WorldWideScience

Sample records for acyl-coenzyme a-dependent lysophosphatidic

  1. Acyl-coenzyme A binding protein (ACBP)

    DEFF Research Database (Denmark)

    Kragelund, B B; Knudsen, J; Poulsen, F M

    1999-01-01

    Acyl-coenzyme A binding proteins are known from a large group of eukaryote species and to bind a long chain length acyl-CoA ester with very high affinity. Detailed biochemical mapping of ligand binding properties has been obtained as well as in-depth structural studies on the bovine apo-protein...... and of the complex with palmitoyl-CoA using NMR spectroscopy. In the four alpha-helix bundle structure, a set of 21 highly conserved residues present in more that 90% of all known sequences of acyl-coenzyme A binding proteins constitutes three separate mini-cores. These residues are predominantly located...... at the helix-helix interfaces. From studies of a large set of mutant proteins the role of the conserved residues has been related to structure, function, folding and stability....

  2. Acyl-coenzyme A binding protein, ACBP

    DEFF Research Database (Denmark)

    Kragelund, Birthe Brandt; Knudsen, J.; Poulsen, Flemming

    1999-01-01

    Acyl-coenzyme A binding proteins are known from a large group of eukaryote species and to bind a long chain length acyl-CoA ester with very high affinity. Detailed biochemical mapping of ligand binding properties has been obtained as well as in-depth structural studies on the bovine apo-protein...... and of the complex with palmitoyl-CoA using NMR spectroscopy. In the four a-helix bundle structure, a set of 21 highly conserved residues present in more that 90% of all known sequences of acyl-coenzyme A binding proteins constitutes three separate mini-cores. These residues are predominantly located at the helix......-helix interfaces. From studies of a large set of mutant proteins the role of the conserved residues has been related to structure, function, folding and stability....

  3. Continuous recording of long-chain acyl-coenzyme A synthetase activity using fluorescently labeled bovine serum albumin

    DEFF Research Database (Denmark)

    Demant, Erland J.F.; Nystrøm, Birthe T.

    2001-01-01

    acyl-Coenzyme A, synthetase, activity assay, fluorescence recording, fatty acid probe, serum albumin, hydroxycoumarin, detergent, micelles, Pseudomonas fragi, rat liver microsomes......acyl-Coenzyme A, synthetase, activity assay, fluorescence recording, fatty acid probe, serum albumin, hydroxycoumarin, detergent, micelles, Pseudomonas fragi, rat liver microsomes...

  4. Adult peroxisomal acyl-coenzyme A oxidase deficiency with cerebellar and brainstem atrophy

    NARCIS (Netherlands)

    S. Ferdinandusse; S. Barker; K. Lachlan; M. Duran; H.R. Waterham; R.J.A. Wanders; S. Hammans

    2010-01-01

    Peroxisomal acyl-coenzyme A oxidase deficiency ( formerly also called pseudoneonatal adrenoleucodystrophy) is a disorder of peroxisomal fatty acid oxidation with a severe presentation. Most patients present at birth or in early infancy, and the mean age of death was 5 years in a recently published c

  5. Very long chain acyl-coenzyme A dehydrogenase deficiency with adult onset

    DEFF Research Database (Denmark)

    Smelt, A H; Poorthuis, B J; Onkenhout, W;

    1998-01-01

    Very long chain acyl-coenzyme A (acyl-CoA) dehydrogenase (VLCAD) deficiency is a severe disorder of mitochondrial beta-oxidation in infants. We report adult onset of attacks of painful rhabdomyolysis. Gas chromatography identified strongly elevated levels of tetradecenoic acid, 14:1(n-9), tetrade...... be due to residual enzyme activity as a consequence of the two missense mutations. Treatment with L-carnitine and medium chain triglycerides in the diet did not reduce the attacks of rhabdomyolysis.......Very long chain acyl-coenzyme A (acyl-CoA) dehydrogenase (VLCAD) deficiency is a severe disorder of mitochondrial beta-oxidation in infants. We report adult onset of attacks of painful rhabdomyolysis. Gas chromatography identified strongly elevated levels of tetradecenoic acid, 14:1(n-9...

  6. Prolonged QTc Interval in Association With Medium-Chain Acyl-Coenzyme A Dehydrogenase Deficiency

    OpenAIRE

    Wiles, Jason R.; Leslie, Nancy; Knilans, Timothy K.; Akinbi, Henry

    2014-01-01

    Medium-chain acyl-coenzyme A dehydrogenase (MCAD) deficiency is the most common disorder of mitochondrial fatty acid oxidation. We report a term male infant who presented at 3 days of age with hypoglycemia, compensated metabolic acidosis, hypocalcemia, and prolonged QTc interval. Pregnancy was complicated by maternal premature atrial contractions and premature ventricular contractions. Prolongation of the QTc interval resolved after correction of metabolic derangements. The newborn screen was...

  7. Prolonged QTc interval in association with medium-chain acyl-coenzyme A dehydrogenase deficiency.

    Science.gov (United States)

    Wiles, Jason R; Leslie, Nancy; Knilans, Timothy K; Akinbi, Henry

    2014-06-01

    Medium-chain acyl-coenzyme A dehydrogenase (MCAD) deficiency is the most common disorder of mitochondrial fatty acid oxidation. We report a term male infant who presented at 3 days of age with hypoglycemia, compensated metabolic acidosis, hypocalcemia, and prolonged QTc interval. Pregnancy was complicated by maternal premature atrial contractions and premature ventricular contractions. Prolongation of the QTc interval resolved after correction of metabolic derangements. The newborn screen was suggestive for MCAD deficiency, a diagnosis that was confirmed on genetic analysis that showed homozygosity for the disease-associated missense A985G mutation in the ACADM gene. This is the first report of acquired prolonged QTc in a neonate with MCAD deficiency, and it suggests that MCAD deficiency should be considered in the differential diagnoses of acute neonatal illnesses associated with electrocardiographic abnormality. We review the clinical presentation and diagnosis of MCAD deficiency in neonates. PMID:24799540

  8. Patients with medium-chain acyl-coenzyme a dehydrogenase deficiency have impaired oxidation of fat during exercise but no effect of L-carnitine supplementation

    DEFF Research Database (Denmark)

    Madsen, K L; Preisler, N; Orngreen, M C;

    2013-01-01

    It is not clear to what extent skeletal muscle is affected in patients with medium-chain acyl-coenzyme A dehydrogenase deficiency (MCADD). l-Carnitine is commonly used as a supplement in patients with MCADD, although its beneficial effect has not been verified....

  9. Involvement of acyl coenzyme A oxidase isozymes in biotransformation of methyl ricinoleate into gamma-decalactone by Yarrowia lipolytica.

    Science.gov (United States)

    Waché, Y; Laroche, C; Bergmark, K; Møller-Andersen, C; Aguedo, M; Le Dall, M T; Wang, H; Nicaud, J M; Belin, J M

    2000-03-01

    We reported previously on the function of acyl coenzyme A (acyl-CoA) oxidase isozymes in the yeast Yarrowia lipolytica by investigating strains disrupted in one or several acyl-CoA oxidase-encoding genes (POX1 through POX5) (H. Wang et al., J. Bacteriol. 181:5140-5148, 1999). Here, these mutants were studied for lactone production. Monodisrupted strains produced similar levels of lactone as the wild-type strain (50 mg/liter) except for Deltapox3, which produced 220 mg of gamma-decalactone per liter after 24 h. The Deltapox2 Deltapox3 double-disrupted strain, although slightly affected in growth, produced about 150 mg of lactone per liter, indicating that Aox2p was not essential for the biotransformation. The Deltapox2 Deltapox3 Deltapox5 triple-disrupted strain produced and consumed lactone very slowly. On the contrary, the Deltapox2 Deltapox3 Deltapox4 Deltapox5 multidisrupted strain did not grow or biotransform methyl ricinoleate into gamma-decalactone, demonstrating that Aox4p is essential for the biotransformation. PMID:10698800

  10. Anesthetic agents in patients with very long-chain acyl-coenzyme A dehydrogenase deficiency: a literature review.

    Science.gov (United States)

    Redshaw, Charlotte; Stewart, Catherine

    2014-11-01

    Very long-chain acyl-coenzyme A dehydrongenase deficiency (VLCADD) is a rare disorder of fatty acid metabolism that renders sufferers susceptible to hypoglycemia, liver failure, cardiomyopathy, and rhabdomyolysis. The literature about the management of these patients is hugely conflicting, suggesting that both propofol and volatile anesthesia should be avoided. We have reviewed the literature and have concluded that the source papers do not support the statements that volatile anesthetic agents are unsafe. The reports on rhabdomyolysis secondary to anesthesia appear to be due to inadequate supply of carbohydrate not volatile agents. Catabolism must be avoided with minimal fasting, glucose infusions based on age and weight, and attenuation of emotional and physical stress. General anesthesia appears to be protective of stress-induced catabolism and may offer benefits in children and anxious patients over regional anesthesia. Propofol has not been demonstrated to be harmful in VLCADD but is presented in an emulsion containing very long-chain fatty acids which can cause organ lipidosis and itself can inhibit mitochondrial fatty acid metabolism. It is therefore not recommended. Suxamethonium-induced myalgia may mimic symptoms of rhabdomyolysis and cause raised CK therefore should be avoided. Opioids, NSAIDS, regional anesthesia, and local anesthetic techniques have all been used without complication. PMID:25069536

  11. Cloning and functional analysis of human acyl coenzyme A: Cholesterol acyltransferase1 gene P1 promoter.

    Science.gov (United States)

    Ge, Jing; Cheng, Bei; Qi, Benling; Peng, Wen; Wen, Hui; Bai, Lijuan; Liu, Yun; Zhai, Wei

    2016-07-01

    Acyl-coenzyme A: cholesterol acyltransferase 1 (ACAT1) catalyzes the conversion of free cholesterol (FC) to cholesterol ester. The human ACAT1 gene P1 promoter has been cloned. However, the activity and specificity of the ACAT1 gene P1 promoter in diverse cell types remains unclear. The P1 promoter fragment was digested with KpnI/XhoI from a P1 promoter cloning vector, and was subcloned into the multiple cloning site of the Firefly luciferase vector pGL3‑Enhancer to obtain the construct P1E‑1. According to the analysis of biological information, the P1E‑1 plasmid was used to generate deletions of the ACAT1 gene P1 promoter with varying 5' ends and an identical 3' end at +65 by polymerase chain reaction (PCR). All the 5'‑deletion constructs of the P1 promoter were identified by PCR, restriction enzyme digestion mapping and DNA sequencing. The transcriptional activity of each construct was detected after transient transfection into THP‑1, HepG2, HEK293 and Hela cells using DEAE‑dextran and Lipofectamine 2000 liposome transfection reagent. Results showed that the transcriptional activity of the ACAT1 gene P1 promoter and deletions of P1 promoter in THP‑1 and HepG2 cells was higher than that in HEK293 and HeLa cells. Moreover, the transcriptional activity of P1E‑9 was higher compared with those of other deletions in THP‑1, HepG2, HEK293 and HeLa cells. These findings indicate that the transcriptional activity of the P1 promoter and the effects of deletions vary with different cell lines. Thus, the P1 promoter may drive ACAT1 gene expression with cell‑type specificity. In addition, the core sequence of ACAT1 gene P1 promoter was suggested to be between -125 and +65 bp. PMID:27220725

  12. Structure of YciA from Haemophilus influenzae (HI0827), a Hexameric Broad Specificity Acyl-Coenzyme A Thioesterase

    Energy Technology Data Exchange (ETDEWEB)

    Willis, Mark A.; Zhuang, Zhihao; Song, Feng; Howard, Andrew; Dunaway-Mariano, Debra; Herzberg, Osnat (UNM); (IIT); (UMBI)

    2008-04-02

    The crystal structure of HI0827 from Haemophilus influenzae Rd KW20, initially annotated 'hypothetical protein' in sequence databases, exhibits an acyl-coenzyme A (acyl-CoA) thioesterase 'hot dog' fold with a trimer of dimers oligomeric association, a novel assembly for this enzyme family. In studies described in the preceding paper [Zhuang, Z., Song, F., Zhao, H., Li, L., Cao, J., Eisenstein, E., Herzberg, O., and Dunaway-Mariano, D. (2008) Biochemistry 47, 2789-2796], HI0827 is shown to be an acyl-CoA thioesterase that acts on a wide range of acyl-CoA compounds. Two substrate binding sites are located across the dimer interface. The binding sites are occupied by two CoA molecules, one with full occupancy and the second only partially occupied. The CoA molecules, acquired from HI0827-expressing Escherichia coli cells, remained tightly bound to the enzyme through the protein purification steps. The difference in CoA occupancies indicates a different substrate affinity for each of the binding sites, which in turn implies that the enzyme might be subject to allosteric regulation. Mutagenesis studies have shown that the replacement of the putative catalytic carboxylate Asp44 with an alanine residue abolishes activity. The impact of this mutation is seen in the crystal structure of D44A HI0827. Whereas the overall fold and assembly of the mutant protein are the same as those of the wild-type enzyme, the CoA ligands are absent. The dimer interface is perturbed, and the channel that accommodates the thioester acyl chain is more open and wider than that observed in the wild-type enzyme. A model of intact substrate bound to wild-type HI0827 provides a structural rationale for the broad substrate range.

  13. Expression of the Acyl-Coenzyme A: Cholesterol Acyltransferase GFP Fusion Protein in Sf21 Insect Cells

    Science.gov (United States)

    Mahtani, H. K.; Richmond, R. C.; Chang, T. Y.; Chang, C. C. Y.; Rose, M. Franklin (Technical Monitor)

    2001-01-01

    The enzyme acyl-coenzyme A:cholesterol acyltransferase (ACAT) is an important contributor to the pathological expression of plaque leading to artherosclerosis n a major health problem. Adequate knowledge of the structure of this protein will enable pharmaceutical companies to design drugs specific to the enzyme. ACAT is a membrane protein located in the endoplasmic reticulum.t The protein has never been purified to homogeneity.T.Y. Chang's laboratory at Dartmouth College provided a 4-kb cDNA clone (K1) coding for a structural gene of the protein. We have modified the gene sequence and inserted the cDNA into the BioGreen His Baculovirus transfer vector. This was successfully expressed in Sf2l insect cells as a GFP-labeled ACAT protein. The advantage to this ACAT-GFP fusion protein (abbreviated GCAT) is that one can easily monitor its expression as a function of GFP excitation at 395 nm and emission at 509 nm. Moreover, the fusion protein GCAT can be detected on Western blots with the use of commercially available GFP antibodies. Antibodies against ACAT are not readily available. The presence of the 6xHis tag in the transfer vector facilitates purification of the recombinant protein since 6xHis fusion proteins bind with high affinity to Ni-NTA agarose. Obtaining highly pure protein in large quantities is essential for subsequent crystallization. The purified GCAT fusion protein can readily be cleaved into distinct GFP and ACAT proteins in the presence of thrombin. Thrombin digests the 6xHis tag linking the two protein sequences. Preliminary experiments have indicated that both GCAT and ACAT are expressed as functional proteins. The ultimate aim is to obtain large quantities of the ACAT protein in pure and functional form appropriate for protein crystal growth. Determining protein structure is the key to the design and development of effective drugs. X-ray analysis requires large homogeneous crystals that are difficult to obtain in the gravity environment of earth

  14. Purification of a jojoba embryo fatty acyl-coenzyme A reductase and expression of its cDNA in high erucic acid rapeseed.

    Science.gov (United States)

    Metz, J G; Pollard, M R; Anderson, L; Hayes, T R; Lassner, M W

    2000-03-01

    The jojoba (Simmondsia chinensis) plant produces esters of long-chain alcohols and fatty acids (waxes) as a seed lipid energy reserve. This is in contrast to the triglycerides found in seeds of other plants. We purified an alcohol-forming fatty acyl-coenzyme A reductase (FAR) from developing embryos and cloned the cDNA encoding the enzyme. Expression of a cDNA in Escherichia coli confers FAR activity upon those cells and results in the accumulation of fatty alcohols. The FAR sequence shows significant homology to an Arabidopsis protein of unknown function that is essential for pollen development. When the jojoba FAR cDNA is expressed in embryos of Brassica napus, long-chain alcohols can be detected in transmethylated seed oils. Resynthesis of the gene to reduce its A plus T content resulted in increased levels of alcohol production. In addition to free alcohols, novel wax esters were detected in the transgenic seed oils. In vitro assays revealed that B. napus embryos have an endogenous fatty acyl-coenzyme A: fatty alcohol acyl-transferase activity that could account for this wax synthesis. Thus, introduction of a single cDNA into B. napus results in a redirection of a portion of seed oil synthesis from triglycerides to waxes.

  15. Characterization of Two Members among the Five ADP-Forming Acyl Coenzyme A (Acyl-CoA) Synthetases Reveals the Presence of a 2-(Imidazol-4-yl)Acetyl-CoA Synthetase in Thermococcus kodakarensis

    OpenAIRE

    Awano, Tomotsugu; Wilming, Anja; Tomita, Hiroya; Yokooji, Yuusuke; Fukui, Toshiaki; Imanaka, Tadayuki; Atomi, Haruyuki

    2014-01-01

    The genome of Thermococcus kodakarensis, along with those of most Thermococcus and Pyrococcus species, harbors five paralogous genes encoding putative α subunits of nucleoside diphosphate (NDP)-forming acyl coenzyme A (acyl-CoA) synthetases. The substrate specificities of the protein products for three of these paralogs have been clarified through studies on the individual enzymes from Pyrococcus furiosus and T. kodakarensis. Here we have examined the biochemical properties of the remaining t...

  16. Shrinking the FadE proteome of Mycobacterium tuberculosis: insights into cholesterol metabolism through identification of an α2β2 heterotetrameric acyl coenzyme A dehydrogenase family.

    Science.gov (United States)

    Wipperman, Matthew F; Yang, Meng; Thomas, Suzanne T; Sampson, Nicole S

    2013-10-01

    The ability of the pathogen Mycobacterium tuberculosis to metabolize steroids like cholesterol and the roles that these compounds play in the virulence and pathogenesis of this organism are increasingly evident. Here, we demonstrate through experiments and bioinformatic analysis the existence of an architecturally distinct subfamily of acyl coenzyme A (acyl-CoA) dehydrogenase (ACAD) enzymes that are α2β2 heterotetramers with two active sites. These enzymes are encoded by two adjacent ACAD (fadE) genes that are regulated by cholesterol. FadE26-FadE27 catalyzes the dehydrogenation of 3β-hydroxy-chol-5-en-24-oyl-CoA, an analog of the 5-carbon side chain cholesterol degradation intermediate. Genes encoding the α2β2 heterotetrameric ACAD structures are present in multiple regions of the M. tuberculosis genome, and subsets of these genes are regulated by four different transcriptional repressors or activators: KstR1 (also known as KstR), KstR2, Mce3R, and SigE. Homologous ACAD gene pairs are found in other Actinobacteria, as well as Proteobacteria. Their structures and genomic locations suggest that the α2β2 heterotetrameric structural motif has evolved to enable catalysis of dehydrogenation of steroid- or polycyclic-CoA substrates and that they function in four subpathways of cholesterol metabolism.

  17. Rosiglitazone inhibits expression of acyl-coenzyme A:cholesterol acyltransferase-1 in THP-1 macrophages induced by advanced glycation end-products

    Institute of Scientific and Technical Information of China (English)

    Yang Qihong; Xu Qiang; Zhang Hong; Si Liangyi

    2008-01-01

    Objective: To investigate the effects of rosiglitazone, a synthetic ligand of peroxisome proliferators-activated receptor gamma (PPARγ), on the expression of acyl-coenzyme A: cholesterol acyltransferase-1 (ACAT-1) in phorbol myristate acetate (PMA)-pretreated THP-1 cells after the inducement of advanced glycation end products (AGEs). Methods: After THP-1 cells were cultured in the presence of 0.1 umol/L PMA for 72 h to induce phagocytic differentiation, the obtained THP-1 macrophages were treated with rosiglitazone for 4 h at different concentrations (1,5 or 10 μmol/L) and then exposed to AGEs-modified bovine serum albumin (AGEs-BSA) for 24 h at a concentration of 200 mg/L. Reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis were performed to detect the mRNA and protein expressions of ACAT-1 respectively. Results: Administration of AGEs-BSA (200 mg/L) into the THP-1 macrophages resulted in up-regulation of ACAT-1 at mRNA and protein levels when compared with the expressions in macrophages incubated with serum-free RPM11640. Pretreatment of rosiglitazone inhibited significantly the increased expression of ACAT-1 induced by AGEs-BSA in a concentration-dependent manner. Conclusion: PPARγ activation by rosiglitazone down-regulates ACAT-1 expression induced by AGEs in THP-1 macrophages, which might provide a new way for treating atherogenesis in diabetic patients.

  18. Acyl coenzyme A synthetase long-chain 1 (ACSL1 gene polymorphism (rs6552828 and elite endurance athletic status: a replication study.

    Directory of Open Access Journals (Sweden)

    Thomas Yvert

    Full Text Available The aim of this study was to determine the association between the rs6552828 polymorphism in acyl coenzyme A synthetase (ACSL1 and elite endurance athletic status. We studied 82 Caucasian (Spanish World/Olympic-class endurance male athletes, and a group of sex and ethnically matched healthy young adults (controls, n=197. The analyses were replicated in a cohort of a different ethnic origin (Chinese of the Han ethnic group, composed of elite endurance athletes (runners [cases, n=241 (128 male] and healthy sedentary adults [controls, n=504 (267 male]. In the Spanish cohort, genotype (P=0.591 and minor allele (A frequencies were similar in cases and controls (P=0.978. In the Chinese cohort, genotype (P=0.973 and minor allele (G frequencies were comparable in female endurance athletes and sedentary controls (P=0.881, whereas in males the frequency of the G allele was higher in endurance athletes (0.40 compared with their controls (0.32, P=0.040. The odds ratio (95%CI for an elite endurance Chinese athlete to carry the G allele compared with ethnically matched controls was 1.381 (1.015-1.880 (P-value=0.04. Our findings suggest that the ACSL1 gene polymorphism rs6552828 is not associated with elite endurance athletic status in Caucasians, yet a marginal association seems to exist for the Chinese (Han male population.

  19. Effect of Tumor Necrosis Factor-α on Acyl Coenzyme A: Cholesteryl Acyltransferase Activity and ACAT1 Gene Expression in THP-1 Macrophages

    Institute of Scientific and Technical Information of China (English)

    HE Ping; CHENG Bei; WANG Yi; WANG Hongxing

    2007-01-01

    In order to explore the effect and mechanisms of tumor necrosis factor-α (TNF-α) on the activity of the acyl coenzyme A: cholesteryl acyltransferase (ACAT), THP-1 monocytes were cultured and induced to differentiate into macrophages with phorbol ester. TNF-α (60 ng/mL) was added at different time points into the macrophage-containing medium and the ACAT enzyme activity was measured by quantifying the incorporation of [1-14C] oleoyl CoA into cholesteryl esters. The expression of ACAT-1 protein and mRNA was respectively detected by Western blotting and RT-PCR in THP-1 macrophages 24 h after treatment with TNF-α (60 ng/mL). The results indicated that ACAT activity in THP-1 macrophages treated with TNF-α was increased in a time-dependent manner. The expression levels of ACAT-1 protein and mRNA were significantly increased in THP-1 macrophages after treatment with TNF-α (P<0.05). It was suggested that TNF-α could increase the activity of ACAT in THP-1 macrophages by up-regulating the expression of ACAT-1 gene.

  20. Mangiferin treatment inhibits hepatic expression of acyl-coenzyme A:diacylglycerol acyltransferase-2 in fructose-fed spontaneously hypertensive rats: a link to amelioration of fatty liver.

    Science.gov (United States)

    Xing, Xiaomang; Li, Danyang; Chen, Dilong; Zhou, Liang; Chonan, Ritsu; Yamahara, Johji; Wang, Jianwei; Li, Yuhao

    2014-10-15

    Mangiferin, a xanthone glucoside, and its associated traditional herbs have been demonstrated to improve abnormalities of lipid metabolism. However, its underlying mechanisms remain largely unclear. This study investigated the anti-steatotic effect of mangiferin in fructose-fed spontaneously hypertensive rat (SHR)s that have a mutation in sterol regulatory element binding protein (SREBP)-1. The results showed that co-administration of mangiferin (15 mg/kg, once daily, by oral gavage) over 7 weeks dramatically diminished fructose-induced increases in hepatic triglyceride content and Oil Red O-stained area in SHRs. However, blood pressure, fructose and chow intakes, white adipose tissue weight and metabolic parameters (plasma concentrations of glucose, insulin, triglyceride, total cholesterol and non-esterified fatty acids) were unaffected by mangiferin treatment. Mechanistically, mangiferin treatment suppressed acyl-coenzyme A:diacylglycerol acyltransferase (DGAT)-2 expression at the mRNA and protein levels in the liver. In contrast, mangiferin treatment was without effect on hepatic mRNA and/or protein expression of SREBP-1/1c, carbohydrate response element binding protein, liver pyruvate kinase, fatty acid synthase, acetyl-CoA carboxylase-1, stearoyl-CoA desaturase-1, DGAT-1, monoacyglycerol acyltransferase-2, microsomal triglyceride transfer protein, peroxisome proliferator-activated receptor-alpha, carnitine palmitoyltransferase-1 and acyl-CoA oxidase. Collectively, our results suggest that mangiferin treatment ameliorates fatty liver in fructose-fed SHRs by inhibiting hepatic DGAT-2 that catalyzes the final step in triglyceride biosynthesis. The anti-steatotic effect of mangiferin may occur independently of the hepatic signals associated with de novo fatty acid synthesis and oxidation.

  1. Acyl-coenzyme A oxidases 1 and 3 in brown trout (Salmo trutta f. fario): Can peroxisomal fatty acid β-oxidation be regulated by estrogen signaling?

    Science.gov (United States)

    Madureira, Tânia Vieira; Castro, L Filipe C; Rocha, Eduardo

    2016-02-01

    Acyl-coenzyme A oxidases 1 (Acox1) and 3 (Acox3) are key enzymes in the regulation of lipid homeostasis. Endogenous and exogenous factors can disrupt their normal expression/activity. This study presents for the first time the isolation and characterization of Acox1 and Acox3 in brown trout (Salmo trutta f. fario). Additionally, as previous data point to the existence of a cross-talk between two nuclear receptors, namely peroxisome proliferator-activated receptors and estrogen receptors, it was here evaluated after in vitro exposures of trout hepatocytes the interference caused by ethynylestradiol in the mRNA levels of an inducible (by peroxisome proliferators) and a non-inducible oxidase. The isolated Acox1 and Acox3 show high levels of sequence conservation compared to those of other teleosts. Additionally, it was found that Acox1 has two alternative splicing isoforms, corresponding to 3I and 3II isoforms of exon 3 splicing variants. Both isoforms display tissue specificity, with Acox1-3II presenting a more ubiquitous expression in comparison with Acox1-3I. Acox3 was expressed in almost all brown trout tissues. According to real-time PCR data, the highest estrogenic stimulus was able to cause a down-regulation of Acox1 and an up-regulation of Acox3. So, for Acox1 we found a negative association between an estrogenic input and a directly activated PPARα target gene. In conclusion, changes in hormonal estrogenic stimulus may impact the mobilization of hepatic lipids to the gonads, with ultimate consequences in reproduction. Further studies using in vivo assays will be fundamental to clarify these issues. PMID:26508171

  2. Acyl-coenzyme A-binding protein regulates Beta-oxidation required for growth and survival of non-small cell lung cancer.

    Science.gov (United States)

    Harris, Fredrick T; Rahman, S M Jamshedur; Hassanein, Mohamed; Qian, Jun; Hoeksema, Megan D; Chen, Heidi; Eisenberg, Rosana; Chaurand, Pierre; Caprioli, Richard M; Shiota, Masakazu; Massion, Pierre P

    2014-07-01

    We identified acyl-coenzyme A-binding protein (ACBP) as part of a proteomic signature predicting the risk of having lung cancer. Because ACBP is known to regulate β-oxidation, which in turn controls cellular proliferation, we hypothesized that ACBP contributes to regulation of cellular proliferation and survival of non-small cell lung cancer (NSCLC) by modulating β-oxidation. We used matrix-assisted laser desorption/ionization-imaging mass spectrometry (MALDI-IMS) and immunohistochemistry (IHC) to confirm the tissue localization of ABCP in pre-invasive and invasive NSCLCs. We correlated ACBP gene expression levels in NSCLCs with clinical outcomes. In loss-of-function studies, we tested the effect of the downregulation of ACBP on cellular proliferation and apoptosis in normal bronchial and NSCLC cell lines. Using tritiated-palmitate ((3)H-palmitate), we measured β-oxidation levels and tested the effect of etomoxir, a β-oxidation inhibitor, on proliferation and apoptosis. MALDI-IMS and IHC analysis confirmed that ACBP is overexpressed in pre-invasive and invasive lung cancers. High ACBP gene expression levels in NSCLCs correlated with worse survival (HR = 1.73). We observed a 40% decrease in β-oxidation and concordant decreases in proliferation and increases in apoptosis in ACBP-depleted NSCLC cells as compared with bronchial airway epithelial cells. Inhibition of β-oxidation by etomoxir in ACBP-overexpressing cells produced dose-dependent decrease in proliferation and increase in apoptosis (P = 0.01 and P oxidation.

  3. Mangiferin treatment inhibits hepatic expression of acyl-coenzyme A:diacylglycerol acyltransferase-2 in fructose-fed spontaneously hypertensive rats: a link to amelioration of fatty liver

    Energy Technology Data Exchange (ETDEWEB)

    Xing, Xiaomang; Li, Danyang; Chen, Dilong; Zhou, Liang [Faculty of Basic Medical Sciences, Chongqing Medical University, Chongqing 400016 China (China); Chonan, Ritsu [Koei Kogyo Co., Ltd., Tokyo, 101-0063 Japan (Japan); Yamahara, Johji [Pharmafood Institute, Kyoto, 602-8136 Japan (Japan); Wang, Jianwei, E-mail: wangjianwei1968@gmail.com [Department of Traditional Chinese Medicine, Chongqing Medical University, Chongqing 400016 China (China); Li, Yuhao, E-mail: yuhao@sitcm.edu.au [Endocrinology and Metabolism Group, Sydney Institute of Health Sciences/Sydney Institute of Traditional Chinese Medicine, NSW 2000 Australia (Australia)

    2014-10-15

    Mangiferin, a xanthone glucoside, and its associated traditional herbs have been demonstrated to improve abnormalities of lipid metabolism. However, its underlying mechanisms remain largely unclear. This study investigated the anti-steatotic effect of mangiferin in fructose-fed spontaneously hypertensive rat (SHR)s that have a mutation in sterol regulatory element binding protein (SREBP)-1. The results showed that co-administration of mangiferin (15 mg/kg, once daily, by oral gavage) over 7 weeks dramatically diminished fructose-induced increases in hepatic triglyceride content and Oil Red O-stained area in SHRs. However, blood pressure, fructose and chow intakes, white adipose tissue weight and metabolic parameters (plasma concentrations of glucose, insulin, triglyceride, total cholesterol and non-esterified fatty acids) were unaffected by mangiferin treatment. Mechanistically, mangiferin treatment suppressed acyl-coenzyme A:diacylglycerol acyltransferase (DGAT)-2 expression at the mRNA and protein levels in the liver. In contrast, mangiferin treatment was without effect on hepatic mRNA and/or protein expression of SREBP-1/1c, carbohydrate response element binding protein, liver pyruvate kinase, fatty acid synthase, acetyl-CoA carboxylase-1, stearoyl-CoA desaturase-1, DGAT-1, monoacyglycerol acyltransferase-2, microsomal triglyceride transfer protein, peroxisome proliferator-activated receptor-alpha, carnitine palmitoyltransferase-1 and acyl-CoA oxidase. Collectively, our results suggest that mangiferin treatment ameliorates fatty liver in fructose-fed SHRs by inhibiting hepatic DGAT-2 that catalyzes the final step in triglyceride biosynthesis. The anti-steatotic effect of mangiferin may occur independently of the hepatic signals associated with de novo fatty acid synthesis and oxidation. - Highlights: • We investigated the anti-steatotic effect of mangiferin (MA) in fructose-fed SHR. • MA (15 mg/kg/day for 7 weeks) ameliorated fructose-induced fatty liver in

  4. SLC1 and SLC4 encode partially redundant acyl-coenzyme A 1-acylglycerol-3-phosphate O-acyltransferases of budding yeast

    DEFF Research Database (Denmark)

    Benghezal, Mohammed; Roubaty, Carole; Veepuri, Vijayanath;

    2007-01-01

    does not eliminate all microsomal 1-acylglycerol-3-phosphate O-acyltransferase activity, suggesting that an additional enzyme may exist. Here we show that SLC4 (Yor175c), a gene of hitherto unknown function, encodes a second 1-acyl-sn-glycerol-3-phosphate acyltransferase. SLC4 harbors a membrane......-bound O-acyltransferase motif and down-regulation of SLC4 strongly reduces 1-acyl-sn-glycerol-3-phosphate acyltransferase activity in microsomes from slc1Delta cells. The simultaneous deletion of SLC1 and SLC4 is lethal. Mass spectrometric analysis of lipids from slc1Delta and slc4Delta cells demonstrates...... that in vivo Slc1p and Slc4p generate almost the same glycerophospholipid profile. Microsomes from slc1Delta and slc4Delta cells incubated with [14C]oleoyl-coenzyme A in the absence of lysophosphatidic acid and without CTP still incorporate the label into glycerophospholipids, indicating that Slc1p and...

  5. Photoaffinity Labeling of Developing Jojoba Seed Microsomal Membranes with a Photoreactive Analog of Acyl-Coenzyme A (Acyl-CoA) (Identification of a Putative Acyl-CoA:Fatty Alcohol Acyltransferase.

    Science.gov (United States)

    Shockey, J. M.; Rajasekharan, R.; Kemp, J. D.

    1995-01-01

    Jojoba (Simmondsia chinensis, Link) is the only plant known that synthesizes liquid wax. The final step in liquid wax biosynthesis is catalyzed by an integral membrane enzyme, fatty acyl-coenzyme A (CoA):fatty alcohol acyltransferase, which transfers an acyl chain from acyl-CoA to a fatty alcohol to form the wax ester. To purify the acyltransferase, we have labeled the enzyme with a radioiodinated, photoreactive analog of acyl-CoA, 12-[N-(4-azidosalicyl)amino] dodecanoyl-CoA (ASD-CoA). This molecule acts as an inhibitor of acyltransferase activity in the dark and as an irreversible inhibitor upon exposure to ultraviolet light. Oleoyl-CoA protects enzymatic activity in a concentration-dependent manner. Photolysis of microsomal membranes with labeled ASD-CoA resulted in strong labeling of two polypeptides of 57 and 52 kD. Increasing concentrations of oleoyl-CoA reduced the labeling of the 57-kD polypeptide dramatically, whereas the labeling of the 52-kD polypeptide was much less responsive to oleoyl-CoA. Also, unlike the other polypeptide, the labeling of the 57-kD polypeptide was enhanced considerably when photolyzed in the presence of dodecanol. These results suggest that a 57-kD polypeptide from jojoba microsomes may be the acyl-CoA:fatty alcohol acyltransferase.

  6. Characterization of two members among the five ADP-forming acyl coenzyme A (Acyl-CoA) synthetases reveals the presence of a 2-(Imidazol-4-yl)acetyl-CoA synthetase in Thermococcus kodakarensis.

    Science.gov (United States)

    Awano, Tomotsugu; Wilming, Anja; Tomita, Hiroya; Yokooji, Yuusuke; Fukui, Toshiaki; Imanaka, Tadayuki; Atomi, Haruyuki

    2014-01-01

    The genome of Thermococcus kodakarensis, along with those of most Thermococcus and Pyrococcus species, harbors five paralogous genes encoding putative α subunits of nucleoside diphosphate (NDP)-forming acyl coenzyme A (acyl-CoA) synthetases. The substrate specificities of the protein products for three of these paralogs have been clarified through studies on the individual enzymes from Pyrococcus furiosus and T. kodakarensis. Here we have examined the biochemical properties of the remaining two acyl-CoA synthetase proteins from T. kodakarensis. The TK0944 and TK2127 genes encoding the two α subunits were each coexpressed with the β subunit-encoding TK0943 gene. In both cases, soluble proteins with an α2β2 structure were obtained and their activities toward various acids in the ADP-forming reaction were examined. The purified TK0944/TK0943 protein (ACS IIITk) accommodated a broad range of acids that corresponded to those generated in the oxidative metabolism of Ala, Val, Leu, Ile, Met, Phe, and Cys. In contrast, the TK2127/TK0943 protein exhibited relevant levels of activity only toward 2-(imidazol-4-yl)acetate, a metabolite of His degradation, and was thus designated 2-(imidazol-4-yl)acetyl-CoA synthetase (ICSTk), a novel enzyme. Kinetic analyses were performed on both proteins with their respective substrates. In T. kodakarensis, we found that the addition of histidine to the medium led to increases in intracellular ADP-forming 2-(imidazol-4-yl)acetyl-CoA synthetase activity, and 2-(imidazol-4-yl)acetate was detected in the culture medium, suggesting that ICSTk participates in histidine catabolism. The results presented here, together with those of previous studies, have clarified the substrate specificities of all five known NDP-forming acyl-CoA synthetase proteins in the Thermococcales. PMID:24163338

  7. LC-quadrupole/Orbitrap high-resolution mass spectrometry enables stable isotope-resolved simultaneous quantification and ¹³C-isotopic labeling of acyl-coenzyme A thioesters.

    Science.gov (United States)

    Frey, Alexander J; Feldman, Daniel R; Trefely, Sophie; Worth, Andrew J; Basu, Sankha S; Snyder, Nathaniel W

    2016-05-01

    Acyl-coenzyme A (acyl-CoA) thioesters are evolutionarily conserved, compartmentalized, and energetically activated substrates for biochemical reactions. The ubiquitous involvement of acyl-CoA thioesters in metabolism, including the tricarboxylic acid cycle, fatty acid metabolism, amino acid degradation, and cholesterol metabolism highlights the broad applicability of applied measurements of acyl-CoA thioesters. However, quantitation of acyl-CoA levels provides only one dimension of metabolic information and a more complete description of metabolism requires the relative contribution of different precursors to individual substrates and pathways. Using two distinct stable isotope labeling approaches, acyl-CoA thioesters can be labeled with either a fixed [(13)C3(15)N1] label derived from pantothenate into the CoA moiety or via variable [(13)C] labeling into the acyl chain from metabolic precursors. Liquid chromatography-hybrid quadrupole/Orbitrap high-resolution mass spectrometry using parallel reaction monitoring, but not single ion monitoring, allowed the simultaneous quantitation of acyl-CoA thioesters by stable isotope dilution using the [(13)C3(15)N1] label and measurement of the incorporation of labeled carbon atoms derived from [(13)C6]-glucose, [(13)C5(15)N2]-glutamine, and [(13)C3]-propionate. As a proof of principle, we applied this method to human B cell lymphoma (WSU-DLCL2) cells in culture to precisely describe the relative pool size and enrichment of isotopic tracers into acetyl-, succinyl-, and propionyl-CoA. This method will allow highly precise, multiplexed, and stable isotope-resolved determination of metabolism to refine metabolic models, characterize novel metabolism, and test modulators of metabolic pathways involving acyl-CoA thioesters. PMID:26968563

  8. Bioconversion of α-linolenic acid to n-3 LCPUFA and expression of PPAR-alpha, acyl Coenzyme A oxidase 1 and carnitine acyl transferase I are incremented after feeding rats with α-linolenic acid-rich oils.

    Science.gov (United States)

    González-Mañán, Daniel; Tapia, Gladys; Gormaz, Juan Guillermo; D'Espessailles, Amanda; Espinosa, Alejandra; Masson, Lilia; Varela, Patricia; Valenzuela, Alfonso; Valenzuela, Rodrigo

    2012-07-01

    High dietary intake of n-6 fatty acids in relation to n-3 fatty acids may generate health disorders, such as cardiovascular and other chronic diseases. Fish consumption rich in n-3 fatty acids is low in Latin America, it being necessary to seek other alternatives to provide α-linolenic acid (ALA), precursor of n-3 LCPUFA (EPA and DHA). Two innovative oils were assayed, chia (Salvia hispanica) and rosa mosqueta (Rosa rubiginosa). This study evaluated hepatic bioconversion of ALA to EPA and DHA, expression of PPAR-α, acyl-Coenzyme A oxidase 1 (ACOX1) and carnitine acyltransferase I (CAT-I), and accumulation of EPA and DHA in plasma and adipose tissue in Sprague-Dawley rats. Three experimental groups were fed 21 days: sunflower oil (SFO, control); chia oil (CO); rosa mosqueta oil (RMO). Fatty acid composition of total lipids and phospholipids from plasma, hepatic and adipose tissue was assessed by gas-liquid chromatography and TLC. Expression of PPAR-α (RT-PCR) and ACOX1 and CAT-I (Western blot). CO and RMO increased plasma, hepatic and adipose tissue levels of ALA, EPA and DHA and decreased n-6:n-3 ratio compared to SFO (p < 0.05, One-way ANOVA and Newman-Keuls test). CO increased levels of ALA and EPA compared to RMO (p < 0.05). No significant differences were observed for DHA levels. CO also increased the expression of PPAR-α, ACOX1 and CAT-I. Only CAT-I levels were increased by RO. CO and RMO may be a nutritional alternative to provide ALA for its bioconversion to EPA and DHA, and to increase the expression of PPAR-α, ACOX1 and CAT-I, especially CO-oil.

  9. Lysophosphatidic acid effects on atherosclerosis and thrombosis

    OpenAIRE

    Cui, Mei-Zhen

    2011-01-01

    Lysophosphatidic acid (LPA) has been found to accumulate in high concentrations in atherosclerotic lesions. LPA is a bioactive phospholipid produced by activated platelets and formed during the oxidation of LDL. Accumulating evidence suggests that this lipid mediator may serve as an important risk factor for development of atherosclerosis and thrombosis. The role of LPA in atherogenesis is supported by the evidence that LPA: stimulates endothelial cells to produce adhesion molecules and chemo...

  10. 正常中国人及内源性高甘油三酯血症患者酰基辅酶A:胆固醇酰基转移酶基因多态性的研究%Analysis of acyl-coenzyme A:cholesterol acyltransferase 1 polymorphism in patients with endogenous hypertriglyceridemia in Chinese population

    Institute of Scientific and Technical Information of China (English)

    李琴; 白怀; 范平; 刘瑞; 刘宇; 刘秉文

    2008-01-01

    目的 研究酰基辅酶A:胆固醇酰基转移酶1(acyl-coenzyme A:cholesterol acyltransferase 1,ACAT1)基因rs1044925多态性是否与正常汉族中国人及内源性高甘油三酯血症(hypertriglyceridemia,HTG)患者血脂及载脂蛋白水平存在关联.方法 应用聚合酶链反应-限制性片段长度多态性分析法,对成都地区372名汉族人(267名正常人和105例内源性高甘油三酯血症患者)ACAT1基因rs1044925多态位点进行分析.结果 中国人ACAT1基因rs1044925多态位点C等位基因频率为0.137,显著低于中部和南部欧洲人的0.354(P<0.05);HTG组和对照组C等位基因频率分别为0.153和0.137,两者之间差异无统计学意义.对照组AA基因型携带者血清低密度脂蛋白胆固醇(low density lipoprotein-cholesterol,LDL-C)和非高密度脂蛋白胆固醇(non-high density lipoprotein cholesterol,nHDL-C)水平均较C等位基因携带者(Ac和CC基因型者)显著升高[(3.25±0.68)mmol/L vs(3.03±0.87)mmol/L,P<0.05;(3.80±0.71)mmol/L vs(3.23±0.82)mmol/L,P<0.05],HTG组AA基因型携带者血清高密度脂蛋白胆固醇(high density lipoprotein-cholesterol,HDL-c)水平较C等位基因携带者显著升高[(1.00±0.28)mmol/L vs(0.87±0.17)mmoL/L,P<0.05].结论 ACAT1 基因rs1044925多态性不仅与正常中国成都地区汉族人血清LDL-C、nHDL-C含量有关,而且还与内源性高甘油三酯血症患者血清HDL-C水平相关联.%Objective To investigate the polymorphism of acyl-coenzyme A:cholesterol acyltransfemse 1(ACAT1)gene and its relationship with endogenous hypertriglyceridemia(HTG)in Chinese population.Methods A total of three hundred and seventy-two subjects(105 endogenous hypertriglyceridemics and 267 healthy controls)from a population of Chinese Han nationality in Chengdu area were studied using PCR-restriction fragment length polymorphism (RFLP).Results The frequency of C allele in normal Chinese at rs1044925 locus was 0.137,which was lower thanthat reported in the

  11. Spiroguanidine rhodamines as fluorogenic probes for lysophosphatidic acid

    OpenAIRE

    Wang, Lei; Sibrian-Vazquez, Martha; Escobedo, Jorge O.; Wang, Jialu; Moore, Richard G.; Strongin, Robert M.

    2015-01-01

    Direct determination of total lysophosphatidic acid (LPA) was accomplished using newly developed spiroguanidines derived from rhodamine B as universal fluorogenic probes. Optimum conditions for the quantitative analysis of total LPA were investigated. The linear range for the determination of total LPA is up to 5 μM with a limit of detection of 0.512 μM.

  12. Simple enrichment and analysis of plasma lysophosphatidic acids

    OpenAIRE

    Wang, Jialu; Sibrian-Vazquez, Martha; Escobedo, Jorge O.; Lowry, Mark; Wang, Lei; Chu, Yu-Hsuan; Moore, Richard G.; Strongin, Robert M.

    2013-01-01

    A simple and highly efficient technique for the analysis of lysophosphatidic acid (LPA) subspecies in human plasma is described. The streamlined sample preparation protocol furnishes the five major LPA subspecies with excellent recoveries. Extensive analysis of the enriched sample reveals only trace levels of other phospholipids. This level of purity not only improves MS analyses, but enables HPLC post-column detection in the visible region with a commercially available fluorescent phospholip...

  13. Lysophosphatidic acid is a chemoattractant for Dictyostelium discoideum amoebae.

    OpenAIRE

    Jalink, K.; Moolenaar, W H; Duijn, B., Van

    1993-01-01

    The naturally occurring phospholipid lysophosphatidic acid (LPA) can induce a number of physiological responses in vertebrate cells, including platelet aggregation, smooth muscle contraction, and fibroblast proliferation. LPA is thought to activate a specific G-protein-coupled receptor, thereby triggering classic second messenger pathways such as stimulation of phospholipase C and inhibition of adenylate cyclase. Here we report that 1-oleoyl-LPA, at submicromolar concentrations, evokes a chem...

  14. Lysophosphatidic Acid Signaling through the Lysophosphatidic Acid-1 Receptor Is Required for Alveolarization.

    Science.gov (United States)

    Funke, Manuela; Knudsen, Lars; Lagares, David; Ebener, Simone; Probst, Clemens K; Fontaine, Benjamin A; Franklin, Alicia; Kellner, Manuela; Kühnel, Mark; Matthieu, Stephanie; Grothausmann, Roman; Chun, Jerold; Roberts, Jesse D; Ochs, Matthias; Tager, Andrew M

    2016-07-01

    Lysophosphatidic acid (LPA) signaling through one of its receptors, LPA1, contributes to both the development and the pathological remodeling after injury of many organs. Because we found previously that LPA-LPA1 signaling contributes to pulmonary fibrosis, here we investigated whether this pathway is also involved in lung development. Quantitative assessment of lung architecture of LPA1-deficient knock-out (KO) and wild-type (WT) mice at 3, 12, and 24 weeks of age using design-based stereology suggested the presence of an alveolarization defect in LPA1 KO mice at 3 weeks, which persisted as alveolar numbers increased in WT mice into adulthood. Across the ages examined, the lungs of LPA1 KO mice exhibited decreased alveolar numbers, septal tissue volumes, and surface areas, and increased volumes of the distal airspaces. Elastic fibers, critical to the development of alveolar septa, appeared less organized and condensed and more discontinuous in KO alveoli starting at P4. Tropoelastin messenger RNA expression was decreased in KO lungs, whereas expression of matrix metalloproteinases degrading elastic fibers was either decreased or unchanged. These results are consistent with the abnormal lung phenotype of LPA1 KO mice, being attributable to reduced alveolar septal formation during development, rather than to increased septal destruction as occurs in the emphysema of chronic obstructive pulmonary disease. Peripheral septal fibroblasts and myofibroblasts, which direct septation in late alveolarization, demonstrated reduced production of tropoelastin and matrix metalloproteinases, and diminished LPA-induced migration, when isolated from LPA1 KO mice. Taken together, our data suggest that LPA-LPA1 signaling is critically required for septation during alveolarization. PMID:27082727

  15. Tetracyclines increase lipid phosphate phosphatase expression on plasma membranes and turnover of plasma lysophosphatidate.

    Science.gov (United States)

    Tang, Xiaoyun; Zhao, Yuan Y; Dewald, Jay; Curtis, Jonathan M; Brindley, David N

    2016-04-01

    Extracellular lysophosphatidate and sphingosine 1-phosphate (S1P) are important bioactive lipids, which signal through G-protein-coupled receptors to stimulate cell growth and survival. The lysophosphatidate and S1P signals are terminated partly by degradation through three broad-specificity lipid phosphate phosphatases (LPPs) on the cell surface. Significantly, the expression of LPP1 and LPP3 is decreased in many cancers, and this increases the impact of lysophosphatidate and S1P signaling. However, relatively little is known about the physiological or pharmacological regulation of the expression of the different LPPs. We now show that treating several malignant and nonmalignant cell lines with 1 μg/ml tetracycline, doxycycline, or minocycline significantly increased the extracellular degradation of lysophosphatidate. S1P degradation was also increased in cells that expressed high LPP3 activity. These results depended on an increase in the stabilities of the three LPPs and increased expression on the plasma membrane. We tested the physiological significance of these results and showed that treating rats with doxycycline accelerated the clearance of lysophosphatidate, but not S1P, from the circulation. However, administering 100 mg/kg/day doxycycline to mice decreased plasma concentrations of lysophosphatidate and S1P. This study demonstrates a completely new property of tetracyclines in increasing the plasma membrane expression of the LPPs.

