Mono-2-ethylhexyl phthalate induces the expression of genes involved in fatty acid synthesis in HepG2 cells.

Mono-2-ethylhexyl phthalate (MEHP) is a major bioactive metabolite in the widely used industrial plasticizer diethylhexyl phthalate (DEHP) that has been found to be toxic to the liver. The aim of this study is to determine whether MEHP exposure can change the expression of fatty acid metabolism-related genes in HepG2 cells, which might be related to non-alcoholic fatty liver disease (NAFLD). The results revealed that exposure to MEHP promoted lipid accumulation in HepG2 cells. The levels of intracellular triglycerides in the hepatocytes increased after exposure to 0.8-100 µM MEHP for 24 h and 48 h. The genetic expressions of SREBP-1c, ChREBP, ACC1, FASN, and SCD significantly increased at 6 h after exposure to MEHP. At 24 h, the expression of the SREBP-1c and ChREBP genes remained increased, while the expression of the FASN and SCD genes decreased. At 48 h, the expression of SREBP-1c, ChREBP, ACC1, FASN, and SCD decreased. Furthermore, the levels of proteins including ACC1, FASN, SCD, and ChREBP (except SREBP-1c) increased at 24 h. These findings suggest that MEHP exposure can promote fatty acid synthesis in hepatocytes by regulating the expression of relevant genes and proteins, contributing to NAFLD.

ACC1 determines memory potential of individual CD4+ T cells by regulating de novo fatty acid biosynthesis.

Immunological memory is central to adaptive immunity and protection from disease. Changing metabolic demands as antigen-specific T cells transition from effector to memory cells have been well documented, but the cell-specific pathways and molecules that govern this transition are poorly defined. Here we show that genetic deletion of ACC1, a rate-limiting enzyme in fatty acid biosynthesis, enhances the formation of CD4+ T memory cells. ACC1-deficient effector helper T (Th) cells have similar metabolic signatures to wild-type memory Th cells, and expression of the gene encoding ACC1, Acaca, was inversely correlated with a memory gene signature in individual cells. Inhibition of ACC1 function enhances memory T cell formation during parasite infection in mice. Using single-cell analyses we identify a memory precursor-enriched population (CCR7hiCD137lo) present during early differentiation of effector CD4+ T cells. Our data indicate that fatty acid metabolism directs cell fate determination during the generation of memory CD4+ T cells.

Inhibition of Acetyl-CoA Carboxylase by Phosphorylation or the Inhibitor ND-654 Suppresses Lipogenesis and Hepatocellular Carcinoma.

The incidence of hepatocellular carcinoma (HCC) is rapidly increasing due to the prevalence of obesity and non-alcoholic fatty liver disease, but the molecular triggers that initiate disease development are not fully understood. We demonstrate that mice with targeted loss-of-function point mutations within the AMP-activated protein kinase (AMPK) phosphorylation sites on acetyl-CoA carboxylase 1 (ACC1 Ser79Ala) and ACC2 (ACC2 Ser212Ala) have increased liver de novo lipogenesis (DNL) and liver lesions. The same mutation in ACC1 also increases DNL and proliferation in human liver cancer cells. Consistent with these findings, a novel, liver-specific ACC inhibitor (ND-654) that mimics the effects of ACC phosphorylation inhibits hepatic DNL and the development of HCC, improving survival of tumor-bearing rats when used alone and in combination with the multi-kinase inhibitor sorafenib. These studies highlight the importance of DNL and dysregulation of AMPK-mediated ACC phosphorylation in accelerating HCC and the potential of ACC inhibitors for treatment.

Etomoxir Actions on Regulatory and Memory T Cells Are Independent of Cpt1a-Mediated Fatty Acid Oxidation.

T cell subsets including effector (Teff), regulatory (Treg), and memory (Tmem) cells are characterized by distinct metabolic profiles that influence their differentiation and function. Previous research suggests that engagement of long-chain fatty acid oxidation (LC-FAO) supports Foxp3+ Treg cell and Tmem cell survival. However, evidence for this is mostly based on inhibition of Cpt1a, the rate-limiting enzyme for LC-FAO, with the drug etomoxir. Using genetic models to target Cpt1a specifically in T cells, we dissected the role of LC-FAO in primary, memory, and regulatory T cell responses. Here we show that the ACC2/Cpt1a axis is largely dispensable for Teff, Tmem, or Treg cell formation, and that the effects of etomoxir on T cell differentiation and function are independent of Cpt1a expression. Together our data argue that metabolic pathways other than LC-FAO fuel Tmem or Treg differentiation and suggest alternative mechanisms for the effects of etomoxir that involve mitochondrial respiration.

