mice). DIO mice were treated for 14 days with (S) -MRI-1867, a peripherally restricted hybrid inhibitor of CB1R and iNOS. (R) -MRI-1867, the CB1R-inactive stereoisomer that retains iNOS inhibitory activity, and JD-5037, a peripherally restricted CB1R antagonist, were used to assess the relative contribution of the two targets to the effects of (S) -MRI-1867. (S) -MRI-1867 reduced hepatic steatosis and the rate of hepatic VLDL secretion, upregulated hepatic LDLR expression, and reduced the circulating levels of proprotein convertase subtilisin/kexin type 9 (PCSK9). The decrease in VLDL secretion could be attributed to CB1R blockade, while the reduction of PCSK9 levels and the related increase in LDLR resulted from iNOS inhibition via an mTOR complex 1-dependent mechanism. In conclusion, this approach based on the concomitant inhibition of CB1R and iNOS represents a promising therapeutic strategy for the treatment of dyslipidemia., (© 2020 by the American Diabetes Association.)

Background: Fish cannot use carbohydrate efficiently and instead utilize protein for energy supply, thus limiting dietary protein storage. Protein deposition is dependent on protein turnover balance, which correlates tightly with cellular energy homeostasis. Mitochondrial fatty acid β-oxidation (FAO) plays a crucial role in energy metabolism. However, the effect of remodeled energy homeostasis caused by inhibited mitochondrial FAO on protein deposition in fish has not been intensively studied., Objectives: This study aimed to identify the regulatory role of mitochondrial FAO in energy homeostasis maintenance and protein deposition by studying lipid, glucose, and protein metabolism in fish., Methods: Carnitine-depleted male Nile tilapia (initial weight: 4.29 ± 0.12 g; 3 mo old) were established by feeding them with mildronate diets (1000 mg/kg/d) for 6 wk. Zebrafish deficient in the carnitine palmitoyltransferase 1b gene (cpt1b) were produced by using CRISPR/Cas9 gene-editing technology, and their males (154 ± 3.52 mg; 3 mo old) were used for experiments. Normal Nile tilapia and wildtype zebrafish were used as controls. We assessed nutrient metabolism and energy homeostasis-related biochemical and molecular parameters, and performed 14C-labeled nutrient tracking and transcriptomic analyses., Results: The mitochondrial FAO decreased by 33.1-88.9% (liver) and 55.6-68.8% (muscle) in carnitine-depleted Nile tilapia and cpt1b-deficient zebrafish compared with their controls (P < 0.05). Notably, glucose oxidation and muscle protein deposition increased by 20.5-24.4% and 6.40-8.54%, respectively, in the 2 fish models compared with their corresponding controls (P < 0.05). Accordingly, the adenosine 5'-monophosphate-activated protein kinase/protein kinase B-mechanistic target of rapamycin (AMPK/AKT-mTOR) signaling was significantly activated in the 2 fish models with inhibited mitochondrial FAO (P < 0.05)., Conclusions: These data show that inhibited mitochondrial FAO in fish induces energy homeostasis remodeling and enhances glucose utilization and protein deposition. Therefore, fish with inhibited mitochondrial FAO could have high potential to utilize carbohydrate. Our results demonstrate a potentially new approach for increasing protein deposition through energy homeostasis regulation in cultured animals., (Copyright © The Author(s) on behalf of the American Society for Nutrition 2020.)

l-Carnitine is essential for mitochondrial β-oxidation and has been used as a lipid-lowering feed additive in humans and farmed animals. d-Carnitine is an optical isomer of l-carnitine and dl-carnitine has been widely used in animal feeds. However, the functional differences between l- and d-carnitine are difficult to study because of the endogenous l-carnitine background. In the present study, we developed a low-carnitine Nile tilapia model by treating fish with a carnitine synthesis inhibitor, and used this model to investigate the functional differences between l- and d-carnitine in nutrient metabolism in fish. l- or d-carnitine (0·4 g/kg diet) was fed to the low-carnitine tilapia for 6 weeks. l-Carnitine feeding increased the acyl-carnitine concentration from 3522 to 10 822 ng/g and alleviated the lipid deposition from 15·89 to 11·97 % in the liver of low-carnitine tilapia. However, as compared with l-carnitine group, d-carnitine feeding reduced the acyl-carnitine concentration from 10 822 to 5482 ng/g, and increased lipid deposition from 11·97 to 20·21 % and the mRNA expression of the genes involved in β-oxidation and detoxification in the liver. d-Carnitine feeding also induced hepatic inflammation, oxidative stress and apoptosis. A metabolomic investigation further showed that d-carnitine feeding increased glycolysis, protein metabolism and activity of the tricarboxylic acid cycle and oxidative phosphorylation. Thus, l-carnitine can be physiologically utilised in fish, whereas d-carnitine is metabolised as a xenobiotic and induces lipotoxicity. d-Carnitine-fed fish demonstrates increases in peroxisomal β-oxidation, glycolysis and amino acid degradation to maintain energy homeostasis. Therefore, d-carnitine is not recommended for use in farmed animals.