  16. Tetracyclines increase lipid phosphate phosphatase expression on plasma membranes and turnover of plasma lysophosphatidate.

    Science.gov (United States)

    Tang, Xiaoyun; Zhao, Yuan Y; Dewald, Jay; Curtis, Jonathan M; Brindley, David N

    2016-04-01

    Extracellular lysophosphatidate and sphingosine 1-phosphate (S1P) are important bioactive lipids, which signal through G-protein-coupled receptors to stimulate cell growth and survival. The lysophosphatidate and S1P signals are terminated partly by degradation through three broad-specificity lipid phosphate phosphatases (LPPs) on the cell surface. Significantly, the expression of LPP1 and LPP3 is decreased in many cancers, and this increases the impact of lysophosphatidate and S1P signaling. However, relatively little is known about the physiological or pharmacological regulation of the expression of the different LPPs. We now show that treating several malignant and nonmalignant cell lines with 1 μg/ml tetracycline, doxycycline, or minocycline significantly increased the extracellular degradation of lysophosphatidate. S1P degradation was also increased in cells that expressed high LPP3 activity. These results depended on an increase in the stabilities of the three LPPs and increased expression on the plasma membrane. We tested the physiological significance of these results and showed that treating rats with doxycycline accelerated the clearance of lysophosphatidate, but not S1P, from the circulation. However, administering 100 mg/kg/day doxycycline to mice decreased plasma concentrations of lysophosphatidate and S1P. This study demonstrates a completely new property of tetracyclines in increasing the plasma membrane expression of the LPPs. PMID:26884614

  17. Lysophosphatidic Acid (LPA Signaling in Human and Ruminant Reproductive Tract

    Directory of Open Access Journals (Sweden)

    Izabela Wocławek-Potocka

    2014-01-01

    Full Text Available Lysophosphatidic acid (LPA through activating its G protein-coupled receptors (LPAR 1–6 exerts diverse cellular effects that in turn influence several physiological processes including reproductive function of the female. Studies in various species of animals and also in humans have identified important roles for the receptor-mediated LPA signaling in multiple aspects of human and animal reproductive tract function. These aspects range from ovarian and uterine function, estrous cycle regulation, early embryo development, embryo implantation, decidualization to pregnancy maintenance and parturition. LPA signaling can also have pathological consequences, influencing aspects of endometriosis and reproductive tissue associated tumors. The review describes recent progress in LPA signaling research relevant to human and ruminant reproduction, pointing at the cow as a relevant model to study LPA influence on the human reproductive performance.

  18. Inhibition of lysophospholipase D activity by unsaturated lysophosphatidic acids or seed extracts containing 1-linoleoyl and 1-oleoyl lysophosphatidic acid.

    Science.gov (United States)

    Liu, Xi-Wen; Sok, Dai-Eun; Yook, Hong-Sun; Sohn, Cheon-Bae; Chung, Young-Jin; Kim, Mee Ree

    2007-10-17

    Lysophospholipase D (lysoPLD), generating lipid mediator lysophosphatidic acid (LPA) from lysophosphatidyclcholine (LPC), is known to be inhibited by lysophosphatidic acids. Meanwhile, some plant lipids are known to contain lysophospholipids as minor components. Therefore, it is interesting to test whether edible seed samples, rich in phospholipids, may contain lysophospholipids, which express a strong inhibition of lysoPLD activity. First, the structural importance of fatty acyl group in LPAs was examined by determining the inhibitory effect of various LPAs on bovine lysoPLD activity. The most potent in the inhibition of lysoPLD activity was linoleoyl-LPA ( K i, 0.21 microM), followed by arachidonoyl-LPA ( K i, 0.55 microM), oleoyl-LPA ( K i, 1.2 microM), and palmitoyl-LPA ( K i, 1.4 microM), based on the fluoresecent assay. The same order of inhibitory potency among LPA analogs with different acyl chains was also found in the spectrophotometric assay. Subsequently, the extracts of 12 edible seeds were screened for the inhibition of lysoPLD activity using both spectrophotometric and fluorescent assays. Among seed extracts tested, the extract from soybean seed, sesame seed, or sunflower seed (30 mg seed weight/mL) was found to exhibit a potent inhibition (>80%) of lysoPLD activity. In further study employing ESI-MS/MS analysis, major LPA components in seed extracts were identified to be 1-linoleoyl LPA, 1-oleoyl LPA, and 1-palmitoyl LPA with 1-linoleoyl LPA being more predominant. Thus, the potent inhibition of lysoPLD activity by seed extracts might be ascribed to the presence of LPA with linoleoyl group rather than other acyl chains. PMID:17887800

  19. Lysophosphatidic acid metabolism and elimination in cardiovascular disease

    Science.gov (United States)

    Salous, Abdelghaffar Kamal

    The bioactive lipids lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are present in human and mouse plasma at a concentration of ~0.1-1 microM and regulate physiological and pathophysiological processes in the cardiovascular system including atherothrombosis, intimal hyperplasia, and immune function, edema formation, and permeability. PPAP2B, the gene encoding LPP3, a broad activity integral membrane enzyme that terminates LPA actions in the vasculature, has a single nucleotide polymorphism that been recently associated with coronary artery disease risk. The synthesis and signaling of LPA and S1P in the cardiovascular system have been extensively studied but the mechanisms responsible for their elimination are less well understood. The broad goal of this research was to examine the role of LPP3 in the termination of LPA signaling in models of cardiovascular disease involving vascular wall cells, investigate the role of LPP3 in the elimination of plasma LPA, and further characterize the elimination of plasma LPA. The central hypothesis is that LPP3 plays an important role in attenuating the pathological responses to LPA signaling and that it mediates the elimination of exogenously applied bioactive lipids from the plasma. These hypotheses were tested using molecular biological approaches, in vitro studies, synthetic lysophospholipid mimetics, modified surgical procedures, and mass spectrometry assays. My results indicated that LPP3 played a critical role in attenuating LPA signaling mediating the pathological processes of intimal hyperplasia and vascular leak in mouse models of disease. Additionally, enzymatic inactivation of lysophospholipids by LPP and PLA enzymes in the plasma was not a primary mechanism for the rapid elimination of plasma LPA and S1P. Instead, evidence strongly suggested a transcellular uptake mechanism by hepatic non-parenchymal cells as the predominant mechanism for elimination of these molecules. These results support a model in

  20. Roles of lysophosphatidic acid in cardiovascular physiology and disease.

    Science.gov (United States)

    Smyth, Susan S; Cheng, Hsin-Yuan; Miriyala, Sumitra; Panchatcharam, Manikandan; Morris, Andrew J

    2008-09-01

    The bioactive lipid mediator lysophosphatidic acid (LPA) exerts a range of effects on the cardiovasculature that suggest a role in a variety of critical cardiovascular functions and clinically important cardiovascular diseases. LPA is an activator of platelets from a majority of human donors identifying a possible role as a regulator of acute thrombosis and platelet function in atherogenesis and vascular injury responses. Of particular interest in this context, LPA is an effective phenotypic modulator of vascular smooth muscle cells promoting the de-differentiation, proliferation and migration of these cells that are required for the development of intimal hyperplasia. Exogenous administration of LPA results in acute and systemic changes in blood pressure in different animal species, suggesting a role for LPA in both normal blood pressure regulation and hypertension. Advances in our understanding of the molecular machinery responsible for the synthesis, actions and inactivation of LPA now promise to provide the tools required to define the role of LPA in cardiovascular physiology and disease. In this review we discuss aspects of LPA signaling in the cardiovasculature focusing on recent advances and attempting to highlight presently unresolved issues and promising avenues for further investigation.

  1. Spontaneous curvature of phosphatidic acid and lysophosphatidic acid.

    Science.gov (United States)

    Kooijman, Edgar E; Chupin, Vladimir; Fuller, Nola L; Kozlov, Michael M; de Kruijff, Ben; Burger, Koert N J; Rand, Peter R

    2005-02-15

    The formation of phosphatidic acid (PA) from lysophosphatidic acid (LPA), diacylglycerol, or phosphatidylcholine plays a key role in the regulation of intracellular membrane fission events, but the underlying molecular mechanism has not been resolved. A likely possibility is that PA affects local membrane curvature facilitating membrane bending and fission. To examine this possibility, we determined the spontaneous radius of curvature (R(0p)) of PA and LPA, carrying oleoyl fatty acids, using well-established X-ray diffraction methods. We found that, under physiological conditions of pH and salt concentration (pH 7.0, 150 mM NaCl), the R(0p) values of PA and LPA were -46 A and +20 A, respectively. Thus PA has considerable negative spontaneous curvature while LPA has the most positive spontaneous curvature of any membrane lipid measured to date. The further addition of Ca(2+) did not significantly affect lipid spontaneous curvature; however, omitting NaCl from the hydration buffer greatly reduced the spontaneous curvature of PA, turning it into a cylindrically shaped lipid molecule (R(0p) of -1.3 x 10(2) A). Our quantitative data on the spontaneous radius of curvature of PA and LPA at a physiological pH and salt concentration will be instrumental in developing future models of biomembrane fission. PMID:15697235

  2. Lysophosphatidate induces chemo-resistance by releasing breast cancer cells from taxol-induced mitotic arrest.

    Directory of Open Access Journals (Sweden)

    Nasser Samadi

    Full Text Available BACKGROUND: Taxol is a microtubule stabilizing agent that arrests cells in mitosis leading to cell death. Taxol is widely used to treat breast cancer, but resistance occurs in 25-69% of patients and it is vital to understand how Taxol resistance develops to improve chemotherapy. The effects of chemotherapeutic agents are overcome by survival signals that cancer cells receive. We focused our studies on autotaxin, which is a secreted protein that increases tumor growth, aggressiveness, angiogenesis and metastasis. We discovered that autotaxin strongly antagonizes the Taxol-induced killing of breast cancer and melanoma cells by converting the abundant extra-cellular lipid, lysophosphatidylcholine, into lysophosphatidate. This lipid stimulates specific G-protein coupled receptors that activate survival signals. METHODOLOGY/PRINCIPAL FINDINGS: In this study we determined the basis of these antagonistic actions of lysophosphatidate towards Taxol-induced G2/M arrest and cell death using cultured breast cancer cells. Lysophosphatidate does not antagonize Taxol action in MCF-7 cells by increasing Taxol metabolism or its expulsion through multi-drug resistance transporters. Lysophosphatidate does not lower the percentage of cells accumulating in G2/M by decreasing exit from S-phase or selective stimulation of cell death in G2/M. Instead, LPA had an unexpected and remarkable action in enabling MCF-7 and MDA-MB-468 cells, which had been arrested in G2/M by Taxol, to normalize spindle structure and divide, thus avoiding cell death. This action involves displacement of Taxol from the tubulin polymer fraction, which based on inhibitor studies, depends on activation of LPA receptors and phosphatidylinositol 3-kinase. CONCLUSIONS/SIGNIFICANCE: This work demonstrates a previously unknown consequence of lysophosphatidate action that explains why autotaxin and lysophosphatidate protect against Taxol-induced cell death and promote resistance to the action of this

  3. The autotaxin-lysophosphatidic acid–lysophosphatidic acid receptor cascade: proposal of a novel potential therapeutic target for treating glioblastoma multiforme

    OpenAIRE

    Tabuchi, Sadaharu

    2015-01-01

    Glioblastoma multiforme (GBM) is the most malignant tumor of the central nervous system (CNS). Its prognosis is one of the worst among all cancer types, and it is considered a fatal malignancy, incurable with conventional therapeutic strategies. As the bioactive multifunctional lipid mediator lysophosphatidic acid (LPA) is well recognized to be involved in the tumorigenesis of cancers by acting on G-protein-coupled receptors, LPA receptor (LPAR) antagonists and LPA synthesis inhibitors have b...

  4. How Chain Length and Charge Affect Surfactant Denaturation of Acyl Coenzyme A Binding Protein (ACBP)

    DEFF Research Database (Denmark)

    Andersen, Kell Kleiner; Otzen, Daniel

    2009-01-01

    . This behavior contrasts with the simplicity of unfolding in chemical denaturants and highlights the changing properties of surfactant micelles. We suggest that the transition from spherical to more elongated micelles leads to inhibition of unfolding kinetics, while weaker binding sites may cause a subsequent...

  5. Regulation of lipolytic activity by long-chain acyl-coenzyme A in islets and adipocytes

    DEFF Research Database (Denmark)

    Hu, Liping; Deeney, Jude T; Nolan, Christopher J;

    2005-01-01

    normal and hormone-sensitive lipase (HSL)-null mice and in phosphatase-treated islets, indicating that the stimulatory effect was neither on HSL nor phosphorylation dependent. In contrast, we reproduced the previously published observations showing inhibition of HSL activity by LC-CoA in adipocytes....... The inhibitory effect of LC-CoA on adipocyte HSL was dependent on phosphorylation and enhanced by acyl-CoA-binding protein (ACBP). In contrast, the stimulatory effect on islet lipase activity was blocked by ACBP, presumably due to binding and sequestration of LC-CoA. These data suggest the following intertissue...

  6. Thermodynamics of ligand binding to acyl-coenzyme A binding protein studied by titration calorimetry

    DEFF Research Database (Denmark)

    Færgeman, Nils J.; Sigurskjold, B W; Kragelund, B B;

    1996-01-01

    Ligand binding to recombinant bovine acyl-CoA binding protein (ACBP) was examined using isothermal microcalorimetry. Microcalorimetric measurements confirm that the binding affinity of acyl-CoA esters for ACBP is strongly dependent on the length of the acyl chain with a clear preference for acyl-...

  7. Polyunsaturated fatty acyl-coenzyme As are inhibitors of cholesterol biosynthesis in zebrafish and mice

    Directory of Open Access Journals (Sweden)

    Santhosh Karanth

    2013-11-01

    Lipid disorders pose therapeutic challenges. Previously we discovered that mutation of the hepatocyte β-hydroxybutyrate transporter Slc16a6a in zebrafish causes hepatic steatosis during fasting, marked by increased hepatic triacylglycerol, but not cholesterol. This selective diversion of trapped ketogenic carbon atoms is surprising because acetate and acetoacetate can exit mitochondria and can be incorporated into both fatty acids and cholesterol in normal hepatocytes. To elucidate the mechanism of this selective diversion of carbon atoms to fatty acids, we fed wild-type and slc16a6a mutant animals high-protein ketogenic diets. We find that slc16a6a mutants have decreased activity of the rate-limiting enzyme of cholesterol biosynthesis, 3-hydroxy-3-methylglutaryl-coenzyme A reductase (Hmgcr, despite increased Hmgcr protein abundance and relative incorporation of mevalonate into cholesterol. These observations suggest the presence of an endogenous Hmgcr inhibitor. We took a candidate approach to identify such inhibitors. First, we found that mutant livers accumulate multiple polyunsaturated fatty acids (PUFAs and PUFA-CoAs, and we showed that human HMGCR is inhibited by PUFA-CoAs in vitro. Second, we injected mice with an ethyl ester of the PUFA eicosapentaenoic acid and observed an acute decrease in hepatic Hmgcr activity, without alteration in Hmgcr protein abundance. These results elucidate a mechanism for PUFA-mediated cholesterol lowering through direct inhibition of Hmgcr.

  8. Lysophosphatidic Acid Receptor Is a Functional Marker of Adult Hippocampal Precursor Cells

    OpenAIRE

    Tara L. Walker; Rupert W. Overall; Steffen Vogler; Alex M. Sykes; Susann Ruhwald; Daniela Lasse; Muhammad Ichwan; Klaus Fabel; Gerd Kempermann

    2016-01-01

    Summary Here, we show that the lysophosphatidic acid receptor 1 (LPA1) is expressed by a defined population of type 1 stem cells and type 2a precursor cells in the adult mouse dentate gyrus. LPA1, in contrast to Nestin, also marks the quiescent stem cell population. Combining LPA1-GFP with EGFR and prominin-1 expression, we have enabled the prospective separation of both proliferative and non-proliferative precursor cell populations. Transcriptional profiling of the isolated proliferative pre...

  9. Lysophosphatidic Acid Stimulates MCP-1 Secretion from C2C12 Myoblast

    OpenAIRE

    Tamotsu Tsukahara; Hisao Haniu

    2012-01-01

    Chemokines are regulatory proteins that play an important role in muscle cell migration and proliferation. In this study, C2C12 cells treated with lysophosphatidic acid (LPA) showed an increase in endogenous monocyte chemotactic protein-1 (MCP-1) expression and secretion. LPA is a naturally occurring bioactive lysophospholipid with hormone- and growth-factor-like activities. LPA is produced by activated platelets, cytokine-stimulated leukocytes, and possibly by other cell types. However, the ...

  10. What makes the bioactive lipids phosphatidic acid and lysophosphatidic acid so special?

    Science.gov (United States)

    Kooijman, Edgar E; Carter, Karen M; van Laar, Emma G; Chupin, Vladimir; Burger, Koert N J; de Kruijff, Ben

    2005-12-27

    Phosphatidic acid and lysophosphatidic acid are minor but important anionic bioactive lipids involved in a number of key cellular processes, yet these molecules have a simple phosphate headgroup. To find out what is so special about these lipids, we determined the ionization behavior of phosphatidic acid (PA) and lysophosphatidic acid (LPA) in extended (flat) mixed lipid bilayers using magic angle spinning 31P NMR. Our data show two surprising results. First, despite identical phosphomonoester headgroups, LPA carries more negative charge than PA when present in a phosphatidylcholine bilayer. Dehydroxy-LPA [1-oleoyl-3-(phosphoryl)propanediol] behaves in a manner identical to that of PA, indicating that the difference in negative charge between LPA and PA is caused by the hydroxyl on the glycerol backbone of LPA and its interaction with the phosphomonoester headgroup. Second, deprotonation of phosphatidic acid and lysophosphatidic acid was found to be strongly stimulated by the inclusion of phosphatidylethanolamine in the bilayer, indicating that lipid headgroup charge depends on local lipid composition and will vary between the different subcellular locations of (L)PA. Our findings can be understood in terms of a hydrogen bond formed within the phosphomonoester headgroup of (L)PA and its destabilization by competing intra- or intermolecular hydrogen bonds. We propose that this hydrogen bonding property of (L)PA is involved in the various cellular functions of these lipids. PMID:16363814

  11. Lysophosphatidic Acid Signaling in Late Cleavage and Blastocyst Stage Bovine Embryos

    OpenAIRE

    Ana Catarina Torres; Dorota Boruszewska; Mariana Batista; Ilona Kowalczyk-Zieba; Patricia Diniz; Emilia Sinderewicz; Jean Sebastian Saulnier-Blache; Izabela Woclawek-Potocka; Luis Lopes-da-Costa

    2014-01-01

    Lysophosphatidic acid (LPA) is a known cell signaling lipid mediator in reproductive tissues. In the cow, LPA is involved in luteal and early pregnancy maintenance. Here, we evaluated the presence and role of LPA in bovine early embryonic development. In relevant aspects, bovine embryos reflect more closely the scenario occurring in human embryos than the mouse model. Transcription of mRNA and protein expression of enzymes involved in LPA synthesis (ATX and cPLA2) and of LPA receptors (LPAR1–...

  12. Mutations of lysophosphatidic acid receptor-1 gene during progression of lung tumors in rats

    International Nuclear Information System (INIS)

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. In this study, mutations of lysophosphatidic acid receptor-1 (LPA1) gene were investigated to clarify the possible molecular mechanisms underlying the development of lung tumors induced by N-nitrosobis(2-hydroxypropyl)amine (BHP) in rats. Male Wistar rats, 6 weeks of age, were given 2000 ppm BHP in their drinking water for 12 weeks and then maintained without further treatment until sacrifice at 25 weeks. Genomic DNAs were extracted from paraffin-embedded tissues and exons 2-4 were examined for mutations, using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) analysis. No LPA1 mutations were detected in 15 hyperplasias, but 2 out of 12 adenomas (16.7%) and 7 out of 17 adenocarcinomas (41.2%). These results suggest that mutations of LPA1 gene may be involved in the acquisition of growth advantage from adenomas to adenocarcinomas in lung carcinogenesis induced in rats by BHP.

  13. Lysophosphatidic Acid Receptor Is a Functional Marker of Adult Hippocampal Precursor Cells

    Directory of Open Access Journals (Sweden)

    Tara L. Walker

    2016-04-01

    Full Text Available Here, we show that the lysophosphatidic acid receptor 1 (LPA1 is expressed by a defined population of type 1 stem cells and type 2a precursor cells in the adult mouse dentate gyrus. LPA1, in contrast to Nestin, also marks the quiescent stem cell population. Combining LPA1-GFP with EGFR and prominin-1 expression, we have enabled the prospective separation of both proliferative and non-proliferative precursor cell populations. Transcriptional profiling of the isolated proliferative precursor cells suggested immune mechanisms and cytokine signaling as molecular regulators of adult hippocampal precursor cell proliferation. In addition to LPA1 being a marker of this important stem cell population, we also show that the corresponding ligand LPA is directly involved in the regulation of adult hippocampal precursor cell proliferation and neurogenesis, an effect that can be attributed to LPA signaling via the AKT and MAPK pathways.

  14. Cloning and identification of the human LPAAT-zeta gene, a novel member of the lysophosphatidic acid acyltransferase family.

    Science.gov (United States)

    Li, Dan; Yu, Long; Wu, Hai; Shan, Yuxi; Guo, Jinhu; Dang, Yongjun; Wei, Youheng; Zhao, Shouyuan

    2003-01-01

    Lysophosphatidic acid (LPA) is a naturally occurring component of phospholipid and plays a critical role in the regulation of many physiological and pathophysiological processes including cell growth, survival, and pro-angiogenesis. LPA is converted to phosphatidic acid by the action of lysophosphatidic acid acyltransferase (LPAAT). Five members of the LPAAT gene family have been detected in humans to date. Here, we report the identification of a novel LPAAT member, which is designated as LPAAT-zeta. LPAAT-zeta was predicted to encode a protein consisting of 456 amino acid residues with a signal peptide sequence and the acyltransferase domain. Northern blot analysis showed that LPAAT-zeta was ubiquitously expressed in all 16 human tissues examined, with levels in the skeletal muscle, heart, and testis being relatively high and in the lung being relatively low. The human LPAAT-zeta gene consisted of 13 exons and is positioned at chromosome 8p11.21. PMID:12938015

  15. Lysophosphatidic Acid Alters the Expression Profiles of Angiogenic Factors, Cytokines, and Chemokines in Mouse Liver Sinusoidal Endothelial Cells

    OpenAIRE

    Chia-Hung Chou; Shou-Lun Lai; Cheng-Maw Ho; Wen-Hsi Lin; Chiung-Nien Chen; Po-Huang Lee; Fu-Chuo Peng; Sung-Hsin Kuo; Szu-Yuan Wu; Hong-Shiee Lai

    2015-01-01

    Background and Aims Lysophosphatidic acid (LPA) is a multi-function glycerophospholipid. LPA affects the proliferation of hepatocytes and stellate cells in vitro, and in a partial hepatectomy induced liver regeneration model, the circulating LPA levels and LPA receptor (LPAR) expression levels in liver tissue are significantly changed. Liver sinusoidal endothelial cells (Lsecs) play an important role during liver regeneration. However, the effects of LPA on Lsecs are not well known. Thus, we ...

  16. Lysophosphatidic acid stimulates gastric cancer cell proliferation via ERK1-dependent upregulation of sphingosine kinase 1 transcription

    OpenAIRE

    Ramachandran, Subramaniam; Shida, Dai; Nagahashi, Masayuki; Fang, Xianjun; Milstien, Sheldon; Takabe, Kazuaki; Spiegel, Sarah

    2010-01-01

    In MKN1 gastric cancer cells, lysophosphatidic acid (LPA) upregulates expression of sphingosine kinase 1 (SphK1) and its downregulation or inhibition suppresses LPA-mediated proliferation. Although LPA activates numerous signaling pathways downstream of its receptors, including ERK1/2, p38, JNK, and Akt, and the transactivation of the EGF receptor, pharmacological and molecular approaches demonstrated that only activation of ERK1, in addition to the CCAAT/enhancer-binding protein β (C/EBPβ) t...

  17. Determination of an ensemble of structures representing the denatured state of the bovine acyl-coenzyme a binding protein

    DEFF Research Database (Denmark)

    Lindorff-Larsen, Kresten; Kristjansdottir, Sigridur; Teilum, Kaare;

    2004-01-01

    is highly heterogeneous. The high sensitivity of the computational method that we present, however, enabled us to identify long-range interactions between two regions, located near the N- and C-termini, that include both native and non-native elements. The preferential formation of these contacts suggests...

  18. Ethylmalonic aciduria is associated with an amino acid variant of short chain acyl-coenzyme A dehydrogenase

    DEFF Research Database (Denmark)

    Corydon, M J; Gregersen, N; Lehnert, W;

    1996-01-01

    population, respectively. One hundred and thirty-five patients from Germany, Denmark, the Czech Republic, Spain, and the United States were selected for this study on the basis of abnormal EMA excretion ranging from 18 to 1185 mmol/mol of creatinine (controls ...Ethylmalonic aciduria is a common biochemical finding in patients with inborn errors of short chain fatty acid beta-oxidation. The urinary excretion of ethylmalonic acid (EMA) may stem from decreased oxidation by short chain acyl-CoA dehydrogenase (SCAD) of butyryl-CoA, which is alternatively...

  19. Genetic basis for correction of very-long-chain acyl-coenzyme A dehydrogenase deficiency by bezafibrate in patient fibroblasts

    DEFF Research Database (Denmark)

    Gobin-Limballe, S; Djouadi, F; Aubey, F;

    2007-01-01

    there is no established treatment. Recent data suggest that bezafibrate could improve the FAO capacities in beta-oxidation-deficient cells, by enhancing the residual level of mutant enzyme activity via gene-expression stimulation. Since VLCAD-deficient patients frequently harbor missense mutations with unpredictable...

  20. A novel highly potent autotaxin/ENPP2 inhibitor produces prolonged decreases in plasma lysophosphatidic acid formation in vivo and regulates urethral tension.

    Directory of Open Access Journals (Sweden)

    Hiroshi Saga

    Full Text Available Autotaxin, also known as ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2, is a secreted enzyme that has lysophospholipase D activity, which converts lysophosphatidylcholine to bioactive lysophosphatidic acid. Lysophosphatidic acid activates at least six G-protein coupled recpetors, which promote cell proliferation, survival, migration and muscle contraction. These physiological effects become dysfunctional in the pathology of cancer, fibrosis, and pain. To date, several autotaxin/ENPP2 inhibitors have been reported; however, none were able to completely and continuously inhibit autotaxin/ENPP2 in vivo. In this study, we report the discovery of a highly potent autotaxin/ENPP2 inhibitor, ONO-8430506, which decreased plasma lysophosphatidic acid formation. The IC50 values of ONO-8540506 for lysophospholipase D activity were 6.4-19 nM for recombinant autotaxin/ENPP2 proteins and 4.7-11.6 nM for plasma from various animal species. Plasma lysophosphatidic acid formation during 1-h incubation was almost completely inhibited by the addition of >300 nM of the compound to human plasma. In addition, when administered orally to rats at a dose of 30 mg/kg, the compound demonstrated good pharmacokinetics in rats and persistently inhibited plasma lysophosphatidic acid formation even at 24 h after administration. Smooth muscle contraction is a known to be promoted by lysophosphatidic acid. In this study, we showed that dosing rats with ONO-8430506 decreased intraurethral pressure accompanied by urethral relaxation. These findings demonstrate the potential of this autotaxin/ENPP2 inhibitor for the treatment of various diseases caused by lysophosphatidic acid, including urethral obstructive disease such as benign prostatic hyperplasia.

  1. Toluene diisocyanate: Induction of the autotaxin-lysophosphatidic acid axis and its association with airways symptoms

    Energy Technology Data Exchange (ETDEWEB)

    Broström, Julia M. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Ye, Zhi-wei [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden); Axmon, Anna; Littorin, Margareta; Tinnerberg, Håkan; Lindh, Christian H. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Zheng, Huiyuan; Ghalali, Aram; Stenius, Ulla [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden); Jönsson, Bo A.G. [Division of Occupational and Environmental Medicine, Lund University, SE 221 85 Lund (Sweden); Högberg, Johan, E-mail: johan.hogberg@ki.se [Institute of Environmental Medicine, Karolinska Institutet, Box 210, SE 171 77 Stockholm (Sweden)

    2015-09-15

    Diisocyanates are industrial chemicals which have a wide range of applications in developed and developing countries. They are notorious lung toxicants and respiratory sensitizers. However, the mechanisms behind their adverse effects are not adequately characterized. Autotaxin (ATX) is an enzyme producing lysophosphatidic acid (LPA), and the ATX-LPA axis has been implicated in lung related inflammatory conditions and diseases, including allergic asthma, but not to toxicity of environmental low-molecular-weight chemicals. We investigated effects of toluene diisocyanate (TDI) on ATX induction in human lung epithelial cell models, and we correlated LPA-levels in plasma to biomarkers of TDI exposure in urine collected from workers exposed to < 5 ppb (parts per billion). Information on workers' symptoms was collected through interviews. One nanomolar TDI robustly induced ATX release within 10 min in vitro. A P2X7- and P2X4-dependent microvesicle formation was implicated in a rapid ATX release and a subsequent protein synthesis. Co-localization between purinergic receptors and ATX was documented by immunofluorescence and confocal microscopy. The release was modulated by monocyte chemoattractant protein-1 (MCP-1) and by extracellular ATP. In workers, we found a dose–response relationship between TDI exposure biomarkers in urine and LPA levels in plasma. Among symptomatic workers reporting “sneezing”, the LPA levels were higher than among non-symptomatic workers. This is the first report indicating induction of the ATX-LPA axis by an environmental low-molecular-weight chemical, and our data suggest a role for the ATX-LPA axis in TDI toxicity. - Highlights: • Human epithelial cells release autotaxin in response to 1 nM toluene diisocyanate (TDI). • The release involves P2X4 and P2X7 receptors and is modulated by ATP and MCP-1. • Lysophosphatidic acid (LPA) was measured in workers exposed to < 5 ppb TDI. • LPA in plasma correlated to TDI exposure

  2. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    International Nuclear Information System (INIS)

    Highlights: → Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. → Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. → VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. → LPA-LPAR1/3 signaling regulated TGFβ1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. → LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGFβ1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor β1 (TGFβ1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1/3 signaling system is involved in the

  3. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Gan, Lu [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Xue, Jian-Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Laboratory of Stem Cell Biology, West China Hospital, Sichuan University, Chengdu (China); Li, Xin [Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Liu, De-Song [Department of Pediatrics, Sichuan Provincial Hospital of Women and Children, Chengdu (China); Ge, Yan; Ni, Pei-Yan; Deng, Lin [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Lu, You, E-mail: radyoulu@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Department of Thoracic Oncology, Cancer Center, West China Hospital, Sichuan University, Chengdu (China); Jiang, Wei, E-mail: wcumsjw72@hotmail.com [State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu (China); Molecular Medicine Research Center, West China Hospital, Sichuan University, Chengdu (China)

    2011-05-27

    Highlights: {yields} Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. {yields} Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. {yields} VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. {yields} LPA-LPAR1/3 signaling regulated TGF{beta}1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. {yields} LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGF{beta}1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor {beta}1 (TGF{beta}1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1

  4. The autotaxin-lysophosphatidic acid-lysophosphatidic acid receptor cascade: proposal of a novel potential therapeutic target for treating glioblastoma multiforme.

    Science.gov (United States)

    Tabuchi, Sadaharu

    2015-01-01

    Glioblastoma multiforme (GBM) is the most malignant tumor of the central nervous system (CNS). Its prognosis is one of the worst among all cancer types, and it is considered a fatal malignancy, incurable with conventional therapeutic strategies. As the bioactive multifunctional lipid mediator lysophosphatidic acid (LPA) is well recognized to be involved in the tumorigenesis of cancers by acting on G-protein-coupled receptors, LPA receptor (LPAR) antagonists and LPA synthesis inhibitors have been proposed as promising drugs for cancer treatment. Six LPARs, named LPA1-6, are currently recognized. Among them, LPA1 is the dominant LPAR in the CNS and is highly expressed in GBM in combination with the overexpression of autotaxin (ATX), the enzyme (a phosphodiesterase, which is a potent cell motility-stimulating factor) that produces LPA.Invasion is a defining hallmark of GBM. LPA is significantly related to cell adhesion, cell motility, and invasion through the Rho family GTPases Rho and Rac. LPA1 is responsible for LPA-driven cell motility, which is attenuated by LPA4. GBM is among the most vascular human tumors. Although anti-angiogenic therapy (through the inhibition of vascular endothelial growth factor (VEGF)) was established, sufficient results have not been obtained because of the increased invasiveness triggered by anti-angiogenesis. As both ATX and LPA play a significant role in angiogenesis, similar to VEGF, inhibition of the ATX/LPA axis may be beneficial as a two-pronged therapy that includes anti-angiogenic and anti-invasion therapy. Conventional approaches to GBM are predominantly directed at cell proliferation. Recurrent tumors regrow from cells that have invaded brain tissues and are less proliferative, and are thus quite resistant to conventional drugs and radiation, which preferentially kill rapidly proliferating cells. A novel approach that targets this invasive subpopulation of GBM cells may improve the prognosis of GBM. Patients with GBM that

  5. Lysophosphatidic Acid Stimulates MCP-1 Secretion from C2C12 Myoblast.

    Science.gov (United States)

    Tsukahara, Tamotsu; Haniu, Hisao

    2012-01-01

    Chemokines are regulatory proteins that play an important role in muscle cell migration and proliferation. In this study, C2C12 cells treated with lysophosphatidic acid (LPA) showed an increase in endogenous monocyte chemotactic protein-1 (MCP-1) expression and secretion. LPA is a naturally occurring bioactive lysophospholipid with hormone- and growth-factor-like activities. LPA is produced by activated platelets, cytokine-stimulated leukocytes, and possibly by other cell types. However, the LPA analog cyclic phosphatidic acid (cPA) had no effect on the expression and secretion of MCP-1. LPA, although similar in structure to cPA, had potent inducing effects on MCP-1 expression in C2C12 cells. In this study, we showed that LPA enhanced MCP-1 mRNA expression and protein secretion in a dose-dependent manner. Taken together, these results suggest that LPA enhances MCP-1 secretion in C2C12 cells and thus may play an important role in cell proliferation. PMID:24049655

  6. Lysophosphatidic acid-functionalised titanium as a superior surface for supporting human osteoblast (MG63 maturation

    Directory of Open Access Journals (Sweden)

    JP Mansell

    2012-05-01

    Full Text Available Covalent modifications of titanium with small molecules known to promote human osteoblast maturation are especially attractive in developing superior biomaterials. An important step in securing competent bone formation at implant sites is promoting the formation of mature osteoblasts, either from committed pre-osteoblasts or from their mesenchymal progenitors. To this end our research has focussed on identifying molecules that enhance human osteoblast formation and maturation and to develop ways of covalently attaching these molecules to implant surfaces so that they are more likely to withstand the rigors of the implantation process whilst still retaining their bioactivity. Herein we report the novel production of lipid-functionalised titanium using lysophosphatidic acid or a related compound, (3S 1-fluoro-3-hydroxy-4-butyl-1-phosphonate. Both lipids were especially effective at co-operating with calcitriol to promote human osteoblast maturation at these modified Ti surfaces in vitro. The novel findings presented offer enticing new developments towards the fabrication of next-generation implant devices with the potential to significantly enhance the osseointegration process and with it improvements in future prosthesis performance and longevity.

  7. Elevated Serum Levels of Arachidonoyl-lysophosphatidic Acid and Sphingosine 1-Phosphate in Systemic Sclerosis

    Directory of Open Access Journals (Sweden)

    Akira Tokumura, Laura D. Carbone, Yasuko Yoshioka, Junichi Morishige, Masaki Kikuchi, Arnold Postlethwaite, Mitchell A. Watsky

    2009-01-01

    Full Text Available Systemic sclerosis (SSc is an often fatal disease characterized by autoimmunity and inflammation, leading to widespread vasculopathy and fibrosis. Lysophosphatidic acid (LPA, a bioactive phospholipid in serum, is generated from lysophospholipids secreted from activated platelets in part by the action of lysophospholipase D (lysoPLD. Sphingosine 1-phosphate (S1P, a member of the bioactive lysophospholipid family, is also released from activated platelets. Because activated platelets are a hallmark of SSc, we wanted to determine whether subjects with SSc have altered serum lysophospholipid levels or lysoPLD activity. Lysophospholipid levels were measured using mass spectrometric analysis. LysoPLD activity was determined by quantifying choline released from exogenous lysophosphatidylcholine (LPC. The major results were that serum levels of arachidonoyl (20:4-LPA and S1P were significantly higher in SSc subjects versus controls. Furthermore, serum LPA:LPC ratios of two different polyunsaturated phospholipid molecular species, and also the ratio of all species combined, were significantly higher in SSc subjects versus controls. No significant differences were found between other lysophospholipid levels or lysoPLD activities. Elevated 20:4 LPA, S1P levels and polyunsaturated LPA:LPC ratios may be markers for and/or play a significant role in the etiology of SSc and may be future pharmacological targets for SSc treatment.

  8. Lysophosphatidic acid mediates myeloid differentiation within the human bone marrow microenvironment.

    Directory of Open Access Journals (Sweden)

    Denis Evseenko

    Full Text Available Lysophosphatidic acid (LPA is a pleiotropic phospholipid present in the blood and certain tissues at high concentrations; its diverse effects are mediated through differential, tissue specific expression of LPA receptors. Our goal was to determine if LPA exerts lineage-specific effects during normal human hematopoiesis. In vitro stimulation of CD34+ human hematopoietic progenitors by LPA induced myeloid differentiation but had no effect on lymphoid differentiation. LPA receptors were expressed at significantly higher levels on Common Myeloid Progenitors (CMP than either multipotent Hematopoietic Stem/Progenitor Cells (HSPC or Common Lymphoid Progenitors (CLP suggesting that LPA acts on committed myeloid progenitors. Functional studies demonstrated that LPA enhanced migration, induced cell proliferation and reduced apoptosis of isolated CMP, but had no effect on either HSPC or CLP. Analysis of adult and fetal human bone marrow sections showed that PPAP2A, (the enzyme which degrades LPA was highly expressed in the osteoblastic niche but not in the perivascular regions, whereas Autotaxin (the enzyme that synthesizes LPA was expressed in perivascular regions of the marrow. We propose that a gradient of LPA with the highest levels in peri-sinusoidal regions and lowest near the endosteal zone, regulates the localization, proliferation and differentiation of myeloid progenitors within the bone marrow marrow.

  9. Lysophosphatidic acid activates Arf6 to promote the mesenchymal malignancy of renal cancer.

    Science.gov (United States)

    Hashimoto, Shigeru; Mikami, Shuji; Sugino, Hirokazu; Yoshikawa, Ayumu; Hashimoto, Ari; Onodera, Yasuhito; Furukawa, Shotaro; Handa, Haruka; Oikawa, Tsukasa; Okada, Yasunori; Oya, Mototsugu; Sabe, Hisataka

    2016-01-01

    Acquisition of mesenchymal properties by cancer cells is critical for their malignant behaviour, but regulators of the mesenchymal molecular machinery and how it is activated remain elusive. Here we show that clear cell renal cell carcinomas (ccRCCs) frequently utilize the Arf6-based mesenchymal pathway to promote invasion and metastasis, similar to breast cancers. In breast cancer cells, ligand-activated receptor tyrosine kinases employ GEP100 to activate Arf6, which then recruits AMAP1; and AMAP1 then binds to the mesenchymal-specific protein EPB41L5, which promotes epithelial-mesenchymal transition and focal adhesion dynamics. In renal cancer cells, lysophosphatidic acid (LPA) activates Arf6 via its G-protein-coupled receptors, in which GTP-Gα12 binds to EFA6. The Arf6-based pathway may also contribute to drug resistance. Our results identify a specific mesenchymal molecular machinery of primary ccRCCs, which is triggered by a product of autotaxin and it is associated with poor outcome of patients. PMID:26854204

  10. Studies on lysophosphatidic acid action during in vitro preimplantation embryo development.

    Science.gov (United States)

    Boruszewska, D; Sinderewicz, E; Kowalczyk-Zieba, I; Grycmacher, K; Woclawek-Potocka, I

    2016-01-01

    Assisted reproductive technologies, including in vitro embryo production (IVP), have been successfully used in animal reproduction to optimize breeding strategies for improved production and health in animal husbandry. Despite the progress in IVP techniques over the years, further improvements in in vitro embryo culture systems are required for the enhancement of oocyte and embryo developmental competence. One of the most important issues associated with IVP procedures is the optimization of the in vitro culture of oocytes and embryos. Studies in different species of animals and in humans have identified important roles for receptor-mediated lysophosphatidic acid (LPA) signaling in multiple aspects of human and animal reproductive tract function. The data on LPA signaling in the ovary and uterus suggest that LPA can directly contribute to embryo-maternal interactions via its influence on early embryo development beginning from the influence of the ovarian environment on the oocyte to the influence of the uterine environment on the preimplantation embryo. This review discusses the current status of LPA as a potential supplement in oocyte maturation, fertilization, and embryo culture media and current views on the potential involvement of the LPA signaling pathway in early embryo development.

  11. The effects of aspirin on platelet function and lysophosphatidic acids depend on plasma concentrations of EPA and DHA

    OpenAIRE

    Block, Robert C.; Abdolahi, Amir; Tu, Xin; Georas, Steve N.; Brenna, J. Thomas; Phipps, Richard P.; Lawrence, Peter; Mousa, Shaker A.

    2014-01-01

    Aspirin’s prevention of cardiovascular disease (CVD) events in individuals with type 2 diabetes mellitus is controversial. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and aspirin all affect the cyclooxygenase enzyme. The relationship between plasma EPA and DHA and aspirin’s effects has not been determined. Thirty adults with type 2 diabetes mellitus ingested aspirin (81 mg/day) for 7 days, then EPA+DHA (2.6 g/day) for 28 days, then both for another 7 days. Lysophosphatidic acid...

  12. Yangxueqingnao particles inhibit rat vascular smooth muscle cell proliferation induced by lysophosphatidic acid

    Institute of Scientific and Technical Information of China (English)

    CAI Wei; XU Yi; CHEN Jun-zhu; HUANG Shu-ru; LU Zhen-ya; WANG Zhan-kun

    2005-01-01

    Objective: To observe the effect of Yangxueqingnao particles on rat vascular smooth muscle cell (VSMC) proliferation induced by lysophosphatidic acid (LPA). Methods: The amount of3H-TdR (3H-thymidine) admixed in cultured rat VSMC was measured and mitogen-activated protein kinase (MAPK) activity and lipid peroxidation end product malondialdehyde (MDA)content of the VSMC were assayed. Results: 1×10-9, 1×10-8, 1×10-7 mol/L LPA in a concentration dependent manner, induced the amount of 3H-TdR admixed, MAP kinase activity, and MDA content of the cultured rat VSMC to increase. However, 5%, 10%,and 15% Yangxueqingnao serum preincubation resulted in a decrease of 23.0%, 42.0%, and 52.0% (P<0.01) respectively in the amount of 3H-TdR admixed, a decline in VSMC MAP kinase activity of 13.9% (P<0.05), 29.6% (P<0.01), and 48.9% (P<0.01)respectively, and also, a decrease in MDA content of VSMC of 19.4%, 24.7%, and 43.2% (P<0.01) respectively, in the 1×10-7mol/L LPA-treated VSMC. Conclusions: LPA activates the proliferation and lipid peroxidation of VSMC in a concentration dependent manner. The LPA-induced VSMC proliferation is related to the activity of MAP kinases, enzymes involved in an intracellular signalling pathway. The results of the present study showed that Yangxueqingnao particles can effectively inhibit LPA-induced VSMC proliferation, MAP kinase activation, and reduce lipid peroxidative lesion.

  13. Lysophosphatidic acid (LPA 18:1 transcriptional regulation of primary human gingival fibroblasts

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    D. Roselyn Cerutis

    2014-12-01

    Full Text Available The pleiotropic, bioactive lipid lysophosphatidic acid [(LPA, 1-acyl-sn-glycerol-3-phosphate] exerts critical regulatory actions in physiology and pathophysiology in many systems. It is present in normal bodily fluids, and is elevated in pathology (1. In vivo, “LPA” exists as distinct molecular species, each having a single fatty acid of varying chain length and degree of unsaturation covalently attached to the glycerol backbone via an acyl, alkyl, or alkenyl link. These species differ in affinities for the individual LPA receptors [(LPARs, LPA1-6] and coupling to G proteins (2. However, LPA 18:1 has been and continues to be the most commonly utilized species in reported studies. The actions of “LPA” remain poorly defined in oral biology and pathophysiology. Our laboratory has addressed this knowledge gap by studying in vitro the actions of the major human salivary LPA species [18:1, 18:0, and 16:0 (3] in human oral cells (4–7. This includes gingival fibroblasts (GF, which our flow cytometry data from multiple donors found that they express LPA1-5 (6. We have also reported that these species are ten-fold elevated to pharmacologic levels in the saliva and gingival crevicular fluid obtained from patients with moderate–severe periodontitis (8. As the potential of LPA to regulate transcriptional activity had not been examined in the oral system, this study used whole human genome microarray analysis to test the hypothesis that LPA 18:1-treated human GF would show significant changes in gene transcripts relevant to their biology, wound-healing, and inflammatory responses. LPA 18:1 was found to significantly regulate a large, complex set of genes critical to GF biology in these categories and to periodontal disease. The raw data has been deposited at NCBI's GEO database as record GSE57496.

  14. Tubular cell phenotype in HIV-associated nephropathy: role of phospholipid lysophosphatidic acid.

    Science.gov (United States)

    Ayasolla, Kamesh R; Rai, Partab; Rahimipour, Shai; Hussain, Mohammad; Malhotra, Ashwani; Singhal, Pravin C

    2015-08-01

    Collapsing glomerulopathy and microcysts are characteristic histological features of HIV-associated nephropathy (HIVAN). We have previously reported the role of epithelial mesenchymal transition (EMT) in the development of glomerular and tubular cell phenotypes in HIVAN. Since persistent tubular cell activation of NFκB has been reported in HIVAN, we now hypothesize that HIV may be contributing to tubular cell phenotype via lysophosphatidic acid (LPA) mediated downstream signaling. Interestingly, LPA and its receptors have also been implicated in the tubular interstitial cell fibrosis (TIF) and cyst formation in autosomal dominant polycystic kidney disease (PKD). Primary human proximal tubular cells (HRPTCs) were transduced with either empty vector (EV/HRPTCs), HIV (HIV/HRPTCs) or treated with LPA (LPA/HRPTC). Immunoelectrophoresis of HIV/HRPTCs and LPA/HRPTCs displayed enhanced expression of pro-fibrotic markers: a) fibronectin (2.25 fold), b) connective tissue growth factor (CTGF; 4.8 fold), c) α-smooth muscle actin (α-SMA; 12 fold), and d) collagen I (5.7 fold). HIV enhanced tubular cell phosphorylation of ILK-1, FAK, PI3K, Akt, ERKs and P38 MAPK. HIV increased tubular cell transcriptional binding activity of NF-κB; whereas, a LPA biosynthesis inhibitor (AACOCF3), a DAG kinase inhibitor, a LPA receptor blocker (Ki16425), a NF-κB inhibitor (PDTC) and NFκB-siRNA not only displayed downregulation of a NFκB activity but also showed attenuated expression of profibrotic/EMT genes in HIV milieu. These findings suggest that LPA could be contributing to HIV-induced tubular cell phenotype via NFκB activation in HIVAN. PMID:26079546

  15. Ginseng pharmacology: a new paradigm based on gintonin-lysophosphatidic acid receptor interactions

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    Seung-Yeol eNah

    2015-10-01

    Full Text Available Ginseng, the root of Panax ginseng, is used as a traditional medicine. Despite the long history of the use of ginseng, there is no specific scientific or clinical rationale for ginseng pharmacology besides its application as a general tonic. The ambiguous description of ginseng pharmacology might be due to the absence of a predominant active ingredient that represents ginseng pharmacology. Recent studies show that ginseng abundantly contains lysophosphatidic acids (LPAs, which are phospholipid-derived growth factor with diverse biological functions including those claimed to be exhibited by ginseng. LPAs in ginseng form a complex with ginseng proteins, which can bind and deliver LPA to its cognate receptors with a high affinity. As a first messenger, gintonin produces second messenger Ca2+ via G protein-coupled LPA receptors. Ca2+ is an intracellular mediator of gintonin and initiates a cascade of amplifications for further intercellular communications by activation of Ca2+-dependent kinases, receptors, gliotransmitter and neurotransmitter release. Ginsenosides, which have been regarded as primary ingredients of ginseng, cannot elicit intracellular [Ca2+]i transients, since they lack specific cell surface receptor. However, ginsenosides exhibit non-specific ion channel and receptor regulations. This is the key characteristic that distinguishes gintonin from ginsenosides. Although the current discourse on ginseng pharmacology is focused on ginsenosides, gintonin can definitely provide a mode of action for ginseng pharmacology that ginsenosides cannot. This review article introduces a novel concept of ginseng ligand-LPA receptor interaction and proposes to establish a paradigm that shifts the focus from ginsenosides to gintonin as a major ingredient representing ginseng pharmacology.

