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1.
Life Sci Alliance ; 3(2)2020 02.
Article in English | MEDLINE | ID: mdl-31992650

ABSTRACT

Obesity-induced inflammation, or meta-inflammation, plays key roles in metabolic syndrome and is a significant risk factor in diabetes and cardiovascular disease. To investigate causal links between obesity, meta-inflammation, and insulin signaling we established a Drosophila model to determine how elevated dietary fat and changes in the levels and balance of saturated fatty acids (SFAs) and polyunsaturated fatty acids (PUFAs) influence inflammation. We observe negligible effect of saturated fatty acid on inflammation but marked enhancement or suppression by omega-6 and omega-3 PUFAs, respectively. Using combined lipidomic and genetic analysis, we show omega-6 PUFA enhances meta-inflammation by producing linoleic acid-derived lipid mediator 9-hydroxy-octadecadienoic acid (9-HODE). Transcriptome analysis reveals 9-HODE functions by regulating FOXO family transcription factors. We show 9-HODE activates JNK, triggering FOXO nuclear localisation and chromatin binding. FOXO TFs are important transducers of the insulin signaling pathway that are normally down-regulated by insulin. By activating FOXO, 9-HODE could antagonise insulin signaling providing a molecular conduit linking changes in dietary fatty acid balance, meta-inflammation, and insulin resistance.


Subject(s)
Drosophila Proteins/metabolism , Drosophila/metabolism , Forkhead Box Protein O3/metabolism , Forkhead Transcription Factors/metabolism , Linoleic Acid/metabolism , Obesity/metabolism , Animals , Animals, Genetically Modified , Cell Nucleus/metabolism , Chromatin/metabolism , Dietary Fats/adverse effects , Disease Models, Animal , Drosophila/genetics , Drosophila Proteins/genetics , Fatty Acids, Omega-3/metabolism , Fatty Acids, Omega-3/pharmacology , Forkhead Box Protein O3/genetics , Forkhead Transcription Factors/genetics , Gene Expression Regulation/drug effects , HeLa Cells , Humans , Inflammation/chemically induced , Inflammation/metabolism , Insulin/metabolism , Linoleic Acid/pharmacology , Linoleic Acids, Conjugated/genetics , Linoleic Acids, Conjugated/metabolism , Linoleic Acids, Conjugated/pharmacology , Obesity/chemically induced , Protein Binding , Signal Transduction/drug effects , Signal Transduction/genetics , Transcriptome , Transfection
2.
Lipids ; 50(5): 469-81, 2015 May.
Article in English | MEDLINE | ID: mdl-25820808

ABSTRACT

cis-9, trans-11 Conjugated linoleic acid (CLA) is one of the most extensively studied CLA isomers due to its multiple isomer-specific effects. However, the molecular mechanisms of cis-9,trans-11 CLA synthesis in ruminant mammary gland are still not clearly understood. This process may be mediated, to a certain extent, by trans-11 C18:1 regulated by stearoyl-CoA desaturase-1 (SCD1) and/or its syntrophic proteins. This study aimed to investigate the effects of TVA on SCD1-mediated cis-9,trans-11 CLA synthesis in MAC-T cells and its potential molecular mechanism. Results showed that trans-11 C18:1 was continually taken up and converted into cis-9,trans-11 CLA in MAC-T cells during the 4-h incubation of 50 µM trans-11 C18:1. SCD1 protein expression increased more than twofold at 2 h (P < 0.01) and 2.5 h (P < 0.05) before decreasing to less than half of the normal level at 4 h (P < 0.05). One up-regulated (RAS guanyl releasing protein 4 isoform 1 [RASGRP4]) and six down-regulated proteins (glucosamine-6-phosphate deaminase 1 [GNPDA1], triosephosphate isomerase [TPI1], phosphoglycerate mutase 1 [PGAM1], heat shock protein beta-1 [HSPB1], annexin A3 [ANXA3], thiopurine S-methyltransferase [TPMT]) were found in MAC-T cells treated with trans-11 C18:1. Of these seven identified proteins, the presence of GNPDA1 and PGAM1 was verified in several models. More trans-11 C18:1 was taken up after PGAM1 knockdown by small interfering RNA (siRNA). In conclusion, our data suggested that PGAM1 may have a negative relationship with SCD1 and seemed to be involved in cis-9, trans-11 CLA synthesis by facilitating the absorption of trans-11 C18:1 in the bovine mammary gland.