Sulforaphane Improves Abnormal Lipid Metabolism via Both ERS-Dependent XBP1/ACC &SCD1 and ERS-Independent SREBP/FAS Pathways.

SCOPE: To investigate the effect of sulforaphane (SFN) on the abnormal lipid metabolism and underlying mechanisms. METHODS AND RESULTS: Models with abnormal lipid metabolism are established both in rats and human hepatocytes. Hepatic steatosis is detected by hematoxylin and eosin and oil red O staining. The structure of endoplasmic reticulum is visualized by transmission electron microscopy. The expressions of X-box binding protein 1 (XBP1), protein kinase-like ER kinase (PERK), sterol regulatory element binding protein-1c (SREBP1c), and lipogenic enzymes are determined by real-time PCR and western blot analysis. SFN lowers the content of triglyceride and cholesterol. SFN alleviates the swelling of ER and decreases the perimeter of ER. SFN significantly decreases the expressions of acetyl CoA carboxylase 1 (ACC1), stearoyl-CoA desaturase 1 (SCD1), and fatty acid synthase. SFN inhibits SREBP1c by blocking the PERK. Meanwhile, SFN suppresses ACC1 and SCD1 via blocking the formation of splicing-type XBP1. The key roles of XBP1 and SREBP1c in SFN-reduced lipid droplets are confirmed by a timed sequence of measurement according to time points. CONCLUSION: SFN improves abnormal lipid metabolism via both ER-stress-dependent and -independent pathways.

Minimal impact of age and housing temperature on the metabolic phenotype of Acc2-/- mice.

An important regulator of fatty acid oxidation (FAO) is the allosteric inhibition of CPT-1 by malonyl-CoA produced by the enzyme acetyl-CoA carboxylase 2 (ACC2). Initial studies suggested that deletion of Acc2 (Acacb) increased fat oxidation and reduced adipose tissue mass but in an independently generated strain of Acc2 knockout mice we observed increased whole-body and skeletal muscle FAO and a compensatory increase in muscle glycogen stores without changes in glucose tolerance, energy expenditure or fat mass in young mice (12-16 weeks). The aim of the present study was to determine whether there was any effect of age or housing at thermoneutrality (29 °C; which reduces total energy expenditure) on the phenotype of Acc2 knockout mice. At 42-54 weeks of age, male WT and Acc2(-/-) mice had similar body weight, fat mass, muscle triglyceride content and glucose tolerance. Consistent with younger Acc2(-/-) mice, aged Acc2(-/-) mice showed increased whole-body FAO (24 h average respiratory exchange ratio=0.95±0.02 and 0.92±0.02 for WT and Acc2(-/-) mice respectively, P<0.05) and skeletal muscle glycogen content (+60%, P<0.05) without any detectable change in whole-body energy expenditure. Hyperinsulinaemic-euglycaemic clamp studies revealed no difference in insulin action between groups with similar glucose infusion rates and tissue glucose uptake. Housing Acc2(-/-) mice at 29 °C did not alter body composition, glucose tolerance or the effects of fat feeding compared with WT mice. These results confirm that manipulation of Acc2 may alter FAO in mice, but this has little impact on body composition or insulin action.

Associations of acetyl-coenzyme A carboxylase α, stearoyl-coenzyme A desaturase, and lipoprotein lipase genes with dairy traits in Alpine goats.