  16. Lysophosphatidic acid acyltransferase β (LPAATβ promotes the tumor growth of human osteosarcoma.

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    Farbod Rastegar

    Full Text Available BACKGROUND: Osteosarcoma is the most common primary malignancy of bone with poorly characterized molecular pathways important in its pathogenesis. Increasing evidence indicates that elevated lipid biosynthesis is a characteristic feature of cancer. We sought to investigate the role of lysophosphatidic acid acyltransferase β (LPAATβ, aka, AGPAT2 in regulating the proliferation and growth of human osteosarcoma cells. LPAATβ can generate phosphatidic acid, which plays a key role in lipid biosynthesis as well as in cell proliferation and survival. Although elevated expression of LPAATβ has been reported in several types of human tumors, the role of LPAATβ in osteosarcoma progression has yet to be elucidated. METHODOLOGY/PRINCIPAL FINDINGS: Endogenous expression of LPAATβ in osteosarcoma cell lines is analyzed by using semi-quantitative PCR and immunohistochemical staining. Adenovirus-mediated overexpression of LPAATβ and silencing LPAATβ expression is employed to determine the effect of LPAATβ on osteosarcoma cell proliferation and migration in vitro and osteosarcoma tumor growth in vivo. We have found that expression of LPAATβ is readily detected in 8 of the 10 analyzed human osteosarcoma lines. Exogenous expression of LPAATβ promotes osteosarcoma cell proliferation and migration, while silencing LPAATβ expression inhibits these cellular characteristics. We further demonstrate that exogenous expression of LPAATβ effectively promotes tumor growth, while knockdown of LPAATβ expression inhibits tumor growth in an orthotopic xenograft model of human osteosarcoma. CONCLUSIONS/SIGNIFICANCE: Our results strongly suggest that LPAATβ expression may be associated with the aggressive phenotypes of human osteosarcoma and that LPAATβ may play an important role in regulating osteosarcoma cell proliferation and tumor growth. Thus, targeting LPAATβ may be exploited as a novel therapeutic strategy for the clinical management of osteosarcoma. This

  17. Lysophosphatidic Acid Up-Regulates Hexokinase II and Glycolysis to Promote Proliferation of Ovarian Cancer Cells

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    Abir Mukherjee

    2015-09-01

    Full Text Available Lysophosphatidic acid (LPA, a blood-borne lipid mediator, is present in elevated concentrations in ascites of ovarian cancer patients and other malignant effusions. LPA is a potent mitogen in cancer cells. The mechanism linking LPA signal to cancer cell proliferation is not well understood. Little is known about whether LPA affects glucose metabolism to accommodate rapid proliferation of cancer cells. Here we describe that in ovarian cancer cells, LPA enhances glycolytic rate and lactate efflux. A real time PCR-based miniarray showed that hexokinase II (HK2 was the most dramatically induced glycolytic gene to promote glycolysis in LPA-treated cells. Analysis of the human HK2 gene promoter identified the sterol regulatory element-binding protein as the primary mediator of LPA-induced HK2 transcription. The effects of LPA on HK2 and glycolysis rely on LPA2, an LPA receptor subtype overexpressed in ovarian cancer and many other malignancies. We further examined the general role of growth factor-induced glycolysis in cell proliferation. Like LPA, epidermal growth factor (EGF elicited robust glycolytic and proliferative responses in ovarian cancer cells. Insulin-like growth factor 1 (IGF-1 and insulin, however, potently stimulated cell proliferation but only modestly induced glycolysis. Consistent with their differential effects on glycolysis, LPA and EGF-dependent cell proliferation was highly sensitive to glycolytic inhibition while the growth-promoting effect of IGF-1 or insulin was more resistant. These results indicate that LPA- and EGF-induced cell proliferation selectively involves up-regulation of HK2 and glycolytic metabolism. The work is the first to implicate LPA signaling in promotion of glucose metabolism in cancer cells.

  18. Expression of lysophosphatidic acid and its receptor in human pancreatic cancer and its clinical evaluation of diagnosis and therapy

    Institute of Scientific and Technical Information of China (English)

    WANG Shao-kai; TAO Chen-jie; WANG Wei-dong; L(U)Guang-mei; GONG Yong-ling

    2011-01-01

    Lysophosphatidic acid(LPA) is a naturally occurring phospholipid with diverse effects in various cells, ranging from immediate morphological alteration to long lasting cellular function changes, such as induction of stimulation of cell proliferation, survival, drug resistance and motility. Like many other biomediators, LPA interacts with cells through specific cell surface receptors(G protein-coupled receptors). LPA1/Edg-2,LPA2/Edg-4 and LPA3/Edg-7, named as Edg/LP subfamily, are the three most common lysophosphatidic acid receptors. LPA plays a critical role as a general growth, survival and pro-angiogenic factor in the regulation of pathophysiological processes in vivo and in vitro. Recent literatures suggest that abnormalities in LPA metabolism and function in pancreatic cancer patients may contribute to the initiation and progression of the disease. Thus, LPA might be a potential target for clinical pancreatic cancer diagnosis and therapy. Herein we review the expression of LPA and its receptors in the carcinogenesis of human malignancies, with focus on human pancreatic cancer, and also clinical diagnosis and treatment has been evaluated.

  19. Clinical significance of plasma lysophosphatidic acid levels in the differential diagnosis of ovarian cancer

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    Yun-Jie Zhang

    2015-01-01

    Full Text Available Objective: To investigate the value of lysophosphatidic acid (LPA in the diagnosis of ovarian cancer. Materials and Methods: We first performed a hospital-based, case-control study involving 123 ovarian cancer patients and 101 benign ovarian tumor patients, and then conducted a meta-analysis with 19 case-control studies to assess the correlation between ovarian cancer and plasma LPA levels. Results: The case-control study results demonstrated that ovarian cancer patients have increased LPA and cancer antigen (CA-125 levels compared to patients with benign ovarian tumor (LPA: Ovarian cancer vs benign ovarian tumor: 5.28 ± 1.52 vs 1.82 ± 0.77 μmol/L; CA-125: Ovarian cancer vs benign ovarian tumor: 87.17 ± 45.81 vs. 14.03 ± 10.14 U/mL, which showed statistically significant differences (both P < 0.05. LPA with advanced sensitivity, specificity, positive predictive value, negative predictive value, and accuracy rate of diagnosis excelled CA-125 in the diagnosis of ovarian cancer (both P < 0.05. The areas under the receiver operating characteristic (ROC curve in the diagnosis of ovarian cancer (LPA: 0.983; CA-125: 0.910 were statistically significant compared with the reference (both P < 0.001 and the difference of the areas of ROC curve between LPA and CA-125 in the diagnosis of ovarian cancer showed statistically significant difference (P < 0.05. The meta-analysis results suggested that plasma LPA levels were higher in ovarian cancer tissues than in benign tissues (standardized mean difference (SMD =2.36, 95% confidence interval (CI: 1.61-3.11, P < 0.001 and normal tissues (SMD = 2.32, 95% CI: 1.77-2.87, P < 0.001. Conclusion: LPA shows greater value in the diagnosis of ovarian cancer compared to CA-125 and may be employed as a biological index to diagnose ovarian cancer.

  20. Golgi membrane fission requires the CtBP1-S/BARS-induced activation of lysophosphatidic acid acyltransferase δ.

    Science.gov (United States)

    Pagliuso, Alessandro; Valente, Carmen; Giordano, Lucia Laura; Filograna, Angela; Li, Guiling; Circolo, Diego; Turacchio, Gabriele; Marzullo, Vincenzo Manuel; Mandrich, Luigi; Zhukovsky, Mikhail A; Formiggini, Fabio; Polishchuk, Roman S; Corda, Daniela; Luini, Alberto

    2016-01-01

    Membrane fission is an essential cellular process by which continuous membranes split into separate parts. We have previously identified CtBP1-S/BARS (BARS) as a key component of a protein complex that is required for fission of several endomembranes, including basolateral post-Golgi transport carriers. Assembly of this complex occurs at the Golgi apparatus, where BARS binds to the phosphoinositide kinase PI4KIIIβ through a 14-3-3γ dimer, as well as to ARF and the PKD and PAK kinases. We now report that, when incorporated into this complex, BARS binds to and activates a trans-Golgi lysophosphatidic acid (LPA) acyltransferase type δ (LPAATδ) that converts LPA into phosphatidic acid (PA); and that this reaction is essential for fission of the carriers. LPA and PA have unique biophysical properties, and their interconversion might facilitate the fission process either directly or indirectly (via recruitment of proteins that bind to PA, including BARS itself). PMID:27401954

  1. Golgi membrane fission requires the CtBP1-S/BARS-induced activation of lysophosphatidic acid acyltransferase δ.

    Science.gov (United States)

    Pagliuso, Alessandro; Valente, Carmen; Giordano, Lucia Laura; Filograna, Angela; Li, Guiling; Circolo, Diego; Turacchio, Gabriele; Marzullo, Vincenzo Manuel; Mandrich, Luigi; Zhukovsky, Mikhail A; Formiggini, Fabio; Polishchuk, Roman S; Corda, Daniela; Luini, Alberto

    2016-07-12

    Membrane fission is an essential cellular process by which continuous membranes split into separate parts. We have previously identified CtBP1-S/BARS (BARS) as a key component of a protein complex that is required for fission of several endomembranes, including basolateral post-Golgi transport carriers. Assembly of this complex occurs at the Golgi apparatus, where BARS binds to the phosphoinositide kinase PI4KIIIβ through a 14-3-3γ dimer, as well as to ARF and the PKD and PAK kinases. We now report that, when incorporated into this complex, BARS binds to and activates a trans-Golgi lysophosphatidic acid (LPA) acyltransferase type δ (LPAATδ) that converts LPA into phosphatidic acid (PA); and that this reaction is essential for fission of the carriers. LPA and PA have unique biophysical properties, and their interconversion might facilitate the fission process either directly or indirectly (via recruitment of proteins that bind to PA, including BARS itself).

  2. Autotaxin activity increases locally following lung injury, but is not required for pulmonary lysophosphatidic acid production or fibrosis.

    Science.gov (United States)

    Black, Katharine E; Berdyshev, Evgeny; Bain, Gretchen; Castelino, Flavia V; Shea, Barry S; Probst, Clemens K; Fontaine, Benjamin A; Bronova, Irina; Goulet, Lance; Lagares, David; Ahluwalia, Neil; Knipe, Rachel S; Natarajan, Viswanathan; Tager, Andrew M

    2016-06-01

    Lysophosphatidic acid (LPA) is an important mediator of pulmonary fibrosis. In blood and multiple tumor types, autotaxin produces LPA from lysophosphatidylcholine (LPC) via lysophospholipase D activity, but alternative enzymatic pathways also exist for LPA production. We examined the role of autotaxin (ATX) in pulmonary LPA production during fibrogenesis in a bleomycin mouse model. We found that bleomycin injury increases the bronchoalveolar lavage (BAL) fluid levels of ATX protein 17-fold. However, the LPA and LPC species that increase in BAL of bleomycin-injured mice were discordant, inconsistent with a substrate-product relationship between LPC and LPA in pulmonary fibrosis. LPA species with longer chain polyunsaturated acyl groups predominated in BAL fluid after bleomycin injury, with 22:5 and 22:6 species accounting for 55 and 16% of the total, whereas the predominant BAL LPC species contained shorter chain, saturated acyl groups, with 16:0 and 18:0 species accounting for 56 and 14% of the total. Further, administration of the potent ATX inhibitor PAT-048 to bleomycin-challenged mice markedly decreased ATX activity systemically and in the lung, without effect on pulmonary LPA or fibrosis. Therefore, alternative ATX-independent pathways are likely responsible for local generation of LPA in the injured lung. These pathways will require identification to therapeutically target LPA production in pulmonary fibrosis.-Black, K. E., Berdyshev, E., Bain, G., Castelino, F. V., Shea, B. S., Probst, C. K., Fontaine, B. A., Bronova, I., Goulet, L., Lagares, D., Ahluwalia, N., Knipe, R. S., Natarajan, V., Tager, A. M. Autotaxin activity increases locally following lung injury, but is not required for pulmonary lysophosphatidic acid production or fibrosis.

  3. Lysophosphatidic acid is a major serum noncytokine survival factor for murine macrophages which acts via the phosphatidylinositol 3-kinase signaling pathway.

    OpenAIRE

    Koh, J. S.; Lieberthal, W; Heydrick, S; Levine, J. S.

    1998-01-01

    Lysophosphatidic acid (LPA) is the smallest and structurally simplest of all the glycerophospholipids. It occurs normally in serum and binds with high affinity to albumin, while retaining its biological activity. The effects of LPA are pleiotropic and range from mitogenesis to stress fiber formation. We show a novel role for LPA: as a macrophage survival factor with potency equivalent to serum. Administration of LPA protects macrophages from apoptosis induced by serum deprivation, and protect...

  4. Regulation of gene expression and subcellular protein distribution in MLO-Y4 osteocytic cells by lysophosphatidic acid: Relevance to dendrite outgrowth

    OpenAIRE

    Waters, Katrina M.; Jon M Jacobs; Gritsenko, Marina A.; Karin, Norman J.

    2011-01-01

    Osteoblastic and osteocytic cells are highly responsive to the lipid growth factor lysophosphatidic acid (LPA) but the mechanisms by which LPA alters bone cell functions are largely unknown. A major effect of LPA on osteocytic cells is the stimulation of dendrite membrane outgrowth, a process that we predicted to require changes in gene expression and protein distribution. We employed DNA microarrays for global transcriptional profiling of MLO-Y4 osteocytic cells grown for 6 and 24 hours in t...

  5. Lipid phosphate phosphatases regulate lysophosphatidic acid production and signaling in platelets: studies using chemical inhibitors of lipid phosphate phosphatase activity.

    Science.gov (United States)

    Smyth, Susan S; Sciorra, Vicki A; Sigal, Yury J; Pamuklar, Zehra; Wang, Zuncai; Xu, Yong; Prestwich, Glenn D; Morris, Andrew J

    2003-10-31

    Blood platelets play an essential role in ischemic heart disease and stroke contributing to acute thrombotic events by release of potent inflammatory agents within the vasculature. Lysophosphatidic acid (LPA) is a bioactive lipid mediator produced by platelets and found in the blood and atherosclerotic plaques. LPA receptors on platelets, leukocytes, endothelial cells, and smooth muscle cells regulate growth, differentiation, survival, motility, and contractile activity. Definition of the opposing pathways of synthesis and degradation that control extracellular LPA levels is critical to understanding how LPA bioactivity is regulated. We show that intact platelets and platelet membranes actively dephosphorylate LPA and identify the major enzyme responsible as lipid phosphate phosphatase 1 (LPP1). Localization of LPP1 to the platelet surface is increased by exposure to LPA. A novel receptor-inactive sn-3-substituted difluoromethylenephosphonate analog of phosphatidic acid that is a potent competitive inhibitor of LPP1 activity potentiates platelet aggregation and shape change responses to LPA and amplifies LPA production by agonist-stimulated platelets. Our results identify LPP1 as a pivotal regulator of LPA signaling in the cardiovascular system. These findings are consistent with genetic and cell biological evidence implicating LPPs as negative regulators of lysophospholipid signaling and suggest that the mechanisms involve both attenuation of lysophospholipid actions at cell surface receptors and opposition of lysophospholipid production. PMID:12909631

  6. Expression and function of lysophosphatidic acid receptors (LPARs) 1 and 3 in human hepatic cancer progenitor cells.

    Science.gov (United States)

    Zuckerman, Valentina; Sokolov, Eugene; Swet, Jacob H; Ahrens, William A; Showlater, Victor; Iannitti, David A; Mckillop, Iain H

    2016-01-19

    Hepatocellular carcinoma (HCC) is the most common primary cancer of the liver and is characterized by rapid tumor expansion and metastasis. Lysophosphatidic acid (LPA) signaling, via LPA receptors 1-6 (LPARs1-6), regulates diverse cell functions including motility, migration, and proliferation, yet the role of LPARs in hepatic tumor pathology is poorly understood. We sought to determine the expression and function of endothelial differentiation gene (EDG) LPARs (LPAR1-3) in human HCC and complimentary in vitro models. Human HCC were characterized by significantly elevated LPAR1/LPAR3 expression in the microenvironment between the tumor and non-tumor liver (NTL), a finding mirrored in human SKHep1 cells. Analysis of human tissue and human hepatic tumor cells in vitro revealed cells that express LPAR3 (HCC-NTL margin in vivo and SKHep1 in vitro) also express cancer stem cell markers in the absence of hepatocyte markers. Treatment of SKHep1 cells with exogenous LPA led to significantly increased cell motility but not proliferation. Using pharmacological agents and cells transfected to knock-down LPAR1 or LPAR3 demonstrated LPA-dependent cell migration occurs via an LPAR3-Gi-ERK-pathway independent of LPAR1. These data suggest cells that stain positive for both LPAR3 and cancer stem cell markers are distinct from the tumor mass per se, and may mediate tumor invasiveness/expansion via LPA-LPAR3 signaling. PMID:26701886

  7. Transgenic Expression of Human Lysophosphatidic Acid Receptor LPA2 in Mouse Intestinal Epithelial Cells Induces Intestinal Dysplasia.

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    Michihiro Yoshida

    Full Text Available Lysophosphatidic acid (LPA acts on LPA2 receptor to mediate multiple pathological effects that are associated with tumorigenesis. The absence of LPA2 attenuates tumor progression in rodent models of colorectal cancer, but whether overexpression of LPA2 alone can lead to malignant transformation in the intestinal tract has not been studied. In this study, we expressed human LPA2 in intestinal epithelial cells (IECs under control of the villin promoter. Less than 4% of F1-generation mice had germline transmission of transgenic (TG human LPA2; as such only 3 F1 mice out of 72 genotyped had TG expression. These TG mice appeared anemic with hematochezia and died shortly after birth. TG mice were smaller in size compared with the wild type mouse of the same age and sex. Morphological analysis showed that TG LPA2 colon had hyper-proliferation of IECs resulting in increased colonic crypt depth. Surprisingly, TG small intestine had villus blunting and decreased IEC proliferation and dysplasia. In both intestine and colon, TG expression of LPA2 compromised the terminal epithelial differentiation, consistent with epithelial dysplasia. Furthermore, we showed that epithelial dysplasia was observed in founder mouse intestine, correlating LPA2 overexpression with epithelial dysplasia. The current study demonstrates that overexpression of LPA2 alone can lead to intestinal dysplasia.

  8. Dual Action of Lysophosphatidate-Functionalised Titanium: Interactions with Human (MG63) Osteoblasts and Methicillin Resistant Staphylococcus aureus.

    Science.gov (United States)

    Skindersoe, Mette Elena; Krogfelt, Karen A; Blom, Ashley; Zhang, Jianxing; Jiang, Guowei; Prestwich, Glenn D; Mansell, Jason Peter

    2015-01-01

    Titanium (Ti) is a widely used material for surgical implants; total joint replacements (TJRs), screws and plates for fixing bones and dental implants are forged from Ti. Whilst Ti integrates well into host tissue approximately 10% of TJRs will fail in the lifetime of the patient through a process known as aseptic loosening. These failures necessitate revision arthroplasties which are more complicated and costly than the initial procedure. Finding ways of enhancing early (osseo)integration of TJRs is therefore highly desirable and continues to represent a research priority in current biomaterial design. One way of realising improvements in implant quality is to coat the Ti surface with small biological agents known to support human osteoblast formation and maturation at Ti surfaces. Lysophosphatidic acid (LPA) and certain LPA analogues offer potential solutions as Ti coatings in reducing aseptic loosening. Herein we present evidence for the successful bio-functionalisation of Ti using LPA. This modified Ti surface heightened the maturation of human osteoblasts, as supported by increased expression of alkaline phosphatase. These functionalised surfaces also deterred the attachment and growth of Staphylococcus aureus, a bacterium often associated with implant failures through sepsis. Collectively we provide evidence for the fabrication of a dual-action Ti surface finish, a highly desirable feature towards the development of next-generation implantable devices.

  9. Dual Action of Lysophosphatidate-Functionalised Titanium: Interactions with Human (MG63 Osteoblasts and Methicillin Resistant Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Mette Elena Skindersoe

    Full Text Available Titanium (Ti is a widely used material for surgical implants; total joint replacements (TJRs, screws and plates for fixing bones and dental implants are forged from Ti. Whilst Ti integrates well into host tissue approximately 10% of TJRs will fail in the lifetime of the patient through a process known as aseptic loosening. These failures necessitate revision arthroplasties which are more complicated and costly than the initial procedure. Finding ways of enhancing early (osseointegration of TJRs is therefore highly desirable and continues to represent a research priority in current biomaterial design. One way of realising improvements in implant quality is to coat the Ti surface with small biological agents known to support human osteoblast formation and maturation at Ti surfaces. Lysophosphatidic acid (LPA and certain LPA analogues offer potential solutions as Ti coatings in reducing aseptic loosening. Herein we present evidence for the successful bio-functionalisation of Ti using LPA. This modified Ti surface heightened the maturation of human osteoblasts, as supported by increased expression of alkaline phosphatase. These functionalised surfaces also deterred the attachment and growth of Staphylococcus aureus, a bacterium often associated with implant failures through sepsis. Collectively we provide evidence for the fabrication of a dual-action Ti surface finish, a highly desirable feature towards the development of next-generation implantable devices.

  10. Lysophosphatidic acid induces reactive oxygen species generation by activating protein kinase C in PC-3 human prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Chu-Cheng; Lin, Chuan-En; Lin, Yueh-Chien [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Ju, Tsai-Kai [Instrumentation Center, National Taiwan University, Taipei, Taiwan, ROC (China); Technology Commons, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Huang, Yuan-Li [Department of Biotechnology, Asia University, Taichung, Taiwan, ROC (China); Lee, Ming-Shyue [Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei, Taiwan, ROC (China); Chen, Jiun-Hong [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Department of Life Science, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Lee, Hsinyu, E-mail: hsinyu@ntu.edu.tw [Institute of Zoology, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Department of Life Science, College of Life Science, National Taiwan University, Taipei, Taiwan, ROC (China); Center for Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Research Center for Developmental Biology and Regenerative Medicine, National Taiwan University, Taipei, Taiwan, ROC (China)

    2013-11-01

    Highlights: •LPA induces ROS generation through LPA{sub 1} and LPA{sub 3}. •LPA induces ROS generation by activating PLC. •PKCζ mediates LPA-induced ROS generation. -- Abstract: Prostate cancer is one of the most frequently diagnosed cancers in males, and PC-3 is a cell model popularly used for investigating the behavior of late stage prostate cancer. Lysophosphatidic acid (LPA) is a lysophospholipid that mediates multiple behaviors in cancer cells, such as proliferation, migration and adhesion. We have previously demonstrated that LPA enhances vascular endothelial growth factor (VEGF)-C expression in PC-3 cells by activating the generation of reactive oxygen species (ROS), which is known to be an important mediator in cancer progression. Using flow cytometry, we showed that LPA triggers ROS generation within 10 min and that the generated ROS can be suppressed by pretreatment with the NADPH oxidase (Nox) inhibitor diphenylene iodonium. In addition, transfection with LPA{sub 1} and LPA{sub 3} siRNA efficiently blocked LPA-induced ROS production, suggesting that both receptors are involved in this pathway. Using specific inhibitors and siRNA, phospholipase C (PLC) and protein kinase C (PKC) were also suggested to participate in LPA-induced ROS generation. Overall, we demonstrated that LPA induces ROS generation in PC-3 prostate cancer cells and this is mediated through the PLC/PKC/Nox pathway.

  11. Involvement of aberrant DNA methylation on reduced expression of lysophosphatidic acid receptor-1 gene in rat tumor cell lines

    International Nuclear Information System (INIS)

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. Recently, it has been reported that alterations of LPA receptor expression might be important in the malignant transformation of tumor cells. Therefore, to assess an involvement of DNA methylation in reduced expression of the LPA receptor-1 (lpa1) gene, we investigated the expression of the lpa1 gene and its DNA methylation patterns in rat tumor cell lines. Both rat brain-derived neuroblastoma B103 and liver-derived hepatoma RH7777 cells used in this study indicated no expression of lpa1. For the analysis of methylation status, bisulfite sequencing was performed with B103 and RH7777 cells, comparing with other lpa1 expressed cells and normal tissues of brain and liver. The lpa1 expressed cells and tissues were all unmethylated in this region of lpa1. In contrast, both B103 and RH7777 cells were highly methylated, correlating with reduced expression of the lpa1. Treatment with 5-aza 2'-deoxycytidine induced expression of lpa1 gene in B103 and RH7777 cells after 24 h. In RH7777 cells treated with 5-aza 2'-deoxycytidine, stress fiber formation was also observed in response to LPA in RH7777 cells, but not in untreated RH7777 cells. These results suggest that aberrant DNA methylation of the lpa1 gene may be involved in its reduced expression in rat tumor cells

  12. Genetic Basis for Correction of Very‐Long‐Chain Acyl-Coenzyme A Dehydrogenase Deficiency by Bezafibrate in Patient Fibroblasts: Toward a Genotype‐Based Therapy

    DEFF Research Database (Denmark)

    Gobin‐Limballe, S.; Djouadi, F.; Aubey, F.;

    2007-01-01

    treatment. Recent data suggest that bezafibrate could improve the FAO capacities in β‐oxidation-deficient cells, by enhancing the residual level of mutant enzyme activity via gene‐expression stimulation. Since VLCAD‐deficient patients frequently harbor missense mutations with unpredictable effects on enzyme...... activity, we investigated the response to bezafibrate as a function of genotype in 33 VLCAD‐deficient fibroblasts representing 45 different mutations. Treatment with bezafibrate (400 μM for 48 h) resulted in a marked increase in FAO capacities, often leading to restoration of normal values, for 21...... genotypes that mainly corresponded to patients with the myopathic phenotype. In contrast, bezafibrate induced no changes in FAO for 11 genotypes corresponding to severe neonatal or infantile phenotypes. This pattern of response was not due to differential inductions of VLCAD messenger RNA, as shown by...

  13. Cardiolipin Molecular Species with Shorter Acyl Chains Accumulate in Saccharomyces cerevisiae Mutants Lacking the Acyl Coenzyme A-binding Protein Acb1p

    Science.gov (United States)

    Rijken, Pieter J.; Houtkooper, Riekelt H.; Akbari, Hana; Brouwers, Jos F.; Koorengevel, Martijn C.; de Kruijff, Ben; Frentzen, Margrit; Vaz, Frédéric M.; de Kroon, Anton I. P. M.

    2009-01-01

    The function of the mitochondrial phospholipid cardiolipin (CL) is thought to depend on its acyl chain composition. The present study aims at a better understanding of the way the CL species profile is established in Saccharomyces cerevisiae by using depletion of the acyl-CoA-binding protein Acb1p as a tool to modulate the cellular acyl chain content. Despite the presence of an intact CL remodeling system, acyl chains shorter than 16 carbon atoms (C16) were found to accumulate in CL in cells lacking Acb1p. Further experiments revealed that Taz1p, a key CL remodeling enzyme, was not responsible for the shortening of CL in the absence of Acb1p. This left de novo CL synthesis as the only possible source of acyl chains shorter than C16 in CL. Experiments in which the substrate specificity of the yeast cardiolipin synthase Crd1p and the acyl chain composition of individual short CL species were investigated, indicated that both CL precursors (i.e. phosphatidylglycerol and CDP-diacylglycerol) contribute to comparable extents to the shorter acyl chains in CL in acb1 mutants. Based on the findings, we conclude that the fatty acid composition of mature CL in yeast is governed by the substrate specificity of the CL-specific lipase Cld1p and the fatty acid composition of the Taz1p substrates. PMID:19656950

  14. Systematic Analysis of Gene Expression Alterations and Clinical Outcomes for Long-Chain Acyl-Coenzyme A Synthetase Family in Cancer.

    Directory of Open Access Journals (Sweden)

    Wei-Ching Chen

    Full Text Available Dysregulated lipid metabolism contributes to cancer progression. Our previous study indicates that long-chain fatty acyl-Co A synthetase (ACSL 3 is essential for lipid upregulation induced by endoplasmic reticulum stress. In this report, we aimed to identify the role of ACSL family in cancer with systematic analysis and in vitro experiment. We explored the ACSL expression using Oncomine database to determine the gene alteration during carcinogenesis and identified the association between ACSL expression and the survival of cancer patient using PrognoScan database. ACSL1 may play a potential oncogenic role in colorectal and breast cancer and play a potential tumor suppressor role in lung cancer. Co-expression analysis revealed that ACSL1 was coexpressed with MYBPH, PTPRE, PFKFB3, SOCS3 in colon cancer and with LRRFIP1, TSC22D1 in lung cancer. In accordance with PrognoScan analysis, downregulation of ACSL1 in colon and breast cancer cell line inhibited proliferation, migration, and anchorage-independent growth. In contrast, increase of oncogenic property was observed in lung cancer cell line by attenuating ACSL1. High ACSL3 expression predicted a better prognosis in ovarian cancer; in contrast, high ACSL3 predicted a worse prognosis in melanoma. ACSL3 was coexpressed with SNUPN, TRIP13, and SEMA5A in melanoma. High expression of ACSL4 predicted a worse prognosis in colorectal cancer, but predicted better prognosis in breast, brain and lung cancer. ACSL4 was coexpressed with SERPIN2, HNRNPCL1, ITIH2, PROCR, LRRFIP1. High expression of ACSL5 predicted good prognosis in breast, ovarian, and lung cancers. ACSL5 was coexpressed with TMEM140, TAPBPL, BIRC3, PTPRE, and SERPINB1. Low ACSL6 predicted a worse prognosis in acute myeloid leukemia. ACSL6 was coexpressed with SOX6 and DARC. Altogether, different members of ACSLs are implicated in diverse types of cancer development. ACSL-coexpressed molecules may be used to further investigate the role of ACSL family in individual type of cancers.

  15. Role of lysophosphatidic acid receptor LPA2 in the development of allergic airway inflammation in a murine model of asthma

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    Chun Jerold

    2009-11-01

    Full Text Available Abstract Background Lysophosphatidic acid (LPA plays a critical role in airway inflammation through G protein-coupled LPA receptors (LPA1-3. We have demonstrated that LPA induced cytokine and lipid mediator release in human bronchial epithelial cells. Here we provide evidence for the role of LPA and LPA receptors in Th2-dominant airway inflammation. Methods Wild type, LPA1 heterozygous knockout mice (LPA1+/-, and LPA2 heterozygous knockout mice (LPA2+/- were sensitized with inactivated Schistosoma mansoni eggs and local antigenic challenge with Schistosoma mansoni soluble egg Ag (SEA in the lungs. Bronchoalveolar larvage (BAL fluids and lung tissues were collected for analysis of inflammatory responses. Further, tracheal epithelial cells were isolated and challenged with LPA. Results BAL fluids from Schistosoma mansoni egg-sensitized and challenged wild type mice (4 days of challenge showed increase of LPA level (~2.8 fold, compared to control mice. LPA2+/- mice, but not LPA1+/- mice, exposed to Schistosoma mansoni egg revealed significantly reduced cell numbers and eosinophils in BAL fluids, compared to challenged wild type mice. Both LPA2+/- and LPA1+/- mice showed decreases in bronchial goblet cells. LPA2+/- mice, but not LPA1+/- mice showed the decreases in prostaglandin E2 (PGE2 and LPA levels in BAL fluids after SEA challenge. The PGE2 production by LPA was reduced in isolated tracheal epithelial cells from LPA2+/- mice. These results suggest that LPA and LPA receptors are involved in Schistosoma mansoni egg-mediated inflammation and further studies are proposed to understand the role of LPA and LPA receptors in the inflammatory process.

  16. Embryo spacing and implantation timing are differentially regulated by LPA3-mediated lysophosphatidic acid signaling in mice.

    Science.gov (United States)

    Hama, Kotaro; Aoki, Junken; Inoue, Asuka; Endo, Tomoko; Amano, Tomokazu; Motoki, Rie; Kanai, Motomu; Ye, Xiaoqin; Chun, Jerold; Matsuki, Norio; Suzuki, Hiroshi; Shibasaki, Masakatsu; Arai, Hiroyuki

    2007-12-01

    In polytocous animals, blastocysts are evenly distributed along each uterine horn and implant. The molecular mechanisms underlying these precise events remain elusive. We recently showed that lysophosphatidic acid (LPA) has critical roles in the establishment of early pregnancy by affecting embryo spacing and subsequent implantation through its receptor, LPA3. Targeted deletion of Lpa3 in mice resulted in delayed implantation and embryo crowding, which is associated with a dramatic decrease in the prostaglandins and prostaglandin-endoperoxide synthase 2 expression levels. Exogenous administration of prostaglandins rescued the delayed implantation but did not rescue the defects in embryo spacing, suggesting the role of prostaglandins in implantation downstream of LPA3 signaling. In the present study, to know how LPA3 signaling regulates the embryo spacing, we determined the time course distribution of blastocysts during the preimplantation period. In wild-type (WT) uteri, blastocysts were distributed evenly along the uterine horns at Embryonic Day 3.8 (E3.8), whereas in the Lpa3-deficient uteri, they were clustered in the vicinity of the cervix, suggesting that the mislocalization and resulting crowding of the embryos are the cause of the delayed implantation. However, embryos transferred singly into E2.5 pseudopregnant Lpa3-deficient uterine horns still showed delayed implantation but on-time implantation in WT uteri, indicating that embryo spacing and implantation timing are two segregated events. We also found that an LPA3-specific agonist induced rapid uterine contraction in WT mice but not in Lpa3-deficient mice. Because the uterine contraction is critical for embryo spacing, our results suggest that LPA3 signaling controls embryo spacing via uterine contraction around E3.5.

  17. Lysophosphatidic acid induced nuclear translocation of nuclear factor-κB in Panc-1 cells by mobilizing cytosolic free calcium

    Institute of Scientific and Technical Information of China (English)

    Yoshiyuki Arita; Tetsuhide Ito; Takamasa Pond; Ken Kawabe; Terumasa Hisano; Ryoichi Takayanagi

    2008-01-01

    AIM: To clarify whether Lysophosphatidic acid (LPA) activates the nuclear translocation of nuclear factor-κB (NF-κB) in pancreatic cancer.METHODS: Panc-1, a human pancreatic cancer cell line, was used throughout the study. The expression of LPA receptors was confirmed by reverse-transcript polymerase chain reaction (RT-PCR). Cytosolic free calcium was measured by fluorescent calcium indicator fura-2, and the localization of NF-κB was visualized by immunofluorescent method with or without various agents, which effect cell signaling.RESULTS: Panc-1 expressed LPA receptors, LPAA1,LPA2 and LPA3. LPA caused the elevation of cytosolic free calcium dose-dependently. LPA also caused the nuclear translocation of NF-κB. Cytosolic free calcium was attenuated by pertussis toxin (PTX) and U73122,an inhibitor of phospholipase C. The translocation of NF-κB was similarly attenuated by PTX and U73122,but phorbol ester, an activator of protein kinase C,alone did not translocate NF-κB. Furthermore, the transtocation of NF-κB was completely blocked by Ca2+ chelator BAPTA-AM. Thapsigargin, an endoplasmic-reticulum Ca2+-ATPase pump inhibitor, also promoted the translocation of NF-κB. Staurosporine, a protein kinase C inhibitor, attenuated translocation of NF-κB induced by LPA.CONCLUSlON: These findings suggest that protein kinase C is activated endogenously in Panc-1, and protein kinase C is essential for activating NF-κB with cytosolic calcium and that LPA induces the nuclear translocation of NF-κB in Panc-1 by mobilizing cytosolic free calcium.

  18. Progesterone produces antinociceptive and neuroprotective effects in rats with microinjected lysophosphatidic acid in the trigeminal nerve root

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    Kim Min

    2012-03-01

    Full Text Available Abstract Background In our present study, we studied the role of demyelination of the trigeminal nerve root in the development of prolonged nociceptive behavior in the trigeminal territory. Results Under anesthesia, the Sprague-Dawley rats were mounted onto a stereotaxic frame and 3 μL of lysophosphatidic acid (LPA, 1 nmol was injected into the trigeminal nerve root to produce demyelination. This treatment decreased the air-puff thresholds, persisted until postoperative day 130, and then returned to the preoperative levels 160 days after LPA injection. The LPA-treated rats also showed a significant hyper-responsiveness to pin-prick stimulation. We further investigated the antinociceptive and neuroprotective effects of progesterone in rats undergoing demyelination of the trigeminal nerve root. Progesterone (8, 16 mg/kg/day was administered subcutaneously, beginning on the operative day, for five consecutive days in the LPA-treated rats. Treatment with progesterone produced significant early anti-allodynic effects and delayed prolonged anti-allodynic effects. The expression of protein zero (P0 and peripheral myelin protein 22 (PMP22 were significantly down-regulated in the trigeminal nerve root on postoperative day 5 following LPA injection. This down-regulation of the P0 and PMP22 levels was blocked by progesterone treatment. Conclusions These results suggest that progesterone produces antinociceptive effects through neuroprotective action in animals with LPA-induced trigeminal neuropathic pain. Moreover, progesterone has potential utility as a novel therapy for trigeminal neuropathic pain relief at an appropriate managed dose and is therefore a possible future treatment strategy for improving the recovery from injury.

  19. Derivatives of Dictyostelium differentiation-inducing factors inhibit lysophosphatidic acid-stimulated migration of murine osteosarcoma LM8 cells.

    Science.gov (United States)

    Kubohara, Yuzuru; Komachi, Mayumi; Homma, Yoshimi; Kikuchi, Haruhisa; Oshima, Yoshiteru

    2015-08-01

    Osteosarcoma is a common metastatic bone cancer that predominantly develops in children and adolescents. Metastatic osteosarcoma remains associated with a poor prognosis; therefore, more effective anti-metastatic drugs are needed. Differentiation-inducing factor-1 (DIF-1), -2, and -3 are novel lead anti-tumor agents that were originally isolated from the cellular slime mold Dictyostelium discoideum. Here we investigated the effects of a panel of DIF derivatives on lysophosphatidic acid (LPA)-induced migration of mouse osteosarcoma LM8 cells by using a Boyden chamber assay. Some DIF derivatives such as Br-DIF-1, DIF-3(+2), and Bu-DIF-3 (5-20 μM) dose-dependently suppressed LPA-induced cell migration with associated IC50 values of 5.5, 4.6, and 4.2 μM, respectively. On the other hand, the IC50 values of Br-DIF-1, DIF-3(+2), and Bu-DIF-3 versus cell proliferation were 18.5, 7.2, and 2.0 μM, respectively, in LM8 cells, and >20, 14.8, and 4.3 μM, respectively, in mouse 3T3-L1 fibroblasts (non-transformed). Together, our results demonstrate that Br-DIF-1 in particular may be a valuable tool for the analysis of cancer cell migration, and that DIF derivatives such as DIF-3(+2) and Bu-DIF-3 are promising lead anti-tumor agents for the development of therapies that suppress osteosarcoma cell proliferation, migration, and metastasis. PMID:26056940

  20. Lysophosphatidic Acid and Sphingosine-1-Phosphate: A Concise Review of Biological Function and Applications for Tissue Engineering.

    Science.gov (United States)

    Binder, Bernard Y K; Williams, Priscilla A; Silva, Eduardo A; Leach, J Kent

    2015-12-01

    The presentation and controlled release of bioactive signals to direct cellular growth and differentiation represents a widely used strategy in tissue engineering. Historically, work in this field has primarily focused on the delivery of large cytokines and growth factors, which can be costly to manufacture and difficult to deliver in a sustained manner. There has been a marked increase over the past decade in the pursuit of lipid mediators due to their wide range of effects over multiple cell types, low cost, and ease of scale-up. Lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) are two bioactive lysophospholipids (LPLs) that have gained attention for use as pharmacological agents in tissue engineering applications. While these lipids can have similar effects on cellular response, they possess distinct chemical backbones, mechanisms of synthesis and degradation, and signaling pathways using a discrete set of G-protein-coupled receptors (GPCRs). LPA and S1P predominantly act extracellularly on their GPCRs and can directly regulate cell survival, differentiation, cytokine secretion, proliferation, and migration--each of the important functions that must be considered in regenerative medicine. In addition to these potent physiological functions, these LPLs play pivotal roles in a number of pathophysiological processes. To capitalize on the promise of these molecules in tissue engineering, these lipids have been incorporated into biomaterials for in vivo delivery. Here, we survey the effects of LPA and S1P on both cellular- and tissue-level phenotypes, with an eye toward regulating stem/progenitor cell growth and differentiation. In particular, we examine work that has translational applications for cell-based tissue engineering strategies in promoting cell survival, bone and cartilage engineering, and therapeutic angiogenesis.

  1. Absence of the lysophosphatidic acid receptor LPA1 results in abnormal bone development and decreased bone mass☆,☆☆

    Science.gov (United States)

    Gennero, Isabelle; Laurencin-Dalicieux, Sara; Conte-Auriol, Françoise; Briand-Mésange, Fabienne; Laurencin, Danielle; Rue, Jackie; Beton, Nicolas; Malet, Nicole; Mus, Marianne; Tokumura, Akira; Bourin, Philippe; Vico, Laurence; Brunel, Gérard; Oreffo, Richard O. C.; Chun, Jerold; Salles, Jean Pierre

    2013-01-01

    Lysophosphatidic acid (LPA) is a lipid mediator that acts in paracrine systems via interaction with a subset of G protein-coupled receptors (GPCRs). LPA promotes cell growth and differentiation, and has been shown to be implicated in a variety of developmental and pathophysiological processes. At least 6 LPA GPCRs have been identified to date: LPA1–LPA6. Several studies have suggested that local production of LPA by tissues and cells contributes to paracrine regulation, and a complex interplay between LPA and its receptors, LPA1 and LPA4, is believed to be involved in the regulation of bone cell activity. In particular, LPA1may activate both osteoblasts and osteoclasts. However, its role has not as yet been examined with regard to the overall status of bone in vivo. We attempted to clarify this role by defining the bone phenotype of LPA1(−/−) mice. These mice demonstrated significant bone defects and low bone mass, indicating that LPA1 plays an important role in osteogenesis. The LPA1(−/−) mice also presented growth and sternal and costal abnormalities, which highlights the specific roles of LPA1 during bone development. Microcomputed tomography and histological analysis demonstrated osteoporosis in the trabecular and cortical bone of LPA1(−/−) mice. Finally, bone marrow mesenchymal progenitors from these mice displayed decreased osteoblastic differentiation. These results suggest that LPA1 strongly influences bone development both qualitatively and quantitatively and that, in vivo, its absence results in decreased osteogenesis with no clear modification of osteoclasis. They open perspectives for a better understanding of the role of the LPA/LPA1 paracrine pathway in bone pathophysiology. PMID:21569876

  2. Absence of the lysophosphatidic acid receptor LPA1 results in abnormal bone development and decreased bone mass.

    Science.gov (United States)

    Gennero, Isabelle; Laurencin-Dalicieux, Sara; Conte-Auriol, Françoise; Briand-Mésange, Fabienne; Laurencin, Danielle; Rue, Jackie; Beton, Nicolas; Malet, Nicole; Mus, Marianne; Tokumura, Akira; Bourin, Philippe; Vico, Laurence; Brunel, Gérard; Oreffo, Richard O C; Chun, Jerold; Salles, Jean Pierre

    2011-09-01

    Lysophosphatidic acid (LPA) is a lipid mediator that acts in paracrine systems via interaction with a subset of G protein-coupled receptors (GPCRs). LPA promotes cell growth and differentiation, and has been shown to be implicated in a variety of developmental and pathophysiological processes. At least 6 LPA GPCRs have been identified to date: LPA1-LPA6. Several studies have suggested that local production of LPA by tissues and cells contributes to paracrine regulation, and a complex interplay between LPA and its receptors, LPA1 and LPA4, is believed to be involved in the regulation of bone cell activity. In particular, LPA1 may activate both osteoblasts and osteoclasts. However, its role has not as yet been examined with regard to the overall status of bone in vivo. We attempted to clarify this role by defining the bone phenotype of LPA1((-/-)) mice. These mice demonstrated significant bone defects and low bone mass, indicating that LPA1 plays an important role in osteogenesis. The LPA1((-/-)) mice also presented growth and sternal and costal abnormalities, which highlights the specific roles of LPA1 during bone development. Microcomputed tomography and histological analysis demonstrated osteoporosis in the trabecular and cortical bone of LPA1((-/-)) mice. Finally, bone marrow mesenchymal progenitors from these mice displayed decreased osteoblastic differentiation. These results suggest that LPA1 strongly influences bone development both qualitatively and quantitatively and that, in vivo, its absence results in decreased osteogenesis with no clear modification of osteoclasis. They open perspectives for a better understanding of the role of the LPA/LPA1 paracrine pathway in bone pathophysiology.

  3. The mouse lp(A3)/Edg7 lysophosphatidic acid receptor gene: genomic structure, chromosomal localization, and expression pattern.

    Science.gov (United States)

    Contos, J J; Chun, J

    2001-04-18

    The extracellular signaling molecule, lysophosphatidic acid (LPA), mediates proliferative and morphological effects on cells and has been proposed to be involved in several biological processes including neuronal development, wound healing, and cancer progression. Three mammalian G protein-coupled receptors, encoded by genes designated lp (lysophospholipid) receptor or edg (endothelial differentiation gene), mediate the effects of LPA, activating similar (e.g. Ca(2+) release) as well as distinct (neurite retraction) responses. To understand the evolution and function of LPA receptor genes, we characterized lp(A3)/Edg7 in mouse and human and compared the expression pattern with the other two known LPA receptor genes (lp(A1)/Edg2 and lp(A2)/Edg4non-mutant). We found mouse and human lp(A3) to have nearly identical three-exon genomic structures, with introns upstream of the coding region for transmembrane domain (TMD) I and within the coding region for TMD VI. This structure is similar to lp(A1) and lp(A2), indicating a common ancestral gene with two introns. We localized mouse lp(A3) to distal Chromosome 3 near the varitint waddler (Va) gene, in a region syntenic with the human lp(A3) chromosomal location (1p22.3-31.1). We found highest expression levels of each of the three LPA receptor genes in adult mouse testes, relatively high expression levels of lp(A2) and lp(A3) in kidney, and moderate expression of lp(A2) and lp(A3) in lung. All lp(A) transcripts were expressed during brain development, with lp(A1) and lp(A2) transcripts expressed during the embryonic neurogenic period, and lp(A3) transcript during the early postnatal period. Our results indicate both overlapping as well as distinct functions of lp(A1), lp(A2), and lp(A3). PMID:11313151

  4. Cancer cell expression of autotaxin controls bone metastasis formation in mouse through lysophosphatidic acid-dependent activation of osteoclasts.

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    Marion David

    Full Text Available BACKGROUND: Bone metastases are highly frequent complications of breast cancers. Current bone metastasis treatments using powerful anti-resorptive agents are only palliative indicating that factors independent of bone resorption control bone metastasis progression. Autotaxin (ATX/NPP2 is a secreted protein with both oncogenic and pro-metastatic properties. Through its lysosphospholipase D (lysoPLD activity, ATX controls the level of lysophosphatidic acid (LPA in the blood. Platelet-derived LPA promotes the progression of osteolytic bone metastases of breast cancer cells. We asked whether ATX was involved in the bone metastasis process. We characterized the role of ATX in osteolytic bone metastasis formation by using genetically modified breast cancer cells exploited on different osteolytic bone metastasis mouse models. METHODOLOGY/PRINCIPAL FINDINGS: Intravenous injection of human breast cancer MDA-B02 cells with forced expression of ATX (MDA-B02/ATX to immunodeficiency BALB/C nude mice enhanced osteolytic bone metastasis formation, as judged by increased bone loss, tumor burden, and a higher number of active osteoclasts at the metastatic site. Mouse breast cancer 4T1 cells induced the formation of osteolytic bone metastases after intracardiac injection in immunocompetent BALB/C mice. These cells expressed active ATX and silencing ATX expression inhibited the extent of osteolytic bone lesions and decreased the number of active osteoclasts at the bone metastatic site. In vitro, osteoclast differentiation was enhanced in presence of MDA-B02/ATX cell conditioned media or recombinant autotaxin that was blocked by the autotaxin inhibitor vpc8a202. In vitro, addition of LPA to active charcoal-treated serum restored the capacity of the serum to support RANK-L/MCSF-induced osteoclastogenesis. CONCLUSION/SIGNIFICANCE: Expression of autotaxin by cancer cells controls osteolytic bone metastasis formation. This work demonstrates a new role for LPA as a

  5. Derivatives of Dictyostelium differentiation-inducing factors inhibit lysophosphatidic acid–stimulated migration of murine osteosarcoma LM8 cells

    International Nuclear Information System (INIS)

    Osteosarcoma is a common metastatic bone cancer that predominantly develops in children and adolescents. Metastatic osteosarcoma remains associated with a poor prognosis; therefore, more effective anti-metastatic drugs are needed. Differentiation-inducing factor-1 (DIF-1), −2, and −3 are novel lead anti-tumor agents that were originally isolated from the cellular slime mold Dictyostelium discoideum. Here we investigated the effects of a panel of DIF derivatives on lysophosphatidic acid (LPA)-induced migration of mouse osteosarcoma LM8 cells by using a Boyden chamber assay. Some DIF derivatives such as Br-DIF-1, DIF-3(+2), and Bu-DIF-3 (5–20 μM) dose-dependently suppressed LPA-induced cell migration with associated IC50 values of 5.5, 4.6, and 4.2 μM, respectively. On the other hand, the IC50 values of Br-DIF-1, DIF-3(+2), and Bu-DIF-3 versus cell proliferation were 18.5, 7.2, and 2.0 μM, respectively, in LM8 cells, and >20, 14.8, and 4.3 μM, respectively, in mouse 3T3-L1 fibroblasts (non-transformed). Together, our results demonstrate that Br-DIF-1 in particular may be a valuable tool for the analysis of cancer cell migration, and that DIF derivatives such as DIF-3(+2) and Bu-DIF-3 are promising lead anti-tumor agents for the development of therapies that suppress osteosarcoma cell proliferation, migration, and metastasis. - Highlights: • LPA induces cell migration (invasion) in murine osteosarcoma LM8 cells. • DIFs are novel lead anti-tumor agents found in Dictyostelium discoideum. • We examined the effects of DIF derivatives on LPA-induced LM8 cell migration in vitro. • Some of the DIF derivatives inhibited LPA-induced LM8 cell migration

  6. Derivatives of Dictyostelium differentiation-inducing factors inhibit lysophosphatidic acid–stimulated migration of murine osteosarcoma LM8 cells

    Energy Technology Data Exchange (ETDEWEB)

    Kubohara, Yuzuru, E-mail: ykuboha@juntendo.ac.jp [Department of Molecular and Cellular Biology, Institute for Molecular and Cellular Regulation (IMCR), Gunma University, Maebashi 371-8512 (Japan); Department of Health Science, Juntendo University Graduate School of Health and Sports Science, Inzai 270-1695 (Japan); Komachi, Mayumi [Department of Molecular and Cellular Biology, Institute for Molecular and Cellular Regulation (IMCR), Gunma University, Maebashi 371-8512 (Japan); Department of Radiation Oncology, Gunma University Graduate School of Medicine, Maebashi 371-8511 (Japan); Homma, Yoshimi [Department of Biomolecular Science, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295 (Japan); Kikuchi, Haruhisa; Oshima, Yoshiteru [Laboratory of Natural Product Chemistry, Tohoku University Graduate School of Pharmaceutical Sciences, Aoba-yama, Aoba-ku, Sendai 980-8578 (Japan)

    2015-08-07

    Osteosarcoma is a common metastatic bone cancer that predominantly develops in children and adolescents. Metastatic osteosarcoma remains associated with a poor prognosis; therefore, more effective anti-metastatic drugs are needed. Differentiation-inducing factor-1 (DIF-1), −2, and −3 are novel lead anti-tumor agents that were originally isolated from the cellular slime mold Dictyostelium discoideum. Here we investigated the effects of a panel of DIF derivatives on lysophosphatidic acid (LPA)-induced migration of mouse osteosarcoma LM8 cells by using a Boyden chamber assay. Some DIF derivatives such as Br-DIF-1, DIF-3(+2), and Bu-DIF-3 (5–20 μM) dose-dependently suppressed LPA-induced cell migration with associated IC{sub 50} values of 5.5, 4.6, and 4.2 μM, respectively. On the other hand, the IC{sub 50} values of Br-DIF-1, DIF-3(+2), and Bu-DIF-3 versus cell proliferation were 18.5, 7.2, and 2.0 μM, respectively, in LM8 cells, and >20, 14.8, and 4.3 μM, respectively, in mouse 3T3-L1 fibroblasts (non-transformed). Together, our results demonstrate that Br-DIF-1 in particular may be a valuable tool for the analysis of cancer cell migration, and that DIF derivatives such as DIF-3(+2) and Bu-DIF-3 are promising lead anti-tumor agents for the development of therapies that suppress osteosarcoma cell proliferation, migration, and metastasis. - Highlights: • LPA induces cell migration (invasion) in murine osteosarcoma LM8 cells. • DIFs are novel lead anti-tumor agents found in Dictyostelium discoideum. • We examined the effects of DIF derivatives on LPA-induced LM8 cell migration in vitro. • Some of the DIF derivatives inhibited LPA-induced LM8 cell migration.