Subject(s)
Linoleic Acids, Conjugated/metabolism , Mammary Glands, Animal/enzymology , Phosphoglycerate Mutase/metabolism , Proteomics/methods , Stearoyl-CoA Desaturase/metabolism , Animals , Cattle , Cells, Cultured , Female , Gene Expression Regulation , Gene Knockdown Techniques , Linoleic Acids, Conjugated/genetics , Mammary Glands, Animal/cytology , Mammary Glands, Animal/metabolism , Trans Fatty Acids/metabolism
3.
Anim Genet ; 45(4): 485-99, 2014 Aug.
Article in English | MEDLINE | ID: mdl-24909189

ABSTRACT

A selective DNA pooling approach was applied to identify QTL for conjugated linoleic acid (CLA), vaccenic acid (VA) and Δ(9) -desaturase (D9D) milk content in Italian Brown Swiss dairy cattle. Milk samples from 60 animals with higher values (after correction for environmental factors) and 60 animals with lower values for each of these traits from each of five half-sib families were pooled separately. The pools were genotyped using the Illumina BovineSNP50 BeadChip. Sire allele frequencies were compared between high and low tails at the sire and marker level for SNPs for which the sires were heterozygous. An r procedure was implemented to perform data analysis in a selective DNA pooling design. A correction for multiple tests was applied using the proportion of false positives among all test results. BTA 19 showed the largest number of markers in association with CLA. Associations between SNPs and the VA and Δ(9) -desaturase traits were found on several chromosomes. A bioinformatics survey identified genes with an important role in pathways for milk fat and fatty acids metabolism within 1 Mb of SNP markers associated with fatty acids contents.


Subject(s)
Cattle/genetics , Linoleic Acids, Conjugated/genetics , Oleic Acids/genetics , Polymorphism, Single Nucleotide , Quantitative Trait Loci , Stearoyl-CoA Desaturase/genetics , Animals , Cattle/metabolism , Female , Gene Frequency , Linoleic Acids, Conjugated/metabolism , Mammary Glands, Animal/enzymology , Mammary Glands, Animal/metabolism , Milk/chemistry , Oleic Acids/metabolism , Oligonucleotide Array Sequence Analysis/veterinary , Stearoyl-CoA Desaturase/metabolism
4.
PLoS One ; 9(1): e86211, 2014.
Article in English | MEDLINE | ID: mdl-24465964

ABSTRACT

In dairy cows the milk associated energy output in early lactation exceeds the input via voluntary feed intake. To spare glucose for mammary lactose synthesis, peripheral insulin sensitivity (IS) is reduced and fat mobilization is stimulated. For these processes a link between IS and the endocrine functions of adipose tissue (AT) is likely; we thus aimed to characterise the mRNA expression from bovine AT derived proteins and receptors that are related to IS according to the literature in metabolically active tissues plus systemic IS throughout lactation. Conjugated linoleic acids (CLA) reduce milk fat thus decreasing the milk drain of energy and potentially dampening lipolysis, but may also affect IS. Subcutaneous (s.c.) AT and liver from pluriparous cows receiving either control fat or CLA supplement (100 g/day from 1 to 182 days in milk each) were biopsied covering week -3 to 36 relative to parturition. In an additional trial with primiparous cows treated analogously and slaughtered on days in milk 1, 42 or 105, samples from liver, udder, skeletal muscle and 3 visceral and 3 s.c. AT were obtained and assayed for mRNA abundance of adiponectin, its receptors, leptin, leptin receptor, PPARγ, PPARγ2, IL-6, and TNF-α. In pluriparous animals, the mRNA abundance of most of the target genes decreased after parturition in s.c. AT but increased in liver. In primiparous cows, AT depot specific differences were mostly related to retroperitoneal AT; adiponectin receptor 1 and TNF-α were affected predominantly. CLA effects in primiparous cows were largely limited to decreased PPARγ2 mRNA abundance in udder tissue. In pluriparous cows, insulin secretion was increased by CLA resulting in decreased systemic IS but without consistent changes in tissue target mRNA abundance. The temporal gene expression profiles from the adipokines and related receptors support their coactive function in adapting to the needs of lactation.