Milk yield and composition are of great economic importance for the dairy goat industry. The identification of genes associated with phenotypic differences for these traits could allow for the implementation of gene-assisted selection programs in goats. Associations between polymorphisms at 3 candidate genes and milk production traits in Alpine goats farmed in Italy were investigated in the present research. Considered genes were acetyl-coenzyme A carboxylase α (ACACA), the major regulatory enzyme of fatty acid biosynthesis; stearoyl-coenzyme A desaturase (SCD), involved in the biosynthesis of monounsaturated fatty acids in the mammary gland; and lipoprotein lipase (LPL), which plays a central role in plasma triglyceride metabolism. An approach somewhat similar to the granddaughter design for detecting quantitative trait loci in dairy cattle was followed. Effects of genotypes of a sample of 59 Alpine bucks on phenotypes of their 946 daughters raised in 75 flocks were investigated. Data comprised 13,331 daily records for milk yields (L/d), fat and protein yields (kg/d), and fat and protein contents (%) of 2,200 lactations. Population genetics parameters were calculated and associations between milk production traits and 10 single nucleotide polymorphisms (SNP) at the 3 genes were tested. Two markers at the ACACA, 1 for the SCD and 1 at the LPL locus, deviated significantly from the Hardy-Weinberg equilibrium, with an observed heterozygosity lower than expected. Flock, age of the goat, kidding season, and stage of lactation affected all traits considered, except fat percentage. Three SNP were found to be significantly associated with milk production traits. The SNP located on the ACACA gene showed an effect on milk yield, with daughters of TT bucks having an average test-day milk yield of about 0.3 to 0.25 L/d lower than the other 2 genotypes. The marker on the LPL locus was highly associated with milk yield, with the largest values for CC daughters (about 0.50L more than GG). The TGT deletion located on the untranslated region of the SCD gene showed significant effects on average milk and protein yields. The homozygote-deleted genotype had values about 0.5 L/d and 16 g/d lower for milk and protein daily yield, respectively, compared with the TGT/TGT genotype. Differences between genotypes were quite constant across most of the lactation. Associations found in the present study, which should be tested in a larger sample, especially for those markers that show rare genotypes, may offer useful indications for the genetic improvement of dairy traits in goats.

Metabolite proofreading, a neglected aspect of intermediary metabolism.

Enzymes of intermediary metabolism are less specific than what is usually assumed: they often act on metabolites that are not their 'true' substrate, making abnormal metabolites that may be deleterious if they accumulate. Some of these abnormal metabolites are reconverted to normal metabolites by repair enzymes, which play therefore a role akin to the proofreading activities of DNA polymerases and aminoacyl-tRNA synthetases. An illustrative example of such repair enzymes is L-2-hydroxyglutarate dehydrogenase, which eliminates a metabolite abnormally made by a Krebs cycle enzyme. Mutations in L-2-hydroxyglutarate dehydrogenase lead to L-2-hydroxyglutaric aciduria, a leukoencephalopathy. Other examples are the epimerase and the ATP-dependent dehydratase that repair hydrated forms of NADH and NADPH; ethylmalonyl-CoA decarboxylase, which eliminates an abnormal metabolite formed by acetyl-CoA carboxylase, an enzyme of fatty acid synthesis; L-pipecolate oxidase, which repairs a metabolite formed by a side activity of an enzyme of L-proline biosynthesis. Metabolite proofreading enzymes are likely quite common, but most of them are still unidentified. A defect in these enzymes may account for new metabolic disorders.

Transcriptional regulation of lipid synthesis in bovine mammary epithelial cells by sterol regulatory element binding protein-1.

The objective of this study was to investigate the transcriptional regulation of lipid synthesis by sterol regulatory element binding protein-1 (SREBP-1) in bovine mammary epithelial cells. In the current study, bovine mammary epithelial (MAC-T) cells cultured in insulin- and prolactin-containing medium were treated with a transfection reagent as control, a nontargeting small interfering (si)RNA sequence (100 nM) as a negative control, or an SREBP-1-specific siRNA (100 nM) for 48 h. The mRNA expression of SREBP-1 was decreased more than 90% by siRNA. Precursor and mature forms of SREBP-1 protein were undetectable in cells treated with SREBP-1 siRNA. Fatty acid synthesis and fatty acid uptake, measured using isotope incorporation, were reduced significantly in cells treated with SREBP-1 siRNA compared with controls. Transcript abundance of acyl-CoA synthetase short-chain family member 2, acetyl-CoA carboxylase, fatty acid synthetase, and isocitrate dehydrogenase 1 (key enzymes of de novo lipogenesis) was decreased by 40 to 65% with SREBP-1 siRNA, in agreement with acetate incorporation data. The mRNA levels of fatty acid binding protein 3 and stearyl-CoA desaturase 1 (proteins responsible for intracellular fatty acid trafficking and long-chain fatty acid modification) were decreased 76 and 60%, respectively, by SREBP-1 siRNA treatment compared with controls. The mRNA expression of mitochondrial glycerol-3-phosphate acyltransferase and lipin 1 (involved in triglyceride synthesis) was significantly decreased in cells treated with SREBP-1 siRNA compared with control cells. However, the expression of milk fat globule membrane proteins measured did not differ among treatments. In conclusion, SREBP-1 plays an important role in integrated regulation of lipid synthesis in bovine mammary epithelial cells through regulation of key enzymes.