  7. Lysophosphatidic acid alters the expression profiles of angiogenic factors, cytokines, and chemokines in mouse liver sinusoidal endothelial cells.

    Directory of Open Access Journals (Sweden)

    Chia-Hung Chou

    Full Text Available Lysophosphatidic acid (LPA is a multi-function glycerophospholipid. LPA affects the proliferation of hepatocytes and stellate cells in vitro, and in a partial hepatectomy induced liver regeneration model, the circulating LPA levels and LPA receptor (LPAR expression levels in liver tissue are significantly changed. Liver sinusoidal endothelial cells (Lsecs play an important role during liver regeneration. However, the effects of LPA on Lsecs are not well known. Thus, we investigated the effects of LPA on the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs.Mouse Lsecs were isolated using CD31-coated magnetic beads. The mRNA expression levels of LPAR's and other target genes were determined by quantitative RT-PCR. The protein levels of angiogenesis factors, cytokines, and chemokines were determined using protein arrays and enzyme immunoassay (EIA. Critical LPAR related signal transduction was verified by using an appropriate chemical inhibitor.LPAR1 and LPAR3 mRNA's were expressed in mouse LPA-treated Lsecs. Treating Lsecs with a physiological level of LPA significantly enhanced the protein levels of angiogenesis related proteins (cyr61 and TIMP-1, cytokines (C5/C5a, M-CSF, and SDF-1, and chemokines (MCP-5, gp130, CCL28, and CXCL16. The LPAR1 and LPAR3 antagonist ki16425 significantly inhibited the LPA-enhanced expression of cyr61, TIMP-1, SDF-1, MCP-5, gp130, CCL28, and CXCL16, but not that of C5/C5a or M-CSF. LPA-induced C5/C5a and M-CSF expression may have been through an indirect regulation mechanism.LPA regulated the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs that was mediated via LPAR1 and LPAR3 signaling. Most of the factors that were enhanced by LPA have been found to play critical roles during liver regeneration. Thus, these results may prove useful for manipulating LPA effects on liver regeneration.

  8. Lysophosphatidic Acid Alters the Expression Profiles of Angiogenic Factors, Cytokines, and Chemokines in Mouse Liver Sinusoidal Endothelial Cells

    Science.gov (United States)

    Chou, Chia-Hung; Lai, Shou-Lun; Ho, Cheng-Maw; Lin, Wen-Hsi; Chen, Chiung-Nien; Lee, Po-Huang; Peng, Fu-Chuo; Kuo, Sung-Hsin; Wu, Szu-Yuan; Lai, Hong-Shiee

    2015-01-01

    Background and Aims Lysophosphatidic acid (LPA) is a multi-function glycerophospholipid. LPA affects the proliferation of hepatocytes and stellate cells in vitro, and in a partial hepatectomy induced liver regeneration model, the circulating LPA levels and LPA receptor (LPAR) expression levels in liver tissue are significantly changed. Liver sinusoidal endothelial cells (Lsecs) play an important role during liver regeneration. However, the effects of LPA on Lsecs are not well known. Thus, we investigated the effects of LPA on the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs. Methods Mouse Lsecs were isolated using CD31-coated magnetic beads. The mRNA expression levels of LPAR’s and other target genes were determined by quantitative RT-PCR. The protein levels of angiogenesis factors, cytokines, and chemokines were determined using protein arrays and enzyme immunoassay (EIA). Critical LPAR related signal transduction was verified by using an appropriate chemical inhibitor. Results LPAR1 and LPAR3 mRNA’s were expressed in mouse LPA-treated Lsecs. Treating Lsecs with a physiological level of LPA significantly enhanced the protein levels of angiogenesis related proteins (cyr61 and TIMP-1), cytokines (C5/C5a, M-CSF, and SDF-1), and chemokines (MCP-5, gp130, CCL28, and CXCL16). The LPAR1 and LPAR3 antagonist ki16425 significantly inhibited the LPA-enhanced expression of cyr61, TIMP-1, SDF-1, MCP-5, gp130, CCL28, and CXCL16, but not that of C5/C5a or M-CSF. LPA-induced C5/C5a and M-CSF expression may have been through an indirect regulation mechanism. Conclusion LPA regulated the expression profiles of angiogenic factors, cytokines, and chemokines in Lsecs that was mediated via LPAR1 and LPAR3 signaling. Most of the factors that were enhanced by LPA have been found to play critical roles during liver regeneration. Thus, these results may prove useful for manipulating LPA effects on liver regeneration. PMID:25822713

  9. Identification of Heparin-Binding EGF-Like Growth Factor (HB-EGF) as a Biomarker for Lysophosphatidic Acid Receptor Type 1 (LPA1) Activation in Human Breast and Prostate Cancers

    OpenAIRE

    David, Marion; Sahay, Debashish; Mege, Florence; Descotes, Françoise; Leblanc, Raphaël; Ribeiro, Johnny; Clézardin, Philippe; Peyruchaud, Olivier

    2014-01-01

    Lysophosphatidic acid (LPA) is a natural bioactive lipid with growth factor-like functions due to activation of a series of six G protein-coupled receptors (LPA1–6). LPA receptor type 1 (LPA1) signaling influences the pathophysiology of many diseases including cancer, obesity, rheumatoid arthritis, as well as lung, liver and kidney fibrosis. Therefore, LPA1 is an attractive therapeutic target. However, most mammalian cells co-express multiple LPA receptors whose co-activation impairs the vali...

  10. Heart-type fatty-acid-binding protein (FABP3 is a lysophosphatidic acid-binding protein in human coronary artery endothelial cells

    Directory of Open Access Journals (Sweden)

    Ryoko Tsukahara

    2014-01-01

    Full Text Available Fatty-acid-binding protein 3, muscle and heart (FABP3, also known as heart-type FABP, is a member of the family of intracellular lipid-binding proteins. It is a small cytoplasmic protein with a molecular mass of about 15 kDa. FABPs are known to be carrier proteins for transporting fatty acids and other lipophilic substances from the cytoplasm to the nucleus, where these lipids are released to a group of nuclear receptors such as peroxisome proliferator-activated receptors (PPARs. In this study, using lysophosphatidic acid (LPA-coated agarose beads, we have identified FABP3 as an LPA carrier protein in human coronary artery endothelial cells (HCAECs. Administration of LPA to HCAECs resulted in a dose-dependent increase in PPARγ activation. Furthermore, the LPA-induced PPARγ activation was abolished when the FABP3 expression was reduced using small interfering RNA (siRNA. We further show that the nuclear fraction of control HCAECs contained a significant amount of exogenously added LPA, whereas FABP3 siRNA-transfected HCAECs had a decreased level of LPA in the nucleus. Taken together, these results suggest that FABP3 governs the transcriptional activities of LPA by targeting them to cognate PPARγ in the nucleus.

  11. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2011-05-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  12. Quantitative Phosphoproteome Analysis of Lysophosphatidic Acid Induced Chemotaxis applying Dual-step ¹⁸O Labeling Coupled with Immobilized Metal-ion Affinity Chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Ding, Shi-Jian; Wang, Yingchun; Jacobs, Jon M.; Qian, Weijun; Yang, Feng; Tolmachev, Aleksey V.; Du, Xiuxia; Wang, Wei; Moore, Ronald J.; Monroe, Matthew E.; Purvine, Samuel O.; Waters, Katrina M.; Heibeck, Tyler H.; Adkins, Joshua N.; Camp, David G.; Klemke, Richard L.; Smith, Richard D.

    2008-10-01

    Reversible protein phosphorylation is a central cellular regulatory mechanism in modulating protein activity and propagating signals within cellular pathways and networks. Development of more effective methods for the simultaneous identification of phosphorylation sites and quantification of temporal changes in protein phosphorylation could provide important insights into molecular signaling mechanisms in a variety of different cellular processes. Here we present an integrated quantitative phosphoproteomics approach and its applications for comparative analysis of Cos-7 cells in response to lysophosphatidic acid (LPA) gradient stimulation. The approach combines trypsin-catalyzed 16O/18O labeling plus 16O/18O-methanol esterification labeling for quantitation, a macro- Immobilized Metal-ion Affinity Chromatography trap for phosphopeptide enrichment, and a monolithic capillary column with integrated electrospray emitter. LC separation and MS/MS is followed by neutral loss-dependent MS/MS/MS for phosphopeptide identification using a linear ion trap (LTQ)-FT mass spectrometer and complementary searching algorithms for interpreting MS/MS spectra. Protein phosphorylation involved in various signaling pathways of cell migration were identified and quantified, such as mitogen-activated protein kinase 1, dual-specificity mitogen-activated protein kinase kinase 2, and dual-specificity tyrosine-phosphorylation regulated kinase 1b, and a number of Rho GTPase-activating proteins. These results demonstrate the efficiency of this quantitative phosphoproteomics approach and its application for rapid discovery of phosphorylation events associated with gradient sensing and cell chemotaxis.

  13. Lysophosphatidic acid transactivates both c-Met and epidermal growth factor receptor, and induces cyclooxygenase-2 expression in human colon cancer LoVo cells

    Institute of Scientific and Technical Information of China (English)

    Dai Shida; Joji Kitayama; Hironori Yamaguchi; Hiroharu Yamashita; Ken Mori; Toshiaki Watanabe; Hirokazu Nagawa

    2005-01-01

    AIM: To examine whether lysophosphatidic acid (LPA)induces phosphorylation of c-Met and epidermal growth factor receptor (EGFR), both of which have been proposed as prognostic markers of colorectal cancer, and whether LPA induces cyclooxygenase-2 (COX-2) expression in human colon cancer cells.METHODS: Using a human colon cancer cell line, LoVo cells, we performed immunoprecipitation analysis,followed by Western blot analysis. We also examined whether LPA induced COX-2 expression, by Western blot analysis.RESULTS: Immunoprecipitation analysis revealed that 10 μmol/L LPA induced tyrosine phosphorylation of c-Met and EGFR in LoVo cells within a few minutes. We found that c-Met tyrosine phosphorylation induced by LPA was not attenuated by pertussis toxin or a matrix metalloproteinase inhibitor, in marked contrast to the results for EGFR. In addition, 0.2-40 μmol/L LPA induced COX-2 expression in a dose-dependent manner.CONCLUSION: Our results suggest that LPA acts upstream of various receptor tyrosine kinases (RTKs) and COX-2,and thus may act as a potent stimulator of colorectal cancer.

  14. Lysophosphatidic acid activates peroxisome proliferator activated receptor-γ in CHO cells that over-express glycerol 3-phosphate acyltransferase-1.

    Directory of Open Access Journals (Sweden)

    Cliona M Stapleton

    Full Text Available Lysophosphatidic acid (LPA is an agonist for peroxisome proliferator activated receptor-γ (PPARγ. Although glycerol-3-phosphate acyltransferase-1 (GPAT1 esterifies glycerol-3-phosphate to form LPA, an intermediate in the de novo synthesis of glycerolipids, it has been assumed that LPA synthesized by this route does not have a signaling role. The availability of Chinese Hamster Ovary (CHO cells that stably overexpress GPAT1, allowed us to analyze PPARγ activation in the presence of LPA produced as an intracellular intermediate. LPA levels in CHO-GPAT1 cells were 6-fold higher than in wild-type CHO cells, and the mRNA abundance of CD36, a PPARγ target, was 2-fold higher. Transactivation assays showed that PPARγ activity was higher in the cells that overexpressed GPAT1. PPARγ activity was enhanced further in CHO-GPAT1 cells treated with the PPARγ ligand troglitazone. Extracellular LPA, phosphatidic acid (PA or a membrane-permeable diacylglycerol had no effect, showing that PPARγ had been activated by LPA generated intracellularly. Transient transfection of a vector expressing 1-acylglycerol-3-phosphate acyltransferase-2, which converts endogenous LPA to PA, markedly reduced PPARγ activity, as did over-expressing diacylglycerol kinase, which converts DAG to PA, indicating that PA could be a potent inhibitor of PPARγ. These data suggest that LPA synthesized via the glycerol-3-phosphate pathway can activate PPARγ and that intermediates of de novo glycerolipid synthesis regulate gene expression.

  15. Chitinase 3-like 1 expression by human (MG63) osteoblasts in response to lysophosphatidic acid and 1,25-dihydroxyvitamin D3.

    Science.gov (United States)

    Mansell, J P; Cooke, M; Read, M; Rudd, H; Shiel, A I; Wilkins, K; Manso, M

    2016-01-01

    Chitinase 3-like 1, otherwise known as YKL-40, is a secreted glycoprotein purported to have a role in extracellular matrix metabolism. The first mammalian cell type found to express YKL-40 was the human osteosarcoma-derived osteoblast, MG63. In that first study the active vitamin D3 metabolite, 1,25-dihydroxycholecalciferol (1,25D), stimulated YKL-40 expression, thereby indicating that a vital factor for skeletal health promoted YKL-40 synthesis by bone forming cells. However, when these MG63 cells were exposed to 1,25D they were also exposed to serum, a rich source of the pleiotropic lipid mediator, lysophosphatidic acid (LPA). Given that 1,25D is now known to co-operate with selected growth factors, including LPA, to influence human osteoblast differentiation we hypothesised that 1,25D and LPA may work together to stimulate osteoblast YKL-40 expression. Herein we report that 1,25D and LPA synergistically promote YKL-40 expression by MG63 cells. Inhibitors targeting AP1, MEK, Sp1 and STAT3 blunted the expression of both alkaline phosphatase and YKL-40 by MG63 cells in response to co-stimulation with 1,25D and LPA. Other ligands of the vitamin D receptor also co-operated with LPA in driving YKL-40 mobilisation. Collectively our findings highlight another important role of 1,25D and LPA in the regulation of human osteoblast function. PMID:27575987

  16. Mechanism of sphingosine 1-phosphate- and lysophosphatidic Acid-induced up-regulation of adhesion molecules and eosinophil chemoattractant in nerve cells.

    LENUS (Irish Health Repository)

    Costello, Richard W

    2012-02-01

    The lysophospholipids sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) act via G-protein coupled receptors S1P(1-5) and LPA(1-3) respectively, and are implicated in allergy. Eosinophils accumulate at innervating cholinergic nerves in asthma and adhere to nerve cells via intercellular adhesion molecule-1 (ICAM-1). IMR-32 neuroblastoma cells were used as an in vitro cholinergic nerve cell model. The G(i) coupled receptors S1P(1), S1P(3), LPA(1), LPA(2) and LPA(3) were expressed on IMR-32 cells. Both S1P and LPA induced ERK phosphorylation and ERK- and G(i)-dependent up-regulation of ICAM-1 expression, with differing time courses. LPA also induced ERK- and G(i)-dependent up-regulation of the eosinophil chemoattractant, CCL-26. The eosinophil granule protein eosinophil peroxidase (EPO) induced ERK-dependent up-regulation of transcription of S1P(1), LPA(1), LPA(2) and LPA(3), providing the situation whereby eosinophil granule proteins may enhance S1P- and\\/or LPA- induced eosinophil accumulation at nerve cells in allergic conditions.

  17. Aromatic hydrocarbon receptor inhibits lysophosphatidic acid-induced vascular endothelial growth factor-A expression in PC-3 prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Pei-Yi; Lin, Yueh-Chien; Lan, Shun-Yan [Institute of Zoology, National Taiwan University, Taipei, Taiwan (China); Huang, Yuan-Li [Department of Biotechnology, Asia University, Taichung, Taiwan (China); Lee, Hsinyu, E-mail: hsinyu@ntu.edu.tw [Institute of Zoology, National Taiwan University, Taipei, Taiwan (China); Department of Life Science, National Taiwan University, Taipei, Taiwan (China)

    2013-08-02

    Highlights: •LPA-induced VEGF-A expression was regulated by HIF-1α and ARNT. •PI3K mediated LPA-induced VEGF-A expression. •AHR signaling inhibited LPA-induced VEGF-A expression in PC-3 cells. -- Abstract: Lysophosphatidic acid (LPA) is a lipid growth factor with multiple biological functions and has been shown to stimulate cancer cell secretion of vascular endothelial growth factor-A (VEGF-A) and trigger angiogenesis. Hypoxia-inducible factor-1 (HIF-1), a heterodimer consisting of HIF-1α and HIF-1β (also known as aromatic hydrocarbon receptor nuclear translocator (ARNT)) subunits, is an important regulator of angiogenesis in prostate cancer (PC) through the enhancement of VEGF-A expression. In this study, we first confirmed the ability of LPA to induce VEGF-A expression in PC-3 cells and then validated that LPA-induced VEGF-A expression was regulated by HIF-1α and ARNT through phosphatidylinositol 3-kinase activation. Aromatic hydrocarbon receptor (AHR), a receptor for dioxin-like compounds, functions as a transcription factor through dimerization with ARNT and was found to inhibit prostate carcinogenesis and vanadate-induced VEGF-A production. Since ARNT is a common dimerization partner of AHR and HIF-1α, we hypothesized that AHR might suppress LPA-induced VEGF-A expression in PC-3 cells by competing with HIF-1α for ARNT. Here we demonstrated that overexpression and ligand activation of AHR inhibited HIF-1-mediated VEGF-A induction by LPA treatment of PC-3 cells. In conclusion, our results suggested that AHR activation may inhibit LPA-induced VEGF-A expression in PC-3 cells by attenuating HIF-1α signaling, and subsequently, suppressing angiogenesis and metastasis of PC. These results suggested that AHR presents a potential therapeutic target for the prevention of PC metastasis.

  18. A novel approach for measuring sphingosine-1-phosphate and lysophosphatidic acid binding to carrier proteins using monoclonal antibodies and the Kinetic Exclusion Assay.

    Science.gov (United States)

    Fleming, Jonathan K; Glass, Thomas R; Lackie, Steve J; Wojciak, Jonathan M

    2016-09-01

    Sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) are bioactive signaling lysophospholipids that activate specific G protein-coupled receptors on the cell surface triggering numerous biological events. In circulation, S1P and LPA associate with specific carrier proteins or chaperones; serum albumin binds both S1P and LPA while HDL shuttles S1P via interactions with apoM. We used a series of kinetic exclusion assays in which monoclonal anti-S1P and anti-LPA antibodies competed with carrier protein for the lysophospholipid to measure the equilibrium dissociation constants (Kd) for these carrier proteins binding S1P and the major LPA species. Fatty acid-free (FAF)-BSA binds these lysophospholipids with the following Kd values: LPA(16:0), 68 nM; LPA(18:1), 130 nM; LPA(18:2), 350 nM; LPA(20:4), 2.2 μM; and S1P, 41 μM. FAF human serum albumin binds each lysophospholipid with comparable affinities. By measuring the apoM concentration and expanding the model to include endogenous ligand, we were able to resolve the Kd values for S1P binding apoM in the context of human HDL and LDL particles (21 nM and 2.4 nM, respectively). The novel competitive assay and analysis described herein enables measurement of Kd values of completely unmodified lysophospholipids binding unmodified carrier proteins in solution, and thus provide insights into S1P and LPA storage in the circulation system and may be useful in understanding chaperone-dependent receptor activation and signaling. PMID:27444045

  19. Regulation of gene expression and subcellular protein distribution in MLO-Y4 osteocytic cells by lysophosphatidic acid: Relevance to dendrite outgrowth.

    Energy Technology Data Exchange (ETDEWEB)

    Waters, Katrina M.; Jacobs, Jon M.; Gritsenko, Marina A.; Karin, Norman J.

    2011-02-26

    Osteoblastic and osteocytic cells are highly responsive to the lipid growth factor lysophosphatidic acid (LPA) but the mechanisms by which LPA alters bone cell functions are largely unknown. A major effect of LPA on osteocytic cells is the stimulation of dendrite membrane outgrowth, a process that we predicted to require changes in gene expression and protein distribution. We employed DNA microarrays for global transcriptional profiling of MLO-Y4 osteocytic cells grown for 6 and 24h in the presence or absence of LPA. We identified 932 transcripts that displayed statistically significant changes in abundance of at least 1.25-fold in response to LPA treatment. Gene ontology (GO) analysis revealed that the regulated gene products were linked to diverse cellular processes, including DNA repair, response to unfolded protein, ossification, protein-RNA complex assembly, and amine biosynthesis. Gene products associated with the regulation of actin microfilament dynamics displayed the most robust expression changes, and LPA-induced dendritogenesis in vitro was blocked by the stress fiber inhibitor cytochalasin D. Mass spectrometry-based proteomic analysis of MLO-Y4 cells revealed significant LPA-induced changes in the abundance of 284 proteins at 6h and 844 proteins at 24h. GO analysis of the proteomic data linked the effects of LPA to cell processes that control of protein distribution and membrane outgrowth, including protein localization, protein complex assembly, Golgi vesicle transport, cytoskeleton-dependent transport, and membrane invagination/endocytosis. Dendrites were isolated from LPA-treated MLO-Y4 cells and subjected to proteomic analysis to quantitatively assess the subcellular distribution of proteins. Sets of 129 and 36 proteins were enriched in the dendrite fraction as compared to whole cells after 6h and 24h of LPA exposure, respectively. Protein markers indicated that membranous organelles were largely excluded from the dendrites. Highly represented among

  20. Cardiolipin molecular species with shorter acyl chains accumulate in Saccharomyces cerevisiae mutants lacking the acyl coenzyme A-binding protein Acb1p: new insights into acyl chain remodeling of cardiolipin.

    Science.gov (United States)

    Rijken, Pieter J; Houtkooper, Riekelt H; Akbari, Hana; Brouwers, Jos F; Koorengevel, Martijn C; de Kruijff, Ben; Frentzen, Margrit; Vaz, Frédéric M; de Kroon, Anton I P M

    2009-10-01

    The function of the mitochondrial phospholipid cardiolipin (CL) is thought to depend on its acyl chain composition. The present study aims at a better understanding of the way the CL species profile is established in Saccharomyces cerevisiae by using depletion of the acyl-CoA-binding protein Acb1p as a tool to modulate the cellular acyl chain content. Despite the presence of an intact CL remodeling system, acyl chains shorter than 16 carbon atoms (C16) were found to accumulate in CL in cells lacking Acb1p. Further experiments revealed that Taz1p, a key CL remodeling enzyme, was not responsible for the shortening of CL in the absence of Acb1p. This left de novo CL synthesis as the only possible source of acyl chains shorter than C16 in CL. Experiments in which the substrate specificity of the yeast cardiolipin synthase Crd1p and the acyl chain composition of individual short CL species were investigated, indicated that both CL precursors (i.e. phosphatidylglycerol and CDP-diacylglycerol) contribute to comparable extents to the shorter acyl chains in CL in acb1 mutants. Based on the findings, we conclude that the fatty acid composition of mature CL in yeast is governed by the substrate specificity of the CL-specific lipase Cld1p and the fatty acid composition of the Taz1p substrates. PMID:19656950

  1. 偏头痛急性期患者溶血磷脂酸与TCD的变化%Changes of lysophosphatidic acid and TCD in patients with migraine during acute stage

    Institute of Scientific and Technical Information of China (English)

    宋叶华; 牛建平; 汤婷; 叶良灶; 何倪靖

    2013-01-01

    Objective Observation the changesof plasma lysophosphatidic acid (LPA) 、lysophosphatidic acid similar levels of phospholipids(AP) and Transcranial Doppler ultrasound (TCD) in acute migraine patients. Methodes Determination plasma LPA and AP in migraine patients with acute period (migraine without aura and migraine with aura group) ,at the same time,were examined by TCD, compared with normal patients. Results Migraine patients with LPA 、AP increased significantly, especial y for migraine with aura group increased more significantly, suggesting that platelet activation in patients with acute migraine attacks, and blood flow velocity in patients with migraine acute stage significantly increased in patients with migraine.The results support functions - vascular nerve disorder in patients with migraine.%目的观察偏头痛急性发作期患者血浆溶血磷脂酸(LPA)及溶血磷脂酸相似磷脂水平(AP)、经颅多普勒超声(TCD)的变化。方法测定偏头痛急性期患者(无先兆偏头痛组及有先兆偏头痛组)血浆LPA及AP水平,同时对患者行TCD检查,与正常患者进行对照。结果偏头痛组血浆LPA水平高于对照组,尤其是有先兆偏头痛组,且偏头痛急性期TCD以血流速度增快为主,尤其是患侧大脑中动脉血流速度。结论偏头痛发作期患者LPA、AP明显升高,尤其是有先兆偏头痛组升高更明显,提示偏头痛急性发作期患者存在血小板活化过程,同时偏头痛急性期患者血流速度常明显增快,支持偏头痛患者存在神经-血管功能紊乱。

  2. Nucleon effective mass and the A dependence of structure functions

    Energy Technology Data Exchange (ETDEWEB)

    Garcia Canal, C.A.; Santangelo, E.M.; Vucetich, H.

    1984-10-08

    The nucleon effective mass was successfully used, as the only free parameter, to adjust the ratio R(A) of structure functions measured in a nucleus of mass number A and in the deuteron, for each A value in the SLAC set of experimental data. The resulting A dependence of the effective mass, being linear in A/sup -1/3/, is consistent with the behavior expected from nuclear structure considerations. The extrapolated value of the effective mass for nuclear matter agrees with previous estimations.

  3. Identification of heparin-binding EGF-like growth factor (HB-EGF as a biomarker for lysophosphatidic acid receptor type 1 (LPA1 activation in human breast and prostate cancers.

    Directory of Open Access Journals (Sweden)

    Marion David

    Full Text Available Lysophosphatidic acid (LPA is a natural bioactive lipid with growth factor-like functions due to activation of a series of six G protein-coupled receptors (LPA₁₋₆. LPA receptor type 1 (LPA₁ signaling influences the pathophysiology of many diseases including cancer, obesity, rheumatoid arthritis, as well as lung, liver and kidney fibrosis. Therefore, LPA₁ is an attractive therapeutic target. However, most mammalian cells co-express multiple LPA receptors whose co-activation impairs the validation of target inhibition in patients because of missing LPA receptor-specific biomarkers. LPA₁ is known to induce IL-6 and IL-8 secretion, as also do LPA₂ and LPA₃. In this work, we first determined the LPA induced early-gene expression profile in three unrelated human cancer cell lines expressing different patterns of LPA receptors (PC3: LPA₁,₂,₆; MDA-MB-231: LPA1,2; MCF-7: LPA₂,₆. Among the set of genes upregulated by LPA only in LPA₁-expressing cells, we validated by QPCR and ELISA that upregulation of heparin-binding EGF-like growth factor (HB-EGF was inhibited by LPA₁-₃ antagonists (Ki16425, Debio0719. Upregulation and downregulation of HB-EGF mRNA was confirmed in vitro in human MDA-B02 breast cancer cells stably overexpressing LPA₁ (MDA-B02/LPA₁ and downregulated for LPA₁ (MDA-B02/shLPA1, respectively. At a clinical level, we quantified the expression of LPA₁ and HB-EGF by QPCR in primary tumors of a cohort of 234 breast cancer patients and found a significantly higher expression of HB-EGF in breast tumors expressing high levels of LPA₁. We also generated human xenograph prostate tumors in mice injected with PC3 cells and found that a five-day treatment with Ki16425 significantly decreased both HB-EGF mRNA expression at the primary tumor site and circulating human HB-EGF concentrations in serum. All together our results demonstrate that HB-EGF is a new and relevant biomarker with potentially high value in

  4. A Dependent Hidden Markov Model of Credit Quality

    Directory of Open Access Journals (Sweden)

    Małgorzata Wiktoria Korolkiewicz

    2012-01-01

    Full Text Available We propose a dependent hidden Markov model of credit quality. We suppose that the "true" credit quality is not observed directly but only through noisy observations given by posted credit ratings. The model is formulated in discrete time with a Markov chain observed in martingale noise, where "noise" terms of the state and observation processes are possibly dependent. The model provides estimates for the state of the Markov chain governing the evolution of the credit rating process and the parameters of the model, where the latter are estimated using the EM algorithm. The dependent dynamics allow for the so-called "rating momentum" discussed in the credit literature and also provide a convenient test of independence between the state and observation dynamics.

  5. 花生溶血磷脂酸酰基转移酶基因的克隆与表达分析%Cloning and Expression Analysis of Lysophosphatidic Acid Acyltransferase (LPAT) Encoding Gene in Peanut

    Institute of Scientific and Technical Information of China (English)

    陈四龙; 黄家权; 雷永; 任小平; 文奇根; 陈玉宁; 姜慧芳; 晏立英; 廖伯寿

    2012-01-01

    Lysophosphatidic acid acyltransferase (LPAT) is a key enzyme in biosynthesis pathway of vegetable oil in plant. It is important for oil crops to improve oil quality and increase seed oil content through genetic engineering. We constructed a full-length cDNA library of peanut (Arachis hypogaea) seed via a large number of sequences of expressed sequence tag (EST) and gene functional annotation, a lysophosphatidic acid acyltransferase gene, designated AhLPAT, and its genomic DNA sequence were isolated from peanut. The sequence of AhLPAT cDN A was 1 629 bp, and its genomic sequence was 5 331 bp. Bioinformatic analysis showed that AhLPAT was composed of 11 exons and 10 introns with typical GT-AG characteristic in comparison of its sequences of genomic DNA and cDNA by Splign in NCBI. A peptide of 387 amino acid residues with protein molecular weight of 43.2 kD and isoelectric point (p7) of 9.42 were deduced from AhLPAT. Conserved domains prediction indicated that AhLPAT comprised a typical conserved acyltransferase domain and a conserved lysophospholipid acyltransferases domain. The deduced amino acid had a high identity with the LPAT proteins reported from other species. Amino acid similarities of LPAT protein be tween peanut and Tropaeolum majus, Brassica napus, Crambe hispanica subsp. Abyssinica, Ricinus communis, and Arabidopsis thaliana were 90%, 89%, 89%, 88%, and 87%, respectively. A phylogenetic tree was constructed by the Neighbor-Joining method using MEGA5.0. The phylogenetic tree suggested that AhLPAT and AtLPAT2 derived from Arabidopsis thaliana were grouped into the same class. Both AhLPAT and AtLPAT2 were endoplasmic reticulum type LPATs. The tissue specific expression analysis by using quantitative RT-PCR assays indicated that AhLPAT was ubiquitously expressed in root, stem, leaf, flower, gynophore, seed of peanut with the highest level in gynophore and seed. The expression level reached a peak in the stage from 50 to 60 days after flowering. The

  6. Bioinformatics analysis of long chain acyl-coenzyme A synthetases homologous genes in Chlamydomonas reinhardtii%莱茵衣藻长链酰基辅酶A合成酶(LACS)同源基因的生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    顾守来; 马忠岩; 谭小力

    2012-01-01

    长链酰基辅酶A合成酶(LACS)能催化游离的脂肪酸形成酰基辅酶A硫脂,在油脂合成及降解途径中起着重要的作用.研究在莱茵衣藻(Chlamydomonas reinhardtii)中发现两个LACS基因,将其命名为CrLACS1和CrLACS2.生物信息学分析表明CrLACS1和CrLACS2在蛋白的理化性质及结构上都具有较高的相似性,而系统进化树分析显示CrLACS1和CrLACS2处于不同分枝,预测亚细胞定位也不相同.综合结果表明,CrLACS1和CrLACS2具有相似的结构,催化相同的反应,但具有不同的生物学功能.他们可能参与油脂代谢的不同途径:CrLACS1参与油脂的合成途径,而CrLACS2参与油脂的降解途径.%Long chain acyl-eoenzyme A synthetases (LACSs) activate free fatty acid to acyl-CoA thioesters, and play important roles in the biosynthesis and degradation of lipids. In this study, two LACS genes were found in Chlamydomonas reinhardtii and designated as CrLACSl and CrLACS2. CrLACSl and CrLACS2 were highly similar in physicochemical characters and structure using bioinformatics methods. But, the phylogenetic tree analysis showed that CrLACSl and CrLACS2 were in different clade, and subcellular localization analysis indicated that they were in different cellular organelles. Taken together, though CrLACSl and CrLACS2 catalyzed the same reaction as their similar structures, they participated in the different pathways. CrLACSl participated in the pathway of lipid biosynthesis , while CrLACS2 played a role in the degradation of lipid.

  7. The expression of lysophosphatidic acid, its receptors, and IL-6 and IL-8 in breast cancer%溶血磷脂酸及其受体和IL-6 IL-8在乳腺癌进展中的表达变化与意义

    Institute of Scientific and Technical Information of China (English)

    涂福平; 黄莉; 王祥财; 许明君; 王钇力; 衷敬华

    2013-01-01

    Objective:This work aimed to investigate the expression level of lysophosphatidic acid (LPA) and its receptors. The paper also discussed the interrelationship among the LPA, the receptors, and IL-6 and IL-8 in breast cancer tissues. Methods:The ex-pressions of the 3 hypo-types of LPA receptor in the breast cancer and paraneoplastic tissues were detected using semi-quantitative re-verse transcription polymerase chain reaction. The plasma levels of LPA, IL-6 and IL-8 were respectively detected in healthy subjects and in patients with benign breast tumor using the LPA biochemistry and enzyme linked immunosorbent assay kits. Results:The plas-ma LPA level was significantly higher in patients with breast cancer metastasis than in those with local breast cancer (P<0.01), benign breast tumor (P<0.01), and healthy volunteers (P<0.01). In addition, the IL-6 and IL-8 plasma levels were higher in the group with me-tastasis compared with the other three groups, too (P<0.01). LPA1 expression level was higher in breast cancer tissue than in benign breast tumor (P<0.05) and in normal breast tissue (P<0.05). There was a significantly positive correlation between the plasma LPA and the plasma IL-6 in patients with breast cancer (P<0.01), and between the plasma LPA and IL-8 (P<0.01). Conclusion:LPA expressions on the endogenous IL-6 and IL-8 in patients with breast cancer may have an up-regulation. Moreover, the detection of the LPA, IL-6, and IL-8 expression levels may have some predictable effects on metastatic breast cancer, especially bone metastases.%  目的:探讨溶血磷脂酸(lysophosphatidic acid,LPA)及其受体和IL-6与IL-8在乳腺癌进展中的表达及临床意义。方法:采用半定量RT-PCR方法检测乳腺肿瘤组织和瘤旁组织中LPA受体的表达水平。采用LPA生化测定法和酶联免疫吸附(ELISA)法分别检测乳腺肿瘤患者和健康妇女的血浆LPA、IL-6和IL-8水平。结果:术后复发转

  8. Angiomodulin is a specific marker of vasculature and regulates VEGF-A dependent neo-angiogenesis

    OpenAIRE

    Hooper, Andrea T.; Shmelkov, Sergey V.; Gupta, Sunny; Milde, Till; Bambino, Kathryn; Gillen, Kelly; Goetz, Mollie; Chavala, Sai; Baljevic, Muhamed; Murphy, Andrew J.; Valenzuela, David M; Gale, Nicholas W.; Thurston, Gavin; Yancopoulos, George D.; Vahdat, Linda

    2009-01-01

    Blood vessel formation is controlled by the balance between pro- and anti-angiogenic pathways. Although much is known about the factors that drive sprouting of neovessels, the factors that stabilize and pattern neovessels are undefined. The expression of angiomodulin (AGM), a VEGF-A binding protein, was increased in the vasculature of several human tumors as compared to normal tissue, raising the hypothesis that AGM may modulate VEGF-A-dependent vascular patterning. To elucidate the expressio...

  9. Relationship between levels of plasma lysophosphatidic acid, matrix metalloproteinase-9 and coronary stenosis%血浆溶血磷脂酸、基质金属蛋白酶-9水平与冠状动脉狭窄程度的相关性研究

    Institute of Scientific and Technical Information of China (English)

    杨波; 林琍; 宗文霞

    2011-01-01

    目的:观察冠心病患者血浆溶血磷脂酸(lysophosphatidic acid,LPA)及基质金属蛋白酶-9(matrix metalloproteinase-9,MMP-9)水平与冠状动脉病变的关系,探讨其在冠心病中的临床意义.方法:140例冠脉造影者根据病情及冠脉造影结果分为急性心肌梗死(AMI)组(n=40)、不稳定性心绞痛(UAP)组(n=35)、稳定性心绞痛(SAP)组(n=35)、对照组(n=30).用Gensini积分评定冠状动脉狭窄程度,根据评分四分位间距分组将患者分为4组:Ⅰ组(0~7分)35例,Ⅱ组(8~25分)36例,Ⅲ组(26~46分)26例及Ⅳ组(>46分)43例.分别用无机磷定量法和酶联免疫吸附法测定血浆LPA、MMP-9水平.结果:冠心病各组血浆LPA、 MMP-9水平及Genisi评分均显著高于对照组(P<0.01),AMI组高于UAP组及SAP组(P<0.01),UAP组高于SAP组(P<0.01).不同Genisi评分各组之间LPA、 MMP-9水平均差异有统计学意义(P<0.01).LPA与MMP-9水平呈正相关(r=0.22,P<0.05).结论:冠心病患者血浆LPA与MMP-9水平显著增高,且与冠心病严重程度及冠脉狭窄程度密切相关.%Objective: To investigate the relationship between levels of plasma lysophosphatidic acid ( LPA), matrix metalloproteinase-9(MMP-9) and the severity of coronary artery disease in patients with acute coronary heart disease (CHD) and to explore the potential clinical significance. Methods: One hundred and forty cases undergone coronary arteriography were divided into 4 groups according to the state of illness and results of coronary angiography: acute myocardial infarction (AMI) group( n= 40 ), unstable angina pectoris (UAP) group ( n=35 ), stable angina pectoris (SAP) group (n= 35) and control group(n=30 ). The degree of coronary artery stenosis was determined by Gensini's scores system, and the patients were redivided into 4 groups based on the interquartile of the Gensini's scores: group Ⅰ (0-7 scores,n= 35), group Ⅱ (8-25 scores,n= 36), group Ⅲ (26-46 scores,n= 26 ) and group Ⅳ (>46 scores

  10. PUMA promotes Bax translocation in FOXO3a-dependent pathway during STS-induced apoptosis

    Science.gov (United States)

    Zhang, Yingjie; Chen, Qun

    2009-08-01

    PUMA (p53 up-regulated modulator of apoptosis, also called Bbc3) was first identified as a BH3-only Bcl-2 family protein that is transcriptionally up-regulated by p53 and activated upon p53-dependent apoptotic stimuli, such as treatment with DNA-damaging drugs or UV irradiation. Recently studies have been shown that Puma is also up-regulated in response to certain p53-independent apoptotic stimuli, such as growth factor deprivation or treatment with glucocorticoids or STS (staurosporine). However, the molecular mechanisms of PUMA up-regulation and how PUMA functions in response to p53-independent apoptotic stimuli remain poorly understood. In this study, based on real-time single cell analysis, flow cytometry and western blotting technique, we investigated the function of PUMA in living human lung adenocarcinoma cells (ASTC-a-1) after STS treatment. Our results show that FOXO3a was activated by STS stimulation and then translocated from cytosol to nucleus. The expression of PUMA was up-regulated via a FOXO3a-dependent manner after STS treatment, while p53 had little function in this process. Moreover, cell apoptosis and Bax translocation induced by STS were not blocked by Pifithrin-α (p53 inhibitor), which suggested that p53 was not involved in this signaling pathway. Taken together, these results indicate that PUMA promoted Bax translocation in a FOXO3a-dependment pathway during STS-induced apoptosis, while p53 was dispensable in this process.

  11. 溶血磷脂酸调控RhoA/ROCK2信号通路对乳腺癌细胞增殖的影响%Influence of lysophosphatidic acid on proliferation of breast cancer cell by adjusted RhoA/ROCK2 signal pathway

    Institute of Scientific and Technical Information of China (English)

    许海; 段刚峰

    2013-01-01

    目的 探讨溶血磷脂酸(LPA)与RhoA/ROCK2信号通路对乳腺癌细胞增殖的影响及其作用机制.方法 以不同浓度LPA干预乳腺癌MDA-MB-231细胞,每隔24 h以细胞计数法观察和记录细胞的增殖.以最佳LPA促增殖浓度作用于MDA-MB-231细胞,观察Rho激酶抑制剂(Y-27632)对癌细胞的影响;以Pull-down及Western blot法检测各组细胞内RhoA活性及RhoA、ROCK2蛋白表达.结果 LPA以时间及剂量依赖性关系显著促进MDA-MB-231细胞的增殖(P<0.05);Y-27632可以显著抑制LPA的促增殖作用;LPA干预后RhoA活性及RhoA、ROCK2蛋白表达显著升高(P<0.05),Y-27632干预后RhoA活性及RhoA、ROCK2蛋白表达显著下降(P<0.05).结论 LPA可能通过调控RhoA/ROCK2信号通路促进乳腺癌细胞的增殖,为乳腺癌的临床治疗提供了新思路.%Objective To investigate the influence and mechanism of lysophosphatidic acid and RhoA/ROCK2 signal pathway on proliferation of breast cancer cell. Methods After treatment with different concentration of LPA, the proliferation of breast cancer cell MDA-MB-231 was observed and recorded by cell count method every of 24 h. MDA-MB-231 treated with optimal concentration of LPA and observed the effect of Rho kinase inhibitor( Y-27632) on LPA-induced proliferation. The activity of RhoA was tested by a pull-down way. The protein expression of RhoA and ROCK2 were determined by Western blot. Results LPA could promote MDA-MB-231 proliferation in a time and dose-dependent manner (P 〈 0. 05). ROCK inhibitor significantly inhibited LPA-induced cell proliferation (P 〈 0. 05 ). The activity of RhoA and expressionof RhoA, ROCK2 were enhanced significantly after LPA intervention (P 〈0. 05). However Y-27632 markedly decreased LPA-induced the increase of RhoA activity and protein expression of RhoA and ROCK2 ( P 〈 0. 05). Conclusions LPA may promote breast cancer cell proliferation through regulating RhoA/ROCK2 signal pathway. It provides a new idea

  12. $A$-dependence of coherent electroproduction of $\\rho^{0}$ mesons on nuclei in forward direction

    CERN Document Server

    Akopov, N; Aslanyan, G; Grigoryan, L

    2007-01-01

    This article presents the $A$-dependence of the differential cross section for the coherent electroproduction of vector mesons on nuclei in forward direction, at fixed values of longitudinal momentum transfer $q_{L}$. It is shown that such cross section has complicated behavior over the atomic mass number $A$ with local minimums and maximums. It is also shown that a ratio of the real to the imaginary parts of the forward coherent amplitude on nuclei $\\alpha_{A} = \\Re e{f_{A}} / \\Im m{f_{A}}$ has breaking points at some values of $A$. Comparison of the behaviors of the normalized cross section $\\Big(\\frac{d\\sigma}{d\\Omega}\\Big)_{A}\\Big/\\Big(\\frac{d\\sigma}{d\\Omega}\\Big)_{N}$ and $\\alpha_{A}$ over $A$ shows that the location of minimums of the cross section are very close to the breaking points of $\\alpha_{A}$.

  13. A-dependence of the Spectra of the F Isotopes from ab initio Calculations

    Science.gov (United States)

    Barrett, Bruce R.; Dikmen, Erdal; Maris, Pieter; Vary, James P.; Shirokov, Andrey M.

    2016-03-01

    Using a succession of Okubo-Lee-Suzuki transformations within the No Core Shell Model (NCSM) formalism, we derive an ab initio, non-perturbative procedure for calculating the input for standard shell-model (SSM) calculations within one major shell. We have used this approach for calculating the spectra of the F isotopes from A=18 to A=25, so as to study the A-dependence of the results. In particular, we are interested in seeing if the theoretical input is weak enough, so that a single set of two-body effective interactions can be used for all of the F isotopes investigated. We will present results from SSM calculations based on input obtained with the JISP16 nucleon-nucleon interaction in an initial 4 ℏΩ NCSM basis space. This work supported in part by TUBITAK-BIDEB, the US DOE, the US NSF, NERSC, and the Russian Ministry of Education and Science.

  14. Measurement of the $Z/A$ dependence of neutrino charged-current total cross-sections

    CERN Document Server

    Kayis-Topaksu, A; Van Dantzig, R; De Jong, M; Konijn, J; Melzer, O; Oldeman, R G C; Pesen, E; Van der Poel, C A F J; Spada, F R; Visschers, J L; Güler, M; Serin-Zeyrek, M; Kama, S; Sever, R; Tolun, P; Zeyrek, M T; Armenise, N; Catanesi, M G; De Serio, M; Ieva, M; Muciaccia, M T; Radicioni, E; Simone, S; Bülte, A; Winter, Klaus; El-Aidi, R; Van de Vyver, B; Vilian, P; Wilquet, G; Saitta, B; Di Capua, E; Ogawa, S; Shibuya, H; Artamonov, A V; Brunner, J; Chizhov, M; Cussans, D G; Doucet, M; Fabre, Jean-Paul; Hristova, I R; Kawamura, T; Kolev, D; Litmaath, M; Meinhard, H; Panman, J; Papadopoulos, I M; Ricciardi, S; Rozanov, A; Saltzberg, D; Tsenov, R V; Uiterwijk, J W E; Zucchelli, P; Goldberg, J; Chikawa, M; Arik, E; Song, J S; Yoon, C S; Kodama, K; Ushida, N; Aoki, S; Hara, T; Delbar, T; Favart, D; Grégoire, G; Kalinin, S; Makhlyoueva, I V; Gorbunov, P; Khovanskii, V D; Shamanov, V V; Tsukerman, I; Bruski, N; Frekers, D; Rondeshagen, D; Wolff, T; Hoshino, K; Kawada, J; Komatsu, M; Miyanishi, M; Nakamura, M; Nakano, T; Narita, K; Niu, K; Niwa, K; Nonaka, N; Sato, O; Toshito, T; Buontempo, S; Cocco, A G; D'Ambrosio, N; De Lellis, G; De Rosa, G; Di Capua, F; Ereditato, A; Fiorillo, G; Marotta, A; Messina, M; Migliozzi, P; Pistillo, C; Santorelli, R; Scotto-Lavina, L; Strolin, P; Tioukov, V; Nakamura, K; Okusawa, T; Dore, U; Loverre, P F; Ludovici, L; Maslennikov, A L; Righini, P; Rosa, G; Santacesaria, R; Satta, A; Barbuto, E; Bozza, C; Grella, G; Romano, G; Sirignano, C; Sorrentino, S; Sato, Y; Tezuka, I

    2003-01-01

    A relative measurement of total cross-sections is reported for polyethylene, marble, iron, and lead targets for the inclusive charged-current reaction nu_mu + N -> mu^- + X. The targets, passive blocks of ~100kg each, were exposed simultaneously to the CERN SPS wide-band muon-neutrino beam over a period of 18 weeks. Systematics effects due to differences in the neutrino flux and detector efficiency for the different target locations were minimised by changing the position of the four targets on their support about every two weeks. The relative neutrino fluxes on the targets were monitored within the same experiment using charged-current interactions in the calorimeter positioned directly downstream of the four targets. From a fit to the Z/A dependence of the total cross-sections a value is deduced for the effective neutron-to-proton cross-section ratio.

  15. Measurement of the Z/A dependence of neutrino charged-current total cross-sections

    CERN Document Server

    Kayis-Topasku, A; Dantzig, R V

    2003-01-01

    A relative measurement of total cross-sections is reported for polyethylene, marble, iron, and lead targets for the inclusive charged-current reaction nu submu + N -> mu sup - + X. The targets, passive blocks of propor to 100 kg each, were exposed simultaneously to the CERN SPS wide-band muon-neutrino beam over a period of 18 weeks. Systematic effects due to differences in the neutrino flux and detector efficiency for the different target locations were minimised by changing the position of the four targets on their support about every two weeks. The relative neutrino fluxes on the targets were monitored within the same experiment using charged-current interactions in the calorimeter positioned directly downstream of the four targets. From a fit to the Z/A dependence of the total cross-sections a value is deduced for the effective neutron-to-proton cross-section ratio. (orig.)