Obesity is one of the major public health issues, and its prevalence is steadily increasing all the world over. The endocannabinoid system (ECS) has been shown to be involved in the intake of palatable food via activation of cannabinoid 1 receptor (CB₁R). However, the involvement of lingual CB₁R in the orosensory perception of dietary fatty acids has never been investigated. In the present study, behavioral tests on CB₁R -/- and wild type (WT) mice showed that the invalidation of Cb₁r gene was associated with low preference for solutions containing rapeseed oil or a long-chain fatty acid (LCFA), such as linoleic acid (LA). Administration of rimonabant, a CB₁R inverse agonist, in mice also brought about a low preference for dietary fat. No difference in CD36 and GPR120 protein expressions were observed in taste bud cells (TBC) from WT and CB₁R -/- mice. However, LCFA induced a higher increase in [Ca 2+ ] i in TBC from WT mice than that in TBC from CB₁R -/- mice. TBC from CB₁R -/- mice also exhibited decreased Proglucagon and Glp-1r mRNA and a low GLP-1 basal level. We report that CB₁R is involved in fat taste perception via calcium signaling and GLP-1 secretion.

Objective- Treatment with liraglutide, a GLP-1 (glucagon-like peptide-1) agonist, has been shown to reduce postprandial lipidemia, an important feature of diabetic dyslipidemia. However, the underlying mechanisms for this effect remain unknown. This prompted us to study the effect of liraglutide on the metabolism of ApoB48 (apolipoprotein B48). Approach and Results- We performed an in vivo kinetic study with stable isotopes (D 8 -valine) in the fed state in 10 patients with type 2 diabetes mellitus before treatment and 6 months after the initiation of treatment with liraglutide (1.2 mg/d). We also evaluated, in mice, the effect of a 1-week liraglutide treatment on postload triglycerides and analysed in vitro on jejunum, the direct effect of liraglutide on the expression of genes involved in the biosynthesis of chylomicron. In diabetic patients, liraglutide treatment induced a dramatic reduction of ApoB48 pool (65±38 versus 162±87 mg; P=0.005) because of a significant decrease in ApoB48 production rate (3.02±1.33 versus 6.14±4.27 mg kg -1 d -1 ; P=0.009) and a significant increase in ApoB48 fractional catabolic rate (5.12±1.35 versus 3.69±0.75 pool d -1 ; P=0.005). One-week treatment with liraglutide significantly reduced postload plasma triglycerides in mice and liraglutide, in vitro, reduced the expression of ApoB48, DGAT1 (diacylglycerol O-acyltransferase 1), and MTP (microsomal transfer protein) genes. Conclusions- We show that treatment with liraglutide induces a significant reduction of the ApoB48 pool because of both a reduction of ApoB48 production and an increase in ApoB48 catabolism. In vitro, liraglutide reduces the expression of genes involved in chylomicron synthesis. These effects might benefit cardiovascular health. Clinical Trial Registration- URL: https://www.clinicaltrials.gov . Unique identifier: NCT02721888.

Impaired mitochondrial fatty acid β-oxidation has been correlated with many metabolic syndromes, and the metabolic characteristics of the mammalian models of mitochondrial dysfunction have also been intensively studied. However, the effects of the impaired mitochondrial fatty acid β-oxidation on systemic metabolism in teleost have never been investigated. In the present study, we established a low-carnitine zebrafish model by feeding fish with mildronate as a specific carnitine synthesis inhibitor [0.05% body weight (BW)/d] for 7 weeks, and the systemically changed nutrient metabolism, including carnitine and triglyceride (TG) concentrations, fatty acid (FA) β-oxidation capability, and other molecular and biochemical assays of lipid, glucose, and protein metabolism, were measured. The results indicated that mildronate markedly decreased hepatic carnitine concentrations while it had no effect in muscle. Liver TG concentrations increased by more than 50% in mildronate-treated fish. Mildronate decreased the efficiency of liver mitochondrial β-oxidation, increased the hepatic mRNA expression of genes related to FA β-oxidation and lipolysis, and decreased the expression of lipogenesis genes. Mildronate decreased whole body glycogen content, increased glucose metabolism rate, and upregulated the expression of glucose uptake and glycolysis genes. Mildronate also increased whole body protein content and hepatic mRNA expression of mechanistic target of rapamycin ( mtor ), and decreased the expression of a protein catabolism-related gene. Liver, rather than muscle, was the primary organ targeted by mildronate. In short, mildronate-induced hepatic inhibited carnitine synthesis in zebrafish caused decreased mitochondrial FA β-oxidation efficiency, greater lipid accumulation, and altered glucose and protein metabolism. This reveals the key roles of mitochondrial fatty acid β-oxidation in nutrient metabolism in fish, and this low-carnitine zebrafish model could also be used as a novel fish model for future metabolism studies.