Metabolic regulation of invadopodia and invasion by acetyl-CoA carboxylase 1 and de novo lipogenesis.

Invadopodia are membrane protrusions that facilitate matrix degradation and cellular invasion. Although lipids have been implicated in several aspects of invadopodia formation, the contributions of de novo fatty acid synthesis and lipogenesis have not been defined. Inhibition of acetyl-CoA carboxylase 1 (ACC1), the committed step of fatty acid synthesis, reduced invadopodia formation in Src-transformed 3T3 (3T3-Src) cells, and also decreased the ability to degrade gelatin. Inhibition of fatty acid synthesis through AMP-activated kinase (AMPK) activation and ACC phosphorylation also decreased invadopodia incidence. The addition of exogenous 16∶0 and 18∶1 fatty acid, products of de novo fatty acid synthesis, restored invadopodia and gelatin degradation to cells with decreased ACC1 activity. Pharmacological inhibition of ACC also altered the phospholipid profile of 3T3-Src cells, with the majority of changes occurring in the phosphatidylcholine (PC) species. Exogenous supplementation with the most abundant PC species, 34∶1 PC, restored invadopodia incidence, the ability to degrade gelatin and the ability to invade through matrigel to cells deficient in ACC1 activity. On the other hand, 30∶0 PC did not restore invadopodia and 36∶2 PC only restored invadopodia incidence and gelatin degradation, but not cellular invasion through matrigel. Pharmacological inhibition of ACC also reduced the ability of MDA-MB-231 breast, Snb19 glioblastoma, and PC-3 prostate cancer cells to invade through matrigel. Invasion of PC-3 cells through matrigel was also restored by 34∶1 PC supplementation. Collectively, the data elucidate the novel metabolic regulation of invadopodia and the invasive process by de novo fatty acid synthesis and lipogenesis.

AMP-activated protein kinase activates neuropeptide Y neurons in the hypothalamic arcuate nucleus to increase food intake in rats.

AMP-activated protein kinase (AMPK) is an energy sensor that is activated by the increase of intracellular AMP:ATP ratio. AMPK in the hypothalamic arcuate nucleus (ARC) is activated during fasting and the activation of AMPK stimulates food intake. To clarify the pathway underlying AMPK-induced feeding, we monitored the activity of single ARC neurons by measuring cytosolic Ca(2+) concentration ([Ca(2+)](i)) with fura-2 fluorescence imaging. An AMPK activator, AICA-riboside (AICAR), at 200 µM increased [Ca(2+)](i) in 24% of ARC neurons. AMPK and acetyl CoA carboxylase were phosphorylated in the neurons with [Ca(2+)](i) responses to AICAR. AICAR-induced [Ca(2+)](i) increases were inhibited by Ca(2+)-free condition but not by thapsigargin, suggesting that AICAR increases [Ca(2+)](i) through Ca(2+) influx from extracellular space. Among AICAR-responding ARC neurons, 38% were neuropeptide Y (NPY)-immunoreactive neurons while no proopiomelanocortin (POMC)-immunoreactive neuron was observed. Intracerebroventricular administration of AICAR increased food intake, and the AICAR-induced food intake was abolished by the co-administration of NPY Y1 receptor antagonist, 1229U91. These results indicate that the activation of AMPK leads to the activation of ARC NPY neurons through Ca(2+) influx, thereby causing NPY-dependent food intake. These mechanisms could be implicated in the stimulation of food intake by physiological orexigenic substances.

A potential role for fatty acid biosynthesis genes during molting and cuticle formation in Caenorhabditis elegans.