  16. Function of the N-terminal segment of the RecA-dependent nuclease Ref.

    Science.gov (United States)

    Gruber, Angela J; Olsen, Tayla M; Dvorak, Rachel H; Cox, Michael M

    2015-02-18

    The bacteriophage P1 Ref (recombination enhancement function) protein is a RecA-dependent, HNH endonuclease. It can be directed to create targeted double-strand breaks within a displacement loop formed by RecA. The 76 amino acid N-terminal region of Ref is positively charged (25/76 amino acid residues) and inherently unstructured in solution. Our investigation of N-terminal truncation variants shows this region is required for DNA binding, contains a Cys involved in incidental dimerization and is necessary for efficient Ref-mediated DNA cleavage. Specifically, Ref N-terminal truncation variants lacking between 21 and 47 amino acids are more effective RecA-mediated targeting nucleases. We propose a more refined set of options for the Ref-mediated cleavage mechanism, featuring the N-terminal region as an anchor for at least one of the DNA strand cleavage events.

  17. Metabolism of β-valine via a CoA-dependent ammonia lyase pathway.

    Science.gov (United States)

    Otzen, Marleen; Crismaru, Ciprian G; Postema, Christiaan P; Wijma, Hein J; Heberling, Matthew M; Szymanski, Wiktor; de Wildeman, Stefaan; Janssen, Dick B

    2015-11-01

    Pseudomonas species strain SBV1 can rapidly grow on medium containing β-valine as a sole nitrogen source. The tertiary amine feature of β-valine prevents direct deamination reactions catalyzed by aminotransferases, amino acid dehydrogenases, and amino acid oxidases. However, lyase- or aminomutase-mediated conversions would be possible. To identify enzymes involved in the degradation of β-valine, a PsSBV1 gene library was prepared and used to complement the β-valine growth deficiency of a closely related Pseudomonas strain. This resulted in the identification of a gene encoding β-valinyl-coenzyme A ligase (BvaA) and two genes encoding β-valinyl-CoA ammonia lyases (BvaB1 and BvaB2). The BvaA protein demonstrated high sequence identity to several known phenylacetate CoA ligases. Purified BvaA enzyme did not convert phenyl acetic acid but was able to activate β-valine in an adenosine triphosphate (ATP)- and CoA-dependent manner. The substrate range of the enzyme appears to be narrow, converting only β-valine and to a lesser extent, 3-aminobutyrate and β-alanine. Characterization of BvaB1 and BvaB2 revealed that both enzymes were able to deaminate β-valinyl-CoA to produce 3-methylcrotonyl-CoA, a common intermediate in the leucine degradation pathway. Interestingly, BvaB1 and BvaB2 demonstrated no significant sequence identity to known CoA-dependent ammonia lyases, suggesting they belong to a new family of enzymes. BLAST searches revealed that BvaB1 and BvaB2 show high sequence identity to each other and to several enoyl-CoA hydratases, a class of enzymes that catalyze a similar reaction with water instead of amine as the leaving group.

  18. RecA-dependent programmable endonuclease Ref cleaves DNA in two distinct steps.

    Science.gov (United States)

    Ronayne, Erin A; Cox, Michael M

    2014-04-01

    The bacteriophage P1 recombination enhancement function (Ref) protein is a RecA-dependent programmable endonuclease. Ref targets displacement loops formed when an oligonucleotide is bound by a RecA filament and invades homologous double-stranded DNA sequences. Mechanistic details of this reaction have been explored, revealing that (i) Ref is nickase, cleaving the two target strands of a displacement loop sequentially, (ii) the two strands are cleaved in a prescribed order, with the paired strand cut first and (iii) the two cleavage events have different requirements. Cutting the paired strand is rapid, does not require RecA-mediated ATP hydrolysis and is promoted even by Ref active site variant H153A. The displaced strand is cleaved much more slowly, requires RecA-mediated ATP hydrolysis and does not occur with Ref H153A. The two cleavage events are also affected differently by solution conditions. We postulate that the second cleavage (displaced strand) is limited by some activity of RecA protein.

  19. Nuclear Envelope Protein SUN2 Promotes Cyclophilin-A-Dependent Steps of HIV Replication

    Directory of Open Access Journals (Sweden)

    Xavier Lahaye

    2016-04-01

    Full Text Available During the early phase of replication, HIV reverse transcribes its RNA and crosses the nuclear envelope while escaping host antiviral defenses. The host factor Cyclophilin A (CypA is essential for these steps and binds the HIV capsid; however, the mechanism underlying this effect remains elusive. Here, we identify related capsid mutants in HIV-1, HIV-2, and SIVmac that are restricted by CypA. This antiviral restriction of mutated viruses is conserved across species and prevents nuclear import of the viral cDNA. Importantly, the inner nuclear envelope protein SUN2 is required for the antiviral activity of CypA. We show that wild-type HIV exploits SUN2 in primary CD4+ T cells as an essential host factor that is required for the positive effects of CypA on reverse transcription and infection. Altogether, these results establish essential CypA-dependent functions of SUN2 in HIV infection at the nuclear envelope.

  20. Arousal effect of orexin A depends on activation of the histaminergic system.

    Science.gov (United States)

    Huang, Z L; Qu, W M; Li, W D; Mochizuki, T; Eguchi, N; Watanabe, T; Urade, Y; Hayaishi, O

    2001-08-14

    Orexin neurons are exclusively localized in the lateral hypothalamic area and project their fibers to the entire central nervous system, including the histaminergic tuberomammillary nucleus (TMN). Dysfunction of the orexin system results in the sleep disorder narcolepsy, but the role of orexin in physiological sleep-wake regulation and the mechanisms involved remain to be elucidated. Here we provide several lines of evidence that orexin A induces wakefulness by means of the TMN and histamine H(1) receptor (H1R). Perfusion of orexin A (5 and 25 pmol/min) for 1 hr into the TMN of rats through a microdialysis probe promptly increased wakefulness for 2 hr after starting the perfusion by 2.5- and 4-fold, respectively, concomitant with a reduction in rapid eye movement (REM) and non-REM sleep. Microdialysis studies showed that application of orexin A to the TMN increased histamine release from both the medial preoptic area and the frontal cortex by approximately 2-fold over the baseline for 80 to 160 min in a dose-dependent manner. Furthermore, infusion of orexin A (1.5 pmol/min) for 6 hr into the lateral ventricle of mice produced a significant increase in wakefulness during the 8 hr after starting infusion to the same level as the wakefulness observed during the active period in wild-type mice, but not at all in H1R gene knockout mice. These findings strongly indicate that the arousal effect of orexin A depends on the activation of histaminergic neurotransmission mediated by H1R.

  1. Charge-dependent and A-dependent effects in isotope shifts of Coulomb displacement energies

    International Nuclear Information System (INIS)

    Coulomb displacement energies in a series of isotopes generally decrease with A. This decrease can arise from an increase with A of the average distance of interaction between pairs of protons. In the shell model a decrease can also result from charge-independence-breaking effects if the neutron-proton interaction for the valence nucleons is more attractive than the neutron-neutron interaction. Using the model recently proposed by Sherr and Talmi for the 1d/sub 3/2/ shell, existing data for this shell and also the 1d/sub 5/2/ and 1f/sub 7/2/ shells were analyzed allowing all matrix elements to vary as A/sup -lambda/3/. Least squares calculations of the rms deviation sigma were carried out for varying values of lambda from -2 to +2. It was found that although there was a minimum in sigma vs lambda it was too shallow to exclude any lambda for -1 to +1 in the 1d/sub 3/2/ and 1f/sub 7/2/ shells or 0 to +1 in the 1d/sub 5/2/ shell. It is therefore not possible to distinguish between A dependence and charge dependence in this model. The magnitude of the latter as expressed in terms of (np-nn) matrix elements depends strongly on the former. As lambda increases from -1 to +1, these (np-nn) matrix elements decrease roughly linearly in absolute magnitude and eventually change sign. For lambda = 0 they have appreciable and reasonable magnitudes for the 1d/sub 3/2/ and 1f/sub 7/2/ shells but for the 1d/sub 5/2/ shell the values are too small to be considered significant

  2. A-dependence of nuclear transparency in quasielastic A(e,e{prime}p) at high Q{sup 2}

    Energy Technology Data Exchange (ETDEWEB)

    O`Neill, T.G.; Lorenzon, W.; Arrington, J. [California Institute of Technology, Pasadena, CA (United States). W.K. Kellogg Radiation Lab.] [and others

    1994-05-01

    The A-dependence of the quasielastic A(e,e{prime}p) reaction has been studied with {sup 3}H, C, Fe, and Au nuclei at momentum transfers Q{sup 2} = 1, 3, 5, and 6.8(GeV/c){sup 2}. The authors extract the nuclear transparency T(A,Q{sup 2}), a measure of the average probability of escape of a proton from a nucleus A. Several calculations predict a significant increase in T with momentum transfer, a phenomenon known as color transparency. No statistically significant rise is seen for any of the nuclei studied.

  3. Group boundary permeability moderates the effect of a dependency meta-stereotype on help-seeking behaviour.

    Science.gov (United States)

    Zhang, Lange; Kou, Yu; Zhao, Yunlong; Fu, Xinyuan

    2016-08-01

    Previous studies have found that when low-status group members are aware that their in-group is stereotyped as dependent by a specific out-group (i.e. a dependency meta-stereotype is salient), they are reluctant to seek help from the high-status out-group to avoid confirming the negative meta-stereotype. However, it is unclear whether low-status group members would seek more help in the context of a salient dependency meta-stereotype when there is low (vs. high) group boundary permeability. Therefore, we conducted two experiments to examine the moderating effect of permeability on meta-stereotype confirmation with a real group. In study 1, we manipulated the salience of the dependency meta-stereotype, measured participants' perceived permeability and examined their help-seeking behaviour in a real-world task. Participants who perceived low permeability sought more help when the meta-stereotype was salient (vs. not salient), whereas participants who perceived high permeability sought the same amount of help across conditions. In study 2, we manipulated the permeability levels and measured the dependency meta-stereotype. Participants who endorsed a high-dependency meta-stereotype sought more help than participants who endorsed a low-dependency meta-stereotype; this effect was particularly strong in the low-permeability condition. The implications of these results for social mobility and intergroup helping are discussed. PMID:25885332

  4. A study of the A dependence of deep-inelastic scattering of leptons and its implications for understanding of the EMC effect

    CERN Document Server

    Smirnov, G I

    1995-01-01

    It is suggested to determine the A dependence of distortions of the nucleon structure function F_2(x) by summing the distortions over an interval (x_1, x_2). It was found from the analysis of data on deep--inelastic scattering of muons and electrons from nuclei that the A dependence of distortion magnitudes obtained in each of three regions under study, namely shadowing, antishadowing and the EMC effect region, follow the same functional form, being different in the normalizing factor only. All the available data give evidence for the saturation of the distortion magnitude with rising A.

  5. Acyl-CoA metabolism and partitioning

    DEFF Research Database (Denmark)

    Grevengoed, Trisha J; Klett, Eric L; Coleman, Rosalind A

    2014-01-01

    Long-chain fatty acyl-coenzyme As (CoAs) are critical regulatory molecules and metabolic intermediates. The initial step in their synthesis is the activation of fatty acids by one of 13 long-chain acyl-CoA synthetase isoforms. These isoforms are regulated independently and have different tissue e...

  6. Neuropsychological Outcomes in Fatty Acid Oxidation Disorders: 85 Cases Detected by Newborn Screening

    Science.gov (United States)

    Waisbren, Susan E.; Landau, Yuval; Wilson, Jenna; Vockley, Jerry

    2013-01-01

    Mitochondrial fatty acid oxidation disorders include conditions in which the transport of activated acyl-Coenzyme A (CoA) into the mitochondria or utilization of these substrates is disrupted or blocked. This results in a deficit in the conversion of fat into energy. Most patients with fatty acid oxidation defects are now identified through…

  7. Safe and unsafe duration of fasting for children with MCAD deficiency

    NARCIS (Netherlands)

    Derks, Terry G J; van Spronsen, Francjan J; Rake, Jan Peter; van der Hilst, Christian S; Span, Mark M; Smit, G Peter A

    2007-01-01

    OBJECTIVE: To study the safe and unsafe duration of fasting in children with medium chain acyl-Coenzyme A dehydrogenase (MCAD) deficiency, the literature and the database on Dutch MCAD-deficient patients were searched for data on fasting studies in patients with MCAD deficiency. MATERIALS AND METHOD

  8. Definition of a Dependent Child

    CERN Multimedia

    Human Resources Department

    2005-01-01

    The Department of Human Resources wishes to remind members of the personnel that, under the provisions of § 6 of Administrative Circular No. 5 “Dependent child”, in the case of a child over 18 years of age the status of dependent child comes to an end once a course of studies is completed. Consequently, the payment of the dependent child allowance and the child's membership of the CERN Health Insurance Scheme terminate with effect from the last day of the month in which the course of study concerned ends. In this connection, members of the personnel are reminded that children who are no longer dependent according to the Staff Rules and Regulations and who are less than 26 years of age can nevertheless opt for membership of the normal health insurance under the terms and conditions laid down in the CERN Health Insurance Rules. The Department of Human Resources also wishes to remind members of the personnel that, pursuant to Article R IV 1.17 of the Staff Regulations, a member of the personnel is requ...

  9. A Dependency Parser for Tweets

    OpenAIRE

    Kong, Lingpeng; Schneider, Nathan; Swayamdipta, Swabha; Bhatia, Archna; Dyer, Chris; Smith, Noah A.

    2014-01-01

    We describe a new dependency parser for English tweets, TWEEBOPARSER. The parser builds on several contributions: new syntactic annotations for a corpus of tweets (TWEEBANK), with conventions informed by the domain; adaptations to a statistical parsing algorithm; and a new approach to exploiting out-of-domain Penn Treebank data. Our experiments show that the parser achieves over 80% unlabeled attachment accuracy on our new, high-quality test set and measure the benefit of our contributions.Ou...

  10. Lysophosphatidic acid (LPA) signaling in neuropathic pain development and Schwann cell biology

    OpenAIRE

    Lin, Mu-En

    2012-01-01

    Neuropathic pain is a chonic pain state caused by lesions or diseases in the nervous system. Unlike acute pain, neuropathic pain persists without obvious injury or stimuli and can severely interfere with normal daily life for those who suffer from it. Despite numerous efforts on studying its mechanism and possible treatments, there is no effective treatment currently available to remove or alleviate this symptom. This dissertation aims to provide further understanding into the relationship be...

  11. Cell Shrinkage is Essential in Lysophosphatidic Acid Signaling in Ehrlich Ascites

    DEFF Research Database (Denmark)

    Pedersen, Susanne; Hoffmann, Else Kay; Hougaard, Charlotte;

    2000-01-01

    ; (ii) a subsequent cell shrinkage and increased polymerization of F-actin, and (iii) activation of a Na(+)/H(+) exchange, resulting in a concentration-dependent intracellular alkalinization. The EC(50) value for the LPA-induced rate of alkalinization was estimated at 0. 37 nm LPA. When cell shrinkage...

  12. Lysophosphatidic acid activates Arf6 to promote the mesenchymal malignancy of renal cancer

    OpenAIRE

    Hashimoto, Shigeru; Mikami, Shuji; Sugino, Hirokazu; Yoshikawa, Ayumu; Hashimoto, Ari; Onodera, Yasuhito; Furukawa, Shotaro; Handa, Haruka; Oikawa, Tsukasa; OKADA, YASUNORI; Oya, Mototsugu; Sabe, Hisataka

    2016-01-01

    Acquisition of mesenchymal properties by cancer cells is critical for their malignant behaviour, but regulators of the mesenchymal molecular machinery and how it is activated remain elusive. Here we show that clear cell renal cell carcinomas (ccRCCs) frequently utilize the Arf6-based mesenchymal pathway to promote invasion and metastasis, similar to breast cancers. In breast cancer cells, ligand-activated receptor tyrosine kinases employ GEP100 to activate Arf6, which then recruits AMAP1; and...

  13. L-histidine inhibits production of lysophosphatidic acid by the tumor-associated cytokine, autotaxin

    Directory of Open Access Journals (Sweden)

    Schiffmann Elliott

    2005-02-01

    Full Text Available Abstract Background Autotaxin (ATX, NPP-2, originally purified as a potent tumor cell motility factor, is now known to be the long-sought plasma lysophospholipase D (LPLD. The integrity of the enzymatic active site, including three crucial histidine moieties, is required for motility stimulation, as well as LPLD and 5'nucleotide phosphodiesterase (PDE activities. Except for relatively non-specific chelation agents, there are no known inhibitors of the ATX LPLD activity. Results We show that millimolar concentrations of L-histidine inhibit ATX-stimulated but not LPA-stimulated motility in two tumor cell lines, as well as inhibiting enzymatic activities. Inhibition is reversed by 20-fold lower concentrations of zinc salt. L-histidine has no significant effect on the Km of LPLD, but reduces the Vmax by greater than 50%, acting as a non-competitive inhibitor. Several histidine analogs also inhibit the LPLD activity of ATX; however, none has greater potency than L-histidine and all decrease cell viability or adhesion. Conclusion L-histidine inhibition of LPLD is not a simple stoichiometric chelation of metal ions but is more likely a complex interaction with a variety of moieties, including the metal cation, at or near the active site. The inhibitory effect of L-histidine requires all three major functional groups of histidine: the alpha amino group, the alpha carboxyl group, and the metal-binding imidazole side chain. Because of LPA's involvement in pathological processes, regulation of its formation by ATX may give insight into possible novel therapeutic approaches.

  14. A liver-specific defect of Acyl-CoA degradation produces hyperammonemia, hypoglycemia and a distinct hepatic Acyl-CoA pattern.

    Directory of Open Access Journals (Sweden)

    Nicolas Gauthier

    Full Text Available Most conditions detected by expanded newborn screening result from deficiency of one of the enzymes that degrade acyl-coenzyme A (CoA esters in mitochondria. The role of acyl-CoAs in the pathophysiology of these disorders is poorly understood, in part because CoA esters are intracellular and samples are not generally available from human patients. We created a mouse model of one such condition, deficiency of 3-hydroxy-3-methylglutaryl-CoA lyase (HL, in liver (HLLKO mice. HL catalyses a reaction of ketone body synthesis and of leucine degradation. Chronic HL deficiency and acute crises each produced distinct abnormal liver acyl-CoA patterns, which would not be predictable from levels of urine organic acids and plasma acylcarnitines. In HLLKO hepatocytes, ketogenesis was undetectable. Carboxylation of [2-(14C] pyruvate diminished following incubation of HLLKO hepatocytes with the leucine metabolite 2-ketoisocaproate (KIC. HLLKO mice also had suppression of the normal hyperglycemic response to a systemic pyruvate load, a measure of gluconeogenesis. Hyperammonemia and hypoglycemia, cardinal features of many inborn errors of acyl-CoA metabolism, occurred spontaneously in some HLLKO mice and were inducible by administering KIC. KIC loading also increased levels of several leucine-related acyl-CoAs and reduced acetyl-CoA levels. Ultrastructurally, hepatocyte mitochondria of KIC-treated HLLKO mice show marked swelling. KIC-induced hyperammonemia improved following administration of carglumate (N-carbamyl-L-glutamic acid, which substitutes for the product of an acetyl-CoA-dependent reaction essential for urea cycle function, demonstrating an acyl-CoA-related mechanism for this complication.

  15. Creating directed double-strand breaks with the Ref protein: a novel RecA-dependent nuclease from bacteriophage P1.

    Science.gov (United States)

    Gruenig, Marielle C; Lu, Duo; Won, Sang Joon; Dulberger, Charles L; Manlick, Angela J; Keck, James L; Cox, Michael M

    2011-03-11

    The bacteriophage P1-encoded Ref protein enhances RecA-dependent recombination in vivo by an unknown mechanism. We demonstrate that Ref is a new type of enzyme; that is, a RecA-dependent nuclease. Ref binds to ss- and dsDNA but does not cleave any DNA substrate until RecA protein and ATP are added to form RecA nucleoprotein filaments. Ref cleaves only where RecA protein is bound. RecA functions as a co-nuclease in the Ref/RecA system. Ref nuclease activity can be limited to the targeted strands of short RecA-containing D-loops. The result is a uniquely programmable endonuclease activity, producing targeted double-strand breaks at any chosen DNA sequence in an oligonucleotide-directed fashion. We present evidence indicating that cleavage occurs in the RecA filament groove. The structure of the Ref protein has been determined to 1.4 Å resolution. The core structure, consisting of residues 77-186, consists of a central 2-stranded β-hairpin that is sandwiched between several α-helical and extended loop elements. The N-terminal 76 amino acid residues are disordered; this flexible region is required for optimal activity. The overall structure of Ref, including several putative active site histidine residues, defines a new subclass of HNH-family nucleases. We propose that enhancement of recombination by Ref reflects the introduction of directed, recombinogenic double-strand breaks.

  16. Creating Directed Double-strand Breaks with the Ref Protein: A Novel Rec A-Dependent Nuclease from Bacteriophage P1

    Energy Technology Data Exchange (ETDEWEB)

    Gruenig, Marielle C.; Lu, Duo; Won, Sang Joon; Dulberger, Charles L.; Manlick, Angela J.; Keck, James L.; Cox, Michael M. (UW)

    2012-03-16

    The bacteriophage P1-encoded Ref protein enhances RecA-dependent recombination in vivo by an unknown mechanism. We demonstrate that Ref is a new type of enzyme; that is, a RecA-dependent nuclease. Ref binds to ss- and dsDNA but does not cleave any DNA substrate until RecA protein and ATP are added to form RecA nucleoprotein filaments. Ref cleaves only where RecA protein is bound. RecA functions as a co-nuclease in the Ref/RecA system. Ref nuclease activity can be limited to the targeted strands of short RecA-containing D-loops. The result is a uniquely programmable endonuclease activity, producing targeted double-strand breaks at any chosen DNA sequence in an oligonucleotide-directed fashion. We present evidence indicating that cleavage occurs in the RecA filament groove. The structure of the Ref protein has been determined to 1.4 {angstrom} resolution. The core structure, consisting of residues 77-186, consists of a central 2-stranded {beta}-hairpin that is sandwiched between several {alpha}-helical and extended loop elements. The N-terminal 76 amino acid residues are disordered; this flexible region is required for optimal activity. The overall structure of Ref, including several putative active site histidine residues, defines a new subclass of HNH-family nucleases. We propose that enhancement of recombination by Ref reflects the introduction of directed, recombinogenic double-strand breaks.

  17. A Dependence Study of $\\Xi^{*0}$ and $\\bar{\\Xi}^{*0}$ in 250 GeV/c $\\pi^-$. $K^-$ -nucleon Interactions

    Energy Technology Data Exchange (ETDEWEB)

    Seixas de Rezende, Fabio Antonio; /Rio de Janeiro, CBPF

    2005-04-01

    A direct measurement of the mass number (A) dependence of the production of the hyperon {Xi}*{sup 0} and its opposite {bar {Xi}}*{sup 0} in {pi}{sup -}, K{sup -} beam-nucleon interactions at 250 GeV/c is reported. The data derive from the experiment E769 at Fermilab. The results were obtained for different targets: Be, Al, Cu and W. It was observed the data are found to be well described by the parametrization {sigma}{sub A} = {sigma}{sub 0}A{sup {alpha}}, {alpha} being calculated for different beams. The results obtained are compared with those results of E769 experiment. The results shown here are preliminary.

  18. A dual role for mycobacterial RecO in RecA-dependent homologous recombination and RecA-independent single-strand annealing.

    Science.gov (United States)

    Gupta, Richa; Ryzhikov, Mikhail; Koroleva, Olga; Unciuleac, Mihaela; Shuman, Stewart; Korolev, Sergey; Glickman, Michael S

    2013-02-01

    Mycobacteria have two genetically distinct pathways for the homology-directed repair of DNA double-strand breaks: homologous recombination (HR) and single-strand annealing (SSA). HR is abolished by deletion of RecA and reduced in the absence of the AdnAB helicase/nuclease. By contrast, SSA is RecA-independent and requires RecBCD. Here we examine the function of RecO in mycobacterial DNA recombination and repair. Loss of RecO elicits hypersensitivity to DNA damaging agents similar to that caused by deletion of RecA. We show that RecO participates in RecA-dependent HR in a pathway parallel to the AdnAB pathway. We also find that RecO plays a role in the RecA-independent SSA pathway. The mycobacterial RecO protein displays a zinc-dependent DNA binding activity in vitro and accelerates the annealing of SSB-coated single-stranded DNA. These findings establish a role for RecO in two pathways of mycobacterial DNA double-strand break repair and suggest an in vivo function for the DNA annealing activity of RecO proteins, thereby underscoring their similarity to eukaryal Rad52. PMID:23295671

  19. Negociando a dependência: relações militares Brasil-Estados Unidos no início da Guerra Fria

    Directory of Open Access Journals (Sweden)

    Eduardo Svartman

    2014-12-01

    Full Text Available Durante a Guerra Fria, os laços estabelecidos entre as forças armadas dos Estados Unidos e os militares de sues aliados em diferentes regiões do mundo foi elemento importante tanto da estratégia anticomunista de Washington quanto da projeção deste país em áreas até então influenciadas pelas potências europeias. O artigo argumenta que o Brasil foi um caso ilustrativo deste processo, no qual apesar da profunda dependência e alinhamento, essas relações militares não foram lineares nem desprovidas de conflitos. O caráter instrumental conferido pelo Brasil a essas relações e a coincidência apenas parcial de objetivos entre os dois países fez com que, em determinadas ocasiões, os militares brasileiros procurassem renegociar a dependência. O impacto político dessa interação proporcionou a criação de espaços de formulação ideológica e de articulação política, polarizou a oficialidade em torno de temas como exploração de petróleo e participação na Guerra da Coreia e reforçou disposições para o protagonismo na política interna.

  20. Unconventional secretion by autophagosome exocytosis

    OpenAIRE

    Pfeffer, Suzanne R

    2010-01-01

    In this issue, Duran et al. (2010. J. Cell Biol. doi: 10.1083/jcb.200911154) and Manjithaya et al. (2010. J. Cell Biol. doi: 10.1083/jcb.200911149) use yeast genetics to reveal a role for autophagosome intermediates in the unconventional secretion of an acyl coenzyme A (CoA)–binding protein that lacks an endoplasmic reticulum signal sequence. Medium-chain acyl CoAs are also required and may be important for substrate routing to this pathway.

  1. Rimonabant is a dual inhibitor of acyl CoA:cholesterol acyltransferases 1 and 2

    OpenAIRE

    Netherland, Courtney; Thewke, Douglas P.

    2010-01-01

    Acyl-coenzymeA:cholesterol acyltransferase (ACAT) catalyzes the intracellular synthesis of cholesteryl esters (CE). Both ACAT isoforms, ACAT1 and ACAT2, play key roles in the pathophysiology of atherosclerosis and ACAT inhibition retards atherosclerosis in animal models. Rimonabant, a type 1 cannabinoid receptor (CB1) antagonist, produces anti-atherosclerotic effects in humans and animals by mechanisms which are not completely understood. Rimonabant is structurally similar to two other cannab...

  2. Lysophosphatidic acid inhibition of the accumulation of Pseudomonas aeruginosa PAO1 alginate, pyoverdin, elastase and LasA

    DEFF Research Database (Denmark)

    Laux, D.C.; Corson, J.M.; Givskov, Michael Christian;

    2002-01-01

    The pathogenesis of Pseudomonas aeruginosa is at least partially attributable to its ability to synthesize and secrete the siderophore pyoverdin and the two zinc metal loproteases elastase and LasA, and its ability to form biofilms in which bacterial cells are embedded in an alginate matrix....... In the present study, a lysophospholipid, 1-paimitoyl-2-hydroxy-sn-glycero-3-phosphate [also called monopalmitoylphosphatidic acid (MPPA)], which accumulates in inflammatory exudates, was shown to inhibit the extracellular accumulation of P. aeruginosa PAO1 alginate, elastase, LasA protease and the siderophore...

  3. [Is psoriasis a dependent cardiovascular risk factor?

    NARCIS (Netherlands)

    Wakkee, M.; Jong, E.M. de

    2013-01-01

    Psoriasis is a chronic inflammatory skin disease that affects approximately 2% of the Dutch population. It has been hypothesized that chronic inflammation occurring in psoriasis patients is more than skin deep, resulting in increased cardiovascular risk. Some observational studies have confirmed thi

  4. The A dependence of dilepton production

    International Nuclear Information System (INIS)

    The discovery in 1982, by the EMC group, that the structure function F2(x) per nucleon is different in iron than deuterium was the first evidence that the structure of the nucleon might be altered in nuclei. Much more extensive and precise and data have been taken since the original discovery. This body of data is now quite consistent and many of the features of the original ''EMC effect'' remain intact. There have been a variety of theoretical explanations of the data, but none are totally compelling. The deep-inelastic-scattering (DIS) data for the ratio of F2Fe(x) in iron to F2D(x) in deuterium are given for the region 0.03 ≤ x ≤ ∼ 1.0

  5. 3-Methylcholanthrene, an AhR agonist, caused cell-cycle arrest by histone deacetylation through a RhoA-dependent recruitment of HDAC1 and pRb2 to E2F1 complex.

    Directory of Open Access Journals (Sweden)

    Chih-Cheng Chang

    Full Text Available We previously showed that treating vascular endothelial cells with 3-methylcholanthrene (3MC caused cell-cycle arrest in the Go/G1 phase; this resulted from the induction of p21 and p27 and a decreased level and activity of the cyclin-dependent kinase, Cdk2. We further investigated the molecular mechanisms that modulate cell-cycle regulatory proteins through the aryl-hydrocarbon receptor (AhR/Ras homolog gene family, member A (RhoA dependent epigenetic modification of histone. AhR/RhoA activation mediated by 3MC was essential for the upregulation of retinoblastoma 2 (pRb2 and histone deacetylase 1 (HDAC1, whereas their nuclear translocation was primarily modulated by RhoA activation. The combination of increased phosphatase and tensin homolog (PTEN activity and decreased phosphatidylinositide 3-kinase (PI3K activation by 3MC led to the inactivation of the Ras-cRaf pathway, which contributed to pRb2 hypophosphorylation. Increased HDAC1/pRb2 recruitment to the E2F1 complex decreased E2F1-transactivational activity and H3/H4 deacetylation, resulting in the downregulation of cell-cycle regulatory proteins (Cdk2/4 and Cyclin D3/E. Co-immunoprecipitation and electrophoretic mobility shift assay (EMSA results showed that simvastatin prevented the 3MC-increased binding activities of E2F1 proteins in their promoter regions. Additionally, RhoA inhibitors (statins reversed the effect of 3MC in inhibiting DNA synthesis by decreasing the nuclear translocation of pRb2/HDAC1, leading to a recovery of the levels of cell-cycle regulatory proteins. In summary, 3MC decreased cell proliferation by the epigenetic modification of histone through an AhR/RhoA-dependent mechanism that can be rescued by statins.

  6. A Influência da Confiança, Dependência e Comprometimento na Orientação de Longo Prazo de Varejistas para com os Fabricantes Líderes do Mercado de Bebidas

    Directory of Open Access Journals (Sweden)

    Ivan Lapuente Garrido

    2013-09-01

    Full Text Available O objetivo principal deste estudo é analisar como a confiança, o comprometimento e a dependência influenciam a orientação de longo prazo de varejistas para com o relacionamento com as empresas fabricantes de cervejas e refrigerantes líderes no mercado nacional. Para atingir o objetivo proposto, fizeram-se necessárias duas etapas de pesquisa. Na primeira etapa de cunho exploratório, foram realizadas entrevistas em profundidade com varejistas que possuíam relacionamento comercial com as empresas líderes do mercado nacional de cervejas e refrigerantes; esta etapa possibilitou aprofundamento sobre o campo de estudo e adequação de escalas aplicadas a segunda etapa de pesquisa. Na segunda etapa de pesquisa, por meio de questionário composto por questões fechadas, 299 varejistas avaliaram seu relacionamento com os fabricantes líderes de mercado. Utilizando-se de regressão múltipla, os resultados indicam que: (a a dependência dos varejistas influencia positivamente o comprometimento para com os fabricantes, (b a confiança influencia positivamente a orientação de longo prazo dos varejistas para com os fabricantes e, (c o comprometimento influencia positivamente a orientação de longo prazo dos varejistas para com os fabricantes. As hipóteses que consideravam a relação entre confiança e comprometimento e entre dependência e orientação de longo prazo não obtiveram significância estatística. DOI: 10.5585/remark.v12i3.2328

  7. Very long-chain acyl CoA dehydrogenase deficiency which was accepted as infanticide.

    Science.gov (United States)

    Eminoglu, Tuba F; Tumer, Leyla; Okur, Ilyas; Ezgu, Fatih S; Biberoglu, Gursel; Hasanoglu, Alev

    2011-07-15

    Very-long-chain acyl-coenzyme A (CoA) dehydrogenase deficiency (VLCADD) (OMIM #201475) is an autosomal recessive disorder of fatty acid oxidation. Major phenotypic expressions are hypoketotic hypoglycemia, hepatomegaly, cardiomyopathy, myopathy, rhabdomyolysis, elevated creatinine kinase, and lipid infiltration of liver and muscle. At the same time, it is a rare cause of Sudden Infant Death Syndrome (SIDS) or unexplained death in the neonatal period [1-4]. We report a patient with VLCADD whose parents were investigated for infanticide because her three previous siblings had suddenly died after normal deliveries.

  8. The nicotine dependence associated with alcohol use and other psychoactive substance A dependência da nicotina associada ao uso de álcool e outras substâncias psicoativas

    Directory of Open Access Journals (Sweden)

    Rafaela Serra Bacchi

    2008-10-01

    Full Text Available To examine an association between nicotine dependence with alcohol, other psychoactive use, and depressive disorder. Smokers were recruited from Centro de Referência de Abordagem e Tratamento do Tabagismo at the Hospital das Clínicas da Universidade Estadual de Londrina (AHC/ UEL. All subjects were informed and gave then written consent for the research as approved by the Ethics Research Committee of Universidade Estadual de Londrina. The measures used were: structured questionnaire, alcohol, smoking, and psychoactive substance involvement screening test (ASSIST v 3.0, the Fagerström test for Nicotine Dependence (FTND, and the Diagnostic Interview for Research on Depressive disorder of the World Health Organization. Smokers presented the following socio-demographic characteristics: prevalence of the female sex and mean age of 47 years old with capacity for domestic activities and work. The mean age of onset of cigarette use for smokers was 16 years old. Fagerström’s test presented a medium punctuation of 6, so much for users of substances psicoativas, as for the ones that they don’t use them. Relationship between serious depression and the of psychoactive substances use was relevant for the research. This study evidenced an association among the use of the tobacco and other psychoactive substances, and depressive disorder. The health professional in smoking cessation intervention would be to identify subgroups of adult smokers, associated with depression, psychoactive substance use, and promote an intervention in both comorbidities and larger effectiveness of the smoking cessation. Analisar a associação entre a dependência de nicotina com o uso de álcool, outras substâncias psicoativas e transtorno depressivo. Os tabagistas foram recrutados a partir do Centro de Referência de Abordagem e Tratamento do Tabagismo no Hospital de Clínicas da Universidade Estadual de Londrina (AHC/UEL. Todos os participantes foram informados e

  9. The Effect of Lysophosphatidic Acid during In Vitro Maturation of Bovine Oocytes: Embryonic Development and mRNA Abundances of Genes Involved in Apoptosis and Oocyte Competence

    OpenAIRE

    Dorota Boruszewska; Ana Catarina Torres; Ilona Kowalczyk-Zieba; Patricia Diniz; Mariana Batista; Luis Lopes-da-Costa; Izabela Woclawek-Potocka

    2014-01-01

    In the present study we examined whether LPA can be synthesized and act during in vitro maturation of bovine cumulus oocyte complexes (COCs). We found transcription of genes coding for enzymes of LPA synthesis pathway (ATX and PLA2) and of LPA receptors (LPAR 1–4) in bovine oocytes and cumulus cells, following in vitro maturation. COCs were matured in vitro in presence or absence of LPA (10−5 M) for 24 h. Supplementation of maturation medium with LPA increased mRNA abundance of FST and GDF9 i...

  10. Dual Action of Lysophosphatidate-Functionalised Titanium: Interactions with Human (MG63) Osteoblasts and Methicillin Resistant Staphylococcus aureus

    DEFF Research Database (Denmark)

    Skindersø, Mette Elena; Krogfelt, Karen Angeliki; Blom, Ashley;

    2015-01-01

    Titanium (Ti) is a widely used material for surgical implants; total joint replacements (TJRs), screws and plates for fixing bones and dental implants are forged from Ti. Whilst Ti integrates well into host tissue approximately 10% of TJRs will fail in the lifetime of the patient through a process...... known as aseptic loosening. These failures necessitate revision arthroplasties which are more complicated and costly than the initial procedure. Finding ways of enhancing early (osseo)integration of TJRs is therefore highly desirable and continues to represent a research priority in current biomaterial...

  11. The multigene family of lysophosphatidate acyltransferase (LPAT)-related enzymes in Ricinus communis: cloning and molecular characterization of two LPAT genes that are expressed in castor seeds.

    Science.gov (United States)

    Arroyo-Caro, José María; Chileh, Tarik; Kazachkov, Michael; Zou, Jitao; Alonso, Diego López; García-Maroto, Federico

    2013-02-01

    The multigene family encoding proteins related to lysophosphatidyl-acyltransferases (LPATs) has been analyzed in the castor plant Ricinus communis. Among them, two genes designated RcLPAT2 and RcLPATB, encoding proteins with LPAT activity and expressed in the developing seed, have been cloned and characterized in some detail. RcLPAT2 groups with well characterized members of the so-called A-class LPATs and it shows a generalized expression pattern in the plant and along seed development. Enzymatic assays of RcLPAT2 indicate a preference for ricinoleoyl-CoA over other fatty acid thioesters when ricinoleoyl-LPA is used as the acyl acceptor, while oleoyl-CoA is the preferred substrate when oleoyl-LPA is employed. RcLPATB groups with B-class LPAT enzymes described as seed specific and selective for unusual fatty acids. However, RcLPATB exhibit a broad specificity on the acyl-CoAs, with saturated fatty acids (12:0-16:0) being the preferred substrates. RcLPATB is upregulated coinciding with seed triacylglycerol accumulation, but its expression is not restricted to the seed. These results are discussed in the light of a possible role for LPAT isoenzymes in the channelling of ricinoleic acid into castor bean triacylglycerol.

  12. Apicoplast-Localized Lysophosphatidic Acid Precursor Assembly Is Required for Bulk Phospholipid Synthesis in Toxoplasma gondii and Relies on an Algal/Plant-Like Glycerol 3-Phosphate Acyltransferase.

    Science.gov (United States)

    Amiar, Souad; MacRae, James I; Callahan, Damien L; Dubois, David; van Dooren, Giel G; Shears, Melanie J; Cesbron-Delauw, Marie-France; Maréchal, Eric; McConville, Malcolm J; McFadden, Geoffrey I; Yamaryo-Botté, Yoshiki; Botté, Cyrille Y

    2016-08-01

    Most apicomplexan parasites possess a non-photosynthetic plastid (the apicoplast), which harbors enzymes for a number of metabolic pathways, including a prokaryotic type II fatty acid synthesis (FASII) pathway. In Toxoplasma gondii, the causative agent of toxoplasmosis, the FASII pathway is essential for parasite growth and infectivity. However, little is known about the fate of fatty acids synthesized by FASII. In this study, we have investigated the function of a plant-like glycerol 3-phosphate acyltransferase (TgATS1) that localizes to the T. gondii apicoplast. Knock-down of TgATS1 resulted in significantly reduced incorporation of FASII-synthesized fatty acids into phosphatidic acid and downstream phospholipids and a severe defect in intracellular parasite replication and survival. Lipidomic analysis demonstrated that lipid precursors are made in, and exported from, the apicoplast for de novo biosynthesis of bulk phospholipids. This study reveals that the apicoplast-located FASII and ATS1, which are primarily used to generate plastid galactolipids in plants and algae, instead generate bulk phospholipids for membrane biogenesis in T. gondii. PMID:27490259

  13. Measuring the impact of a dependence among insured life lengths

    NARCIS (Netherlands)

    M. Denuit; J. Dhaene; C. Le Bailly De Tilleghem; S. Teghem

    2001-01-01

    Actuaries usually compute multiple life premiums based on the unrealistic assumption of independence of the lifelengths of insured persons. Many clinical studies, however, have demonstrated dependence of the lifetimes of paired lives such as husband and wife. In this respect, the present article tri

  14. Design of a dependable Interlock System for linear colliders

    CERN Document Server

    Nouvel, Patrice

    For high energy accelerators, the interlock system is a key part of the machine protection. The interlock principle is to inhibit the beam either on failure of critical equipment and/or on low beam quality evaluation. The dependability of such a system is the most critical parameter. This thesis presents the design of an dependable interlock system for linear collider with an application to the CLIC (Compact Linear Collider) project. This design process is based on the IEEE 1220 standard and is is divided in four steps. First, the specifications are established, with a focus on the dependability, more precisely the reliability and the availability of the system. The second step is the design proposal based on a functional analysis, the CLIC and interfaced systems architecture. Third, the feasibility study is performed, applying the concepts in an accelerator facility. Finally, the last step is the hardware verification. Its aim is to prove that the proposed design is able to reach the requirements.

  15. Treatment of a dependent geriatric pacient in domestic environment

    OpenAIRE

    ŠULISTOVÁ, Veronika

    2016-01-01

    The issue of the treatment of geriatric patients in a home environment is due to the more current demographic trends. The treatment of these patients is not easy and brings a number of problematic situations. Therefore, it is necessary to think, what the quality of care is in a home environment and how the family of geriatric patients on changes in his health will adapt. It is also important the education and the support of the lay family caregivers in the theoretical and practical knowledge ...

  16. The Avalon Beowulf Cluster: A Dependable Tool for Scientific Simulation

    Science.gov (United States)

    Warren, Michael

    2000-03-01

    Avalon is a 140 processor Alpha/Linux Beowulf cluster constructed entirely from commodity personal computer technology and freely available software. Computational Physics simulations performed on Avalon resulted in the award of a 1998 Gordon Bell price/performance prize for significant achievement in parallel processing. Avalon ranked as the 113th fastest computer in the world on the November 1998 TOP500 list, obtaining a result of 48.6 Gigaflops on the parallel Linpack benchmark. The price of hardware and final assembly labor for Avalon totalled 313,000 dollars in the fall of 1998. Avalon currently provides over 15,000 node-hours of production computing time per week, split among about 10 production users. Obtaining an equivalent amount of computing through Los Alamos institutional sources would cost a minimicrons of 30,000 per week. The machine also supports code development for another 60 users. Significant simulations have been performed on Avalon in fields of astrophysics, molecular dynamics, nonlinear dynamics as well as other areas. The largest single simulation performed on Avalon computed a total of over 10^16 floating point operations. We will describe some of the applications which have obtained good performance on Avalon, and their characteristics. Our goal has been to provide dependable cycles for computational physics, and not to perform research into clustered computing systems. One of the main lessons learned from the Avalon project is that the details of the hardware are not nearly as important as the attitudes and expectations of the users and managers of the hardware.

  17. Assessment of Surfactant Protein A (SP-A dependent agglutination

    Directory of Open Access Journals (Sweden)

    Griese Matthias

    2010-11-01

    Full Text Available Abstract Background Monomers of the collectin surfactant associated protein-A (SP-A are arranged in trimers and higher oligomers. The state of oligomerization differs between individuals and likely affects SP-A's functional properties. SP-A can form aggregates together with other SP-A molecules. Here we report and assess a test system for the aggregate forming properties of SP-A in serum and broncho-alveolar lavage samples. Methods Anti-SP-A antibodies fixed to latex beads bound SP-A at its N-terminal end and allowed the interaction with other SP-A molecules in a given sample by their C-terminal carbohydrate recognition domain (CRD to agglutinate the beads to aggregates, which were quantified by light microscopy. Results SP-A aggregation was dependent on its concentration, the presence of calcium, and was dose-dependently inhibited by mannose. Unaffected by the presence of SP-D no aggregation was observed in absence of SP-A. The more complex the oligomeric structure of SP-A present in a particular sample, the better was its capability to induce aggregation at a given total concentration of SP-A. SP-A in serum agglutinated independently of the pulmonary disease; in contrast SP-A in lung lavage fluid was clearly inferior in patients with chronic bronchitis and particularly with cystic fibrosis compared to controls. Conclusions The functional status of SP-A with respect to its aggregating properties in serum and lavage samples can be easily assessed. SP-A in lung lavage fluid in patients with severe neutrophilic bronchitis was inferior.

  18. Separating Surface Order and Syntactic Relations in a Dependency Grammar

    CERN Document Server

    Broeker, N

    1998-01-01

    This paper proposes decoupling the dependency tree from word order, such that surface ordering is not determined by traversing the dependency tree. We develop the notion of a \\emph{word order domain structure}, which is linked but structurally dissimilar to the syntactic dependency tree. The proposal results in a lexicalized, declarative, and formally precise description of word order; features which lack previous proposals for dependency grammars. Contrary to other lexicalized approaches to word order, our proposal does not require lexical ambiguities for ordering alternatives.

  19. A-dependence for the charmed meson production

    International Nuclear Information System (INIS)

    A report is presented of a recent direct measurement of the nucleon number (λ) dependence of the production cross sections for the charmed mesons D0 and D+ using π+- beams incident on a segmented target of Be, Al, Cu and W. The data derive from the experiment E769 - Hadroproduction of Charm - at Fermilab. The experimental apparatus is described together with the following analysis. Starting from a sample of ∼ 1500 D mesons in the range of O F A = σO Aα, with α = 0.99 ± 0.03. The XF dependence of α is examined and the results obtained are compared with those of other experiments and with theoretical expectations based on perturbative QCD and on an EMC like model of nuclear shadowing. (author). 85 refs, 61 figs, 22 tabs

  20. Developing a dependable approach for evaluating waste treatment data

    International Nuclear Information System (INIS)

    Decision makers involved with hazardous waste treatment issues are faced with the challenge of making objective evaluations concerning treatment formulations. This work utilizes an effectiveness factor (denoted as η) as the basis for waste treatment evaluations, which was recently developed for application to mixed waste treatability studies involving solidification and stabilization at the Idaho National Engineering and Environmental Laboratory. The effectiveness factor incorporates an arbitrary treatment criterion Φ, which in practice could be the Toxicity Characteristic Leaching Procedure, Unconfined Compressive Strength, Leachability Index, or any other criterion used to judge treatment performance. Three values for Φ are utilized when assessing a given treatment formulation: before treatment, after treatment, and a reference value (typically a treatment standard). The expression for η also incorporates the waste loading as the prime experimental parameter, and accounts for the contribution that each hazard has upon the overall treatment performance. Also discussed are general guidelines for numerical boundaries and statistical interpretations of treatment data. Case studies are presented that demonstrate the usefulness of the effectiveness factor and related numerical methods, where the typical hazards encountered are toxic metals within mixed waste

  1. A-dependence of weak nuclear structure functions

    Energy Technology Data Exchange (ETDEWEB)

    Haider, H.; Athar, M. Sajjad [Department of Physics, Aligarh Muslim University, Aligarh-202 002 (India); Simo, I. Ruiz [Dipartimento di Fisica, Universitá degli studi di Trento Via Sommarive 14, Povo (Trento) I-38123 (Italy)

    2015-05-15

    Effect of nuclear medium on the weak structure functions F{sub 2}{sup A}(x, Q{sup 2}) and F{sub 3}{sup A}(x, Q{sup 2}) have been studied using charged current (anti)neutrino deep inelastic scattering on various nuclear targets. Relativistic nuclear spectral function which incorporate Fermi motion, binding and nucleon correlations are used for the calculations. We also consider the pion and rho meson cloud contributions calculated from a microscopic model for meson-nucleus self-energies. Using these structure functions, F{sub i}{sup A}/F{sub i}{sup proton} and F{sub i}{sup A}/F{sub i}{sup deuteron}(i=2,3, A={sup 12}C, {sup 16}O, CH and H{sub 2}O) are obtained.

  2. relA-dependent RNA polymerase activity in Escherichia coli.

    OpenAIRE

    Ryals, J; Bremer, H

    1982-01-01

    Parameters relating to RNA synthesis were measured after a temperature shift from 30 to 42 degrees C, in a relA+ and relA- isogenic pair of Escherichia coli strains containing a temperature-sensitive valyl tRNA synthetase. The following results were obtained: (i) the rRNA chain growth rate increased 2-fold in both strains; (ii) newly synthesized rRNA became unstable in both strains; (iii) the stable RNA gene activity (rRNA and tRNA, measured as stable RNA synthesis rate relative to the total ...

  3. The Integrated periodogram of a dependent extremal event sequence

    DEFF Research Database (Denmark)

    Mikosch, Thomas Valentin; Zhao, Yuwei

    2015-01-01

    We investigate the asymptotic properties of the integrated periodogram calculated from a sequence of indicator functions of dependent extremal events. An event in Euclidean space is extreme if it occurs far away from the origin. We use a regular variation condition on the underlying stationary.......i.d. case a Brownian bridge appears. In the general case, we propose a stationary bootstrap procedure for approximating the distribution of the limiting process. The developed theory can be used to construct classical goodness-of-fit tests such as the Grenander–Rosenblatt and Cramér–von Mises tests which...