Subject(s)
Adipose Tissue/metabolism , Fats/metabolism , Insulin Resistance/genetics , Lactation/genetics , Lactation/metabolism , Adiponectin/genetics , Adiponectin/metabolism , Animals , Cattle , Intra-Abdominal Fat/metabolism , Linoleic Acids, Conjugated/genetics , Linoleic Acids, Conjugated/metabolism , Liver/metabolism , Longitudinal Studies , Milk/metabolism , PPAR gamma/genetics , PPAR gamma/metabolism , Parturition/genetics , Parturition/metabolism , RNA, Messenger/genetics , Receptors, Adiponectin/genetics , Receptors, Adiponectin/metabolism , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolism
5.
J Lipid Res ; 55(6): 1139-49, 2014 06.
Article in English | MEDLINE | ID: mdl-24343898

ABSTRACT

Inflammatory activity is evident in patients with chronic kidney disease with limited data available in autosomal dominant polycystic kidney disease (ADPKD). We hypothesized that inflammation is an upstream event in the pathogenesis of ADPKD and may be a contributing factor in the disease severity and progression. Serum samples from 61 HALT study A group patients were compared with samples from 49 patients from HALT study B group with moderately advanced disease. Targeted MS analysis of bioactive lipid mediators as markers of inflammation was performed and correlated with eGFR and total kidney volume (TKV) normalized to the body surface area (BSAR) to assess if these markers are predictive of ADPKD severity. ADPKD patients with eGFR >60 ml/min/1.73 m(2) showed higher levels of 5- and 12/15-lipoxygenase (LOX) and cyclooxygenase, and generated higher levels of hydroxy-octadecadienoic acids 9-HODE and 13-HODE and HETEs 8-HETE, 11-HETE, 12-HETE, and 15-HETE as compared with healthy subjects. Linear regression of 9-HODE and 13-HODE revealed a significant relationship with eGFR and TKV, while 15-HETE significantly correlated with TKV/BSAR. Production of 20-HETE, a P450-produced metabolite of arachidonic acid, was higher in ADPKD patients as compared with healthy subjects and significantly correlated with eGFR and TKV/BSAR. Perturbation in fatty acid metabolism is evident early in ADPKD patients, even in those with preserved kidney function. The identified LOX pathways may be potential therapeutic targets for slowing down ADPKD progression.


Subject(s)
Linoleic Acids, Conjugated/metabolism , Polycystic Kidney, Autosomal Dominant/metabolism , Adult , Arachidonate 12-Lipoxygenase/genetics , Arachidonate 12-Lipoxygenase/metabolism , Arachidonate 15-Lipoxygenase/genetics , Arachidonate 15-Lipoxygenase/metabolism , Female , Humans , Linoleic Acids, Conjugated/genetics , Male , Middle Aged , Polycystic Kidney, Autosomal Dominant/genetics , Polycystic Kidney, Autosomal Dominant/pathology , Polycystic Kidney, Autosomal Dominant/therapy
6.
Meat Sci ; 93(1): 98-104, 2013 Jan.
Article in English | MEDLINE | ID: mdl-22938774

ABSTRACT

Physicochemical properties and fatty acid profiles of meat from Bos indicus, Bos taurus and crossbred B. taurus×B. indicus bullocks (n=216), finished on pasture or grain, were used to estimate the effects of heterosis. Meat quality and fatty acid profiles generally benefited with crossbreeding, but the advantages from heterosis differed among finishing systems. The Warner-Bratzler shear-force in fresh and aged meat was reduced due to heterosis in pasture-finishing, but the effect was minor under grain-finishing. With pasture-finishing, heterosis caused an increase of 5% in CLA concentration, but few other changes in fatty acid profiles. In grain-finishing, heterosis caused a reduction in intramuscular fat and cholesterol, increased amounts of PUFA, n-6 fatty acids and PUFA/SFA ratio, and a decline in atherogenic index. The Δ(9) desaturase estimated activity in crossbreds showed a behavior close to B. indicus, suggesting the existence of few loci and a dominance genetic effect on enzymes involved in fatty acid synthesis and metabolism.