Energy metabolism plays important roles in stress resistance and immunity in mammals, however, such functions have not been established in fish. In the present study, Nile tilapia (Oreochromis niloticus) was fed with mildronate, an inhibitor of mitochondrial fatty acid (FA) β-oxidation, for six weeks subsequently challenged with Aeromonas hydrophila and ammonia nitrogen exposure. Mildronate treatment reduced significantly l-carnitine concentration and mitochondrial FA β-oxidation efficiency, while it increased lipid accumulation in liver. The fish with inhibited hepatic FA catabolism had lower survival rate when exposed to Aeromonas hydrophila and ammonia nitrogen. Moreover, fish fed mildronate supplemented diet had lower immune enzymes activities and anti-inflammatory cytokine genes expressions, but had higher pro-inflammatory cytokine genes expressions. However, the oxidative stress-related biochemical indexes were not significantly affected by mildronate treatment. Taken together, inhibited mitochondrial FA β-oxidation impaired stress resistance ability in Nile tilapia mainly through inhibiting immune functions and triggering inflammation. This is the first study showing the regulatory effects of lipid catabolism on stress resistance and immune functions in fish., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Evidence has accumulated that obesity-related metabolic dysregulation is associated with overactivation of the endocannabinoid system (ECS), which involves cannabinoid receptor 1 (CB1R), in peripheral tissues, including adipose tissue (AT). The functional consequences of CB1R activation on AT metabolism remain unclear. Since excess fat mobilization is considered an important primary event contributing to the onset of insulin resistance, we combined in vivo and in vitro experiments to investigate whether activation of ECS could alter the lipolytic rate. For this purpose, the appearance of plasma glycerol was measured in wild-type and CB1R -/- mice after acute anandamide administration or inhibition of endocannabinoid degradation by JZL195. Additional experiments were conducted on rat AT explants to evaluate the direct consequences of ECS activation on glycerol release and signaling pathways. Treatments stimulated glycerol release in mice fasted for 6 h and injected with glucose but not in 24-h fasted mice or in CB1R -/- , suggesting that the effect was dependent on plasma insulin levels and mediated by CB1R. We concomitantly observed that Akt cascade activity was decreased, indicating an alteration of the antilipolytic action of insulin. Similar results were obtained with tissue explants exposed to anandamide, thus identifying CB1R of AT as a major target. This study indicates the existence of a functional interaction between CB1R and lipolysis regulation in AT. Further investigation is needed to test if the elevation of ECS tone encountered in obesity is associated with excess fat mobilization contributing to ectopic fat deposition and related metabolic disorders., (Copyright © 2017 the American Physiological Society.)

Evidence suggests that alterations of glucose and lipid homeostasis induced by obesity are associated with the elevation of endocannabinoid tone. The biosynthesis of the two main endocannabinoids, N-arachidonoylethanolamine and 2-arachidonoyl-glycerol, which derive from arachidonic acid, is influenced by dietary fatty acids (FAs). We investigated whether exposure to n-3 FA at a young age may decrease tissue endocannabinoid levels and prevent metabolic disorders induced by a later high-fat diet (HFD) challenge. Three-week-old mice received a 5% lipid diet containing lard, lard plus safflower oil, or lard plus linseed oil for 10 weeks. Then, mice were challenged with a 30% lard diet for 10 additional weeks. A low n-6/n-3 FA ratio in the early diet induces a marked decrease in liver endocannabinoid levels. A similar reduction was observed in transgenic Fat-1 mice, which exhibit high tissue levels of n-3 FA compared with wild-type mice. Hepatic expression of key enzymes involved in carbohydrate and lipid metabolism was concomitantly changed. Interestingly, some gene modifications persisted after HFD challenge and were associated with improved glycemic control. These findings indicate that early dietary interventions based on n-3 FA may represent an alternative strategy to drugs for reducing endocannabinoid tone and improving metabolic parameters in the metabolic syndrome., (© 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

The endocannabinoid system (ECS) is associated with an alteration of glucose homeostasis dependent on cannabinoid receptor-1 (CB1R) activation. However, very little information is available concerning the consequences of ECS activation on intestinal glucose absorption. Mice were injected intraperitoneally with anandamide, an endocannabinoid binding both CB1R and CB2R. We measured plasma glucose and xylose appearance after oral loading, gastrointestinal motility, and glucose transepithelial transport using the everted sac method. Anandamide improved hyperglycemia after oral glucose charge whereas glucose clearance and insulin sensitivity were impaired, pointing out some gastrointestinal events. Plasma xylose appearance was delayed in association with a strong decrease in gastrointestinal transit, while anandamide did not alter transporter-mediated glucose absorption. Interestingly, transit was nearly normalized by coinjection of SR141716 and AM630 (CB1R and CB2R antagonist, respectively), and AM630 also reduced the delay of plasma glucose appearance induced by anandamide. When gastric emptying was bypassed by direct glucose administration in the duodenum, anandamide still reduced plasma glucose appearance in wild-type but not in CB1R(-/-) mice. In conclusion, our findings demonstrated that acute activation of intestinal ECS reduced postprandial glycemia independently on intestinal glucose transport but rather inhibiting gastric emptying and small intestine motility and strongly suggest the involvement of both CB1R and CB2R., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

The peroxisomal 3-ketoacyl-CoA thiolase B (ThB) catalyzes the thiolytic cleavage of straight chain 3-ketoacyl-CoAs. Up to now, the ability of ThB to interfere with lipid metabolism was studied in mice fed a laboratory chow enriched or not with the synthetic agonist Wy14,643, a pharmacological activator of the nuclear hormone receptor PPARα. The aim of the present study was therefore to determine whether ThB could play a role in obesity and lipid metabolism when mice are chronically fed a synthetic High Fat Diet (HFD) or a Low Fat Diet (LFD) as a control diet. To investigate this possibility, wild-type (WT) mice and mice deficient for Thb (Thb(-/-)) were subjected to either a synthetic LFD or a HFD for 25 weeks, and their responses were compared. First, when fed a normal regulatory laboratory chow, Thb(-/-) mice displayed growth retardation as well as a severe reduction in the plasma level of Growth Hormone (GH) and Insulin Growth Factor-I (IGF-I), suggesting alterations in the GH/IGF-1 pathway. When fed the synthetic diets, the corrected energy intake to body mass was significantly higher in Thb(-/-) mice, yet those mice were protected from HFD-induced adiposity. Importantly, Thb(-/-) mice also suffered from hypoglycemia, exhibited reduction in liver glycogen stores and circulating insulin levels under the LFD and the HFD. Thb deficiency was also associated with higher levels of plasma HDL (High Density Lipoproteins) cholesterol and increased liver content of cholesterol under both the LFD and the HFD. As shown by the plasma lathosterol to cholesterol ratio, a surrogate marker for cholesterol biosynthesis, whole body cholesterol de novo synthesis was increased in Thb(-/-) mice. By comparing liver RNA from WT mice and Thb(-/-) mice using oligonucleotide microarray and RT-qPCR, a coordinated decrease in the expression of critical cholesterol synthesizing genes and an increased expression of genes involved in bile acid synthesis (Cyp7a1, Cyp17a1, Akr1d1) were observed in Thb(-/-) mice. In parallel, the elevation of the lathosterol to cholesterol ratio as well as the increased expression of cholesterol synthesizing genes were observed in the kidney of Thb(-/-) mice fed the LFD and the HFD. Overall, the data indicate that ThB is not fully interchangeable with the thiolase A isoform. The present study also reveals that modulating the expression of the peroxisomal ThB enzyme can largely reverberate not only throughout fatty acid metabolism but also cholesterol, bile acid and glucose metabolism., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