  4. Care plan for the patient with a dependent personality disorder

    Directory of Open Access Journals (Sweden)

    Ana María Ruiz Galán

    2010-11-01

    Full Text Available Personality is unique for each individual and can be defined as the dynamic collection of characteristics relative to emotions, thought and behaviour.Personality trout’s only mean a Personality Disorder (PD when they are inflexible and maladjusted and cause notable functional deterioration or uneasiness.According to Bermudez personality is “the enduring organization of structural and functional features, innate and acquired under the special conditions of each one’s development that shape the particular and specific collection of behaviour to face different situations”.According to the Diagnostic a Statistical Manual of Mental Disorders (DSM-IV, a Personality Disorder is “an enduring pattern of inner experience and behavior that deviates markedly from the expectations of the person’s culture is pervasive and an inflexible, is stable over time and leads to distress or impairment. The onset of these patterns of behaviour is the beginning of the adulthood and, in rare instances, early adolescence”.There are several types of Personality Disorders (paranoid, schizoid, borderline, antisocial, dependent…. Dependent Personality Disorder is one of the most frequent in the Mental Health Services.People who suffer from this disorder are unable to take a decision by themselves because they don’t have confidence in themselves. They need a lot of social support and affection until the point of deny their individuality by subordinating their desires to other person’s desires and permitting these persons to manage their lives. Maybe they feel desolated by separation and loss and can support any situation, even maltreatment to keep a relationship.As we a deduce this diagnosis is sensible to cultural influences. This work aims to elaborate an standarized plan of cares for the patient with Dependent Personality Disorder by using nursing Diagnosis of NANDA II, Outcomes Criteria (NOC and Interventions Criteria (NIC.

  5. Study on inhibitory elements of a putative PrfA-dependent promoter ParoA2 in Listeria monocytogenes%单核细胞增生李斯特菌毒力基因aroA启动子上阻碍PrfA转录调控元件的研究

    Institute of Scientific and Technical Information of China (English)

    罗勤; 周青春

    2008-01-01

    Objective To investigate the relationship between PrfA-dependent promoters and PrfA regulation. Methods LacZ reporter gene fusions used to investigate the inhibitory elements for PrfA-dependent transcription were carried on two promoters of Listeria monocytogenes: a PrfA-dependent promoter of the phospholipase gene pica (PplcA) and a putative promoter of the aroA gene (ParoA2) which contains a similar PrfA-binding site (PrfA-box) and a similar-10 box as PplcA but does not function as PrfA-dependent promoter. A series of hybrid plcA-aroA promoters by exchanging corresponding sequence elements of these two "promoters" were constructed and incorporated into upstream of a promoterless lacZ gene. The variant promoter-lacZ transcriptional fusions were then electroporated into L. monocytogenes wild-type strain P14, prfA mutant P14a and prfA deletion mutant A42, respectively. The expression level of PrfA is the highest in the P14a and the lowest in A42. The corresponding transcription activities of hybrid promoters were measured by the β-galactosidase assay. Results The two critical elements of PrfA-dependent promoters, the PrfA-box and the-10 box, can be functionally exchanged as long as the distance in between is maintained 22 or 23 bp. However, the interspace sequence and the sequence downstream of the -10 box of ParoA2 were strongly inhibitory for PrfA-dependent transcription. Conclusion Downstream sequence together -10 box of ParoA2 might fold into a hairpin structure when present in a single stranded DNA and possibly block the formation of the transcriptional initiation open complex, hence, inhibit the PrfA-dependent transcription from ParoA2.%目的 研究食源性致病菌单核细胞增生李斯特菌(Listeria monocytogenes,简称LM)毒力基因启动子的结构特点及其与转录调控因子PrfA(positive regulatory factor A)蛋白之间的关系.方法 选取plcA和aroA两个毒力基因启动子作为研究对象,plcA基因启动子(PplcA)上含有一个

  6. 常用英文缩略语名词解释

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    AMD:年龄相关性黄斑变性(age—related macular degeneration)DR:糖尿病视网膜病变(diabetic retinopathy)OIR:氧诱导视网膜病变(oxygen—induced retinopathy)Nrf2:核因子E2相关因子2(nuclear factor—E2-related factor 2)ARE:抗氧化反应元件(antioxidant response element)Hrd1:羟甲基戊二酰辅酶A还原酶降解蛋白1(hydroxymethyl glutaric acyl coenzyme A reductase degradation protein 1)。

  7. Dicty_cDB: Contig-U10831-1 [Dicty_cDB

    Lifescience Database Archive (English)

    Full Text Available mplet... 43 0.033 CP000615_24( CP000615 |pid:none) Burkholderia vietnamiensis G4 chr... 43 0.0... ) RecName: Full=Peroxisomal acyl-coenzyme A oxidase 1; ... 87 2e-15 T20569( T20569 ) hypothetical protein F08A8.3 - Caenorhabdi...icus su... 51 1e-04 AF142581_1( AF142581 |pid:none) Streptomyces coelicolor acyl-CoA d... 51 1e-04 CP000903_2301( CP000903 |pid...i rw*isf*s**ikriiskfkyi**kifngtiin**ts*tnatsmfrdysitigpie*ygke skeisimv*idinv*sisiieicssi*f...re E Sequences producing significant alignments: (bits) Value N ( U87813 ) Dictyostelium discoideum AcoA (aco

  8. Transient intermediary states with high and low folding probabilities in the apparent two-state folding equilibrium of ACBP at low pH

    DEFF Research Database (Denmark)

    Thomsen, Jens K; Kragelund, Birthe B; Teilum, Kaare;

    2002-01-01

    Measurements of the stability as a function of pH for the acyl-coenzyme A binding protein (ACBP) has shown a significant difference in the pH transition midpoint measured by NMR spectroscopy at pH 3.12 and the transition midpoint measured at pH 2.92 and 2.97 by circular dichroism and by...... indicates strongly the existence of short-lived and transient helical structures at pH 2.3. Magnetization transfer studies have been applied to study the equilibrium between folded and unfolded ACBP near the pH transition point measured by NMR. This study has shown that there are two categories of...

  9. Green tea extract suppresses adiposity and affects the expression of lipid metabolism genes in diet-induced obese zebrafish

    Directory of Open Access Journals (Sweden)

    Hasumura Takahiro

    2012-08-01

    Full Text Available Abstract Background Visceral fat accumulation is one of the most important predictors of mortality in obese populations. Administration of green tea extract (GTE can reduce body fat and reduce the risk of obesity-related diseases in mammals. In this study, we investigated the effects and mechanisms of GTE on adiposity in diet-induced obese (DIO zebrafish. Methods Zebrafish at 3.5 to 4.5 months post-fertilization were allocated to four groups: non-DIO, DIO, DIO + 0.0025%GTE, and DIO + 0.0050%GTE. The non-DIO group was fed freshly hatched Artemia once daily (5 mg cysts/fish daily for 40 days. Zebrafish in the three DIO groups were fed freshly hatched Artemia three times daily (60 mg cysts/fish daily. Zebrafish in the DIO + 0.0025%GTE and DIO + 0.0050%GTE groups were exposed to GTE after the start of feeding three times daily for 40 days. Results Three-dimensional microcomputed tomography analysis showed that GTE exposure significantly decreased the volume of visceral but not subcutaneous fat tissue in DIO zebrafish. GTE exposure increased hepatic expression of the lipid catabolism genes ACOX1 (acyl-coenzyme A oxidase 1, palmitoyl, ACADM (acyl-coenzyme A dehydrogenase, c-4 to c-12 straight chain, and PPARA (peroxisome proliferator-activated receptor alpha. GTE exposure also significantly decreased the visceral fat expression of SOCS3 (suppressor of cytokine signaling 3b which inhibits leptin signaling. Conclusions The present results are consistent with those seen in mammals treated with GTE, supporting the validity of studying the effects of GTE in DIO zebrafish. Our results suggest that GTE exerts beneficial effects on adiposity, possibly by altering the expression of lipid catabolism genes and SOCS3.

  10. Famílias com um idoso dependente: avaliação da coesão e adaptação Las familias con un adulto mayor dependiente: evaluación de la cohesión y de la adaptación Families with a dependent elderly member: assessment of adaptation and cohesion

    Directory of Open Access Journals (Sweden)

    Isabel Araújo

    2012-03-01

    final del desarrollo del ciclo vital. Se les consideraron familias cohesivas y adaptadas aunque idealizan una mayor cohesión y adaptación.The aim of this study was to describe the typological profile and developmental stage of families that take care of an elderly dependent person; to identify the degree of dependency of the elderly person in the family context; and to evaluate cohesion and adaptation in families with a dependent elder. Methodologically, the study is positioned in the quantitative paradigm, and is a descriptive exploratory study. We administered a questionnaire, which included FACES III and the Barthel Index, to a group of families registered at health centers or family health clinics in a district in the North of Portugal. We chose a purposive sample of 108 families with an elderly dependent member. Data collection occurred from October 2007 to June 2008. From data analysis, we verified that families that include an elderly person at a high level of dependence are nuclear families and are in the final phase of development of the life cycle. They consider themselves cohesive and adapted families but ideally would like more cohesion and adaptation.

  11. A-dependence in dilepton rapidity distributions: parton model and dipole approach analysis

    Energy Technology Data Exchange (ETDEWEB)

    Gay Ducati, M.B., E-mail: beatriz.gay@ufrgs.b [Instituto de Fisica, Universidade Federal do Rio Grande do Sul P.O. Box 15051, 91501-970 Porto Alegre, Rio Grande do Sul (Brazil); Oliveira, E.G. de, E-mail: emmanuel.deoliveira@ufrgs.b [Instituto de Fisica, Universidade Federal do Rio Grande do Sul P.O. Box 15051, 91501-970 Porto Alegre, Rio Grande do Sul (Brazil)

    2010-02-15

    We calculate dilepton production cross section at RHIC using the improved parton model and color dipole approach in p-A collisions at backward rapidities. Our results are given as nuclear modification factors. We test three nuclear parton distribution functions: EKS, EPS08 and EPS09; we found no changes in the results. In the color dipole approach, we use GBW, DHJ, and BUW color dipole cross section models; again, no relevant difference is found in the nuclear modification factor, in spite of that DHJ breaks geometric scaling and that the p-p cross sections show relevant differences. The presence of intrinsic transverse momentum in the improved parton model is investigated, showing that nuclear effects are sensitive to this assumption. The color dipole approach lacks this intrinsic k{sub T} and thus nuclear effects can discriminate among both analysis. We extend our improved parton model results to forward rapidities and compare with CGC results, obtaining agreement between the models.

  12. Mass Media Content as a Dependent Variable: Five Media Sociology Theories.

    Science.gov (United States)

    Shoemaker, Pamela J.; Mayfield, Elizabeth Kay

    To better understand the effects of the media on audiences, five media sociology theoretical approaches to the study of influences on media were investigated by looking at tests of the approaches in three journals over the last 10 years. The mirror approach predicted that the media would accurately represent reality, but, while accurate, the media…

  13. Measurements of the A-dependence of deep-inelastic electron scattering from nuclei

    International Nuclear Information System (INIS)

    The deep inelastic electron scattering cross sections per nucleon sigma/sub A/ for d, He, Be, C, Al, Ca, Fe, Ag, and Au were measured in kinematic range 0.09 less than or equal to x less than or equal to 0.9 and 2 less than or equal to Q2 less than or equal to 15 (GeV/c)2 using electrons with energies ranging from 8 to 24.5 GeV. The ratio sigma/sub A//sigma/sub d/ is consistent with unity in the range 0.1 2 dependence in the ratio was observed over the kinematic range of the data. This has been interpreted as evidence for a change in the quark momentum distribution in the nucleus due to the presence of 6 quark clusters or a larger nucleon bag size. 13 references

  14. A-dependence of deep-inelastic electron scattering from nuclei

    International Nuclear Information System (INIS)

    The deep inelastic electron scattering cross sections per nucleon sigma/sub A/ for d, He, Be, C, Al, Ca, Fe, Ag, and Au were measured in the kinematic range 0.09 less than or equal to x less than or equal to 0.9 and 2 less than or equal to Q2 less than or equal to 15 (GEV/c)2 using electrons with energies ranging from 8 to 24.5 GeV. The ratio sigma/sub A//sigma/sub d/ is consistent with unity in the range 0.1 2 dependence in the ratio was observed over the kinematic range of the data. 12 references

  15. FOXO3a-dependent regulation of Puma in response to cytokine/growth factor withdrawal

    OpenAIRE

    You, Han; Pellegrini, Marc; Tsuchihara, Katsuya; Yamamoto, Kazuo; Hacker, Georg; Erlacher, Miriam; Villunger, Andreas; Mak, Tak W.

    2006-01-01

    Puma is an essential mediator of p53-dependent and -independent apoptosis in vivo. In response to genotoxic stress, Puma is induced in a p53-dependent manner. However, the transcription factor driving Puma up-regulation in response to p53-independent apoptotic stimuli has yet to be identified. Here, we show that FOXO3a up-regulates Puma expression in response to cytokine or growth factor deprivation. Importantly, dysregulated Akt signaling in lymphoid cells attenuated Puma induction upon cyto...

  16. Towards a dependable homogeneous many-processor system-on-chip

    NARCIS (Netherlands)

    Zhang, Xiao

    2014-01-01

    Nowadays, dependable computing systems are widely required in mission-critical applications. While the advance in CMOS technology enables smaller and faster circuits, the dependability of modern ICs has worsened as a result of the shrinking dimensions of MOS transistors and the increasing complexity

  17. Translating a Dependently-Typed Logic to First-Order Logic

    Science.gov (United States)

    Sojakova, Kristina; Rabe, Florian

    DFOL is a logic that extends first-order logic with dependent types. We give a translation from DFOL to FOL formalized as an institution comorphism and show that it admits the model expansion property. This property together with the borrowing theorem implies the soundness of borrowing — a result that enables us to reason about entailment in DFOL by using automated tools for FOL. In addition, the translation permits us to deduce properties of DFOL such as completeness, compactness, and existence of free models from the corresponding properties of FOL, and to regard DFOL as a fragment of FOL. We give an example that shows how problems about DFOL can be solved by using the automated FOL prover Vampire. Future work will focus on the integration of the translation into the specification and translation tool HeTS.

  18. RNA G-quadruplexes cause eIF4A-dependent oncogene translation in cancer

    OpenAIRE

    Wolfe, Andrew L.; Singh, Kamini; Zhong, Yi; Drewe, Philipp; Vinagolu K. Rajasekhar; Sanghvi, Viraj R.; Mavrakis, Konstantinos J; Jiang, Man; Roderick, Justine E.; Van Der Meulen, Joni; Schatz, Jonathan H.; Rodrigo, Christina M.; Zhao, Chunying; Rondou, Pieter; de Stanchina, Elisa

    2014-01-01

    The translational control of oncoprotein expression is implicated in many cancers. Here we report an eIF4A/DDX2 RNA helicase-dependent mechanism of translational control that contributes to oncogenesis and underlies the anticancer effects of Silvestrol and related compounds. For example, eIF4A promotes T-ALL development in vivo and is required for leukaemia maintenance. Accordingly, inhibition of eIF4A with Silvestrol has powerful therapeutic effects in vitro and in vivo. We use transcriptome...

  19. Metabolism of β-valine via a CoA-dependent ammonia lyase pathway

    NARCIS (Netherlands)

    Otzen, Marleen; Crismaru, Ciprian G.; Postema, Christiaan P.; Wijma, Hein J.; Heberling, Matthew M.; Szymanski, Wiktor; de Wildeman, Stefaan; Janssen, Dick B.

    2015-01-01

    Pseudomonas species strain SBV1 can rapidly grow on medium containing β-valine as a sole nitrogen source. The tertiary amine feature of β-valine prevents direct deamination reactions catalyzed by aminotransferases, amino acid dehydrogenases, and amino acid oxidases. However, lyase- or aminomutase-me

  20. Inhibition of AKT promotes FOXO3a-dependent apoptosis in prostate cancer.

    Science.gov (United States)

    Das, T P; Suman, S; Alatassi, H; Ankem, M K; Damodaran, C

    2016-01-01

    Growth factor-induced activation of protein kinase-B (PKB), also known as AKT, induces pro-survival signaling and inhibits activation of pro-apoptotic signaling molecules including the Forkhead box O-3a (FOXO3a) transcription factor and caspase in transformed prostate cells in vitro. Earlier we reported that Withaferin-A (WA), a small herbal molecule, induces pro-apoptotic response-4 (Par-4) mediated apoptosis in castration-resistant prostate cancer (CRPC) cells. In the present study, we demonstrate that inhibition of AKT facilitates nuclear shuttling of FOXO3a where it regulates Par-4 transcription in CRPC cells. FOXO3a is upstream of Par-4 signaling, which is required for induction of apoptosis in CRPC cells. Promoter bashing studies and Ch-IP analysis confirm a direct interaction of FOXO3a and Par-4; a sequential deletion of FOXO3a-binding sites in the Par-4 promoter fails to induce Par-4 activation. To confirm these observations, we either overexpressed AKT or silenced FOXO3a activation in CRPC cells. Both methods inhibit Par-4 function and apoptosis is significantly compromised. In xenograft tumors derived from AKT-overexpressed CRPC cells, FOXO3a and Par-4 expression is downregulated, leading to aggressive tumor growth. Oral administration of WA to mice with xenograft tumors restores FOXO3a-mediated Par-4 functions and results in inhibited tumor growth. Finally, an inverse correlation of nuclear localization of AKT expression corresponds to cytoplasmic Par-4 localization in human prostate tissue array. Our studies suggest that Par-4 is one of the key transcriptional targets of FOXO3a, and Par-4 activation is required for induction of apoptosis in CRPC cells. Activation of FOXO3a appears to be an attractive target for the treatment of CRPC and molecules such as WA can be explored further for the treatment of CRPC. PMID:26913603

  1. Accountants and economic governance in a dependent country, : Conflicting legacies and new professional issues in Lebanon

    OpenAIRE

    Longuenesse, Elisabeth

    2006-01-01

    Purpose: The purpose of this study was to sort out what was at stake in the fierce debates that raged after 1995 among Lebanese accountants about the conditions of membership to their new association.Approach and methodology: It fits within the recent development of the sociology of professions. But because of the specific nature of accountants expertise, it is also concerned with broader societal issues and socio-political negotiation of interests, which underlie professional battles. Lastly...

  2. FINAL REMINDER EXTENSION/SUPPRESSION OF ALLOWANCE FOR A DEPENDENT CHILD AGED 18 AND ABOVE

    CERN Multimedia

    Human Resources Division

    2001-01-01

    Members of the personnel with dependent children aged 18 or above (or reaching 18 during the 2001/2002 school year) who have not yet provided a SCHOOL CERTIFICATE must do so as soon as possible. If we have not received this certificate by December 11, 2001 at the latest, the child allowance will be withdrawn retroactively as from September 1, 2001.

  3. Forbidding undesirable agreements: a dependence-based approach to the regulation of multi-agent systems

    NARCIS (Netherlands)

    P Turrini; D. Grossi; J. Broersen; J.-J.C. Meyer

    2010-01-01

    The purpose of this contribution is to set up a language to evaluate the results of concerted action among interdependent agents against predetermined properties that we can recognise as desirable from a deontic point of view. Unlike the standard view of logics to reason about coalitionally rational

  4. THE MEANING OF BEING-A-CAREGIVER OF A DEPENDENT RELATIVE SUFFERING FROM CANCER: PALLIATIVE CONTRIBUTIONS

    Directory of Open Access Journals (Sweden)

    Joisy Aparecida Marchi

    2016-01-01

    Full Text Available Se tuvo como objetivo comprender el significado de ser-cuidador de un familiar con cáncer y con gran dependencia para las actividades diarias. Estudio fenomenológico fundamentado en Martín Heidegger realizado junto a tres núcleos integrados de salud en un municipio del noroeste de Paraná. La entrevista sucedió entre noviembre de 2012 y febrero de 2013 con 17 cuidadores familiares. Del análisis propuesto surgieron dos temáticas: “El ser-cuidador vivenciando distintos modos de disposición” y “Siendo-con-el: de la ocupación cotidiana a la preocupación libertadora”. Significó para el ser-cuidador aterrarse con el diagnóstico, horrorizarse con el tratamiento, aterrorizarse con los cuidados paliativos y ser-con-el-otro en la enfermedad. Se mostró ocupado con las cosas, pero también estuvo preocupado, evidenciando la solicitud en sus acciones. Esta base para un cuidado paliativo efectivo, debe permear la labor del enfermero visando que este profesional sea un verdadero ser-del-cuidado.

  5. Computer optimization program finds values for several independent variables that minimize a dependent variable

    Science.gov (United States)

    Warech, E. J.

    1967-01-01

    Computer program finds values of independent variables which minimize the dependent variable. This optimization program has been used on the F-1 and J-2 engine programs to establish minimum film coolant requirements.

  6. FINAL REMINDER EXTENSION/SUPPRESSION OF ALLOWANCE FOR A DEPENDENT CHILD AGED 18 AND ABOVE

    CERN Multimedia

    Social and Statutory Conditions; Tel. 72862-74474

    2000-01-01

    Members of the personnel with dependent children aged 18 or above (or reaching 18 during the 2000/2001 school year) who have not yet provided a SCHOOL CERTIFICATE must do so as soon as possible. If we have not received this certificate by November 28, 2000 at the latest, the child allowance will be withdrawn retroactively as from September 1,2000.

  7. Domestic Violence Screening and Service Acceptance among Adult Victims in a Dependency Court Setting

    Science.gov (United States)

    Rivers, James E.; Maze, Candice L.; Hannah, Stefanie A.; Lederman, Cindy S.

    2007-01-01

    Many child welfare systems are unable to effectively identify and address co-occurring domestic violence and child maltreatment. In response, the Dependency Court Intervention Program for Family Violence implemented a protocol to identify indicators of domestic violence in families involved with child protection proceedings. This article…

  8. Environmental Degradation in a Dependent Region: The Rio Grande Valley of Mexico and Texas.

    Science.gov (United States)

    Jones, Richard C.

    1999-01-01

    Traces the interrelationships among dependence, environmental degradation, and human health in the Rio Grande Valley of Mexico and Texas. Presents a case study on environmental factors threatening family health in households located on both sides of the border; the health problems can be overcome by addressing restrictive zoning, health services,…

  9. Toward a Dependable Peace: A Proposal for an Appropriate Security System.

    Science.gov (United States)

    Johansen, Robert C.

    This booklet proposes that citizens and governments think imaginatively about national and international security and take action for comprehensive arms reductions. The document is presented in eight chapters. Chapter I reports that global insecurity exists despite continuous arms control negotiations since World War II. Chapter II discusses…

  10. Economic Value Added as a Dependence on the Corporate- and Market-life Cycle

    Directory of Open Access Journals (Sweden)

    Konečný Zdeněk

    2011-06-01

    Full Text Available Economic value added (EVA is an indicator which is widely used as the main tool for financial analysis. There are two methods of calculating it. The original method which was made by Stern & Stewart is defined as the net operating profit after taxes minus the cost of capital. The second method which was developed and used by the “Czech Ministry of Industry and Trade” indicates that, the economic value added is the difference between return on equity and the alternate cost of equity that is composed of separate risk rewards, and this “spread” is consequently multiplied by the equity. Economic value added depends on many factors. Whereas some of them are controllable by the company, others are not. This article is focused on the relationship between economic value added and the corporate- vs. market life cycle. This is because, there is an assumption that conditions for developing EVA changes depending on the actual phase of corporate- and market life cycle. In this research, the model by Reiners (2004 is used to identify the phases of corporate- and market life cycle and the method provided by the “Czech Ministry of Industry and Trade” is used to calculate EVA. However, there is a consideration of the relativity of EVA in the form of “spread” because of the intercompany comparison. The study found that, the highest spread is achieved by companies that are in the phase of expansion and phase of market expansion. On the contrary, companies in the phase of declension during market declension achieved the lowest and negative spread.

  11. Choice as a dependent measure in autoshaping: sensitivity to frequency and duration of food presentation.

    OpenAIRE

    Picker, M; Poling, A.

    1982-01-01

    Previous investigations have shown that rate, latency, and percentage of trials with at least one response are somewhat insensitive measures of the strength of autoshaped responding. In the present studies, these measures were contrasted with the allocation of responding during simultaneous choice tests, a measure of response strength frequently used in operant paradigms. In two experiments, nine pigeons were exposed to a forward pairing autoshaping procedure. Training sessions consisted of t...

  12. Adoption and Business Value of Mobile Retail Channel: A Dependency Perspective on Mobile Commerce

    Science.gov (United States)

    Chou, Yen-Chun

    2013-01-01

    Forrest Research estimated that revenues derived from mobile devices will grow at an annual rate of 39% to reach $31 billion by 2016. With the tremendous market growth, mobile banking, mobile marketing, and mobile retailing have been recently introduced to satisfy customer needs. Academic and practical articles have widely discussed unique…

  13. A-dependence in dilepton rapidity distributions: parton model and dipole approach analysis

    International Nuclear Information System (INIS)

    We calculate dilepton production cross section at RHIC using the improved parton model and color dipole approach in p-A collisions at backward rapidities. Our results are given as nuclear modification factors. We test three nuclear parton distribution functions: EKS, EPS08 and EPS09; we found no changes in the results. In the color dipole approach, we use GBW, DHJ, and BUW color dipole cross section models; again, no relevant difference is found in the nuclear modification factor, in spite of that DHJ breaks geometric scaling and that the p-p cross sections show relevant differences. The presence of intrinsic transverse momentum in the improved parton model is investigated, showing that nuclear effects are sensitive to this assumption. The color dipole approach lacks this intrinsic kT and thus nuclear effects can discriminate among both analysis. We extend our improved parton model results to forward rapidities and compare with CGC results, obtaining agreement between the models.

  14. Improving Regression Testing through Modified Ant Colony Algorithm on a Dependency Injected Test Pattern

    Directory of Open Access Journals (Sweden)

    G.Keerthi Lakshmi

    2012-03-01

    Full Text Available Performing regression testing on a pre production environment is often viewed by software practitioners as a daunting task since often the test execution shall by-pass the stipulated downtime or the test coverage would be non linear. Choosing the exact test cases to match this type of complexity not only needs prior knowledge of the system, but also a right use of calculations to set the goals right. On systems that are just entering the production environment after getting promoted from the staging phase, trade-offs are often needed to between time and the test coverage to ensure the maximum test cases are covered within the stipulated time. There arises a need to refine the test cases to accommodate the maximum test coverage it makes within the stipulated period of time since at most of the times, the most important test cases are often not deemed to qualify under the sanity test suite and any bugs that creped in them would go undetected until it is found out by the actual user at firsthand. Hence An attempt has been made in the paper to layout a testing framework to address the process of improving the regression suite by adopting a modified version of the Ant Colony Algorithm over and thus dynamically injecting dependency over the best route encompassed by the ant colony.

  15. A Rab3a-dependent complex essential for lysosome positioning and plasma membrane repair.

    Science.gov (United States)

    Encarnação, Marisa; Espada, Lília; Escrevente, Cristina; Mateus, Denisa; Ramalho, José; Michelet, Xavier; Santarino, Inês; Hsu, Victor W; Brenner, Michael B; Barral, Duarte; Vieira, Otília V

    2016-06-20

    Lysosome exocytosis plays a major role in resealing plasma membrane (PM) disruptions. This process involves two sequential steps. First, lysosomes are recruited to the periphery of the cell and then fuse with the damaged PM. However, the trafficking molecular machinery involved in lysosome exocytosis and PM repair (PMR) is poorly understood. We performed a systematic screen of the human Rab family to identify Rabs required for lysosome exocytosis and PMR. Rab3a, which partially localizes to peripheral lysosomes, was one of the most robust hits. Silencing of Rab3a or its effector, synaptotagmin-like protein 4a (Slp4-a), leads to the collapse of lysosomes to the perinuclear region and inhibition of PMR. Importantly, we have also identified a new Rab3 effector, nonmuscle myosin heavy chain IIA, as part of the complex formed by Rab3a and Slp4-a that is responsible for lysosome positioning at the cell periphery and lysosome exocytosis. PMID:27325790

  16. A-dependence of hadron-nucleus massive lepton pair production

    Energy Technology Data Exchange (ETDEWEB)

    Ayala Filho, A.L.; Ducati, M.B. Gay [Rio Grande do Sul Univ., Porto Alegre, RS (Brazil). Inst. de Fisica; Epele, L.N.; Canal, C.A. Garcia [La Plata Univ. Nacional (Argentina). Lab. de Fisica Teorica

    1995-12-31

    The nuclear effects in the small x region of hadron-nucleus Drell-Yan processes at 800 GeV analyzed. We employ the parton recombination model to describe the suppression in the R{sup pA}{sub DY} (x{sub 2}) ratio as compared with R{sub EMC} (x), in the same kinematical region. A good agreement with experimental results is obtained. (author) 9 refs., 2 figs., 1 tab.

  17. [ital A] dependence of hadron-nucleus massive lepton pair production

    Energy Technology Data Exchange (ETDEWEB)

    Ayala F, A.L.; Gay Ducati, M.B. (Instituto de Fisica, Universidade Federal do Rio Grande do Sul, Caixa Postal 15051, 91500 Porto Alegre, RS (Brazil)); Epele, L.N.; Garcia Canal, C.A. (Laboratorio de Fisica Teorica, Universidad Nacional de La Plata, CC 67, 1900 La Plata (Argentina))

    1994-01-01

    Nuclear effects in the small [ital x] region of hadron-nucleus Drell-Yan processes at 800 GeV are analyzed. We employ the parton recombination model to describe the suppression in the [ital R][sub DY][sup [ital p][ital A

  18. Molecular Underpinnings of Nitrite Effect on CymA-Dependent Respiration in Shewanella oneidensis

    Science.gov (United States)

    Jin, Miao; Fu, Huihui; Yin, Jianhua; Yuan, Jie; Gao, Haichun

    2016-01-01

    Shewanella exhibit a remarkable versatility of respiration, with a diverse array of electron acceptors (EAs). In environments where these bacteria thrive, multiple EAs are usually present. However, we know little about strategies by which these EAs and their interaction affect ecophysiology of Shewanella. In this study, we demonstrate in the model strain, Shewanella oneidensis MR-1, that nitrite, not through nitric oxide to which it may convert, inhibits respiration of fumarate, and probably many other EAs whose reduction depends on quinol dehydrogenase CymA. This is achieved via the repression of cyclic adenosine monophosphate (cAMP) production, a second messenger required for activation of cAMP-receptor protein (Crp) which plays a primary role in regulation of respiration. If nitrite is not promptly removed, intracellular cAMP levels drop, and this impairs Crp activity. As a result, the production of nitrite reductase NrfA, CymA, and fumarate reductase FccA is substantially reduced. In contrast, nitrite can be simultaneously respired with trimethylamine N-oxide, resulting in enhanced biomass. PMID:27493647

  19. Performance analysis of a dependable scheduling strategy based on a fault-tolerant grid model

    Institute of Scientific and Technical Information of China (English)

    WANG Yuanzhuo; LIN Chuang; YANG Yang; SHAN Zhiguang

    2007-01-01

    The grid provides an integrated computer platform composed of differentiated and distributed systems.These resources are dynamic and heterogeneous.In this paper,a novel fault-tolerant grid-scheduling model is pre sented based on Stochastic Petri Nets (SPN) to assure the heterogeneity and dynamism of the grid system.Also,a new grid-scheduling strategy,the dependable strategy for the shortest expected accomplishing time (DSEAT),is put forward,in which the dependability factor is introduced in the task-dispatching strategy.In the end,the performance of the scheduling strategy based on the fault-tolerant gridscheduling model is analyzed by an software package,named SPNP.The numerical results show that dynamic resources will increase the response time for all classes of tasks in differing degrees.Compared with shortest expected accomplishing time (SEAT) strategy,the DSEAT strategy can reduce the negative effects of dynamic and autonomic resources to some extent so as to guarantee a high quality of service (QoS).

  20. História da enfermagem psiquiátrica e a dependência química no Brasil: atravessando a história para reflexão Historia de la enfermería psiquiátrica y la dependencia química en el Brasil: atravesando la historia para la reflexión History of the psychiatric nursing and chemical dependency in Brazil: crossing the history for reflection

    Directory of Open Access Journals (Sweden)

    Amanda Márcia dos Santos Reinaldo

    2007-12-01

    Full Text Available A formação do enfermeiro em Enfermagem Psiquiátrica e na área de dependências químicas norteia a discussão desse artigo, tendo em vista a complexidade dos problemas relacionados ao ensino de enfermagem, saúde mental, psiquiatria e álcool e drogas. Trata-se de um artigo de revisão de literatura onde as autoras compilaram fontes primárias e secundárias sobre o tema e, a partir da leitura do material bibliográfico, fizeram análises e reflexões acerca dos atravessamentos históricos que permeiam a história da Enfermagem Psiquiátrica e a dependência química no Brasil. Os resultados apontam para uma evolução do tema álcool e drogas dada a magnitude do problema na sociedade contemporânea. Em relação à Enfermagem Psiquiátrica, o ensino apresenta mudanças devidas à evolução histórica da psiquiatria que devem ser consideradas durante a formação do profissional enfermeiro. Ambas as temáticas encontram pontos de aproximação e distanciamento conforme o contexto em que são analisadas.La formación del enfermero en enfermería psiquiátrica y en el área de dependencias químicas, orienta la discusión de este artículo teniendo a la vista la complejidad de los problemas relacionados con la enseñanza de enfermería, salud mental, psiquiátrica, alcohol y drogas. Se trata de un artículo de revisión de la literatura donde las autoras compilaron fuentes primarias y secundarias sobre el tema y a partir de la lectura del material bibliográfico se realizaron análisis y reflexiones acerca de los acontecimientos históricos que permean la historia de la enfermería psiquiátrica y la dependencia química en el Brasil. Los resultados apuntan para una evolución del tema alcohol y drogas debido a la magnitud del problema en la sociedad contemporánea. En relación a la enfermería psiquiátrica, la enseñanza presenta cambios debido a la evolución histórica de la psiquiátrica que deben ser consideradas durante la formaci

  1. 小肠胆固醇吸收相关蛋白的研究进展%Several proteins involved in absorption of cholesterol in small intestine

    Institute of Scientific and Technical Information of China (English)

    袁敏; 徐东刚

    2015-01-01

    多种蛋白参与了小肠胆固醇的吸收,其中尼曼-匹克C1型类似蛋白1(Niemann-Pick C1 like 1,NPC1L1)主要介导小肠对胆固醇的吸收;小肠吸收的游离胆固醇在酰基辅酶A-胆固醇酰基转移酶2[acyl-coenzyme A(CoA)∶cholesterol acyltransferase 2,ACAT2]的催化下形成胆固醇酯并经淋巴系统进入血液循环,而未被酯化的胆固醇则通过ATP结合盒转运蛋白G5/G8[ATP-binding cassette(ABC) transporters G5/G8,ABCG5/ABCG8]分泌入肠腔,转录因子肝X受体( liver X receptor,LXR)在小肠胆固醇吸收过程中发挥了重要的调节作用。该文主要对小肠胆固醇吸收相关蛋白NPC1 L1、ABCG5/ABCG8、ACAT2和LXR的研究进展进行了综述。%Several proteins are involved in the absorption of cholesterol in small intestine.Niemann-Pick C1 like 1 (NPC1L1) mainly mediates the absorption of cholesterol, and acyl-coenzyme A ( CoA)∶cholesterol acyltransferase 2 (ACAT2) catalyzes the free cholesterol absorpted by intestine into cholesterol ester,while unesterified free cholesterol is secreted into intestinal lumen by ATP-binding cassette(ABC) transporters G5/G8(ABCG5/ABCG8).Transcription factor liver X receptor( LXR) plays an important role in the process of intestinal cholesterol absorption.The research progress in NPC1L1,ABCG5/ABCG8,ACAT2 and LXR is reviewed in this article.

  2. The minor C-allele of rs2014355 in ACADS is associated with reduced insulin release following an oral glucose load

    Directory of Open Access Journals (Sweden)

    Pisinger Charlotta

    2011-01-01

    Full Text Available Abstract Background A genome-wide association study (GWAS using metabolite concentrations as proxies for enzymatic activity, suggested that two variants: rs2014355 in the gene encoding short-chain acyl-coenzyme A dehydrogenase (ACADS and rs11161510 in the gene encoding medium-chain acyl-coenzyme A dehydrogenase (ACADM impair fatty acid β-oxidation. Chronic exposure to fatty acids due to an impaired β-oxidation may down-regulate the glucose-stimulated insulin release and result in an increased risk of type 2 diabetes (T2D. We aimed to investigate whether the two variants associate with altered insulin release following an oral glucose load or with T2D. Methods The variants were genotyped using KASPar® PCR SNP genotyping system and investigated for associations with estimates of insulin release and insulin sensitivity following an oral glucose tolerance test (OGTT in a random sample of middle-aged Danish individuals (nACADS = 4,324; nACADM = 4,337. The T2D-case-control study involved a total of ~8,300 Danish individuals (nACADS = 8,313; nACADM = 8,344. Results In glucose-tolerant individuals the minor C-allele of rs2014355 of ACADS associated with reduced measures of serum insulin at 30 min following an oral glucose load (per allele effect (β = -3.8% (-6.3%;-1.3%, P = 0.003, reduced incremental area under the insulin curve (β = -3.6% (-6.3%;-0.9%, P = 0.009, reduced acute insulin response (β = -2.2% (-4.2%;0.2%, P = 0.03, and with increased insulin sensitivity ISIMatsuda (β = 2.9% (0.5%;5.2%, P = 0.02. The C-allele did not associate with two other measures of insulin sensitivity or with a derived disposition index. The C-allele was not associated with T2D in the case-control analysis (OR 1.07, 95% CI 0.96-1.18, P = 0.21. rs11161510 of ACADM did not associate with any indices of glucose-stimulated insulin release or with T2D. Conclusions In glucose-tolerant individuals the minor C-allele of rs2014355 of ACADS was associated with reduced

  3. Serum autotaxin is increased in pruritus of cholestasis, but not of other origin, and responds to therapeutic interventions

    NARCIS (Netherlands)

    Kremer, Andreas E.; van Dijk, Remco; Leckie, Pamela; Schaap, Frank G.; Kuiper, Edith M. M.; Mettang, Thomas; Reiners, Katrin S.; Raap, Ulrike; van Buuren, Henk R.; van Erpecum, Karel J.; Davies, Nathan A.; Rust, Christian; Engert, Andreas; Jalan, Rajiv; Elferink, Ronald P. J. Oude; Beuers, Ulrich

    2012-01-01

    Pruritus is a seriously disabling symptom accompanying many cholestatic liver disorders. Recent experimental evidence implicated the lysophospholipase, autotaxin (ATX), and its product, lysophosphatidic acid (LPA), as potential mediators of cholestatic pruritus. In this study, we highlight that incr

  4. Role of beta-oxidation enzymes in gamma-decalactone production by the yeast Yarrowia lipolytica.

    Science.gov (United States)

    Waché, Y; Aguedo, M; Choquet, A; Gatfield, I L; Nicaud, J M; Belin, J M

    2001-12-01

    Some microorganisms can transform methyl ricinoleate into gamma-decalactone, a valuable aroma compound, but yields of the bioconversion are low due to (i) incomplete conversion of ricinoleate (C(18)) to the C(10) precursor of gamma-decalactone, (ii) accumulation of other lactones (3-hydroxy-gamma-decalactone and 2- and 3-decen-4-olide), and (iii) gamma-decalactone reconsumption. We evaluated acyl coenzyme A (acyl-CoA) oxidase activity (encoded by the POX1 through POX5 genes) in Yarrowia lipolytica in lactone accumulation and gamma-decalactone reconsumption in POX mutants. Mutants with no acyl-CoA oxidase activity could not reconsume gamma-decalactone, and mutants with a disruption of pox3, which encodes the short-chain acyl-CoA oxidase, reconsumed it more slowly. 3-Hydroxy-gamma-decalactone accumulation during transformation of methyl ricinoleate suggests that, in wild-type strains, beta-oxidation is controlled by 3-hydroxyacyl-CoA dehydrogenase. In mutants with low acyl-CoA oxidase activity, however, the acyl-CoA oxidase controls the beta-oxidation flux. We also identified mutant strains that produced 26 times more gamma-decalactone than the wild-type parents. PMID:11722925

  5. Endogenous Bioactive Peptides as Potential Biomarkers for Atherosclerotic Coronary Heart Disease

    Directory of Open Access Journals (Sweden)

    Tsutomu Hirano

    2012-04-01

    Full Text Available Cardiovascular disease is the leading cause of death worldwide, with high medical costs and rates of disability. It is therefore important to evaluate the use of cardiovascular biomarkers in the early diagnosis of coronary artery disease (CAD. We have screened a variety of recently identified bioactive peptides candidates in anticipation that they would allow detection of atherosclerotic CAD. Especially, we have focused on novel anti-atherogenic peptides as indicators and negative risk factors for CAD. In vitro, in vivo and clinical studies indicated that human adiponectin, heregulin-β1, glucagon-like peptide-1 (GLP-1, and salusin-α, peptides of 244, 71, 30, and 28 amino acids, respectively, attenuate the development and progression of atherosclerotic lesions by suppressing macrophage foam cell formation via down-regulation of acyl-coenzyme A: cholesterol acyltransferase-1. Circulating levels of these peptides in the blood are significantly decreased in patients with CAD compared to patients without CAD. Receiver operating characteristic analyses showed that salusin-α is a more useful biomarker, with better sensitivity and specificity, compared with the others for detecting CAD. Therefore, salusin-α, heregulin-β1, adiponectin, and/or GLP-1, alone or in various combinations, may be useful as biomarkers for atherosclerotic CAD.

  6. Novel O-palmitolylated beta-E1 subunit of pyruvate dehydrogenase is phosphorylated during ischemia/reperfusion injury

    Directory of Open Access Journals (Sweden)

    Barr Amy J

    2010-07-01

    Full Text Available Abstract Background During and following myocardial ischemia, glucose oxidation rates are low and fatty acids dominate as a source of oxidative metabolism. This metabolic phenotype is associated with contractile dysfunction during reperfusion. To determine the mechanism of this reliance on fatty acid oxidation as a source of ATP generation, a functional proteomics approach was utilized. Results 2-D gel electrophoresis of mitochondria from working rat hearts subjected to 25 minutes of global no flow ischemia followed by 40 minutes of aerobic reperfusion identified 32 changes in protein abundance compared to aerobic controls. Of the five proteins with the greatest change in abundance, two were increased (long chain acyl-coenzyme A dehydrogenase (48 ± 1 versus 39 ± 3 arbitrary units, n = 3, P In silico analysis identified the putative kinases as the insulin receptor kinase for the more basic form and protein kinase Cζ or protein kinase A for the more acidic form. These modifications of pyruvate dehydrogenase are associated with a 35% decrease in glucose oxidation during reperfusion. Conclusions Cardiac ischemia/reperfusion induces significant changes to a number of metabolic proteins of the mitochondrial proteome. In particular, ischemia/reperfusion induced the post-translational modification of pyruvate dehydrogenase, the rate-limiting step of glucose oxidation, which is associated with a 35% decrease in glucose oxidation during reperfusion. Therefore these post-translational modifications may have important implications in the regulation of myocardial energy metabolism.

  7. Testing models of fatty acid transfer and lipid synthesis in spinach leaf using in vivo oxygen-18 labeling

    Energy Technology Data Exchange (ETDEWEB)

    Pollard, M.; Ohlrogge, J.

    1999-12-01

    Oxygen-18 labeling has been applied to the study of plant lipid biosynthesis for the first time. [{sup 13}C{sub 2}{sup 18}O{sub 2}]Acetate was incubated with spinach (Spinacia oleracea) leaves and the {sup 18}O content in fatty acid methyl esters isolated from different lipid classes measured by gas chromatography-mass spectrometry. Fatty acids isolated from lipids synthesized within the plastid, such as monogalactosyldiacylglycerol, show an {sup 18}O content consistent with the exogenous acetate undergoing a single activation step and with the direct utilization of acyl-acyl carrier protein by the acyl transferases of the chloroplast. In contrast, fatty acids isolated from lipids assembled in the cytosol, such as phosphatidylcholine, show a 50% reduction in the {sup 18}O content. This is indicative of export of the fatty acyl groups from the plastid via a free carboxylate anion, and is consistent with the acyl-acyl carrier protein thioesterase:acyl-coenzyme A (CoA) synthetase mediated export mechanism. If this were not the case and the acyl group was transferred directly from acyl-acyl carrier protein to an acyl acceptor on the cytosolic side, there would be either complete retention of {sup 18}O or, less likely, complete loss of {sup 18}O, but not a 50% loss of {sup 18}O. Thus, existing models for fatty acid transfer from the plastid and for spatially separate synthesis of prokaryotic and eukaryotic lipids have both been confirmed.

  8. Testing Models of Fatty Acid Transfer and Lipid Synthesis in Spinach Leaf Using in Vivo Oxygen-18 Labeling1

    Science.gov (United States)

    Pollard, Mike; Ohlrogge, John

    1999-01-01

    Oxygen-18 labeling has been applied to the study of plant lipid biosynthesis for the first time. [13C218O2]Acetate was incubated with spinach (Spinacia oleracea) leaves and the 18O content in fatty acid methyl esters isolated from different lipid classes measured by gas chromatography-mass spectometry. Fatty acids isolated from lipids synthesized within the plastid, such as monogalactosyldiacylglycerol, show an 18O content consistent with the exogenous acetate undergoing a single activation step and with the direct utilization of acyl-acyl carrier protein by the acyl transferases of the chloroplast. In contrast, fatty acids isolated from lipids assembled in the cytosol, such as phosphatidylcholine, show a 50% reduction in the 18O content. This is indicative of export of the fatty acyl groups from the plastid via a free carboxylate anion, and is consistent with the acyl-acyl carrier protein thioesterase:acyl-coenzyme A (CoA) synthetase mediated export mechanism. If this were not the case and the acyl group was transferred directly from acyl-acyl carrier protein to an acyl acceptor on the cytosolic side, there would be either complete retention of 18O or, less likely, complete loss of 18O, but not a 50% loss of 18O. Thus, existing models for fatty acid transfer from the plastid and for spatially separate synthesis of “prokaryotic” and “eukaryotic” lipids have both been confirmed. PMID:10594108

  9. Testing models of fatty acid transfer and lipid synthesis in spinach leaf using in vivo oxygen-18 labeling.

    Science.gov (United States)

    Pollard, M; Ohlrogge, J

    1999-12-01

    Oxygen-18 labeling has been applied to the study of plant lipid biosynthesis for the first time. [(13)C(2)(18)O(2)]Acetate was incubated with spinach (Spinacia oleracea) leaves and the (18)O content in fatty acid methyl esters isolated from different lipid classes measured by gas chromatography-mass spectometry. Fatty acids isolated from lipids synthesized within the plastid, such as monogalactosyldiacylglycerol, show an (18)O content consistent with the exogenous acetate undergoing a single activation step and with the direct utilization of acyl-acyl carrier protein by the acyl transferases of the chloroplast. In contrast, fatty acids isolated from lipids assembled in the cytosol, such as phosphatidylcholine, show a 50% reduction in the (18)O content. This is indicative of export of the fatty acyl groups from the plastid via a free carboxylate anion, and is consistent with the acyl-acyl carrier protein thioesterase:acyl-coenzyme A (CoA) synthetase mediated export mechanism. If this were not the case and the acyl group was transferred directly from acyl-acyl carrier protein to an acyl acceptor on the cytosolic side, there would be either complete retention of (18)O or, less likely, complete loss of (18)O, but not a 50% loss of (18)O. Thus, existing models for fatty acid transfer from the plastid and for spatially separate synthesis of "prokaryotic" and "eukaryotic" lipids have both been confirmed. PMID:10594108

  10. Altering small and medium alcohol selectivity in the wax ester synthase.

    Science.gov (United States)

    Barney, Brett M; Ohlert, Janet M; Timler, Jacobe G; Lijewski, Amelia M

    2015-11-01

    The bifunctional wax ester synthase/acyl-coenzyme A:diacylglycerol acyltransferase (WS/DGAT or wax ester synthase) catalyzes the terminal reaction in the bacterial wax ester biosynthetic pathway, utilizing a range of alcohols and fatty acyl-CoAs to synthesize the corresponding wax ester. The wild-type wax ester synthase Maqu_0168 from Marinobacter aquaeolei VT8 exhibits a preference for longer fatty alcohols, while applications with smaller alcohols would yield products with desired biotechnological properties. Small and medium chain length alcohol substrates are much poorer substrates for the native enzyme, which may hinder broad application of the wax ester synthase in many proposed biosynthetic schemes. Developing approaches to improve enzyme activity toward specific smaller alcohol substrates first requires a clear understanding of which amino acids of the primary sequences of these enzymes contribute to substrate specificity in the native enzyme. In this report, we surveyed a range of potential residues and identified the leucine at position 356 and methionine at position 405 in Maqu_0168 as residues that affected selectivity toward small, branched, and aromatic alcohols when substituted with different amino acids. This analysis provides evidence of residues that line the binding site for wax ester synthase, which will aid rational approaches to improve this enzyme with specific substrates.

  11. Structure of a bifunctional alcohol dehydrogenase involved in bioethanol generation in Geobacillus thermoglucosidasius.

    Science.gov (United States)

    Extance, Jonathan; Crennell, Susan J; Eley, Kirstin; Cripps, Roger; Hough, David W; Danson, Michael J

    2013-10-01

    Bifunctional alcohol/aldehyde dehydrogenase (ADHE) enzymes are found within many fermentative microorganisms. They catalyse the conversion of an acyl-coenzyme A to an alcohol via an aldehyde intermediate; this is coupled to the oxidation of two NADH molecules to maintain the NAD(+) pool during fermentative metabolism. The structure of the alcohol dehydrogenase (ADH) domain of an ADHE protein from the ethanol-producing thermophile Geobacillus thermoglucosidasius has been determined to 2.5 Å resolution. This is the first structure to be reported for such a domain. In silico modelling has been carried out to generate a homology model of the aldehyde dehydrogenase domain, and this was subsequently docked with the ADH-domain structure to model the structure of the complete ADHE protein. This model suggests, for the first time, a structural mechanism for the formation of the large multimeric assemblies or `spirosomes' that are observed for this ADHE protein and which have previously been reported for ADHEs from other organisms.