Subject(s)
Animal Feed , Breeding , Diet , Dietary Fats/metabolism , Fatty Acids/genetics , Hybrid Vigor , Meat/analysis , Animal Husbandry/methods , Animals , Atherosclerosis/chemically induced , Cattle , Cholesterol, Dietary/metabolism , Edible Grain , Fatty Acids/metabolism , Fatty Acids, Unsaturated/genetics , Fatty Acids, Unsaturated/metabolism , Genetic Loci , Linoleic Acids, Conjugated/genetics , Linoleic Acids, Conjugated/metabolism , Meat/standards , Muscle, Skeletal/metabolism , Poaceae , Species Specificity , Stearoyl-CoA Desaturase/genetics , Stearoyl-CoA Desaturase/metabolism , Stress, Mechanical
7.
J Dairy Sci ; 95(8): 4286-97, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22818443

ABSTRACT

This study was conducted to identify proteins associated with the endogenous synthesis of conjugated linoleic acid (CLA) from trans-vaccenic acid (TVA; trans-11 C18:1, a precursor for CLA endogenous synthesis) in mammary tissues. Six lactating goats were divided into 2 groups. One group was given an intravenous bolus injection of TVA (150mg) twice daily over 4 d; the other group received saline injections. Treatment with TVA increased the concentration of cis-9,trans-11 CLA and TVA in goat milk. Additionally, TVA treatment increased the expression of stearoyl-CoA desaturase (SCD) in mammary tissue. Using 2-dimensional gel electrophoresis and electrospray ionization quadrupole time-of-flight mass spectrometry, 3 proteins affected by infusions of TVA were identified. Proteasome (prosome, macropain) subunit α type 5 (PSMA5) was upregulated, whereas peroxiredoxin-1 and translationally controlled tumor protein 1 were downregulated in TVA-treated animals compared with the vehicle-injected controls. Only the effect of TVA on PSMA5 could be confirmed by Western blot analysis. To further explore the regulation of PSMA5 in mammary epithelial cells when TVA is converted into CLA, we used a differentiated bovine mammary epithelial cell line treated with TVA for 6h. Changes in cis-9,trans-11 CLA concentrations and mRNA expression patterns of both SCD and PSMA5 were monitored. The concentration of cis-9,trans-11 CLA increased after TVA treatment. The mRNA expression level of PSMA5 was significantly elevated to 6h, but SCD mRNA expression only increased in 2h after TVA treatment. These results indicate that PSMA5 is highly expressed in goat mammary tissue and bovine mammary epithelial cells when TVA is converted into CLA. Our data suggest that PSMA5 protein is associated with CLA biosynthesis in mammary tissue.


Subject(s)
Goats/metabolism , Linoleic Acids, Conjugated/metabolism , Mammary Glands, Animal/metabolism , Milk/chemistry , Oleic Acids/pharmacology , Animals , Blotting, Western , Cattle , Cell Line , Electrophoresis, Gel, Two-Dimensional/veterinary , Female , Linoleic Acids, Conjugated/analysis , Linoleic Acids, Conjugated/genetics , RNA/chemistry , RNA/genetics , Real-Time Polymerase Chain Reaction , Spectrometry, Mass, Electrospray Ionization/veterinary
8.
J Nutr ; 140(5): 919-24, 2010 May.
Article in English | MEDLINE | ID: mdl-20220207

ABSTRACT

The biological activities and mechanisms of action of individual transoctadecenoic acids (trans-18:1 FA) have not been completely elucidated. We examined the effects of several individual trans-18:1 FA isomers and trans-10, cis-12 conjugated linoleic acid (CLA) on fat synthesis, and expression of lipogenic genes in mammary and liver tissue in lactating mice. From d 6 to 10 postpartum, 30 lactating C57BL/6J mice were randomly assigned to either a control (CTR) diet containing 20 g/kg oleic acid or diets in which the oleic acid was either completely replaced by partially hydrogenated vegetable oil (PHVO), trans-7 18:1 (T7), trans-9 18:1 (T9), or trans-11 18:1 (T11) or partially replaced with 6.66 g/kg trans-10, cis-12 CLA. Milk fat percentage was decreased by CLA (44%), T7 (27%), and PHVO (23%), compared with CTR. In the mammary gland, CLA decreased the expression of genes related to de novo FA synthesis, desaturation, triacylglycerol formation, and transcriptional regulation. PHVO and T7 diets decreased the expression of 1-acylglycerol-3-phosphate O-acyltransferase and thyroid hormone responsive SPOT14 homolog (THRSP) mRNA. In contrast, dietary trans FA (tFA) did not affect hepatic lipogenic gene expression. However, mice fed CLA, T7, and PHVO diets had increased liver weights due to hepatic steatosis. Trans-7 18:1 was extensively desaturated to trans-7, cis-9 CLA in mammary and liver tissues. Dietary trans-7 18:1 could lead to milk fat depression in lactating mice, possibly through its desaturation product trans-7, cis-9 CLA. Also, the differences between the effects of trans-10, cis-12 CLA and other tFA could be attributed to its effects on carbohydrate response element binding protein and PPARgamma, in addition to sterol regulatory element binding transcription factor 1c and THRSP.