Reduced mitochondrial fatty acid (FA) β-oxidation can cause accumulation of triglyceride in liver, while intake of eicosapentaenoic acid (EPA) has been recommended as a promising novel therapy to decrease hepatic triglyceride content. However, reduced mitochondrial FA β-oxidation also facilitates accumulation of EPA. To investigate the interplay between EPA administration, mitochondrial activity and hepatic triglyceride accumulation, we investigated the effects of EPA administration to carnitine-deficient mice with impaired mitochondrial FA β-oxidation. C57BL/6J mice received a high-fat diet supplemented or not with 3% EPA in the presence or absence of 500 mg mildronate/kg/day for 10 days. Liver mitochondrial and peroxisomal oxidation, lipid classes and FA composition were determined. Histological staining was performed and mRNA level of genes related to lipid metabolism and inflammation in liver and adipose tissue was determined. Levels of pro-inflammatory eicosanoids and cytokines were measured in plasma. The results showed that mildronate treatment decreased hepatic carnitine concentration and mitochondrial FA β-oxidation and induced severe triglyceride accumulation accompanied by elevated systemic inflammation. Surprisingly, inclusion of EPA in the diet exacerbated the mildronate-induced triglyceride accumulation. This was accompanied by a considerable increase of EPA accumulation while decreased total n-3/n-6 ratio in liver. However, inclusion of EPA in the diet attenuated the mildronate-induced mRNA expression of inflammatory genes in adipose tissue. Taken together, dietary supplementation with EPA exacerbated the triglyceride accumulation induced by impaired mitochondrial FA β-oxidation. Thus, further thorough evaluation of the potential risk of EPA supplementation as a therapy for NAFLD associated with impaired mitochondrial FA oxidation is warranted., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Unlabelled: It is well established that inactivation of the central endocannabinoid system (ECS) through antagonism of cannabinoid receptor 1 (CB1R) reduces food intake and improves several pathological features associated with obesity, such as dyslipidemia and liver steatosis. Nevertheless, recent data indicate that inactivation of peripheral CB1R could also be directly involved in the control of lipid metabolism independently of central CB1R. To further investigate this notion, we tested the direct effect of the specific CB1R antagonist, SR141716, on hepatic carbohydrate and lipid metabolism using cultured liver slices. CB1R messenger RNA expression was strongly decreased by SR141716, whereas it was increased by the CB1R agonist, arachidonic acid N-hydroxyethylamide (AEA), indicating the effectiveness of treatments in modulating ECS activity in liver explants both from lean or ob/ob mice. The measurement of O(2) consumption revealed that SR141716 increased carbohydrate or fatty acid utilization, according to the cellular hormonal environment. In line with this, SR141716 stimulated ß-oxidation activity, and the role of CB1R in regulating this pathway was particularly emphasized when ECS was hyperactivated by AEA and in ob/ob tissue. SR141716 also improved carbohydrate and lipid metabolism, blunting the AEA-induced increase in gene expression of proteins related to lipogenesis. In addition, we showed that SR141716 induced cholesterol de novo synthesis and high-density lipoprotein uptake, revealing a relationship between CB1R and cholesterol metabolism., Conclusion: These data suggest that blocking hepatic CB1R improves both carbohydrate and lipid metabolism and confirm that peripheral CB1R should be considered as a promising target to reduce cardiometabolic risk in obesity., (Copyright © 2011 American Association for the Study of Liver Diseases.)

The membrane glycoprotein CD36 binds nanomolar concentrations of long chain fatty acids (LCFA) and is highly expressed on the luminal surface of enterocytes. CD36 deficiency reduces chylomicron production through unknown mechanisms. In this report, we provide novel insights into some of the underlying mechanisms. Our in vivo data demonstrate that CD36 gene deletion in mice does not affect LCFA uptake and subsequent esterification into triglycerides by the intestinal mucosa exposed to the micellar LCFA concentrations prevailing in the intestine. In rodents, the CD36 protein disappears early from the luminal side of intestinal villi during the postprandial period, but only when the diet contains lipids. This drop is significant 1 h after a lipid supply and associates with ubiquitination of CD36. Using CHO cells expressing CD36, it is shown that the digestion products LCFA and diglycerides trigger CD36 ubiquitination. In vivo treatment with the proteasome inhibitor MG132 prevents the lipid-mediated degradation of CD36. In vivo and ex vivo, CD36 is shown to be required for lipid activation of ERK1/2, which associates with an increase of the key chylomicron synthesis proteins, apolipoprotein B48 and microsomal triglyceride transfer protein. Therefore, intestinal CD36, possibly through ERK1/2-mediated signaling, is involved in the adaptation of enterocyte metabolism to the postprandial lipid challenge by promoting the production of large triglyceride-rich lipoproteins that are rapidly cleared in the blood. This suggests that CD36 may be a therapeutic target for reducing the postprandial hypertriglyceridemia and associated cardiovascular risks.