  12. Hybrid Structure of a Dynamic Single-Chain Carboxylase from Deinococcus radiodurans.

    Science.gov (United States)

    Hagmann, Anna; Hunkeler, Moritz; Stuttfeld, Edward; Maier, Timm

    2016-08-01

    Biotin-dependent acyl-coenzyme A (CoA) carboxylases (aCCs) are involved in key steps of anabolic pathways and comprise three distinct functional units: biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and carboxyl transferase (CT). YCC multienzymes are a poorly characterized family of prokaryotic aCCs of unidentified substrate specificity, which integrate all functional units into a single polypeptide chain. We employed a hybrid approach to study the dynamic structure of Deinococcus radiodurans (Dra) YCC: crystal structures of isolated domains reveal a hexameric CT core with extended substrate binding pocket and a dimeric BC domain. Negative-stain electron microscopy provides an approximation of the variable positioning of the BC dimers relative to the CT core. Small-angle X-ray scattering yields quantitative information on the ensemble of Dra YCC structures in solution. Comparison with other carrier protein-dependent multienzymes highlights a characteristic range of large-scale interdomain flexibility in this important class of biosynthetic enzymes.

  13. Orthogonal Fatty Acid Biosynthetic Pathway Improves Fatty Acid Ethyl Ester Production in Saccharomyces cerevisiae.

    Science.gov (United States)

    Eriksen, Dawn T; HamediRad, Mohammad; Yuan, Yongbo; Zhao, Huimin

    2015-07-17

    Fatty acid ethyl esters (FAEEs) are a form of biodiesel that can be microbially produced via a transesterification reaction of fatty acids with ethanol. The titer of microbially produced FAEEs can be greatly reduced by unbalanced metabolism and an insufficient supply of fatty acids, resulting in a commercially inviable process. Here, we report on a pathway engineering strategy in Saccharomyces cerevisiae for enhancing the titer of microbially produced FAEEs by providing the cells with an orthogonal route for fatty acid synthesis. The fatty acids generated from this heterologous pathway would supply the FAEE production, safeguarding endogenous fatty acids for cellular metabolism and growth. We investigated the heterologous expression of a Type-I fatty acid synthase (FAS) from Brevibacterium ammoniagenes coupled with WS/DGAT, the wax ester synthase/acyl-coenzyme that catalyzes the transesterification reaction with ethanol. Strains harboring the orthologous fatty acid synthesis yielded a 6.3-fold increase in FAEE titer compared to strains without the heterologous FAS. Variations in fatty acid chain length and degree of saturation can affect the quality of the biodiesel; therefore, we also investigated the diversity of the fatty acid production profile of FAS enzymes from other Actinomyces organisms. PMID:25594225

  14. Purification of a jojoba embryo wax synthase, cloning of its cDNA, and production of high levels of wax in seeds of transgenic arabidopsis.

    Science.gov (United States)

    Lardizabal, K D; Metz, J G; Sakamoto, T; Hutton, W C; Pollard, M R; Lassner, M W

    2000-03-01

    Wax synthase (WS, fatty acyl-coenzyme A [coA]: fatty alcohol acyltransferase) catalyzes the final step in the synthesis of linear esters (waxes) that accumulate in seeds of jojoba (Simmondsia chinensis). We have characterized and partially purified this enzyme from developing jojoba embryos. A protein whose presence correlated with WS activity during chromatographic fractionation was identified and a cDNA encoding that protein was cloned. Seed-specific expression of the cDNA in transgenic Arabidopsis conferred high levels of WS activity on developing embryos from those plants. The WS sequence has significant homology with several Arabidopsis open reading frames of unknown function. Wax production in jojoba requires, in addition to WS, a fatty acyl-CoA reductase (FAR) and an efficient fatty acid elongase system that forms the substrates preferred by the FAR. We have expressed the jojoba WS cDNA in Arabidopsis in combination with cDNAs encoding the jojoba FAR and a beta-ketoacyl-CoA synthase (a component of fatty acid elongase) from Lunaria annua. (13)C-Nuclear magnetic resonance analysis of pooled whole seeds from transgenic plants indicated that as many as 49% of the oil molecules in the seeds were waxes. Gas chromatography analysis of transmethylated oil from individual seeds suggested that wax levels may represent up to 70% (by weight) of the oil present in those seeds.

  15. Changes in cardiac substrate transporters and metabolic proteins mirror the metabolic shift in patients with aortic stenosis.

    Directory of Open Access Journals (Sweden)

    Lisa C Heather

    Full Text Available In the hypertrophied human heart, fatty acid metabolism is decreased and glucose utilisation is increased. We hypothesized that the sarcolemmal and mitochondrial proteins involved in these key metabolic pathways would mirror these changes, providing a mechanism to account for the modified metabolic flux measured in the human heart. Echocardiography was performed to assess in vivo hypertrophy and aortic valve impairment in patients with aortic stenosis (n = 18. Cardiac biopsies were obtained during valve replacement surgery, and used for western blotting to measure metabolic protein levels. Protein levels of the predominant fatty acid transporter, fatty acid translocase (FAT/CD36 correlated negatively with levels of the glucose transporters, GLUT1 and GLUT4. The decrease in FAT/CD36 was accompanied by decreases in the fatty acid binding proteins, FABPpm and H-FABP, the β-oxidation protein medium chain acyl-coenzyme A dehydrogenase, the Krebs cycle protein α-ketoglutarate dehydrogenase and the oxidative phosphorylation protein ATP synthase. FAT/CD36 and complex I of the electron transport chain were downregulated, whereas the glucose transporter GLUT4 was upregulated with increasing left ventricular mass index, a measure of cardiac hypertrophy. In conclusion, coordinated downregulation of sequential steps involved in fatty acid and oxidative metabolism occur in the human heart, accompanied by upregulation of the glucose transporters. The profile of the substrate transporters and metabolic proteins mirror the metabolic shift from fatty acid to glucose utilisation that occurs in vivo in the human heart.

  16. 3,5-Diiodo-L-Thyronine Modifies the Lipid Droplet Composition in a Model of Hepatosteatosis

    Directory of Open Access Journals (Sweden)

    Elena Grasselli

    2014-02-01

    Full Text Available Background/Aims: Fatty acids are the main energy stores and the major membrane components of the cells. In the hepatocyte, fatty acids are esterified to triacylglycerols (TAGs and stored in lipid droplets (LDs. The lipid lowering action of 3,5-diiodo-L-thyronine (T2 on an in vitro model of hepatosteatosis was investigated in terms of fatty acid and protein content of LDs, lipid oxidation and secretion. Methods: FaO cells were exposed to oleate/palmitate, then treated with T2. Results: T2 reduced number and size of LDs, and modified their acyl composition by decreasing the content of saturated (SFA vs monounsaturated (MUFA fatty acids thus reversing the SFA/MUFA ratio. The expression of the LD-associated proteins adipose differentiation-related protein (ADRP, oxidative tissue-enriched PAT protein (OXPAT, and adipose triglyceride lipase (ATGL was increased in ‘steatotic' cells and further up-regulated by T2. Moreover, T2 stimulated the mitochondrial oxidation by up-regulating carnitine-palmitoyl-transferase (CPT1, uncoupling protein 2 (UCP2 and very long-chain acyl-coenzyme A dehydrogenase (VLCAD. Conclusions: T2 leads to mobilization of TAGs from LDs and stimulates mitochondrial oxidative metabolism of fatty acids, in particular of SFAs, and thus enriches of MUFAs the LDs. This action may protect the hepatocyte from excess of SFAs that are more toxic than MUFAs.

  17. An overview of the new frontiers in the treatment of atherogenic dyslipidemias.

    Science.gov (United States)

    Rached, F H; Chapman, M J; Kontush, A

    2014-07-01

    Cardiovascular diseases (CVDs) are the leading cause of morbidity/mortality worldwide. Dyslipidemia is a major risk factor for premature atherosclerosis and CVD. Lowering low-density-lipoprotein cholesterol (LDL-C) levels is well established as an intervention for the reduction of CVDs. Statins are the first-line drugs for treatment of dyslipidemia, but they do not address all CVD risk. Development of novel therapies is ongoing and includes the following: (i) reduction of LDL-C concentrations using antibodies to proprotein convertase subtilisin/kexin-9, antisense oligonucleotide inhibitors of apolipoprotein B production, microsomal transfer protein (MTP) inhibitors, and acyl-coenzyme A cholesterol acyl transferase inhibitors; (ii) reduction in levels of triglyceride-rich lipoproteins with ω-3 fatty acids, MTP inhibitors, and diacylglycerol acyl transferase-1 inhibitors; and (iii) increase of high-density-lipoprotein (HDL) cholesterol levels, HDL particle numbers, and/or HDL functionality using cholesteryl ester transfer protein inhibitors, HDL-derived agents, apolipoprotein AI mimetic peptides, and microRNAs. Large prospective outcome trials of several of these emerging therapies are under way, and thrilling progress in the field of lipid management is anticipated.

  18. A novel RET rearrangement (ACBD5/RET) by pericentric inversion, inv(10)(p12.1;q11.2), in papillary thyroid cancer from an atomic bomb survivor exposed to high-dose radiation.

    Science.gov (United States)

    Hamatani, Kiyohiro; Eguchi, Hidetaka; Koyama, Kazuaki; Mukai, Mayumi; Nakachi, Kei; Kusunoki, Yoichiro

    2014-11-01

    During analysis of RET/PTC rearrangements in papillary thyroid cancer (PTC) among atomic bomb survivors, a cDNA fragment of a novel type of RET rearrangement was identified in a PTC patient exposed to a high radiation dose using the improved 5' RACE method. This gene resulted from the fusion of the 3' portion of RET containing tyrosine kinase domain to the 5' portion of the acyl-coenzyme A binding domain containing 5 (ACBD5) gene, by pericentric inversion inv(10)(p12.1;q11.2); expression of the fusion gene was confirmed by RT-PCR. ACBD5 gene is ubiquitously expressed in various human normal tissues including thyroid. Full-length cDNA of the ACBD5-RET gene was constructed and then examined for tumorigenicity. Enhanced phosphorylation of ERK proteins in the MAPK pathway was observed in NIH3T3 cells transfected with expression vector encoding the full-length ACBD5/RET cDNA, while this was not observed in the cells transfected with empty expression vector. Stable NIH3T3 transfectants with ACBD5-RET cDNA induced tumor formation after their injection into nude mice. These findings suggest that the ACBD5-RET rearrangement is causatively involved in the development of PTC. PMID:25175022

  19. The effect of chronic exposure to high palmitic acid concentrations on the aerobic metabolism of human endothelial EA.hy926 cells.

    Science.gov (United States)

    Broniarek, Izabela; Koziel, Agnieszka; Jarmuszkiewicz, Wieslawa

    2016-09-01

    A chronic elevation of circulating free fatty acids (FFAs) is associated with diseases like obesity or diabetes and can lead to lipotoxicity. The goals of this study were to assess the influence of chronic exposure to high palmitic acid (PAL) levels on mitochondrial respiratory functions in endothelial cells and isolated mitochondria. Human umbilical vein endothelial cells (EA.hy926 line) were grown for 6 days in a medium containing either 100 or 150 μM PAL. Growth at high PAL concentrations induced a considerable increase in fatty acid-supplied respiration and a reduction of mitochondrial respiration during carbohydrate and glutamine oxidation. High PAL levels elevated intracellular and mitochondrial superoxide generation; increased inflammation marker, acyl-coenzyme A (CoA) dehydrogenase, uncoupling protein 2 (UCP2), and superoxide dismutase 2 expression; and decreased hexokinase I and pyruvate dehydrogenase expression. No change in aerobic respiration capacity was observed, while fermentation was decreased. In mitochondria isolated from high PAL-treated cells, an increase in the oxidation of palmitoylcarnitine, a decrease in the oxidation of pyruvate, and an increase in UCP2 activity were observed. Our results demonstrate that exposure to high PAL levels induces a shift in endothelial aerobic metabolism toward the oxidation of fatty acids. Increased levels of PAL caused impairment and uncoupling of the mitochondrial oxidative phosphorylation system. Our data indicate that FFAs significantly affect endothelial oxidative metabolism, reactive oxygen species (ROS) formation, and cell viability and, thus, might contribute to endothelial and vascular dysfunction. PMID:27417103

  20. De novo biosynthesis of biodiesel by Escherichia coli in optimized fed-batch cultivation.

    Directory of Open Access Journals (Sweden)

    Yangkai Duan

    Full Text Available Biodiesel is a renewable alternative to petroleum diesel fuel that can contribute to carbon dioxide emission reduction and energy supply. Biodiesel is composed of fatty acid alkyl esters, including fatty acid methyl esters (FAMEs and fatty acid ethyl esters (FAEEs, and is currently produced through the transesterification reaction of methanol (or ethanol and triacylglycerols (TAGs. TAGs are mainly obtained from oilseed plants and microalgae. A sustainable supply of TAGs is a major bottleneck for current biodiesel production. Here we report the de novo biosynthesis of FAEEs from glucose, which can be derived from lignocellulosic biomass, in genetically engineered Escherichia coli by introduction of the ethanol-producing pathway from Zymomonas mobilis, genetic manipulation to increase the pool of fatty acyl-CoA, and heterologous expression of acyl-coenzyme A: diacylglycerol acyltransferase from Acinetobacter baylyi. An optimized fed-batch microbial fermentation of the modified E. coli strain yielded a titer of 922 mg L(-1 FAEEs that consisted primarily of ethyl palmitate, -oleate, -myristate and -palmitoleate.

  1. Beneficial effects of curcumin on hyperlipidemia and insulin resistance in high-fat-fed hamsters.

    Science.gov (United States)

    Jang, Eun-Mi; Choi, Myung-Sook; Jung, Un Ju; Kim, Myung-Joo; Kim, Hye-Jin; Jeon, Seon-Min; Shin, Su-Kyung; Seong, Chi-Nam; Lee, Mi-Kyung

    2008-11-01

    This study investigated the effect of curcumin (0.05-g/100-g diet) supplementation on a high-fat diet (10% coconut oil, 0.2% cholesterol, wt/wt) fed to hamsters, one of the rodent species that are most closely related to humans in lipid metabolism. Curcumin significantly lowered the levels of free fatty acid, total cholesterol, triglyceride, and leptin and the homeostasis model assessment of insulin resistance index, whereas it elevated the levels of high-density lipoprotein cholesterol and apolipoprotein (apo) A-I and paraoxonase activity in plasma, compared with the control group. The levels of hepatic cholesterol and triglyceride were also lower in the curcumin group than in the control group. In the liver, fatty acid beta-oxidation activity was significantly higher in the curcumin group than in the control group, whereas fatty acid synthase, 3-hydroxy-3-methylglutaryl coenzyme A reductase, and acyl coenzyme A:cholesterol acyltransferase activities were significantly lower. Curcumin significantly lowered the lipid peroxide levels in the erythrocyte and liver compared with the control group. These results indicate that curcumin exhibits an obvious hypolipidemic effect by increasing plasma paraoxonase activity, ratios of high-density lipoprotein cholesterol to total cholesterol and of apo A-I to apo B, and hepatic fatty acid oxidation activity with simultaneous inhibition of hepatic fatty acid and cholesterol biosynthesis in high-fat-fed hamsters.

  2. Transcriptional analysis of sex pheromone biosynthesis signal genes in Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Shi-Heng An; Meng-Fang Du; Li-Juan Su; Xin-Ming Yin

    2012-01-01

    Six sex pheromone synthesis signal genes,including acyl coenzyme A (acylCoA) desaturase (desatl),fatty acyl reductase (FAR),pheromone biosynthesis activating neuropeptide receptor (PBANR),fatty acid transport protein (FATP),acyl-CoA binding protein (ACBP) and store-operated channel protein (OrailA),were studied for their transcriptional regulations.The expression profiles of these transcripts at different developmental stages (from-96 to 48 h) revealed that the genes are expressed in an age-dependent manner.The transcripts of these genes continued to increase despite decapitation,and compared with normally developmental females,decapitation significantly inhibited their expression.Further experiments with a methoprene,a juvenile hormone (JH) analogue,challenge showed that JH was not a key inhibiting factor in the expression of these genes,and mating was found to significantly inhibit the expression of these marker genes.Altogether,the results provide a reference for understanding the mechanism of sex pheromone synthesis.

  3. Rimonabant is a dual inhibitor of acyl CoA:cholesterol acyltransferases 1 and 2.

    Science.gov (United States)

    Netherland, Courtney; Thewke, Douglas P

    2010-08-01

    Acyl coenzyme A:cholesterol acyltransferase (ACAT) catalyzes the intracellular synthesis of cholesteryl esters (CE). Both ACAT isoforms, ACAT1 and ACAT2, play key roles in the pathophysiology of atherosclerosis and ACAT inhibition retards atherosclerosis in animal models. Rimonabant, a type 1 cannabinoid receptor (CB1) antagonist, produces anti-atherosclerotic effects in humans and animals by mechanisms which are not completely understood. Rimonabant is structurally similar to two other cannabinoid receptor antagonists, AM251 and SR144528, recently identified as potent inhibitors of ACAT. Therefore, we examined the effects of Rimonabant on ACAT using both in vivo cell-based assays and in vitro cell-free assays. Rimonabant dose-dependently reduced ACAT activity in Raw 264.7 macrophages (IC(50)=2.9+/-0.38 microM) and isolated peritoneal macrophages. Rimonabant inhibited ACAT activity in intact CHO-ACAT1 and CHO-ACAT2 cells and in cell-free assays with approximately equal efficiency (IC(50)=1.5+/-1.2 microM and 2.2+/-1.1 microM for CHO-ACAT1 and CHO-ACAT2, respectively). Consistent with ACAT inhibition, Rimonabant treatment blocked ACAT-dependent processes in macrophages, oxysterol-induced apoptosis and acetylated-LDL induced foam cell formation. From these results we conclude that Rimonabant is an ACAT1/2 dual inhibitor and suggest that some of the atherosclerotic beneficial effects of Rimonabant are, at least partly, due to inhibition of ACAT. PMID:20609360

  4. Taurine reduces the secretion of apolipoprotein B100 and lipids in HepG2 cells

    Directory of Open Access Journals (Sweden)

    Nagao Koji

    2008-10-01

    Full Text Available Abstract Background Higher concentrations of serum lipids and apolipoprotein B100 (apoB are major individual risk factors of atherosclerosis and coronary heart disease. Therefore ameliorative effects of food components against the diseases are being paid attention in the affluent countries. The present study was undertaken to investigate the effect of taurine on apoB secretion and lipid metabolism in human liver model HepG2 cells. Results The results demonstrated that an addition of taurine to the culture media reduces triacylglycerol (TG-mass in the cells and the medium. Similarly, cellular cholesterol-mass was decreased. Taurine inhibited the incorporation of [14C] oleate into cellular and medium TG, suggesting the inhibition of TG synthesis. In addition, taurine reduced the synthesis of cellular cholesterol ester and its secretion, suggesting the inhibition of acyl-coenzyme A:cholesterol acyltransferase activity. Furthermore, taurine reduced the secretion of apoB, which is a major protein component of very low-density lipoprotein. Conclusion This is a first report to demonstrate that taurine inhibits the secretion of apoB from HepG2 cells.

  5. Cyclic vomiting syndrome masking a fatal metabolic disease.

    LENUS (Irish Health Repository)

    Fitzgerald, Marianne

    2013-05-01

    Disorders of fatty acid oxidation are rare but can be fatal. Hypoglycaemia with acidosis is a cardinal feature. Cases may present during early childhood or can be delayed into adolescence or beyond. We present a case of multiple acyl-coenzyme A dehydrogenase deficiency (MADD), an extremely rare disorder of fatty acid oxidation. Our 20-year-old patient presented with cardiovascular collapse, raised anion gap metabolic acidosis and non-ketotic hypoglycaemia. She subsequently developed multi-organ failure and sadly died. She had a previous diagnosis of cyclic vomiting syndrome (CVS) for more than 10 years, warranting frequent hospital admissions. The association between CVS and MADD has been made before though the exact relationship is unclear. All patients with persistent severe CVS should have metabolic investigations to exclude disorders of fatty acid oxidation. In case of non-ketotic hypoglycaemia with acidosis, the patient should be urgently referred to a specialist in metabolic diseases. All practitioners should be aware of these rare disorders as a cause of unexplained acidosis.

  6. Carnitine deficiency and its related diseases%肉碱缺乏及其相关疾病的研究进展

    Institute of Scientific and Technical Information of China (English)

    房玥晖; 蔡美琴

    2009-01-01

    L-carnitine plays an essential role in the beta-oxidation of fatty acids by transporting long chain acyl-coenzyme A into the mitochondrial matrix.Carnitine deficiency may lead to various diseases,including lipid storage myopathies,systemic carnitine deficiency syndrome,cardiomyopathy,obesity,and infertility.This article summarizes the causes of carnitine deficiency and elucidates the clinical features and treatment strategies of its related diseases.%肉碱是脂肪β-氧化过程中长链脂酰辅酶A透过线粒体内膜时的转运体,其缺乏导致脂质沉积性肌病、全身肉碱缺乏综合征、心肌病、肥胖、男性不育等疾病.本文主要综述肉碱缺乏的原因及所导致的相关疾病的主要临床表现和治疗手段.

  7. Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli.

    Energy Technology Data Exchange (ETDEWEB)

    Lennen, Rebecca M.; Kruziki, Max A.; Kumar, Kritika; Zinkel, Robert A.; Burnum, Kristin E.; Lipton, Mary S.; Hoover, Spencer W.; Ranatunga, Don Ruwan; Wittkopp, Tyler M.; Marner II, Wesley D.; Pfleger, Brian F.

    2011-11-01

    Microbially produced fatty acids are potential precursors to high energy density biofuels, including alkanes and alkyl ethyl esters by either catalytic conversion of free fatty acids (FFAs) or enzymatic conversions of acyl-acyl carrier protein or acyl-coenzyme A intermediates. Metabolic engineering efforts aimed at overproducing FFAs in Escherichia coli have achieved less than 30% of the maximum theoretical yield on the supplied carbon source. In this work, the viability, morphology, transcript levels, and protein levels of a strain of E. coli that overproduces medium chain length FFAs was compared to an engineered control strain. By early stationary phase, an 85% reduction in viable cell counts and exacerbated loss of inner membrane integrity were observed in the FFA overproducing strain. These effects were enhanced in strains endogenously producing FFAs compared to strains exposed to exogenously fed FFAs. Under two sets of cultivation conditions, long chain unsaturated fatty acid content greatly increased and the expression of genes and proteins required for unsaturated fatty acid biosynthesis were significantly decreased. Membrane stresses were further implicated by increased expression of genes and proteins of the phage shock response, the MarA/Rob/SoxS regulon, and the nuo and cyo operons of aerobic respiration. Gene deletion studies confirmed the importance of the phage shock proteins and Rob for maintaining cell viability, however little to no change in FFA titers was observed after 24 h cultivation. The results of this study serve as a baseline for future targeted attempts to improve FFA yields and titers in E. coli.

  8. Statins Activate Human PPAR Promoter and Increase PPAR mRNA Expression and Activation in HepG2 Cells

    Directory of Open Access Journals (Sweden)

    Makoto Seo

    2008-01-01

    Full Text Available Statins increase peroxisome proliferator-activated receptor (PPAR mRNA expression, but the mechanism of this increased PPAR production remains elusive. To examine the regulation of PPAR production, we examined the effect of 7 statins (atorvastatin, cerivastatin, fluvastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin on human PPAR promoter activity, mRNA expression, nuclear protein levels, and transcriptional activity. The main results are as follows. (1 Majority of statins enhanced PPAR promoter activity in a dose-dependent manner in HepG2 cells transfected with the human PPAR promoter. This enhancement may be mediated by statin-induced HNF-4. (2 PPAR mRNA expression was increased by statin treatment. (3 The PPAR levels in nuclear fractions were increased by statin treatment. (4 Simvastatin, pravastatin, and cerivastatin markedly enhanced transcriptional activity in 293T cells cotransfected with acyl-coenzyme A oxidase promoter and PPAR/RXR expression vectors. In summary, these data demonstrate that PPAR production and activation are upregulated through the PPAR promoter activity by statin treatment.

  9. ATP-binding cassette transporters and sterol O-acyltransferases interact at membrane microdomains to modulate sterol uptake and esterification.

    Science.gov (United States)

    Gulati, Sonia; Balderes, Dina; Kim, Christine; Guo, Zhongmin A; Wilcox, Lisa; Area-Gomez, Estela; Snider, Jamie; Wolinski, Heimo; Stagljar, Igor; Granato, Juliana T; Ruggles, Kelly V; DeGiorgis, Joseph A; Kohlwein, Sepp D; Schon, Eric A; Sturley, Stephen L

    2015-11-01

    A key component of eukaryotic lipid homeostasis is the esterification of sterols with fatty acids by sterol O-acyltransferases (SOATs). The esterification reactions are allosterically activated by their sterol substrates, the majority of which accumulate at the plasma membrane. We demonstrate that in yeast, sterol transport from the plasma membrane to the site of esterification is associated with the physical interaction of the major SOAT, acyl-coenzyme A:cholesterol acyltransferase (ACAT)-related enzyme (Are)2p, with 2 plasma membrane ATP-binding cassette (ABC) transporters: Aus1p and Pdr11p. Are2p, Aus1p, and Pdr11p, unlike the minor acyltransferase, Are1p, colocalize to sterol and sphingolipid-enriched, detergent-resistant microdomains (DRMs). Deletion of either ABC transporter results in Are2p relocalization to detergent-soluble membrane domains and a significant decrease (53-36%) in esterification of exogenous sterol. Similarly, in murine tissues, the SOAT1/Acat1 enzyme and activity localize to DRMs. This subcellular localization is diminished upon deletion of murine ABC transporters, such as Abcg1, which itself is DRM associated. We propose that the close proximity of sterol esterification and transport proteins to each other combined with their residence in lipid-enriched membrane microdomains facilitates rapid, high-capacity sterol transport and esterification, obviating any requirement for soluble intermediary proteins.

  10. The role of lipotoxicity in smoke cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Priscila P Santos

    Full Text Available BACKGROUND/AIMS: Experimental and clinical studies have shown the direct toxic effects of cigarette smoke (CS on the myocardium, independent of vascular effects. However, the underlying mechanisms are not well known. METHODS: Wistar rats were allocated to control (C and cigarette smoke (CS groups. CS rats were exposed to cigarette smoke for 2 months. RESULTS: After that morphometric, functional and biochemical parameters were measured. The echocardiographic study showed enlargement of the left atria, increase in the left ventricular systolic volume and reduced systolic function. Within the cardiac metabolism, exposure to CS decreased beta hydroxy acyl coenzyme A dehydrogenases and citrate synthases and increased lactate dehydrogenases. Peroxisome proliferator-activated receptor alpha (PPARα and peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α were expressed similarly in both groups. CS increased serum lipids and myocardial triacylglycerols (TGs. These data suggest that impairment in fatty acid oxidation and the accumulation of cardiac lipids characterize lipotoxicity. CS group exhibited increased oxidative stress and decreased antioxidant defense. Finally, the myocyte cross-sectional area and active Caspase 3 were increased in the CS group. CONCLUSION: The cardiac remodeling that was observed in the CS exposure model may be explained by abnormalities in energy metabolism, including lipotoxicity and oxidative stress.

  11. Functional PDF Signaling in the Drosophila Circadian Neural Circuit Is Gated by Ral A-Dependent Modulation.

    Science.gov (United States)

    Klose, Markus; Duvall, Laura B; Li, Weihua; Liang, Xitong; Ren, Chi; Steinbach, Joe Henry; Taghert, Paul H

    2016-05-18

    The neuropeptide PDF promotes the normal sequencing of circadian behavioral rhythms in Drosophila, but its signaling mechanisms are not well understood. We report daily rhythmicity in responsiveness to PDF in critical pacemakers called small LNvs. There is a daily change in potency, as great as 10-fold higher, around dawn. The rhythm persists in constant darkness and does not require endogenous ligand (PDF) signaling or rhythmic receptor gene transcription. Furthermore, rhythmic responsiveness reflects the properties of the pacemaker cell type, not the receptor. Dopamine responsiveness also cycles, in phase with that of PDF, in the same pacemakers, but does not cycle in large LNv. The activity of RalA GTPase in s-LNv regulates PDF responsiveness and behavioral locomotor rhythms. Additionally, cell-autonomous PDF signaling reversed the circadian behavioral effects of lowered RalA activity. Thus, RalA activity confers high PDF responsiveness, providing a daily gate around the dawn hours to promote functional PDF signaling. PMID:27161526

  12. A-dependence for the charmed meson production; Dependencia em A para a producao de mesons charmosos

    Energy Technology Data Exchange (ETDEWEB)

    Alves, Gilvan Augusto

    1992-03-01

    A report is presented of a recent direct measurement of the nucleon number ({lambda}) dependence of the production cross sections for the charmed mesons D{sup 0} and D{sup +} using {pi}{sup +-} beams incident on a segmented target of Be, Al, Cu and W. The data derive from the experiment E769 - Hadroproduction of Charm - at Fermilab. The experimental apparatus is described together with the following analysis. Starting from a sample of {approx} 1500 D mesons in the range of O < X{sub F} < 1, the data are found to be well described by the parameterization {sigma}{sub A} = {sigma}{sub O} A{sup {alpha}}, with {alpha} = 0.99 {+-} 0.03. The X{sub F} dependence of {alpha} is examined and the results obtained are compared with those of other experiments and with theoretical expectations based on perturbative QCD and on an EMC like model of nuclear shadowing. (author). 85 refs, 61 figs, 22 tabs.

  13. A condition-based maintenance of a dependent degradation-threshold-shock model in a system with multiple degradation processes

    International Nuclear Information System (INIS)

    This paper proposes a condition-based maintenance strategy for a system subject to two dependent causes of failure: degradation and sudden shocks. The internal degradation is reflected by the presence of multiple degradation processes in the system. Degradation processes start at random times following a Non-homogeneous Poisson process and their growths are modelled by using a gamma process. When the deterioration level of a degradation process exceeds a predetermined value, we assume that a degradation failure occurs. Furthermore, the system is subject to sudden shocks that arrive at the system following a Doubly Stochastic Poisson Process. A sudden shock provokes the total breakdown of the system. Thus, the state of the system is evaluated at inspection times and different maintenance tasks can be carried out. If the system is still working at an inspection time, a preventive maintenance task is performed if the deterioration level of a degradation process exceeds a certain threshold. A corrective maintenance task is performed if the system is down at an inspection time. A preventive (corrective) maintenance task implies the replacement of the system by a new one. Under this maintenance strategy, the expected cost rate function is obtained. A numerical example illustrates the analytical results. - Highlights: • A condition-based maintenance model is proposed. • Two dependent causes of failure are considered: deterioration and external shocks. • Deterioration is given by multiple degradation processes growing by a gamma process. • The initiation of degradation processes follows a Non-homogeneous Poisson process. • External shocks arrive at the system by using a Doubly Stochastic Poisson Process

  14. SecA localization and SecA-dependent secretion occurs at new division septa in group B Streptococcus.

    Directory of Open Access Journals (Sweden)

    Sara Brega

    Full Text Available Exported proteins of Streptococcus agalactiae (GBS, which include proteins localized to the bacterial surface or secreted into the extracellular environment, are key players for commensal and pathogenic interactions in the mammalian host. These proteins are transported across the cytoplasmic membrane via the general SecA secretory pathway and those containing the so-called LPXTG sorting motif are covalently attached to the peptidoglycan by sortase A. How SecA, sortase A, and LPXTG proteins are spatially distributed in GBS is not known. In the close relative Streptococcus pyogenes, it was shown that presence of the YSIRKG/S motif (literally YSIRKX3Gx2S in the signal peptide (SP constitutes the targeting information for secretion at the septum. Here, using conventional and deconvolution immunofluorescence analyses, we have studied in GBS strain NEM316 the localization of SecA, SrtA, and the secreted protein Bsp whose signal peptide contains a canonical YSIRKG/S motif (YSLRKykfGlaS. Replacing the SP of Bsp with four other SPs containing or not the YSIRKG/S motif did not alter the localized secretion of Bsp at the equatorial ring. Our results indicate that secretion and cell wall-anchoring machineries are localized at the division septum. Cell wall- anchored proteins displayed polar (PilB, Gbs0791, punctuate (CspA or uniform distribution (Alp2 on the bacterial surface. De novo secretion of Gbs0791 following trypsin treatment indicates that it is secreted at the septum, then redistributed along the lateral sides, and finally accumulated to the poles. We conclude that the ±YSIRK SP rule driving compartimentalized secretion is not true in S. agalactiae.

  15. Early Loss of Telomerase Action in Yeast Creates a Dependence on the DNA Damage Response Adaptor Proteins.

    Science.gov (United States)

    Jay, Kyle A; Smith, Dana L; Blackburn, Elizabeth H

    2016-07-15

    Telomeres cap the ends of chromosomes, protecting them from degradation and inappropriate DNA repair processes that can lead to genomic instability. A short telomere elicits increased telomerase action on itself that replenishes telomere length, thereby stabilizing the telomere. In the prolonged absence of telomerase activity in dividing cells, telomeres eventually become critically short, inducing a permanent cell cycle arrest (senescence). We recently showed that even early after telomerase inactivation (ETI), yeast cells have accelerated mother cell aging and mildly perturbed cell cycles. Here, we show that the complete disruption of DNA damage response (DDR) adaptor proteins in ETI cells causes severe growth defects. This synthetic-lethality phenotype was as pronounced as that caused by extensive DNA damage in wild-type cells but showed genetic dependencies distinct from such damage and was completely alleviated by SML1 deletion, which increases deoxynucleoside triphosphate (dNTP) pools. Our results indicated that these deleterious effects in ETI cells cannot be accounted for solely by the slow erosion of telomeres due to incomplete replication that leads to senescence. We propose that normally occurring telomeric DNA replication stress is resolved by telomerase activity and the DDR in two parallel pathways and that deletion of Sml1 prevents this stress. PMID:27161319

  16. A dependência da política: Fernando Henrique Cardoso e a sociologia no Brasil

    Directory of Open Access Journals (Sweden)

    Afrânio Garcia Jr.

    2004-06-01

    Full Text Available Este artigo analisa a trajetória social, intelectual e profissional de Fernando Henrique Cardoso para entender os diferentes recursos sociais e disposições pessoais utilizados em sua carreira como sociólogo e em suas atividades como especialista da política. Busca demonstrar que os capitais sociais e as disposições responsáveis pelo prestígio como pesquisador e professor de ciências sociais foram distintos dos aplicados no domínio da política, permitindo sua rápida ascensão à presidência da República. Depois de estudar suas origens familiares, focaliza seus investimentos escolares e a escolha do ofício de sociólogo, a carreira promissora sendo bloqueada pelo golpe de 1964. O exílio permitiu a extensão das atividades e o reconhecimento internacional, reinvestidos em novo concurso para a USP; o AI-5 o conduzirá à dupla condição de cientista social e de um dos líderes da frente de oposições aos militares. Por fim, analisa-se a reconversão de seus recursos sociais e pessoais na profissão política.This article analyzes Fernando Henrique Cardoso's social, intellectual and professional trajectory in order to understand the different social resources and personal dispositions carried out in his career as sociologist or in his activities as a professional politician. It seeks to prove that the social capitals and the dispositions that might explain his prestige as researcher and professor of social sciences were very different from those required in the political domain, the ones allowing his fast rise to be the president of the Republic. After having examined his family origins, it focuses his scholar investments and the choice of the sociologist's occupation, a promising career suddenly blocked by the 1964 military coup. The exile encouraged new initiatives and brought him international appraisal, this moment being crowned with his access to the chair of political science at the University of São Paulo; the AI-5 enforced by the military rulers will enable him to assume a double condition, as social scientist and as an important opposition leader facing the military. Finally, it analyzes how he was able to reconvert his social and personal resources into the political profession.

  17. The RecA-Dependent SOS Response Is Active and Required for Processing of DNA Damage during Bacillus subtilis Sporulation.

    Science.gov (United States)

    Ramírez-Guadiana, Fernando H; Barajas-Ornelas, Rocío Del Carmen; Corona-Bautista, Saúl U; Setlow, Peter; Pedraza-Reyes, Mario

    2016-01-01

    The expression of and role played by RecA in protecting sporulating cells of Bacillus subtilis from DNA damage has been determined. Results showed that the DNA-alkylating agent Mitomycin-C (M-C) activated expression of a PrecA-gfpmut3a fusion in both sporulating cells' mother cell and forespore compartments. The expression levels of a recA-lacZ fusion were significantly lower in sporulating than in growing cells. However, M-C induced levels of ß-galactosidase from a recA-lacZ fusion ~6- and 3-fold in the mother cell and forespore compartments of B. subtilis sporangia, respectively. Disruption of recA slowed sporulation and sensitized sporulating cells to M-C and UV-C radiation, and the M-C and UV-C sensitivity of sporangia lacking the transcriptional repair-coupling factor Mfd was significantly increased by loss of RecA. We postulate that when DNA damage is encountered during sporulation, RecA activates the SOS response thus providing sporangia with the repair machinery to process DNA lesions that may compromise the spatio-temporal expression of genes that are essential for efficient spore formation.

  18. The RecA-Dependent SOS Response Is Active and Required for Processing of DNA Damage during Bacillus subtilis Sporulation.

    Directory of Open Access Journals (Sweden)

    Fernando H Ramírez-Guadiana

    Full Text Available The expression of and role played by RecA in protecting sporulating cells of Bacillus subtilis from DNA damage has been determined. Results showed that the DNA-alkylating agent Mitomycin-C (M-C activated expression of a PrecA-gfpmut3a fusion in both sporulating cells' mother cell and forespore compartments. The expression levels of a recA-lacZ fusion were significantly lower in sporulating than in growing cells. However, M-C induced levels of ß-galactosidase from a recA-lacZ fusion ~6- and 3-fold in the mother cell and forespore compartments of B. subtilis sporangia, respectively. Disruption of recA slowed sporulation and sensitized sporulating cells to M-C and UV-C radiation, and the M-C and UV-C sensitivity of sporangia lacking the transcriptional repair-coupling factor Mfd was significantly increased by loss of RecA. We postulate that when DNA damage is encountered during sporulation, RecA activates the SOS response thus providing sporangia with the repair machinery to process DNA lesions that may compromise the spatio-temporal expression of genes that are essential for efficient spore formation.

  19. Shear stress stimulates phosphorylation of eNOS at Ser(635) by a protein kinase A-dependent mechanism

    Science.gov (United States)

    Boo, Yong Chool; Hwang, Jinah; Sykes, Michelle; Michell, Belinda J.; Kemp, Bruce E.; Lum, Hazel; Jo, Hanjoong

    2002-01-01

    Shear stress stimulates nitric oxide (NO) production by phosphorylating endothelial NO synthase (eNOS) at Ser(1179) in a phosphoinositide-3-kinase (PI3K)- and protein kinase A (PKA)-dependent manner. The eNOS has additional potential phosphorylation sites, including Ser(116), Thr(497), and Ser(635). Here, we studied these potential phosphorylation sites in response to shear, vascular endothelial growth factor (VEGF), and 8-bromocAMP (8-BRcAMP) in bovine aortic endothelial cells (BAEC). All three stimuli induced phosphorylation of eNOS at Ser(635), which was consistently slower than that at Ser(1179). Thr(497) was rapidly dephosphorylated by 8-BRcAMP but not by shear and VEGF. None of the stimuli phosphorylated Ser(116). Whereas shear-stimulated Ser(635) phosphorylation was not affected by phosphoinositide-3-kinase inhibitors wortmannin and LY-294002, it was blocked by either treating the cells with a PKA inhibitor H89 or infecting them with a recombinant adenovirus-expressing PKA inhibitor. These results suggest that shear stress stimulates eNOS by two different mechanisms: 1) PKA- and PI3K-dependent and 2) PKA-dependent but PI3K-independent pathways. Phosphorylation of Ser(635) may play an important role in chronic regulation of eNOS in response to mechanical and humoral stimuli.

  20. Long-chain acyl-CoA-dependent regulation of gene expression in bacteria, yeast and mammals

    DEFF Research Database (Denmark)

    Black, P N; Færgeman, Nils J.; DiRusso, C C

    2000-01-01

    Fatty acyl-CoA thioesters are essential intermediates in lipid metabolism. For many years there have been numerous conflicting reports concerning the possibility that these compounds also serve regulatory functions. In this review, we examine the evidence that long-chain acyl-CoA is a regulatory ...

  1. The effect of growth conditions on inducible, recA-dependent resistance to X rays in Escherichia coli

    International Nuclear Information System (INIS)

    Escherichia coli cells grown to logarithmic phase in, and plated on, rich medium (yeast extract-nutrient broth) were more resistant to X rays, ultraviolet (uv) radiation, and methyl methanesulfonate (MMS) than cells grown in, and plated on, minimal medium. We have called this enhanced survival capability medium-dependent resistance (MDR). The magnitude of MDR observed after oxic X irradiation was greater than that observed after anoxic X irradiation, uv irradiation, or MMS treatment. MDR was not observed in stationary-phase cells with X or uv radiation. MDR was associated with an increased ability to repair X-ray-induced DNA single-strand breaks, and with reduced X-ray-induced DNA degradation and protein synthesis retardation. Postirradiation protein synthesis was concluded to be critical in allowing the high X-ray survival associated with MDR, because of the large radiosensitization caused by a postirradiation growth medium shift down or treatment with rifampicin (RIF), recA protein must be at least one of the proteins whose synthesis is critical to MDR, as judged by the absence of MDR or a RIF effect in X-irradiated recA and lexA mutants. The results with X-irradiated temperature-conditional recA cells suggest that it is only after cells have been damaged that the recA gene plays a role in MDR

  2. The effect of growth conditions on inducible, recA-dependent resistance to x rays in Escherichia coli

    International Nuclear Information System (INIS)

    Escherichia coli cells grown to logarithmic phase in, and plated on, rich medium (yeast extract-nutrient broth) were more resistant to X rays, ultraviolet (uv) radiation, and methyl methanesulfonate (MMS) than cells grown in, and plated on, minimal medium. We have called this enhanced survival capability medium-dependent resistance (MDR). The magnitude of MDR observed after oxic X irradiation was greater than that observed after anoxic X irradiation, uv irradiation, or MMS treatment. MDR was not observed in stationary-phase cells with X or uv radiation. MDR was associated with an increased ability to repair X-ray-induced DNA single-strand breaks, and with reduced X-ray-induced DNA degradation and protein synthesis retardation. Postirradiation protein synthesis was concluded to be critical in allowing the high X-ray survival associated with MDR, because of the large radiosensitization caused by a postirradiation growth medium shift down or treatment with rifampicin (RIF). recA protein must be at least one of the proteins whose synthesis is critical to MDR, as judged by the absence of MDR or a RIF effect in X-irradiated recA and lexA mutants. The results with X-irradiated temperature-conditional recA cells suggest that it is only after cells have been damaged that the recA gene plays a role in MDR

  3. The Mass-Metallicity relation explored with CALIFA: I. Is there a dependence on the star formation rate?

    CERN Document Server

    Sanchez, S F; Jungwiert, B; Iglesias-Paramo1, J; Vilchez, J M; Marino, R A; Walcher, C J; Husemann, B; Mast, D; Monreal-Ibero, A; Fernandes, R Cid; Perez, E; Delgado, R Gonzalez; Garcia-Benito, R; Galbany, L; van de Ven, G; Jahnke, K; Flores, H; Bland-Hawthorn, J; Lopez-Sánchez, A R; Stanishev, V; Miralles-Caballero, D; Diaz, A I; Sanchez-Blazquez, P; Molla, M; Gallazzi1, A; Papaderos, P; Gomes, J M; Gruel, N; Pérez, I; Ruiz-Lara, T; Florido, E; de Lorenzo-Cáceres, A; Mendez-Abreu, J; Kehrig, C; Roth, M M; Ziegler, B; Alves, J; Wisotzki, L; Kupko, D; Quirrenbach, A; Bomans, D

    2013-01-01

    We present the results on the study of the global and local M-Z relation based on the first data available from the CALIFA survey (150 galaxies). This survey provides integral field spectroscopy of the complete optical extent of each galaxy (up to 2-3 effective radii), with enough resolution to separate individual HII regions and/or aggregations. Nearly $\\sim$3000 individual HII regions have been detected. The spectra cover the wavelength range between [OII]3727 and [SII]6731, with a sufficient signal-to-noise to derive the oxygen abundance and star-formation rate associated with each region. In addition, we have computed the integrated and spatially resolved stellar masses (and surface densities), based on SDSS photometric data. We explore the relations between the stellar mass, oxygen abundance and star-formation rate using this dataset. We derive a tight relation between the integrated stellar mass and the gas-phase abundance, with a dispersion smaller than the one already reported in the literature ($\\sig...

  4. Is a mean machine better than a dependable drive? It’s geared towards your regulatory focus

    Directory of Open Access Journals (Sweden)

    Graham G. Scott

    2012-08-01

    Full Text Available While many studies have investigated the role of message-level valence in persuasive messages (i.e., how positive or negative message content affects attitudes, none of these have examined whether word-level valence can modulate such effects. We investigated whether emotional language used within persuasive messages influenced attitudes and whether the processing of such communications could be modulated by regulatory focus. Using a 2 (Message: Positive, Negative × 2 (Words: Positive, Negative design, participants read car reviews and rated each on a series of semantic differentials and product recommendations. While positive messages were always rated higher than negative ones, the valence of a message’s component words differentially impacted attitudes toward distinct aspects of the product. On promotion-focus features, messages containing negative words produced higher ratings; for prevention-focus aspects, those with positive words resulted in higher ratings. We argue that adopting a prevention- or promotion-focused stance can influence the interpretation of emotion words in relation to overall message comprehension.

  5. Human plasminogen kringle 1-5 inhibits angiogenesis and induces thrombomodulin degradation in a protein kinase A-dependent manner.

    Science.gov (United States)

    Cho, Chia-Fong; Chen, Po-Ku; Chang, Po-Chiao; Wu, Hau-Lin; Shi, Guey-Yueh

    2013-10-01

    Kringle 1-5 (K1-5), an endogenous proteolytic fragment of human plasminogen (Plg), is an angiostatin-related protein that inhibits angiogenesis. Many angiostatin-related proteins have been identified, but the detailed molecular mechanisms underlying their antiangiogenic effects remain unclear. Thrombomodulin (TM) is a transmembrane glycoprotein that plays a major role in the anticoagulation process in endothelial cells. Previously, we demonstrated that recombinant TM could interact with Plg to enhance Plg activation. In the present study, we investigated the interaction between TM and K1-5, and their functions in endothelial cells. We found that K1-5 colocalized with TM and directly interacted with TM through the TM lectin-like domain. After K1-5 interacted with TM, it induced TM internalization and degradation. In addition, the K1-5-induced TM internalization and degradation in proteasomes after ubiquitin modification were dependent on protein kinase A (PKA). Moreover, a PKA-specific inhibitor reversed the effects of K1-5 on cell migration and tube formation. Consistent with these findings, TM overexpression resulted in increased cell migration; moreover, K1-5 inhibited the increase of TM-mediated cell migration in a PKA-dependent manner. We determined that TM acts as a K1-5 receptor and that K1-5 induces TM internalization, ubiquitination, and degradation through the PKA pathway, by which K1-5 may inhibit endothelial cell migration and tube formation. PMID:23880609

  6. Thanks, but No Thanks: Women's Avoidance of Help-Seeking in the Context of a Dependency-Related Stereotype

    Science.gov (United States)

    Wakefield, Juliet R. H.; Hopkins, Nick; Greenwood, Ronni M.

    2012-01-01

    The stereotype that women are dependent on men is a commonly verbalized, potentially damaging aspect of benevolent sexism. We investigated how women may use behavioral disconfirmation of the personal applicability of the stereotype to negotiate such sexism. In an experiment (N = 86), we manipulated female college students' awareness that women may…

  7. A dependence of perturbative predictions of extended standard model on Z-Z' mixing and QCD renormalization scheme parameters

    International Nuclear Information System (INIS)

    In the framework of the minimal extended electroweak models inspired by superstrings, we evaluate the Zj boson hadronic and total decay widths, the values R ≡ σ(e+e- → hadrons)/σpt and R' ≡ σ(e+e- → hadrons)/σ(e+e- → l+l-) dependences on Z - Z' mixing parameter and on renormalization scheme parameters. All these values are measured now at LEP. Popular models are discussed for comparison. (author). 23 refs, 6 figs

  8. The electron transfer flavoprotein fixABCX gene products from Azospirillum brasilense show a NifA-dependent promoter regulation.

    Science.gov (United States)

    Sperotto, Raul Antonio; Gross, Jeferson; Vedoy, Cleber; Passaglia, Luciane Maria Pereira; Schrank, Irene Silveira

    2004-10-01

    The complete nucleotide sequence of the A. brasilense fixA, fixB, fixC, and fixX genes is reported here. Sequence similarities between the protein sequences deduced from fixABCX genes and many electron transfer flavoproteins (ETFs) have been noted. Comparison of the amino acid sequences of both subunits of ETF with the A. brasilense fixA and fixB gene products exhibits an identity of 30%. The amino acid sequence of the other two genes, fixC and fixX, revealed similarity with the membrane-bound electron transfer flavoprotein ubiquinone oxidoreductase (ETF-QO). Using site-directed mutagenesis, mutations were introduced in the fixA promoter element of the A. brasilense fixABCX operon and chimeric p fixA-lacZ reporter gene fusions were constructed. The activation of the fixA promoter of A. brasilense is dependent upon the presence of the NifA protein being approximately 7 times less active than the A. brasilense nifH promoter. These results indicate that NifA from Klebsiella pneumoniae activates the fix promoter of A. brasilense and provide further evidence in support of the regulatory model of NifA activation in A. brasilense. Although no specific function has been assigned to the fixABCX gene products they are apparently required for symbiotic nitrogen fixation. An electron-transferring capacity in the nitrogen fixation pathway has been suggested for the fix gene products based on sequence homologies to the ETFs and ETF-QO proteins and by the absence of orthologous electron transfer proteins NifJ and NifF in A. brasilense.