Subject(s)
Dietary Fats/administration & dosage , Linoleic Acids, Conjugated/metabolism , Lipid Metabolism/physiology , Lipogenesis/genetics , Mammary Glands, Animal/metabolism , Milk/metabolism , Trans Fatty Acids/metabolism , 1-Acylglycerol-3-Phosphate O-Acyltransferase/genetics , 1-Acylglycerol-3-Phosphate O-Acyltransferase/metabolism , Animals , Fatty Liver , Female , Gene Expression Regulation , Isomerism , Lactation , Linoleic Acids, Conjugated/genetics , Lipid Metabolism/genetics , Liver/metabolism , Mice , Mice, Inbred Strains , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Oleic Acid , Organ Size , Plant Oils , RNA, Messenger/metabolism , Random Allocation , Stereoisomerism , Trans Fatty Acids/genetics , Transcription Factors/genetics , Transcription Factors/metabolism
9.
J Dairy Sci ; 92(1): 392-400, 2009 Jan.
Article in English | MEDLINE | ID: mdl-19109297

ABSTRACT

The objective of this study was to estimate genetic parameters for conjugated linoleic acid (CLA) and other selected milk fatty acid (FA) content and for unsaturation ratios in the Italian Holstein Friesian population. Furthermore, the relationship of milk FA with milk fat and protein content was considered. One morning milk sample was collected from 990 Italian Holstein Friesian cows randomly sampled from 54 half-sib families, located in 34 commercial herds in the North-eastern part of Italy. Each sample was analyzed for milk percentages of fat and protein, and for single FA percentages (computed as FA weight as a proportion of total fat weight). Heritabilities were moderate for unsaturated FA, ranging from 0.14 for C16:1 to 0.19 for C14:1. Less than 10% of heritability was estimated for each saturated FA content. Heritability for index of desaturation, monounsaturated FA and CLA/trans-11 18:1 ratio were 0.15, 0.14, and 0.15, respectively. Standard errors of the heritability values ranged from 0.02 to 0.06. Genetic correlations were high and negative between C16:0 and C18:0, as well as between C14:0 and C18:0. Genetic correlations of index of desaturation were high and negative with C14:0 and C16:0 (-0.70 and -0.72, respectively), and close to zero (0.03) with C18:0. The genetic correlation of C16:0 with fat percentage was positive (0.74), implying that selection for fat percentage should result in a correlated increase of C16:0, whereas trans-11 C18:1 and cis-9, trans-11 C18:2 contents decreased with increasing fat percentage (-0.69 and -0.55, respectively). Genetic correlations of fat percentage with 14:1/14 and 16:1/16 ratios were positive, whereas genetic correlations of fat percentage with 18:1/18 and CLA/trans-11 18:1 ratios were negative. These results suggest that it is possible to change the milk FA composition by genetic selection, which offers opportunities to meet consumer demands regarding health aspects of milk and dairy products.