Caenorhabditis elegans undergoes a developmental molting process that involves a coordinated interplay among diverse intracellular pathways. Here, we investigated the functions of two fatty acid biosynthesis genes; pod-2, encoding acetyl-CoA carboxylase, and fasn-1, encoding fatty acid synthase, in the C. elegans molting process. Although both the pod-2 and fasn-1 genes were expressed at constant levels throughout C. elegans development, knockdown of the proteins encoded by these genes using RNA interference produced severe defects in triglyceride production, molting, and reproduction that were coupled to suppression of NAS-37, a metalloprotease. An assessment of the structure and integrity of the cuticle using a COL-19::GFP marker and Hoechst 33258 staining showed that downregulation of either pod-2 or fasn-1 impaired cuticle formation and disrupted the integrity of the cuticle and the hypodermal membrane.

Exercise training in ovariectomized rats stimulates estrogenic-like effects on expression of genes involved in lipid accumulation and subclinical inflammation in liver.

We hypothesized that the reduction in liver fat accumulation known to occur with exercise training in ovariectomized (Ovx) rats is associated with reduced expression of genes involved in lipogenesis while favoring the expression of transcription factors regulating lipid oxidation. We also tested the hypothesis that liver fat accumulation in Ovx rats is associated with an increased gene expression of several inflammatory biomarkers and that exercise training would attenuate this response. Sprague-Dawley female rats (14 weeks of age) were randomly divided into 4 groups of sedentary sham-operated (Sham), Ovx, Ovx with 17ß-estradiol (E2) supplementation using a pellet (0.72 mg; 0.012 mg/d) with a biodegradable carrier binder, and Ovx trained with endurance exercise. Endurance exercise training consisted of continuous running on a motor-driven rodent treadmill 5 times per week for 5 weeks. Fat accumulation in liver as well as in adipose fat depots was higher (P < .01) in Ovx than in Sham rats. This response was prevented in Ovx animals with 17ß-estradiol supplementation and with endurance exercise training. Liver gene expressions of sterol regulatory element-binding protein 1-c, stearoyl coenzyme A desaturase 1 (and its protein content), carbohydrate response element binding protein, and acetyl-coenzyme A carboxylase were increased with estrogen withdrawal (P < .01). These responses were corrected with E2 supplementation alone as well as with training alone. Conversely, hepatic peroxisome proliferator-activated receptor α messenger RNA levels were lower (P < .01) after estrogen removal compared with Sham rats. The lower hepatic peroxisome proliferator-activated receptor α messenger RNA levels in Ovx rats were reincreased by E2 replacement or by exercise training. Gene expression of proinflammatory cytokines including inhibitor-κB kinase ß and interleukin-6, as well as protein content of nuclear factor-κB, was higher (P < .01) in Ovx than in Sham animals. E2 supplementation or exercise training prevented the expression of the proinflammatory markers. It is concluded that exercise training reduces fat accumulation in liver of Ovx rats possibly through regulation of key molecules involved in lipogenesis and lipid oxidation. Exercise training also acts as estrogens in properly regulating the expression of inflammatory biomarkers in liver of Ovx rats.

Lipotoxicity in diabetic nephropathy: the potential role of fatty acid oxidation.

Cellular toxicity mediated by lipids (lipotoxicity) has been implicated in the pathophysiology of metabolic syndrome and diabetes mellitus. Genetic analyses now implicate lipotoxicity in susceptibility to type 2 diabetes mellitus-associated nephropathy (T2DN), a pathway that had previously been unexplored. A genome-wide association study in Japanese patients identified a single nucleotide polymorphism in the acetyl-CoA carboxylase ß (ACACB) gene associated with T2DN. Replication analyses suggest that this same polymorphism may be a diabetic nephropathy risk allele in other ethnic groups. The ACACB gene (also called ACC2 or acetyl-CoA carboxylase 2) plays a critical role in intracellular fatty acid (FA) oxidation. This manuscript reviews the physiology of FA metabolism and adverse cellular effects that can result from dysregulation of this process. It is hypothesized that glomerular and tubular dysfunction can be induced by increases in intracellular FA concentrations, a process that may be enabled by genetic risk variants. This novel glucolipotoxicity hypothesis in T2DN warrants further investigation.

Macrophage migration inhibitory factor deficiency augments cardiac dysfunction in Type 1 diabetic murine cardiomyocytes.