Elaidic acid (trans-9-C₁₈:₁ or trans-9) is assumed to exert atherogenic effects due to its double bond configuration. The possibility that trans-9 and vaccenic acid (trans-11-C₁₈:₁ or trans-11), its positional isomer, were biochemically equivalent and interchangeable compounds, was investigated by reference to their cis-isomers through esterification-related activities using rat liver cells and subcellular fractions. In hepatocytes, both trans-C₁₈:₁ were incorporated to the same extent in triacylglycerols, but trans-9 was more esterified than trans-11 into phospholipids (P < 0.05). Glycerol-3-phosphate acyltransferase activity in microsomes was lower with trans-11 than with trans-9, while this activity in mitochondria was ~40% greater with trans-11 than with trans-9 (P < 0.05). Activity of 2-lysophosphatidic acid acyltransferase in microsomes was of comparable extent with both trans isomers, but activity of 2-lysophosphatidylcholine acyltransferase was significantly greater with trans-9 than with trans-11 at P < 0.01. Lipoproteins secreted by hepatocytes reached equivalent levels in the presence of any isomers, but triacylglycerol production was more elevated with trans-11 than with trans-9 at P < 0.05. Cholesterol efflux from previously labelled hepatocytes was lower with trans-11 than with trans-9. When these cells were exposed to either trans-C₁₈:₁, the gene expression of proteins involved in fatty acid esterification and lipoprotein synthesis was unaffected, which indicates that the biochemical differences essentially depended on enzyme/substrate affinities. On the whole, vaccenic and elaidic acid were shown to incorporate cell phospholipids unequally, at least in vitro, which suggests they can differently affect lipid metabolic pathways in normal cells.

Peroxisomal 3-ketoacyl-CoA thiolase B (Thb) catalyzes the final step in the peroxisomal β-oxidation of straight-chain acyl-CoAs and is under the transcription control of the nuclear hormone receptor PPARα. PPARα binds to and is activated by the synthetic compound Wy14,643 (Wy). Here, we show that the magnitude of Wy-mediated induction of peroxisomal β-oxidation of radiolabeled (1-(14)C) palmitate was significantly reduced in mice deficient for Thb. In contrast, mitochondrial β-oxidation was unaltered in Thb(-/-) mice. Given that Wy-treatment induced Acox1 and MFP-1/-2 activity at a similar level in both genotypes, we concluded that the thiolase step alone was responsible for the reduced peroxisomal β-oxidation of fatty acids. Electron microscopic analysis and cytochemical localization of catalase indicated that peroxisome proliferation in the liver after Wy-treatment was normal in Thb(-/-) mice. Intriguingly, micro-array analysis revealed that mRNA levels of genes encoding cholesterol biosynthesis enzymes were upregulated by Wy in Wild-Type (WT) mice but not in Thb(-/-) mice, which was confirmed at the protein level for the selected genes. The non-induction of genes encoding cholesterol biosynthesis enzymes by Wy in Thb(-/-) mice appeared to be unrelated to defective SREBP-2 or PPARα signaling. No difference was observed in the plasma lathosterol/cholesterol ratio (a marker for de novo cholesterol biosynthesis) between Wy-treated WT and Thb(-/-) mice, suggesting functional compensation. Overall, we conclude that ThA and SCPx/SCP2 thiolases cannot fully compensate for the absence of ThB. In addition, our data indicate that ThB is involved in the regulation of genes encoding cholesterol biosynthesis enzymes in the liver, suggesting that the peroxisome could be a promising candidate for the correction of cholesterol imbalance in dyslipidemia., (Copyright © 2011 Elsevier Masson SAS. All rights reserved.)

Vaccenic acid (trans-11-C(18:1)) chemically resembles elaidic acid (trans-9-C(18:1)) which is assumed to increase the risk of cardiovascular diseases, and thus could exert similar effects. Possible different oxidation rates of vaccenic versus elaidic acid were checked in muscles and liver, and through related gene expression in normal rat liver cells. In hepatic mitochondria, carnitine palmitoyltransferase (CPT) I exhibited comparable activity rates with both trans-isomers. CPT II activity was 30% greater (P < 0.05) with vaccenic than with elaidic acid as nonesterified fatty acids (NEFAs) or acyl-CoAs. Activity of the first beta-oxidation step was similar between the isomers in all the tissue slices and liver extracts assayed. Respiration rates were comparable with both trans-isomers as NEFAs in various liver extracts, but were 30% greater (P < 0.05) with vaccenoyl-CoA than with elaidoyl-CoA in liver mitochondria. Vaccenic acid was oxidised 25% more (P < 0.05) by liver peroxisomes than elaidic acid. In hepatocytes cultured with trans- and corresponding cis-C(18:1) isomers, gene expression of CPT I, hydroxyacyl-CoA dehydrogenase and hydroxymethylglutaryl-CoA synthase was at least 100% increased (P < 0.05), but was unchanged with vaccenic acid, relative to controls. In conclusion, the position and geometry of the double bonds in acyl chains are suggested to confer on vaccenic and elaidic acid specific biochemical properties that might differently affect their fates in tissues.