Subject(s)
Cattle/genetics , Fatty Acids, Unsaturated/genetics , Fatty Acids/genetics , Lactation/genetics , Linoleic Acids, Conjugated/genetics , Milk/chemistry , Animals , Fatty Acids/analysis , Fatty Acids, Unsaturated/analysis , Female , Least-Squares Analysis , Linoleic Acids, Conjugated/analysis , Male , Milk Proteins/analysis
10.
Biochemistry ; 47(36): 9618-26, 2008 Sep 09.
Article in English | MEDLINE | ID: mdl-18702504

ABSTRACT

An alternative pathway of beta-oxidation for unsaturated fatty acids was studied in Escherichia coli. 9- cis,11- trans-Octadecadienoic acid (conjugated linoleic acid), a potential substrate of this pathway, was shown to support growth of E. coli in the absence of any other carbon source. The identification of 3,5-dodecadienoic acid in the growth medium revealed the partial beta-oxidation of conjugated linoleic acid to 3,5-dodecadienoyl-CoA, which was hydrolyzed to 3,5-dodecadienoic acid and released from cells. The involvement of acyl-CoA thioesterases in this process was evaluated by determining the substrate specificity of thioesterase II and comparing it with that of a novel thioesterase (thioesterase III) and by assessing mutant strains devoid of one or both of these thioesterases for growth on conjugated linoleic acid. Both thioesterases were highly active with 3,5-dodecadienoyl-CoA as substrate. A deficiency of either thioesterase decreased the growth rate of cells on conjugated linoleic acid but not on palmitic acid. The absence of both thioesterases reduced the cellular growth in a cumulative manner but did not abolish it. It is concluded that thioesterases II and III and at least one other thioesterase function in the partial degradation of conjugated linoleic acid via the thioesterase-dependent pathway of beta-oxidation, which provides all energy and carbon precursors required for the growth of E. coli.


Subject(s)
Escherichia coli Proteins/metabolism , Escherichia coli/enzymology , Escherichia coli/growth & development , Fatty Acid Synthases/metabolism , Linoleic Acids, Conjugated/metabolism , Linoleic Acids, Conjugated/pharmacology , Thiolester Hydrolases/metabolism , Escherichia coli/genetics , Escherichia coli Proteins/genetics , Fatty Acid Synthases/genetics , Linoleic Acids, Conjugated/genetics , Lipid Metabolism/drug effects , Lipid Metabolism/physiology , Mutation , Oxidation-Reduction/drug effects , Palmitic Acid/metabolism , Palmitic Acid/pharmacology , Thiolester Hydrolases/genetics
11.
J Appl Microbiol ; 103(2): 365-71, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17650196

ABSTRACT

AIMS: To characterize the cause for the lack of conjugated linoleic acid (CLA) reductase (CLA-R) activity in the Butyrivibrio fibrisolvens MDT-5 strain that rapidly isomerizes linoleic acid (LA) to CLA without hydrogenation, the CLA-R was purified and its gene (cla-r) sequence was determined. METHODS AND RESULTS: CLA-R was purified to near homogeneity as a 53-kDa monomeric protein from the high CLA-R activity-expressing strain MDT-10. The purified CLA-R recognized conjugated double bonds. Unsaturated fatty acids containing 18 carbons markedly increased the CLA-R expression at the transcriptional level. Complete sequencing of the cla-r gene revealed that the CLA-R is a novel protein. Sequence analysis of the cla-r gene from the MDT-5 strain revealed that the MDT-5 CLA-R protein sequence differed from that of the MDT-10 at four consecutive amino acids. Northern and Western blotting analyses confirmed that the cla-r mRNA and protein are expressed normally in MDT-5. CONCLUSIONS: Strain MDT-5 expresses the CLA-R protein that lacks enzyme activity because of mutation, which explains why MDT-5 exclusively produces CLA from LA. SIGNIFICANCE AND IMPACT OF THE STUDY: The cla-r gene was sequenced for the first time. Exogenous fatty acids affected the cla-r transcription. These results will provide additional knowledge on biohydrogenation, and may also augment the CLA production in the gastrointestinal tract.


Subject(s)
Butyrivibrio/genetics , Linoleic Acids, Conjugated/genetics , Oxidoreductases Acting on CH-CH Group Donors/genetics , Amino Acid Sequence/genetics , Bacterial Proteins/genetics , Base Sequence , Blotting, Northern/methods , Blotting, Western/methods , Butyrivibrio/enzymology , Cloning, Molecular/methods , Culture Media , Fatty Acids, Unsaturated/genetics , Gene Expression Regulation, Bacterial/genetics , Genes, Bacterial/genetics , Oxidoreductases Acting on CH-CH Group Donors/isolation & purification , RNA, Bacterial/genetics , RNA, Messenger/genetics , Substrate Specificity/genetics , Transcription, Genetic/genetics
12.
Biochem Biophys Res Commun ; 344(1): 233-40, 2006 May 26.
Article in English | MEDLINE | ID: mdl-16603123