BACKGROUND: It has become evident that macrophage migration inhibitory factor (MIF) is associated with the development of Type 1 diabetes mellitus. The aim of the present study was to determine whether MIF plays a role in cardiac contractile dysfunction in T1DM mice. METHODS: Mechanical and intracellular Ca(2+) properties were measured in cardiomyocytes isolated from wild-type (WT) and MIF-knockout (MIF-KO) mice administrated or not streptozotocin (200 mg/kg, i.p.). Relative stress signaling was evaluated using western blot analysis. RESULTS: Peak shortening (PS) and maximal velocity of shortening/relengthening (±dL/dt) were reduced and the duration of relengthening (TR90) was prolonged in both WT and MIF-KO cardiomyocytes treated with STZ (P < 0.01 vs control), which may be associated with reduced intracellular Ca(2+) decay in both groups. However, STZ-treated WT cardiomyocytes demonstrated significantly better contractile function and intracellular Ca(2+) properties compared with STZ-treated MIF-KO cardiomyocytes (all P < 0.05). Interestingly, the physiological data clearly showed that blood glucose levels were significantly higher in STZ-treated MIF-KO mice than STZ-treated WT mice (P < 0.01). Moreover, phosphorylation of AMP-activated protein kinase (AMPK) and its direct downstream target acetyl-CoA carboxylase (ACC) was markedly lower in hearts from STZ-treated MIF-KO mice than STZ-treated WT mice (P < 0.05). There were no significant differences between untreated WT and MIF-KO control groups. CONCLUSIONS: There is a beneficial action of MIF in the management of cardiac dysfunction in T1DM. The cardioprotective effect of MIF may be associated with AMPK signaling.

Gene knockout of Acc2 has little effect on body weight, fat mass, or food intake.

Deletion of acetyl CoA carboxylase-2 (Acc2) reportedly causes leanness in the setting of hyperphagia. To determine the cellular basis for these effects, we generated a mouse model in which Acc2 can be selectively deleted by the action of Cre recombinase. Deletion of Acc2 from skeletal muscle, the predominant site of Acc2 expression, had no effect on body weight, food intake, or body composition. When Acc2 was inactivated in the germline, Acc2 knockout (Acc2KO) mice displayed no differences in body weight, food intake, body composition, or glucose homeostasis as compared to controls on chow or high fat diet. Total malonyl CoA content and fatty acid oxidation rates in skeletal muscle of Acc2KO mice were unchanged, suggesting metabolic compensation in response to the loss of Acc2. The limited impact of Acc2 deletion on energy balance raises the possibility that selective pharmacological inhibition of Acc2 for the treatment of obesity may be ineffective.

SIRT3 reduces lipid accumulation via AMPK activation in human hepatic cells.

OBJECTIVE: Sirtuin 3 (SIRT3) is a nicotinamide adenine dinucleotide (NAD)(+)-dependent protein deacetylase localized on mitochondria and regulates the adaptive thermogenesis in brown adipocytes. This study aims to investigate the role of SIRT3 in hepatic lipid accumulation, and whether the activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK) is required. METHODS: A retroviral system was used for overexpressing of SIRT3 in HepG2 cells, whereas a lentivirus-mediated vector encoding SIRT3 small interfering RNA (siRNA) was used to infect these cells for knocking down endogenous SIRT3 expression. The cells were treated with oleate to induce lipid accumulation and Nile red staining was used to assess the number of lipid droplets in HepG2 cells. The AMPK signaling pathway was facilitated with the administrating of isoproterenol and an immunoblot analysis was performed to assess the phosphorylation of AMPK and acetyl coenzyme A carboxylase (ACC). Compound C was adopted to inhibit AMPK activity. RESULTS: The number of lipid droplets in HepG2 cells overexpressing SIRT3 was significantly lower than that in the control cells (P < 0.05). SIRT3-infected cells exhibited significantly more phosphorylation of AMPK and ACC (P < 0.05), which was reversed by the treatment of compound C, an inhibitor of AMPK. Knocking down SIRT3 downregulated phosphorylation of AMPK and ACC by 60-80% (P < 0.05) and promoted lipid accumulation. The activation of AMPK by SIRT3 was dependent on SIRT3 deacetylase activity. CONCLUSION: SIRT3 reduces lipid accumulation via AMPK activation in human hepatic cells.

A single nucleotide polymorphism of chicken acetyl-CoA carboxylase A gene associated with fatness traits.