Background: Pioglitazone (PIO) and rosiglitazone (ROSI) are widely used as oral antidiabetic agents for treatment of type 2 diabetes. Although these medications exert similar effects on blood glucose, recent clinical studies indicated that PIO has a more pronounced beneficial effect on lipid parameters than ROSI. In order to get further insight into the lipid effects of both drugs, we tested whether PIO, compared to ROSI, could exert direct effects on lipid liver metabolism in relation with plasma lipids., Methods: We performed in vitro studies using mice liver slices incubated 21 h either with ROSI (1 micromol/L) or PIO (7.5 micromol/L)., Results: We showed that both glitazones slightly reduced HMG-CoA reductase mRNA levels at the same degree but only PIO reduced intracellular cholesterol content, suggesting an alteration of cholesterol uptake rather than an inhibition of cholesterol biosynthesis. This concept was supported by the reduction of scavenger receptor class B type I expression, hepatic lipase activity and high-density lipoprotein cholesterol uptake in PIO-treated liver explants. Conversely, hepatic lipase mRNA levels were increased 3.5-fold. ROSI, but not PIO, induced acetyl-CoA carboxylase and fatty acid synthase gene expression and increased apoB secretion suggesting a stimulation of lipogenesis. Concurrently, peroxisome proliferator-activated receptor-gamma mRNA levels were induced by ROSI and not significantly changed by PIO. Besides, PIO appeared to be a more potent activator of AMP-Activated Protein Kinase than ROSI., Conclusions: PIO and ROSI exert specific direct effects on liver and extrapolating these data to humans could explain the significant improvements in plasma lipids observed in diabetic patients treated with PIO.

Objective: The beneficial effects of the inactivation of endocannabinoid system (ECS) by administration of antagonists of the cannabinoid receptor (CB) 1 on several pathological features associated with obesity is well demonstrated, but the relative contribution of central versus peripheral mechanisms is unclear. We examined the impact of CB1 antagonism on liver and adipose tissue lipid metabolism in a mouse model of diet-induced obesity., Research Design and Methods: Mice were fed either with a standard diet or a high-sucrose high-fat (HSHF) diet for 19 weeks and then treated with the CB1-specific antagonist SR141716 (10 mg x kg(-1) x day(-1)) for 6 weeks., Results: Treatment with SR141716 reduced fat mass, insulin levels, and liver triglycerides primarily increased by HSHF feeding. Serum adiponectin levels were restored after being reduced in HSHF mice. Gene expression of scavenger receptor class B type I and hepatic lipase was induced by CB1 blockade and associated with an increase in HDL-cholesteryl ether uptake. Concomitantly, the expression of CB1, which was strongly increased in the liver and adipose tissue of HSHF mice, was totally normalized by the treatment. Interestingly, in visceral but not subcutaneous fat, genes involved in transport, synthesis, oxidation, and release of fatty acids were upregulated by HSHF feeding, while this effect was counteracted by CB1 antagonism., Conclusions: A reduction in the CB1-mediated ECS activity in visceral fat is associated with a normalization of adipocyte metabolism, which may be a determining factor in the reversion of liver steatosis induced by treatment with SR141716.

Feeding mice the trans-10, cis-12 (t10c12) conjugated linoleic acid (CLA) isomer is associated with lipodystrophy, insulin resistance, hyperinsulinemia, and liver steatosis. It has been hypothesized that CLA-induced liver steatosis is the result of increased hepatic lipogenesis stimulated by high insulin levels. We studied the effects of a 12-d t10c12CLA treatment (1 g/100 g diet) on liver carbohydrate and lipid metabolism in control and streptozotocin (STZ)-injected mice. STZ mice were characterized by insulin deficiency, hypertriglyceridemia, and depletion of liver triglyceride and glycogen. Remarkably, feeding t10c12CLA to diabetic mice (STZ-CLA) normalized these variables. Reconstitution of fat stores in the livers of STZ-CLA mice was associated with lower fatty acid (FA) oxidation rates and greater malonyl-CoA concentration than in STZ mice. FA translocase and VLDL receptor mRNA levels were greater in STZ-CLA than in STZ mice, suggesting that t10c12CLA increased liver lipid uptake. Phosphoenolpyruvate carboxykinase mRNA levels and AMP kinase phosphorylation were lower in STZ-CLA than in STZ mice, indicating that t10c12CLA may reduce glucogenic activity and promote glycogenesis in diabetic mice. Because glycemia and glucokinase expression were not modified by t10c12CLA treatment, we postulated that glycogen accumulation is likely not the result of an effect of t10c12CLA on plasma glucose utilization, but rather is due to the contribution of lactate, the concentration of which was higher in muscle of STZ-CLA mice. The results demonstrate that t10c12CLA stimulates liver lipid accumulation in the absence of insulin and, thus, suggest that t10c12CLA can improve liver carbohydrate and lipid metabolism in type I diabetic mice.