ABSTRACT

The bovine stearoyl-CoA desaturase (Scd) gene plays an important role in the bovine mammary gland where substrates such as stearic and vaccenic acids are converted to oleic acid and conjugated linoleic acid (CLA), respectively. Up to 90% of the CLA in bovine milk is formed due to the action of this enzyme in the mammary gland. The areas of the bovine promoter of importance in regulating this key enzyme were examined and an area of 36bp in length was identified as having a critical role in transcriptional activation and is designated the Scd transcriptional enhancer element (STE). Electrophoretic mobility shift assay detected three binding complexes on this area in Mac-T cell nuclear extracts. Treatment of cells with CLA caused a significant reduction in transcriptional activity, with this effect being mediated through the STE region. The bovine Scd gene promoter was up-regulated by insulin and down-regulated by oleic acid.


Subject(s)
Cattle/genetics , Enhancer Elements, Genetic/genetics , Gene Expression Regulation, Enzymologic , Promoter Regions, Genetic/genetics , Stearoyl-CoA Desaturase/genetics , Animals , Cattle/metabolism , Electrophoretic Mobility Shift Assay , Female , Linoleic Acids, Conjugated/biosynthesis , Linoleic Acids, Conjugated/genetics , Linoleic Acids, Conjugated/pharmacology , Nuclear Proteins/metabolism , Transcription, Genetic/drug effects
13.
Gastroenterology ; 127(3): 777-91, 2004 Sep.
Article in English | MEDLINE | ID: mdl-15362034

ABSTRACT

BACKGROUND & AIMS: The molecular targets for the protective actions of conjugated linoleic acid (CLA) on experimental inflammatory bowel disease (IBD) are unknown. We used a loss-of-function approach to investigate whether CLA ameliorated colitis through a peroxisome proliferator-activated receptor gamma (PPAR gamma)-dependent mechanism. METHODS: The expression of PPAR gamma, delta, and their target genes in the colon of mice fed control or CLA-supplemented diets was assayed after a 7-day dextran sodium sulfate (DSS) challenge by quantitative real-time polymerase chain reaction (PCR). Additionally, nuclear factor-kappa B (NF-kappaB) p65 activation was quantified in the colon. To determine the involvement of PPAR gamma in the mechanism of action of CLA directly, specific deletions of PPAR gamma in the colon were performed in mice by using the Cre-lox recombination system. Colonic PPAR gamma null mice and wild-type littermates were fed either a CLA-supplemented or a control diet for 42 days and challenged with 2.5% DSS. The therapeutic efficacy of CLA also was examined by using the CD4 + CD45RB hi transfer colitis model. RESULTS: CLA induced PPAR gamma and delta, transcriptionally modulated PPAR gamma and delta-responsive gene clusters involved in lipid metabolism (uncoupling protein [UCP]1, UCP3, PPAR gamma coactivator 1alpha [PGC-1alpha], and CD36) and epithelial cell maturation (Gob-4 and Keratin 20). Additionally, CLA repressed tumor necrosis factor alpha (TNF-alpha) expression and NF-kappaB activation while inducing the immunoregulatory cytokine transforming growth factor beta 1 (TGF-beta 1 ). Clinically, CLA ameliorated DSS- and CD4 + -induced colitis. Loss of the PPAR gamma gene in the colon abrogated the beneficial effects of CLA in DSS colitis. CONCLUSIONS: Our studies provide molecular evidence in vivo, suggesting that CLA ameliorates colitis through a PPAR gamma-dependent mechanism.


Subject(s)
Inflammatory Bowel Diseases/immunology , Linoleic Acids, Conjugated/immunology , Receptors, Cytoplasmic and Nuclear/biosynthesis , Transcription Factors/biosynthesis , Animals , Carrier Proteins/immunology , Gene Expression Regulation/immunology , Inflammatory Bowel Diseases/diet therapy , Linoleic Acids, Conjugated/genetics , Mice , Mice, Inbred C57BL , Models, Animal , NF-kappa B/immunology , Neoplasm Proteins/immunology , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Cytoplasmic and Nuclear/immunology , Transcription Factor RelA , Transcription Factors/genetics , Transcription Factors/immunology
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