Acetyl-CoA carboxylase alpha (ACCalpha) is a major rate-limiting enzyme in the biogenesis of long-chain fatty acids. It can catalyze the carboxylation of acetyl-CoA to form malonyl-CoA that plays a key role in the regulation of fatty acid metabolism. The objective of the present study was to investigate the associations of ACCalpha gene polymorphisms with chicken growth and body composition traits. The Northeast Agricultural University broiler lines divergently selected for abdominal fat content and the Northeast Agricultural University F(2) Resource Population were used in the current study. Body weight and body composition traits were measured in the aforementioned two populations. A synonymous mutation was detected in the exon 19 region of ACCalpha gene, then polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was developed to genotype all the individuals derived from the aforementioned populations. Association analysis revealed that the polymorphism was associated with abdominal fat weight and percentage of abdominal fat in the two populations. The results suggested that ACCalpha gene could be a candidate locus or linked to a major gene that affects abdominal fat content in the chicken.

Intracerebroventricular injection of citrate inhibits hypothalamic AMPK and modulates feeding behavior and peripheral insulin signaling.

We hypothesized that citrate might modulate the AMP-activated protein kinase/acetyl-CoA carboxylase (AMPK)/(ACC) pathway and participate in neuronal feeding control and glucose homeostasis. To address this issue, we injected citrate into the lateral ventricle of rats. Intracerebroventricular (ICV) injection of citrate diminished the phosphorylation of hypothalamic AMPK/ACC, increased the expression of anorexigenic neuropeptide (pro-opiomelanocortin and corticotropin-releasing hormone), elevated the level of malonyl-CoA in the hypothalamus, and reduced food intake. No change was observed in the concentration of blood insulin after the injection of citrate. With a euglycemic-hyperinsulinemic clamp, the glucose infusion rate was higher in the citrate group than in the control group (28.6+/-0.8 vs 19.3+/-0.2 mU/kg body weight/min respectively), and so was glucose uptake in skeletal muscle and the epididymal fat pad. Concordantly, insulin receptor (IR), IR substrate type 1 (IRS1), IRS2, and protein kinase B (AKT) phosphorylation in adipose tissue and skeletal muscle was improved by citrate ICV treatment. Moreover, the treatment with citrate for 7 days promoted body weight loss and decreased the adipose tissue. Our results suggest that citrate and glucose may serve as signals of energy and nutrient availability to hypothalamic cells.

Copper deficiency results in AMP-activated protein kinase activation and acetylCoA carboxylase phosphorylation in rat cerebellum.

Copper (Cu) deficiency impairs cerebellar development including biosynthetic processes like myelination and synaptogenesis. The activity of cerebellar mitochondrial cuproenzyme cytochrome c oxidase is markedly lower in Cu deficient rat pups and is accompanied by higher lactate levels indicating mitochondrial inhibition. Cu deficiency impaired energy metabolism is thought to contribute to developmental delays, but specific mechanisms linking these phenomena have remained unexplored. AMP-activated protein kinase (AMPK) is a cellular energy sensor that is activated during mitochondrial inhibition and shuts down biosynthetic processes to help conserve cellular ATP levels. Activated AMPK phosphorylates and inhibits acetylCoA carboxylase (ACC), the first enzyme in fatty acid biosynthesis. We hypothesize that AMPK is activated and ACC inhibited in Cu deficient cerebella. Perinatal copper deficiency was studied in young rats in rapidly frozen cerebella. Compared to copper-adequate (Cu+) pups, copper-deficient (Cu-) pups were hypothermic, had lower brain copper levels and markedly higher cerebellar lactate. Concentration of phosphorylated AMPK (pAMPK), indicating AMPK activation, was robustly higher in Cu- cerebella of rat pups at two ages and in two separate experiments. Compared to Cu+ cerebella, pACC content was significantly higher in all Cu- samples. Mechanisms leading to AMPK activation remain elusive. Higher AMP/ATP ratios and increased reactive nitrogen species (RNS) can lead to AMPK activation. ATP and AMP concentrations were unaltered and nitric oxide metabolites and 3-nitrotyrosine peptide levels remained unchanged in Cu- cerebella. AMPK activation may explain how ATP levels can be maintained even with a severe mitochondrial loss of CCO function.