The study was undertaken to determine whether eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3), esterified in phospholipids (PL) as liposomes or in triglycerides (TG) as oil, exhibited comparable fates in liver lipids and whether these fates were associated with gene expressions related to fatty acid (FA) metabolism. PL and TG mixtures with close contents in EPA and DHA were administered to rats over 2 weeks. Most relevant events occurred after 3 days for both treatments. At that time, liposomes, compared with oil, increased the liver content in PL with a FA composition enriched in n-6 FA, comparable in DHA and much lower in EPA. Moreover, liposomes increased the activity and mRNA levels of carnitine palmitoyltransferase (CPT) I. In contrast, fish oil exerted opposite effects on CPT I and increased the genic expression of lipogenic enzymes. Liposomes, unlike fish oil, apparently increased the mRNA levels of acyl-CoA oxidase and the activity of the peroxisomal FA-oxidising system. Concomitantly, mRNA levels of hepatic lipoprotein receptors were increased with both diets, but intracellular proteins involved in free FA uptake and lipid synthesis were up-regulated only with liposome-treated rats. The quasi absence of EPA in hepatic PL of liposome-treated rats on the short term could result from increased beta-oxidation activities through metabolic regulations induced by more available free EPA and other PUFA.

We investigated whether the hypolipidaemic effect of fenofibrate and fasting observed in most omnivorous mammals may also apply to herbivorous fish. Grass carp (Ctenopharyngodon idella) fed a high-fat (8 %) diet exhibited a marked increase in blood lipids and body fat after 6 weeks. They were then treated with fenofibrate (100 mg/kg body weight) in the same high-fat diet for 2 weeks, followed by fasting for 1 week. Plasma lipid concentration, body fat amount, fatty acid composition, plasma thiobarbituric acid-reactive substances and some parameters related to hepatic fatty acid oxidation were measured, and liver samples were stained for histological examination. Fenofibrate treatment decreased TAG and cholesterol concentrations in plasma, total lipids of the whole body and liver, and EPA and DHA contents in tissues. Further, a mobilisation of mesenteric fat concomitant with an increase in hepatic peroxisomal fatty acid oxidation and lipid peroxidation was observed. Compared with fenofibrate treatment, fasting decreased body weight and plasma TAG, but not plasma cholesterol. It also reduced the fat content of the whole body and increased the EPA and DHA contents in the liver and other tissues. Fatty acid oxidation was stimulated by fasting in mitochondria, but not in peroxisomes. These data suggest that fenofibrate and fasting regulate the lipid metabolism in grass carp through different metabolic pathways. The grass carp is moderately responsive to a fibrate derivative in comparison with the well-known excess responsiveness of the rat model, and so it could be used for the study of lipid abnormalities as a herbivorous model.

Rats with carnitine deficiency due to trimethylhydrazinium propionate (mildronate) administered at 80 mg/100 g body weight per day for 10 days developed liver steatosis only upon fasting. This study aimed to determine whether the transient steatosis resulted from triglyceride accumulation due to the amount of fatty acids preserved through impaired fatty acid oxidation and/or from up-regulation of lipid exchange between liver and adipose tissue. In liver, mildronate decreased the carnitine content by approximately 13-fold and, in fasted rats, lowered the palmitate oxidation rate by 50% in the perfused organ, increased 9-fold the triglyceride content, and doubled the hepatic very low density lipoprotein secretion rate. Concomitantly, triglyceridemia was 13-fold greater than in controls. Hepatic carnitine palmitoyltransferase I activity and palmitate oxidation capacities measured in vitro were increased after treatment. Gene expression of hepatic proteins involved in fatty acid oxidation, triglyceride formation, and lipid uptake were all increased and were associated with increased hepatic free fatty acid content in treated rats. In periepididymal adipose tissue, mildronate markedly increased lipoprotein lipase and hormone-sensitive lipase activities in fed and fasted rats, respectively. On refeeding, carnitine-depleted rats exhibited a rapid decrease in blood triglycerides and free fatty acids, then after approximately 2 h, a marked drop of liver triglycerides and a progressive decrease in liver free fatty acids. Data show that up-regulation of liver activities, peripheral lipolysis, and lipoprotein lipase activity were likely essential factors for excess fat deposit and release alternately occurring in liver and adipose tissue of carnitine-depleted rats during the fed/fasted transition.

This study was designed to address the effects of a moderate consumption of beer on serum and liver lipid parameters and on the development of aortic lesions in a mouse model associated with a human atherogenic lipoprotein profile. LDLr(-/-) apoB(100/100) mice received each day during 12 weeks either water, mild beer (0.570g of ethanol/kg of body weight) or ethanol-free beer in a single pure dose. Serum and liver lipid parameters were analyzed and atherosclerotic lesions were estimated in heart and aorta through their total cholesterol content. mRNA levels of enzymes and receptors involved in lipoprotein uptake, in fatty acid esterification and oxidation, and in reverse cholesterol transport were also measured in the liver. Serum glucose, triglyceride (TG) and cholesterol levels were altered neither by ethanol-free beer nor by mild beer. Nevertheless, both beer treatments significantly increased HDL-cholesterol (HDL-C) and VLDL-C levels by reference to controls with no change in LDL-C levels. Liver TG contents were significantly decreased by either beer treatment. Cholesterol accumulation was attenuated in the whole aorta of mice treated with mild beer at p<0.05 and not significantly with ethanol-free beer. Heart cholesterol contents were comparable in the three series. Among the genes studied, only scavenger receptor-B1 was downregulated by both beer-based beverages. LDL receptor related protein, lecithin-cholesterol acyltransferase and sterol regulatory element-binding protein 2 were downregulated only by mild beer. The expression of other genes assayed was not altered. When administered in chronic and moderate dose, unidentified components of beer may exert beneficial effects towards atherosclerosis development through alteration of lipoprotein metabolism in LDLr(-/-) apoB(100/100) mice. This effect was slightly amplified by the presence of ethanol in beer.

This study explores the mechanisms responsible for the fatty liver setup in mice fed trans-10,cis-12 conjugated linoleic acid (t10c12 CLA), hypothesizing that an induction of low density lipoprotein receptor (LDLR) expression is associated with lipid accumulation. To this end, the effects of t10c12 CLA treatment on lipid parameters, serum lipoproteins, and expression of liver lipid receptors were measured in LDLR(-/-) apoB(100/100) mice as a model of human familial hypercholesterolemia itself depleted of LDLR. Mice were fed t10c12 CLA over 2 or 4 weeks. We first observed that the treatment induced liver steatosis, even in the absence of LDLR. Mice treated for 2 weeks exhibited hypertriglyceridemia with high levels of VLDL and HDL, whereas a 4 week treatment inversely induced a reduction of serum triglycerides (TGs), essentially through a decrease in VLDL levels. In the absence of LDLR, the mRNA levels of other proteins, such as VLDL receptor, lipoprotein lipase, and fatty acid translocase, usually not expressed in the liver, were upregulated, suggesting their involvement in the steatosis setup and lipoprotein clearance. The data also suggest that the TG-lowering effect induced by t10c12 CLA treatment was attributable to both the reduction of circulating free fatty acids in response to the severe lipoatrophy and the high capacity of liver to clear off plasma lipids.

High-fat diets may have favourable effects on growth of some carnivorous fish because of the protein-sparing effect of lipids, but high-fat diets also exert some negative impacts on flesh quality. The goal of the study was therefore to determine the effects of fat-enriched diets in juvenile grass carp (Ctenopharyngodon idella) as a typical herbivorous fish on growth and possible lipid metabolism alterations. Three isonitrogenous diets containing 2, 6 or 10 % of a mixture of lard, maize oil and fish oil (1:1:1, by weight) were applied to fish for 8 weeks in a recirculation system. Data show that feeding diets with increasing lipid levels resulted in lowered feed intake, decreased growth and feed efficiency, and increased mesenteric fat tissue weight. Concomitantly, alteration of lipoprotein synthesis and greater level of lipid peroxidation were apparent in blood. In liver, muscle and mesenteric fat tissue, the percentages of alpha-linolenic acid and DHA were significantly increased or tended to increase with higher dietary lipid levels. Biochemical activity measurements performed on liver showed that, with the increase in dietary lipid level, there was a decrease in both mitochondrial and peroxisomal fatty acid oxidation capacities, which might contribute, at least in part, to the specific accumulation of alpha-linolenic acid and DHA into cells more active in membrane building. On the whole, grass carp have difficulty in energetically utilising excess dietary fat, especially when enriched in n-3 PUFA that are susceptible to peroxidation.

Fish easily accumulate n-3 PUFA of exogenous origin, but the underlying mechanisms are not well established in the whole animal. This study was undertaken to investigate whether this feature was physiologically associated with mitochondrial and peroxisomal capacities that differentially affect FA oxidation. For this purpose, peroxisomal FA oxidation was increased by treating rainbow trout with fenofibrate, which strongly stimulates the peroxisome proliferator-activated receptor-a in rodents. Diets containing EPA and DHA, with or without fenofibrate added, were administered to male trout for 12 d. After treatment, neither liver hypertrophy nor accumulation of fat was apparent within the liver and muscle cells. However, fenofibrate treatment decreased the contents of EPA and DHA in the liver, white muscle, and intraperitoneal fat tissue, which represented (per whole body) at least 280 mg less than in controls. Carnitine-dependent palmitate oxidation rates, expressed per gram of liver, were slightly increased by fenofibrate when measured from tissue homogenates and were unchanged when calculated from isolated mitochondria, relative to control fish. The treatment altered neither carnitine palmitoyltransferase I activity rates, expressed per gram of liver, nor the sensitivity of the enzyme to malonyl-CoA inhibition, but did increase the malonyl-CoA content (+45%). Meanwhile, fenofibrate increased (by about 30%) the peroxisome-related activities, i.e., catalase, carnitine-independent palmitate oxidation, acyl-CoA oxidase, and the peroxisomal FA-oxidizing system, relative to the control group. The data strongly suggest that the induction of peroxisomal activities, some of which being able to oxidize very long chain FA, was responsible for the lower contents of EPA and DHA in the body lipids of fenofibrate-treated trout.

The metabolic and genic effects induced by a 20-fold lowering of carnitine content in the heart were studied in mildronate-treated rats. In the perfused heart, the proportion of palmitate taken up then oxidized was 5-10% lower, while the triacylglycerol (TAG) formation was 100% greater than in controls. The treatment was shown to increase the maximal capacity of heart homogenates to oxidize palmitate, the mRNA level of carnitine palmitoyltransferase I (CPT-I) isoforms, the specific activity of CPT-I in subsarcolemmal mitochondria and the total carnitine content of isolated mitochondria. Concomitantly, the increased mRNA expression of lipoprotein lipase, fatty acid translocase and enzymes of TAG synthesis was associated with a 5- and 2-times increase in serum TAG and free fatty acid contents, respectively. The compartmentation of carnitine at its main functional location was expected to allow the increased CPT-I activity to ensure in vivo correct fatty acid oxidation rates. All the inductions related to fatty acid transport, oxidation and esterification most likely stem from the abundance of blood lipids providing cardiomyocytes with more fatty acids.

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.

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.