  9. RelA-Dependent (p)ppGpp Production Controls Exoenzyme Synthesis in Erwinia carotovora subsp. atroseptica▿

    OpenAIRE

    Wang, Jinhong; Gardiol, Noemie; Burr, Tom; Salmond, George P. C.; Welch, Martin

    2007-01-01

    In this report, we investigate the link between nutrient limitation, RelA-mediated (p)ppGpp production, and virulence in the phytopathogen Erwinia carotovora subsp. atroseptica. A relA null mutant (JWC7) was constructed by allelic exchange, and we confirmed that, unlike the wild-type progenitor, this mutant did not produce elevated levels of (p)ppGpp upon nutrient downshift. However, (p)ppGpp production could be restored in strain JWC7 during nutrient limitation by supplying relA in trans. Du...

  10. On the A-dependence of {sigma}{sub L}/{sigma}{sub T}: Skeletons in the shadow

    Energy Technology Data Exchange (ETDEWEB)

    Milana, J. [Univ. of Maryland, College Park (United States)

    1994-04-01

    A most counter-intuitive dependence in the differential cross-section in the shadowing regime is shown to result from a higher-twist nuclear enhancement in R = {sigma}{sub L}/{sigma}{sub T} which severely complicates the unravelling from present data the corresponding dependence in Q{sup 2} of the nuclear structure functions, F{sub 2}{sup A}(x,Q{sup 2}). Indeed, until precision measurements close this loophole, the extrapolations of the structure functions to either higher Q{sup 2} or other processes (as is necessary to address present data at FNAL or future experiments at RHIC) must be considered problematic. The contribution CEBAF can make in this regard by providing systematic determination of R{sub A}(x, Q{sup 2}) is thus emphasized. The purpose of this talk is to motivate an experiment CEBAF can and should do, especially if upgraded to higher energies. While providing information on nuclear structure that is interesting in itself, the added motivation is that precision results will have important impact on other high-energy experiments involving nuclear targets that have been, and will continue to be done all over the world.

  11. Functional Promiscuity of Two Divergent Paralogs of Type III Plant Polyketide Synthases.

    Science.gov (United States)

    Pandith, Shahzad A; Dhar, Niha; Rana, Satiander; Bhat, Wajid Waheed; Kushwaha, Manoj; Gupta, Ajai P; Shah, Manzoor A; Vishwakarma, Ram; Lattoo, Surrinder K

    2016-08-01

    Plants effectively defend themselves against biotic and abiotic stresses by synthesizing diverse secondary metabolites, including health-protective flavonoids. These display incredible chemical diversity and ubiquitous occurrence and confer impeccable biological and agricultural applications. Chalcone synthase (CHS), a type III plant polyketide synthase, is critical for flavonoid biosynthesis. It catalyzes acyl-coenzyme A thioesters to synthesize naringenin chalcone through a polyketidic intermediate. The functional divergence among the evolutionarily generated members of a gene family is pivotal in driving the chemical diversity. Against this backdrop, this study was aimed to functionally characterize members of the CHS gene family from Rheum emodi, an endangered and endemic high-altitude medicinal herb of northwestern Himalayas. Two full-length cDNAs (1,179 bp each), ReCHS1 and ReCHS2, encoding unique paralogs were isolated and characterized. Heterologous expression and purification in Escherichia coli, bottom-up proteomic characterization, high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry analysis, and enzyme kinetic studies using five different substrates confirmed their catalytic potential. Phylogenetic analysis revealed the existence of higher synonymous mutations in the intronless divergents of ReCHS. ReCHS2 displayed significant enzymatic efficiency (Vmax/Km) with different substrates. There were significant spatial and altitudinal variations in messenger RNA transcript levels of ReCHSs correlating positively with metabolite accumulation. Furthermore, the elicitations in the form of methyl jasmonate, salicylic acid, ultraviolet B light, and wounding, chosen on the basis of identified cis-regulatory promoter elements, presented considerable differences in the transcript profiles of ReCHSs. Taken together, our results demonstrate differential propensities of CHS paralogs in terms of the accumulation of flavonoids and

  12. FadD Is Required for Utilization of Endogenous Fatty Acids Released from Membrane Lipids ▿ †

    Science.gov (United States)

    Pech-Canul, Ángel; Nogales, Joaquina; Miranda-Molina, Alfonso; Álvarez, Laura; Geiger, Otto; Soto, María José; López-Lara, Isabel M.

    2011-01-01

    FadD is an acyl coenzyme A (CoA) synthetase responsible for the activation of exogenous long-chain fatty acids (LCFA) into acyl-CoAs. Mutation of fadD in the symbiotic nitrogen-fixing bacterium Sinorhizobium meliloti promotes swarming motility and leads to defects in nodulation of alfalfa plants. In this study, we found that S. meliloti fadD mutants accumulated a mixture of free fatty acids during the stationary phase of growth. The composition of the free fatty acid pool and the results obtained after specific labeling of esterified fatty acids with a Δ5-desaturase (Δ5-Des) were in agreement with membrane phospholipids being the origin of the released fatty acids. Escherichia coli fadD mutants also accumulated free fatty acids released from membrane lipids in the stationary phase. This phenomenon did not occur in a mutant of E. coli with a deficient FadL fatty acid transporter, suggesting that the accumulation of fatty acids in fadD mutants occurs inside the cell. Our results indicate that, besides the activation of exogenous LCFA, in bacteria FadD plays a major role in the activation of endogenous fatty acids released from membrane lipids. Furthermore, expression analysis performed with S. meliloti revealed that a functional FadD is required for the upregulation of genes involved in fatty acid degradation and suggested that in the wild-type strain, the fatty acids released from membrane lipids are degraded by β-oxidation in the stationary phase of growth. PMID:21926226

  13. FadD is required for utilization of endogenous fatty acids released from membrane lipids.

    Science.gov (United States)

    Pech-Canul, Ángel; Nogales, Joaquina; Miranda-Molina, Alfonso; Álvarez, Laura; Geiger, Otto; Soto, María José; López-Lara, Isabel M

    2011-11-01

    FadD is an acyl coenzyme A (CoA) synthetase responsible for the activation of exogenous long-chain fatty acids (LCFA) into acyl-CoAs. Mutation of fadD in the symbiotic nitrogen-fixing bacterium Sinorhizobium meliloti promotes swarming motility and leads to defects in nodulation of alfalfa plants. In this study, we found that S. meliloti fadD mutants accumulated a mixture of free fatty acids during the stationary phase of growth. The composition of the free fatty acid pool and the results obtained after specific labeling of esterified fatty acids with a Δ5-desaturase (Δ5-Des) were in agreement with membrane phospholipids being the origin of the released fatty acids. Escherichia coli fadD mutants also accumulated free fatty acids released from membrane lipids in the stationary phase. This phenomenon did not occur in a mutant of E. coli with a deficient FadL fatty acid transporter, suggesting that the accumulation of fatty acids in fadD mutants occurs inside the cell. Our results indicate that, besides the activation of exogenous LCFA, in bacteria FadD plays a major role in the activation of endogenous fatty acids released from membrane lipids. Furthermore, expression analysis performed with S. meliloti revealed that a functional FadD is required for the upregulation of genes involved in fatty acid degradation and suggested that in the wild-type strain, the fatty acids released from membrane lipids are degraded by β-oxidation in the stationary phase of growth.

  14. Computational analysis of a novel mutation in ETFDH gene highlights its long-range effects on the FAD-binding motif

    Directory of Open Access Journals (Sweden)

    Chang Jan-Gowth

    2011-10-01

    Full Text Available Abstract Background Multiple acyl-coenzyme A dehydrogenase deficiency (MADD is an autosomal recessive disease caused by the defects in the mitochondrial electron transfer system and the metabolism of fatty acids. Recently, mutations in electron transfer flavoprotein dehydrogenase (ETFDH gene, encoding electron transfer flavoprotein:ubiquinone oxidoreductase (ETF:QO have been reported to be the major causes of riboflavin-responsive MADD. To date, no studies have been performed to explore the functional impact of these mutations or their mechanism of disrupting enzyme activity. Results High resolution melting (HRM analysis and sequencing of the entire ETFDH gene revealed a novel mutation (p.Phe128Ser and the hotspot mutation (p.Ala84Thr from a patient with MADD. According to the predicted 3D structure of ETF:QO, the two mutations are located within the flavin adenine dinucleotide (FAD binding domain; however, the two residues do not have direct interactions with the FAD ligand. Using molecular dynamics (MD simulations and normal mode analysis (NMA, we found that the p.Ala84Thr and p.Phe128Ser mutations are most likely to alter the protein structure near the FAD binding site as well as disrupt the stability of the FAD binding required for the activation of ETF:QO. Intriguingly, NMA revealed that several reported disease-causing mutations in the ETF:QO protein show highly correlated motions with the FAD-binding site. Conclusions Based on the present findings, we conclude that the changes made to the amino acids in ETF:QO are likely to influence the FAD-binding stability.

  15. The mechanisms underlying the hypolipidaemic effects of Grifola frondosa in the liver of rats

    Directory of Open Access Journals (Sweden)

    Yinrun Ding

    2016-08-01

    Full Text Available The present study investigated the hypolipidaemic effects of Grifola frondosa and its regulation mechanism involved in lipid metabolism in liver of rats fed a high-cholesterol diet. The body weights and serum lipid levels of control rats, of hyperlipidaemic rats and of hyperlipidaemic rats treated with oral Grifola frondosa were determined. mRNA expression and concentration of key lipid metabolism enzymes were investigated. Serum cholesterol, triacylglycerol and low-density lipoprotein cholesterol levels were markedly decreased in hyperlipidaemic rats treated with Grifola frondosa compared with untreated hyperlipidaemic rats. mRNA expression of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR, acyl-coenzyme A: cholesterol acyltransferase (ACAT2, apolipoprotein B (ApoB, fatty acid synthase (FAS and acetyl-CoA carboxylase (ACC1 were significantly down-regulated, while expression of cholesterol 7-alpha-hydroxylase (CYP7A1 was significantly up-regulated in the livers of treated rats compared with untreated hyperlipidaemic rats. The concentrations of these enzymes also paralleled the observed changes in mRNA expression. Two-dimensional polyacrylamide gel electrophoresis (2-DE and Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS were used to identify twenty proteins differentially expressed in livers of rats treated with Grifola frondosa compared with untreated hyperlipidemic rats. Of these twenty proteins, seven proteins were down-regulated and thirteen proteins were up-regulated. These findings indicate that the hypolipidaemic effects of Grifola frondosa reflected its modulation of key enzymes involved in cholesterol and triacylglycerol biosynthesis, absorption and catabolic pathways. Grifola frondosa may exert anti-atherosclerotic effects by inhibiting LDL oxidation through down-regulation and up-regulating proteins expression in the liver of rats. Therefore, Grifola frondosa may produce both hypolipidaemic

  16. Neutral lipid biosynthesis in engineered Escherichia coli: jojoba oil-like wax esters and fatty acid butyl esters.

    Science.gov (United States)

    Kalscheuer, Rainer; Stöveken, Tim; Luftmann, Heinrich; Malkus, Ursula; Reichelt, Rudolf; Steinbüchel, Alexander

    2006-02-01

    Wax esters are esters of long-chain fatty acids and long-chain fatty alcohols which are of considerable commercial importance and are produced on a scale of 3 million tons per year. The oil from the jojoba plant (Simmondsia chinensis) is the main biological source of wax esters. Although it has a multitude of potential applications, the use of jojoba oil is restricted, due to its high price. In this study, we describe the establishment of heterologous wax ester biosynthesis in a recombinant Escherichia coli strain by coexpression of a fatty alcohol-producing bifunctional acyl-coenzyme A reductase from the jojoba plant and a bacterial wax ester synthase from Acinetobacter baylyi strain ADP1, catalyzing the esterification of fatty alcohols and coenzyme A thioesters of fatty acids. In the presence of oleate, jojoba oil-like wax esters such as palmityl oleate, palmityl palmitoleate, and oleyl oleate were produced, amounting to up to ca. 1% of the cellular dry weight. In addition to wax esters, fatty acid butyl esters were unexpectedly observed in the presence of oleate. The latter could be attributed to solvent residues of 1-butanol present in the medium component, Bacto tryptone. Neutral lipids produced in recombinant E. coli were accumulated as intracytoplasmic inclusions, demonstrating that the formation and structural integrity of bacterial lipid bodies do not require specific structural proteins. This is the first report on substantial biosynthesis and accumulation of neutral lipids in E. coli, which might open new perspectives for the biotechnological production of cheap jojoba oil equivalents from inexpensive resources employing recombinant microorganisms.

  17. Toxicity of Carboxylic Acid-Containing Drugs: The Role of Acyl Migration and CoA Conjugation Investigated.

    Science.gov (United States)

    Lassila, Toni; Hokkanen, Juho; Aatsinki, Sanna-Mari; Mattila, Sampo; Turpeinen, Miia; Tolonen, Ari

    2015-12-21

    Many carboxylic acid-containing drugs are associated with idiosyncratic drug toxicity (IDT), which may be caused by reactive acyl glucuronide metabolites. The rate of acyl migration has been earlier suggested as a predictor of acyl glucuronide reactivity. Additionally, acyl Coenzyme A (CoA) conjugates are known to be reactive. Here, 13 drugs with a carboxylic acid moiety were incubated with human liver microsomes to produce acyl glucuronide conjugates for the determination of acyl glucuronide half-lives by acyl migration and with HepaRG cells to monitor the formation of acyl CoA conjugates, their further conjugate metabolites, and trans-acylation products with glutathione. Additionally, in vitro cytotoxicity and mitochondrial toxicity experiments were performed with HepaRG cells to compare the predictability of toxicity. Clearly, longer acyl glucuronide half-lives were observed for safe drugs compared to drugs that can cause IDT. Correlation between half-lives and toxicity classification increased when "relative half-lives," taking into account the formation of isomeric AG-forms due to acyl migration and eliminating the effect of hydrolysis, were used instead of plain disappearance of the initial 1-O-β-AG-form. Correlation was improved further when a daily dose of the drug was taken into account. CoA and related conjugates were detected primarily for the drugs that have the capability to cause IDT, although some exceptions to this were observed. Cytotoxicity and mitochondrial toxicity did not correlate to drug safety. On the basis of the results, the short relative half-life of the acyl glucuronide (high acyl migration rate), high daily dose and detection of acyl CoA conjugates, or further metabolites derived from acyl CoA together seem to indicate that carboxylic acid-containing drugs have a higher probability to cause drug-induced liver injury (DILI). PMID:26558897

  18. Enhancement of human ACAT1 gene expression to promote the macrophage-derived foam cell formation by dexamethasone

    Institute of Scientific and Technical Information of China (English)

    Li YANG; Ta Yuan CHANG; Bo Liang LI; Jin Bo YANG; Jia CHEN; Guang Yao YU; Pei ZHOU; Lei LEI; Zhen Zhen WANG; Catherine CY CHANG; XinYing YANG

    2004-01-01

    In macrophages, the accumulation of cholesteryl esters synthesized by the activated acyl-coenzyme A:cholesterol acyltransferase-1 (ACAT1) results in the foam cell formation, a hallmark of early atherosclerotic lesions. In this study,with the treatment of a glucocorticoid hormone dexamethasone (Dex), lipid staining results clearly showed the large accumulation of lipid droplets containing cholesteryl esters in THP- 1-derived macrophages exposed to lower concentration of the oxidized low-density lipoprotein (ox-LDL). More notably, when treated together with specific anti-ACAT inhibitors, the abundant cholesteryl ester accumulation was markedly diminished in THP-l-derived macrophages, confirming that ACAT is the key enzyme responsible for intracellular cholesteryl ester synthesis. RT-PCR and Western blot results indicated that Dex caused up-regulation of human ACAT1 expression at both the mRNA and protein levels in THP-1 and THP- 1-derived macrophages. The luciferase activity assay demonstrated that Dex could enhance the activity of human ACAT1 gene P1 promoter, a major factor leading to the ACAT1 activation, in a cell-specific manner.Further experimental evidences showed that a glucocorticoid response element (GRE) located within human ACAT1gene P1 promoter to response to the elevation of human ACAT1 gene expression by Dex could be functionally bound with glucocorticoid receptor (GR) proteins. These data supported the hypothesis that the clinical treatment with Dex,which increased the incidence of atherosclerosis, may in part due to enhancing the ACAT1 expression to promote the accumulation of cholesteryl esters during the macrophage-derived foam cell formation, an early stage of atherosclerosis.

  19. Fatty acid biosynthesis in Pseudomonas aeruginosa is initiated by the FabY class of β-ketoacyl acyl carrier protein synthases.

    Science.gov (United States)

    Yuan, Yanqiu; Sachdeva, Meena; Leeds, Jennifer A; Meredith, Timothy C

    2012-10-01

    The prototypical type II fatty acid synthesis (FAS) pathway in bacteria utilizes two distinct classes of β-ketoacyl synthase (KAS) domains to assemble long-chain fatty acids, the KASIII domain for initiation and the KASI/II domain for elongation. The central role of FAS in bacterial viability and virulence has stimulated significant effort toward developing KAS inhibitors, particularly against the KASIII domain of the β-acetoacetyl-acyl carrier protein (ACP) synthase FabH. Herein, we show that the opportunistic pathogen Pseudomonas aeruginosa does not utilize a FabH ortholog but rather a new class of divergent KAS I/II enzymes to initiate the FAS pathway. When a P. aeruginosa cosmid library was used to rescue growth in a fabH downregulated strain of Escherichia coli, a single unannotated open reading frame, PA5174, complemented fabH depletion. While deletion of all four KASIII domain-encoding genes in the same P. aeruginosa strain resulted in a wild-type growth phenotype, deletion of PA5174 alone specifically attenuated growth due to a defect in de novo FAS. Siderophore secretion and quorum-sensing signaling, particularly in the rhl and Pseudomonas quinolone signal (PQS) systems, was significantly muted in the absence of PA5174. The defect could be repaired by intergeneric complementation with E. coli fabH. Characterization of recombinant PA5174 confirmed a preference for short-chain acyl coenzyme A (acyl-CoA) substrates, supporting the identification of PA5174 as the predominant enzyme catalyzing the condensation of acetyl coenzyme A with malonyl-ACP in P. aeruginosa. The identification of the functional role for PA5174 in FAS defines the new FabY class of β-ketoacyl synthase KASI/II domain condensation enzymes.

  20. Unexpected functional diversity in the fatty acid desaturases of the flour beetle Tribolium castaneum and identification of key residues determining activity.

    Science.gov (United States)

    Haritos, Victoria S; Horne, Irene; Damcevski, Katherine; Glover, Karen; Gibb, Nerida

    2014-08-01

    Desaturases catalyse modifications to fatty acids which are essential to homeostasis and for pheromone and defensive chemical production. All desaturases of the flour beetle Tribolium castaneum were investigated via query of the sequenced genome which yielded 15 putative acyl-Coenzyme A genes. Eleven desaturase mRNA were obtained in full length and functionally expressed in yeast. Phylogenetic analysis separated the desaturases into 4 distinct clades; one clade contained conserved beetle Δ9 desaturases, second clade was Tribolium-specific having diverse activities including Δ5, Δ9 and Δ12 desaturation and the other 2 clades had mixed insect representatives. Three members of this clade contained unusual inserted sequences of ∼20 residues in the C-terminal region and were related to desaturases that all contained similar inserts. Deletion of the entirety of the insert in the flour beetle Δ12 desaturase abolished its activity but this was partially restored by the reintroduction of two histidine residues, suggesting the histidine(s) are required for activity but the full length insert is not. Five new desaturase activities were discovered: Δ9 desaturation of C12:0-C16:0 substrates; two unprecedented Δ5 enzymes acting on C18:0 and C16:0; Δ9 activity exclusively on C16:0 and a further stearate Δ9 desaturase. qPCR analysis ruled out a role in sex pheromone synthesis for the Δ5 and Δ9/C16:0 desaturases. The flour beetle genome has underpinned an examination of all transcribed desaturases in the organism and revealed a diversity of novel and unusual activities, an improved understanding of the evolutionary relationships among insect desaturases and sequence determinants of activity.

  1. Two Human ACAT2 mRNA Variants Produced by Alternative Splicing and Coding for Novel Isoenzymes

    Institute of Scientific and Technical Information of China (English)

    Xiao-Min YAO; Bo-Liang LI; Can-Hua WANG; Bao-Liang SONG; Xin-Ying YANG; Zhen-Zhen WANG; Wei QI; Zhi-Xin LIN; Catherine C. Y. CHANG; Ta-Yuan CHANG

    2005-01-01

    Acyl coenzyme A:cholesterol acyltransferase 2 (ACAT2) plays an important role in cholesterol absorption. Human ACAT2 is highly expressed in small intestine and fetal liver, but its expression is greatly diminished in adult liver. The full-length human ACAT2 mRNA encodes a protein, designated ACAT2a, with 522 amino acids. We have previously reported the organization of the human ACAT2 gene and the differentiation-dependent promoter activity in intestinal Caco-2 cells. In the current work, two human ACAT2 mRNA variants produced by alternative splicing are cloned and predicted to encode two novel ACAT2 isoforms,named ACAT2b and ACAT2c, with 502 and 379 amino acids, respectively. These mRNA variants differ from ACAT2a mRNA by lack of the exon 4 (ACAT2b mRNA) and exons 4-5 plus 8-9-10 (ACAT2c mRNA).Significantly, comparable amounts of the alternatively spliced ACAT2 mRNA variants were detected by RTPCR, and Western blot analysis confirmed the presence of their corresponding proteins in human liver and intestine cells. Furthermore, phosphorylation and enzymatic activity analyses demonstrated that the novel isoenzymes ACAT2b and ACAT2c lacked the phosphorylatable site SLLD, and their enzymatic activities reduced to 25%-35% of that of ACAT2a. These evidences indicate that alternative splicing produces two human ACAT2 mRNA variants that encode the novel ACAT2 isoenzymes. Our findings might help to understand the regulation of the ACAT2 gene expression under certain physiological and pathological conditions.

  2. Lipidomic and spatio-temporal imaging of fat by mass spectrometry in mice duodenum during lipid digestion.

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    Alexandre Seyer

    Full Text Available Intestinal absorption of dietary fat is a complex process mediated by enterocytes leading to lipid assembly and secretion of circulating lipoproteins as chylomicrons, vLDL and intestinal HDL (iHDL. Understanding lipid digestion is of importance knowing the correlation between excessive fat absorption and atherosclerosis. By using time-of-flight secondary ion mass spectrometry (TOF-SIMS, we illustrated a spatio-temporal localization of fat in mice duodenum, at different times of digestion after a lipid gavage, for the first time. Fatty acids progressively increased in enterocytes as well as taurocholic acid, secreted by bile and engaged in the entero-hepatic re-absorption cycle. Cytosolic lipid droplets (CLD from enterocytes were originally purified separating chylomicron-like, intermediate droplets and smaller HDL-like. A lipidomic quantification revealed their contents in triglycerides, free and esterified cholesterol, phosphatidylcholine, sphingomyelin and ceramides but also in free fatty acids, mono- and di-acylglycerols. An acyl-transferase activity was identified and the enzyme monoacylglycerol acyl transferase 2 (MGAT2 was immunodetected in all CLD. The largest droplets was also shown to contain the microsomal triglyceride transfer protein (MTTP, the acyl-coenzyme A-cholesterol acyltransferases (ACAT 1 and 2, hormone sensitive lipase (HSL and adipose triglyceride lipase (ATGL. This highlights the fact that during the digestion of fats, enterocyte CLD contain some enzymes involved in the different stages of the metabolism of diet fatty acids and cholesterol, in anticipation of the crucial work of endoplasmic reticulum in the process. The data further underlines the dual role of chylomicrons and iHDL in fat digestion which should help to efficiently complement lipid-lowering therapy.

  3. Orphan drugs in development for long-chain fatty acid oxidation disorders: challenges and progress

    Directory of Open Access Journals (Sweden)

    Sun A

    2015-04-01

    Full Text Available Angela Sun, J Lawrence Merritt II Department of Pediatrics, University of Washington, Seattle, WA, USA Abstract: Fatty acid oxidation disorders are inborn errors of metabolism resulting in failure of ß-oxidation within or transport of fatty acids into the mitochondria. The long-chain fatty acid oxidation disorders are characterized by variable presentations ranging from newborn cardiomyopathy, to infantile hypoketotic hypoglycemia resulting from liver involvement, to skeletal myopathy often resulting in rhabdomyolysis in adolescents and adults. Treatments for these long-chain fatty acid oxidation disorders have typically focused upon avoidance of fasting with dietary fat restriction and medium-chain triglyceride supplementation. These treatments have resulted in only a partial response with improvements in hypoglycemia, reduction in frequency of rhabdomyolysis, and improvement in cardiomyopathy with early therapy, but significant risk remains. Recent advances in therapies for long-chain fatty acid oxidation disorders are reviewed in this article. These include sodium D,L-3-hydroxybutyrate, triheptanoin, gene therapy, and bezafibrates. Sodium D,L-3-hydroxybutyrate has shown clinical effect, with improvements in muscle tone, neurological abnormalities, and some cases of cardiomyopathy and leukodystrophy. Triheptanoin has been used as an alternative medium-chain triglyceride in a number of fatty acid oxidation disorders and has shown promising findings in the treatment of cardiomyopathy and hypoglycemia. However, it does not significantly reduce episodes of rhabdomyolysis. Gene therapy has been shown to improve acylcarnitine levels in very-long-chain acyl-coenzyme A dehydrogenase deficiency mouse models, with preservation of glucose levels. Bezafibrates have shown improvements in acylcarnitine concentrations in fibroblast studies, but clinical observations have not demonstrated consistent effects. Together, these treatments have shown some

  4. PPARα Protein Expression Was Increased by Four Weeks of Intermittent Hypoxic Training via AMPKα2-Dependent Manner in Mouse Skeletal Muscle.

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    Ge Li

    Full Text Available Peroxisome proliferator-activated receptor α (PPARα is critical for muscle endurance due to its role in the regulation of fatty acid oxidation. The 5'-AMP-activated protein kinase (AMPK is an energy sensor in cells, but its role in PPARα regulation in vivo remains unknown. In this study, we examined PPARα expression in the skeletal muscle of AMPKα2 overexpression (OE, knockout (KO and wild-type (WT mice after four weeks of exercise under intermittent hypoxia. WT, OE and KO mice were used at 40 mice/strain and randomly subdivided into four subgroups: control (C, running (R, hypoxia (H, and running plus hypoxia (R+H at 10 mice/group. The treadmill running was performed at the speed of 12 m/min, 60 min/day with a slope of 0 degree for four weeks. The hypoxia treatment was performed in daytime with normobaric hypoxia (11.20% oxygen, 8 hours/day. In the R+H group, the treadmill running was conducted in the hypoxic condition. AMPKα2, phosphor-AMPKα (p-AMPKα (Thr172, nuclear PPARα proteins were measured by Western blot and the medium chain acyl coenzyme A dehydrogenase (MCAD mRNA, the key enzyme for fatty acid oxidation and one of the PPARα target genes, was also measured in skeletal muscles after the interventions. The results showed that nuclear PPARα protein was significantly increased by R+H in WT muscles, the increase was enhanced by 41% (p<0.01 in OE mice, but was reduced by 33% (p<0.01 in KO mice. The MCAD mRNA expression was increased after four weeks of R+H intervention. The change in MCAD mRNA followed a similar trend as that of PPARα protein in OE and KO mice. Our data suggest that the increase in nuclear PPARα protein by four-week exercise training under the intermittent hypoxia was dependent on AMPK activation.

  5. Metabolic Regulation as a Consequence of Anaerobic 5-Methylthioadenosine Recycling in Rhodospirillum rubrum

    Science.gov (United States)

    North, Justin A.; Sriram, Jaya; Chourey, Karuna; Ecker, Christopher D.; Sharma, Ritin; Wildenthal, John A.; Hettich, Robert L.

    2016-01-01

    ABSTRACT Rhodospirillum rubrum possesses a novel oxygen-independent, aerobic methionine salvage pathway (MSP) for recycling methionine from 5-methylthioadenosine (MTA), the MTA-isoprenoid shunt. This organism can also metabolize MTA as a sulfur source under anaerobic conditions, suggesting that the MTA-isoprenoid shunt may also function anaerobically as well. In this study, deep proteomics profiling, directed metabolite analysis, and reverse transcriptase quantitative PCR (RT-qPCR) revealed metabolic changes in response to anaerobic growth on MTA versus sulfate as sole sulfur source. The abundance of protein levels associated with methionine transport, cell motility, and chemotaxis increased in the presence of MTA over that in the presence of sulfate. Purine salvage from MTA resulted primarily in hypoxanthine accumulation and a decrease in protein levels involved in GMP-to-AMP conversion to balance purine pools. Acyl coenzyme A (acyl-CoA) metabolic protein levels for lipid metabolism were lower in abundance, whereas poly-β-hydroxybutyrate synthesis and storage were increased nearly 10-fold. The known R. rubrum aerobic MSP was also shown to be upregulated, to function anaerobically, and to recycle MTA. This suggested that other organisms with gene homologues for the MTA-isoprenoid shunt may also possess a functioning anaerobic MSP. In support of our previous findings that ribulose-1,5-carboxylase/oxygenase (RubisCO) is required for an apparently purely anaerobic MSP, RubisCO transcript and protein levels both increased in abundance by over 10-fold in cells grown anaerobically on MTA over those in cells grown on sulfate, resulting in increased intracellular RubisCO activity. These results reveal for the first time global metabolic responses as a consequence of anaerobic MTA metabolism compared to using sulfate as the sulfur source. PMID:27406564

  6. Thiolation-enhanced substrate recognition by D-alanyl carrier protein ligase DltA from Bacillus cereus [v1; ref status: indexed, http://f1000r.es/3dx

    Directory of Open Access Journals (Sweden)

    Liqin Du

    2014-05-01

    Full Text Available D-alanylation of the lipoteichoic acid on Gram-positive cell wall is dependent on dlt gene-encoded proteins DltA, DltB, DltC and DltD. The D-alanyl carrier protein ligase DltA, as a remote homolog of acyl-(coenzyme A (CoA synthetase, cycles through two active conformations for the catalysis of adenylation and subsequent thiolation of D-alanine (D-Ala. The crystal structure of DltA in the absence of any substrate was observed to have a noticeably more disordered pocket for ATP which would explain why DltA has relatively low affinity for ATP in the absence of any D-alanyl carrier. We have previously enabled the thiolation of D-alanine in the presence of CoA as the mimic of D-alanyl carrier protein DltC which carries a 4’-phosphopantetheine group on a serine residue. Here we show that the resulting Michaelis constants in the presence of saturating CoA for both ATP and D-alanine were reduced more than 10 fold as compared to the values obtained in the absence of CoA. The presence of CoA also made DltA ~100-fold more selective on D-alanine over L-alanine. The CoA-enhanced substrate recognition further implies that the ATP and D-alanine substrates of the adenylation reaction are incorporated when the DltA enzyme cycles through its thiolation conformation.

  7. Three TaFAR genes function in the biosynthesis of primary alcohols and the response to abiotic stresses in Triticum aestivum.

    Science.gov (United States)

    Wang, Meiling; Wang, Yong; Wu, Hongqi; Xu, Jing; Li, Tingting; Hegebarth, Daniela; Jetter, Reinhard; Chen, Letian; Wang, Zhonghua

    2016-01-01

    Cuticular waxes play crucial roles in protecting plants against biotic and abiotic stresses. They are complex mixtures of very-long-chain fatty acids and their derivatives, including C20-C32 fatty alcohols. Here, we report the identification of 32 FAR-like genes and the detailed characterization of TaFAR2, TaFAR3 and TaFAR4, wax biosynthetic genes encoding fatty acyl-coenzyme A reductase (FAR) in wheat leaf cuticle. Heterologous expression of the three TaFARs in wild-type yeast and mutated yeast showed that TaFAR2, TaFAR3 and TaFAR4 were predominantly responsible for the accumulation of C18:0, C28:0 and C24:0 primary alcohols, respectively. Transgenic expression of the three TaFARs in tomato fruit and Arabidopsis cer4 mutant led to increased production of C22:0-C30:0 primary alcohols. GFP-fusion protein injection assay showed that the three encoded TaFAR proteins were localized to the endoplasmic reticulum (ER), the site of wax biosynthesis. The transcriptional expression of the three TaFAR genes was induced by cold, salt, drought and ABA. Low air humidity led to increased expression of TaFAR genes and elevated wax accumulation in wheat leaves. Collectively, these data suggest that TaFAR2, TaFAR3 and TaFAR4 encode active alcohol-forming FARs involved in the synthesis of primary alcohol in wheat leaf and the response to environmental stresses. PMID:27112792

  8. First report about the mode of action of combined butafosfan and cyanocobalamin on hepatic metabolism in nonketotic early lactating cows.

    Science.gov (United States)

    Kreipe, L; Deniz, A; Bruckmaier, R M; van Dorland, H A

    2011-10-01

    The primary aim was to investigate the effect of combined butafosfan and cyanocobalamin on liver metabolism in early lactating cows through mRNA expression measurements of genes encoding 31 enzymes and transport proteins of major metabolic processes in the liver using 16 multiparous early lactating dairy cows. The treatments included i.v. injection of 10 mL/100 kg of body weight combined butafosfan and cyanocobalamin (TG, n = 8) on 3 d consecutively at 25 ± 3 d in milk or injection with physiological saline solution similarly applied (CG, n = 8). Results include a higher daily milk production for TG cows (41.1 ± 0.9 kg, mean ± SEM) compared with CG cows (39.5 ± 0.7 kg). In plasma, the concentration of inorganic phosphorus was lower in the TG cows (1.25 ± 0.08 mmol/L) after the treatment than in the CG cows (1.33 ± 0.07 mmol/L). The plasma β-hydroxybutyrate concentration was 0.65 ± 0.13 mmol/L for all cows before the treatment, and remained unaffected post treatment. The unique result was that in the liver, the mRNA abundance of acyl-coenzyme A synthetase long-chain family member 1, involved in fatty acid oxidation and biosynthesis, was lower across time points after the treatment for TG compared with CG cows (17.5 ± 0.15 versus 18.1 ± 0.24 cycle threshold, log(2), respectively). In conclusion, certain effects of combined butafosfan and cyanocobalamin were observed on mRNA abundance of a gene in the liver of nonketotic early lactating cows. PMID:21943742

  9. Human urotensin II promotes hypertension and atherosclerotic cardiovascular diseases.

    Science.gov (United States)

    Watanabe, Takuya; Arita, Shigeko; Shiraishi, Yuji; Suguro, Toshiaki; Sakai, Tetsuo; Hongo, Shigeki; Miyazaki, Akira

    2009-01-01

    Human urotensin II (U-II), the most potent vasoconstrictor undecapeptide identified to date, and its receptor (UT) are involved in the pathogenesis of systemic and pulmonary hypertension. Here, we review recent advances in our understanding of the pathophysiology of U-II with particular reference to its role in atherosclerotic cardiovascular diseases. Single-nucleotide polymorphisms of U-II gene (S89N) are associated with onset of essential hypertension, type II diabetes mellitus, and insulin resistance in the Asian population. Plasma U-II levels are elevated in patients with vascular endothelial dysfunction-related diseases such as essential hypertension, diabetes mellitus, atherosclerosis, ischemic heart disease, and heart failure. Chronic infusion of U-II enhances atherosclerotic lesions in the aorta in apolipoprotein E-knockout mice. In human atherosclerotic plaques from the aorta and coronary and carotid arteries, U-II is expressed at high levels in endothelial cells (ECs) and lymphocytes, whereas UT is expressed at high levels in vascular smooth muscle cells (VSMCs), ECs, monocytes, and macrophages. U-II stimulates vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 expression in human ECs as chemoattractant for monocytes, and accelerates foam cell formation by up-regulation of acyl-coenzyme A:cholesterol acyltransferase-1 in human monocyte-derived macrophages. U-II produces reactive oxygen species (ROS) via nicotinamide adenine dinucleotide phosphate oxidase activation in human VSMCs, and stimulates VSMC proliferation with synergistic effects when combined with ROS, oxidized LDL, and serotonin. Clinical studies demonstrated increased plasma U-II levels in accordance with the severity of carotid atherosclerosis in patients with essential hypertension and that of coronary artery lesions in patients with ischemic heart disease. Here, we summarize the key roles of U-II in progression of hypertension and atherosclerotic cardiovascular diseases

  10. Systematic Mutational Analysis of Histidine Kinase Genes in the Nosocomial Pathogen Stenotrophomonas maltophilia Identifies BfmAK System Control of Biofilm Development.

    Science.gov (United States)

    Zheng, Liu; Wang, Fang-Fang; Ren, Bao-Zhen; Liu, Wei; Liu, Zhong; Qian, Wei

    2016-04-01

    The Gram-negative bacterium Stenotrophomonas maltophilialives in diverse ecological niches. As a result of its formidable capabilities of forming biofilm and its resistance to multiple antibiotic agents, the bacterium is also a nosocomial pathogen of serious threat to the health of patients whose immune systems are suppressed or compromised. Besides the histidine kinase RpfC, the two-component signal transduction system (TCS), which is the canonical regulatory machinery used by most bacterial pathogens, has never been experimentally investigated inS. maltophilia Here, we annotated 62 putative histidine kinase genes in the S. maltophilia genome and successfully obtained 51 mutants by systematical insertional inactivation. Phenotypic characterization identified a series of mutants with deficiencies in bacterial growth, swimming motility, and biofilm development. A TCS, named here BfmA-BfmK (Smlt4209-Smlt4208), was genetically confirmed to regulate biofilm formation inS. maltophilia Together with interacting partner prediction and chromatin immunoprecipitation screens, six candidate promoter regions bound by BfmA in vivo were identified. We demonstrated that, among them, BfmA acts as a transcription factor that binds directly to the promoter regions of bfmA-bfmK and Smlt0800(acoT), a gene encoding an acyl coenzyme A thioesterase that is associated with biofilm development, and positively controls their transcription. Genome-scale mutational analyses of histidine kinase genes and functional dissection of BfmK-BfmA regulation in biofilm provide genetic information to support more in-depth studies on cellular signaling inS. maltophilia, in the context of developing novel approaches to fight this important bacterial pathogen. PMID:26873318

  11. Structural and Functional Studies of Fatty Acyl Adenylate Ligases from E. coli and L. pneumophila

    Energy Technology Data Exchange (ETDEWEB)

    Z Zhang; R Zhou; J Sauder; P Tonge; S Burley; S Swaminathan

    2011-12-31

    Fatty acyl-AMP ligase (FAAL) is a new member of a family of adenylate-forming enzymes that were recently discovered in Mycobacterium tuberculosis. They are similar in sequence to fatty acyl-coenzyme A (CoA) ligases (FACLs). However, while FACLs perform a two-step catalytic reaction, AMP ligation followed by CoA ligation using ATP and CoA as cofactors, FAALs produce only the acyl adenylate and are unable to perform the second step. We report X-ray crystal structures of full-length FAAL from Escherichia coli (EcFAAL) and FAAL from Legionella pneumophila (LpFAAL) bound to acyl adenylate, determined at resolution limits of 3.0 and 1.85 {angstrom}, respectively. The structures share a larger N-terminal domain and a smaller C-terminal domain, which together resemble the previously determined structures of FAAL and FACL proteins. Our two structures occur in quite different conformations. EcFAAL adopts the adenylate-forming conformation typical of FACLs, whereas LpFAAL exhibits a unique intermediate conformation. Both EcFAAL and LpFAAL have insertion motifs that distinguish them from the FACLs. Structures of EcFAAL and LpFAAL reveal detailed interactions between this insertion motif and the interdomain hinge region and with the C-terminal domain. We suggest that the insertion motifs support sufficient interdomain motions to allow substrate binding and product release during acyl adenylate formation, but they preclude CoA binding, thereby preventing CoA ligation.

  12. Separation of isomeric short-chain acyl-CoAs in plant matrices using ultra-performance liquid chromatography coupled with tandem mass spectrometry.

    Science.gov (United States)

    Purves, Randy W; Ambrose, Stephen J; Clark, Shawn M; Stout, Jake M; Page, Jonathan E

    2015-02-01

    Acyl coenzyme A (acyl-CoA) thioesters are important intermediates in cellular metabolism and being able to distinguish among them is critical to fully understanding metabolic pathways in plants. Although significant advances have been made in the identification and quantification of acyl-CoAs using liquid chromatography tandem mass spectrometry (LC-MS/MS), separation of isomeric species such as isobutyryl- and n-butyrl-CoA has remained elusive. Here we report an ultra-performance liquid chromatography (UPLC)-MS/MS method for quantifying short-chain acyl-CoAs including isomeric species n-butyryl-CoA and isobutyryl-CoA as well as n-valeryl-CoA and isovaleryl-CoA. The method was applied to the analysis of extracts of hop (Humulus lupulus) and provided strong evidence for the existence of an additional structural isomer of valeryl-CoA, 2-methylbutyryl-CoA, as well as an unexpected isomer of hexanoyl-CoA. The results showed differences in the acyl-CoA composition among varieties of Humulus lupulus, both in glandular trichomes and cone tissues. When compared with the analysis of hemp (Cannabis sativa) extracts, the contribution of isobutyryl-CoAs in hop was greater as would be expected based on the downstream polyketide products. Surprisingly, branched chain valeryl-CoAs (isovaleryl-CoA and 2-methylbutyryl-CoA) were the dominant form of valeryl-CoAs in both hop and hemp. The capability to separate these isomeric forms will help to understand biochemical pathways leading to specialized metabolites in plants.

  13. Genome-wide screening and transcriptional profile analysis of desaturase genes in the European corn borer moth

    Institute of Scientific and Technical Information of China (English)

    Bingye Xue; Alejandro P. Rooney; Wendell L. Roelofs

    2012-01-01

    Acyl-coenzyme A (Acyl-CoA) desaturases play a key role in the biosynthesis of female moth sex pheromones.Desaturase genes are encoded by a large multigene family,and they have been divided into five subgroups on the basis of biochemical functionality and phylogenetic affinity.In this study both copy numbers and transcriptional levels of desaturase genes in the European corn borer (ECB),Ostrinia nubilalis,were investigated.The results from genome-wide screening of ECB bacterial artificial chromosome (BAC)library indicated there are many copies of some desaturase genes in the genome.An open reading frame (ORF) has been isolated for the novel desaturase gene ECB ezi-△11β from ECB gland complementary DNA and its functionality has been analyzed by two yeast expression systems.No functional activities have been detected for it.The expression levels of the four desaturase genes both in the pheromone gland and fat body of ECB and Asian corn borer (ACB),O.furnacalis,were determined by real-time polymerase chain reaction.In the ECB gland,△ 11 is the most abundant,although the amount of △14 is also considerable.In the ACB gland,△14 is the most abundant and is 100 times more abundant than all the other three combined.The results from the analysis of evolution of desaturase gene transcription in the ECB,ACB and other moths indicate that the pattern of △ 11 gene transcription is significantly different from the transcriptional patterns of other desaturase genes and this difference is tied to the underlying nucleotide composition bias of the genome.

  14. High ACSL5 transcript levels associate with systemic lupus erythematosus and apoptosis in Jurkat T lymphocytes and peripheral blood cells.

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    Antonio Catalá-Rabasa

    Full Text Available BACKGROUND: Systemic lupus erythematosus (SLE is a prototypical autoimmune disease in which increased apoptosis and decreased apoptotic cells removal has been described as most relevant in the pathogenesis. Long-chain acyl-coenzyme A synthetases (ACSLs have been involved in the immunological dysfunction of mouse models of lupus-like autoimmunity and apoptosis in different in vitro cell systems. The aim of this work was to assess among the ACSL isoforms the involvement of ACSL2, ACSL4 and ACSL5 in SLE pathogenesis. FINDINGS: With this end, we determined the ACSL2, ACSL4 and ACSL5 transcript levels in peripheral blood mononuclear cells (PBMCs of 45 SLE patients and 49 healthy controls by quantitative real time-PCR (q-PCR. We found that patients with SLE had higher ACSL5 transcript levels than healthy controls [median (range, healthy controls = 16.5 (12.3-18.0 vs. SLE = 26.5 (17.8-41.7, P = 3.9×10 E-5] but no differences were found for ACSL2 and ACSL4. In in vitro experiments, ACSL5 mRNA expression was greatly increased when inducing apoptosis in Jurkat T cells and PBMCs by Phorbol-Myristate-Acetate plus Ionomycin (PMA+Io. On the other hand, short interference RNA (siRNA-mediated silencing of ACSL5 decreased induced apoptosis in Jurkat T cells up to the control levels as well as decreased mRNA expression of FAS, FASLG and TNF. CONCLUSIONS: These findings indicate that ACSL5 may play a role in the apoptosis that takes place in SLE. Our results point to ACSL5 as a potential novel functional marker of pathogenesis and a possible therapeutic target in SLE.

  15. Sex differences in global mRNA content of human skeletal muscle.

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    Amy C Maher

    Full Text Available Women oxidize more fat as compared to men during endurance exercise and several groups have shown that the mRNA content of selected genes related to fat oxidation are higher in women (e.g. hormone sensitive lipase, beta-hydroxyacyl-CoA dehydrogenase, CD36. One of the possible mechanisms is that women tend to have a higher area percentage of type I skeletal muscle fibers as compared with men. Consequently, we hypothesized that sex would influence the basal mRNA and protein content for genes involved in metabolism and the determination of muscle fiber type. Muscle biopsies from the vastus lateralis were collected from healthy men and women. We examined mRNA content globally using Affymetrix GeneChips, and selected genes were examined and/or confirmed by RT-PCR. Furthermore, we examined protein content by Western blot analysis. Stringent gene array analysis revealed 66 differentially expressed genes representing metabolism, mitochondrial function, transport, protein biosynthesis, cell proliferation, signal transduction pathways, transcription and translation. Stringent gene array analysis and RT-PCR confirmed that mRNA for; acyl-coenzyme A acyltransferase 2 (ACAA2, trifunctional protein beta (HADHB, catalase, lipoprotein lipase (LPL, and uncoupling protein-2 (UCP-2 were higher in women. Targeted gene analysis revealed that myosin heavy chain I (MHCI, peroxisome proliferator-activated receptor (PPARdelta were higher in women compared with men. Surprisingly, there were no significant sex based differences in protein content for HADHB, ACAA2, catalase, PPARdelta, and MHC1. In conclusion, the differences in the basal mRNA content in resting skeletal muscle suggest that men and women are transcriptionally "primed" for known physiological differences in metabolism however the mechanism behind sex differences in fiber type remains to be determined.

  16. c-fos,bax和p53的反义寡聚核苷酸(ASOs)对溶血磷脂酸所致的培养小鼠大脑皮层神经元凋亡的影响%Effects of antisense oligodeoxynucleotides against c-fos,bax,and p53 mRNAs upon apoptosis induced by lysophosphatidic acid in cultured cortical neurons

    Institute of Scientific and Technical Information of China (English)

    郑肇青; 方羡君; 胥显民; 李建国; 乔健天

    2009-01-01

    性,不同于b淀粉样蛋白片段31-35的致凋亡特性.%three genes are indispensable for the LPA-induced apoptosis,showing a different genomic basis as compared to that for AbP31-35-induced apoptosis.

  17. Salmonella enterica serotype Typhimurium DT104 ArtA-dependent modification of pertussis toxin-sensitive G proteins in the presence of [32P]NAD.

    Science.gov (United States)

    Uchida, Ikuo; Ishihara, Ryoko; Tanaka, Kiyoshi; Hata, Eiji; Makino, Sou-ichi; Kanno, Toru; Hatama, Shinichi; Kishima, Masato; Akiba, Masato; Watanabe, Atsushi; Kubota, Takayuki

    2009-11-01

    Salmonella enterica serotype Typhimurium (S. Typhimurium) definitive phage type (DT) 104 has become a widespread cause of human and other animal infections worldwide. The severity of clinical illness in S. Typhimurium DT104 outbreaks suggests that this strain possesses enhanced virulence. ArtA and ArtB - encoded by a prophage in S. Typhimurium DT104 - are homologues of components of pertussis toxin (PTX), including its ADP-ribosyltransferase subunit. Here, we show that exposing DT104 to mitomycin C, a DNA-damaging agent, induced production of prophage-encoded ArtA/ArtB. Pertussis-sensitive G proteins were labelled in the presence of [(32)P]NAD and ArtA, and the label was released by HgCl(2), which is known to cleave cysteine-ADP-ribose bonds. ADP-dependent modification of G proteins was markedly reduced in in vitro-synthesized ArtA(6Arg-Ala) and ArtA(115Glu-Ala), in which alanine was substituted for the conserved arginine at position 6 (necessary for NAD binding) and the predicted catalytic glutamate at position 115, respectively. A cellular ADP-ribosylation assay and two-dimensional electrophoresis showed that ArtA- and PTX-induced ADP-ribosylation in Chinese hamster ovary (CHO) cells occur with the same type of G proteins. Furthermore, exposing CHO cells to the ArtA/ArtB-containing culture supernatant of DT104 resulted in a clustered growth pattern, as is observed in PTX-exposed CHO cells. Hydrogen peroxide, an oxidative stressor, also induced ArtA/ArtB production, suggesting that these agents induce in vivo synthesis of ArtA/ArtB. These results, taken together, suggest that ArtA/ArtB is an active toxin similar to PTX.

  18. Measurements of observables in the pion-nucleon system, nuclear a- dependence of heavy quark production and rare decays of D and B mesons

    Energy Technology Data Exchange (ETDEWEB)

    Sadler, M.E.; Isenhower, L.D.

    1992-02-15

    This report discusses research on the following topics: pion-nucleon interactions; detector tomography facility; nuclear dependence of charm and beauty quark production and a study of two-prong decays of neutral D and B mesons; N* collaboration at CEBAF; and pilac experiments. (LSP)

  19. In vivo damage and recA-dependent repair of plasmid and chromosomal DNA in the radiation-resistant bacterium Deinococcus radiodurans.

    OpenAIRE

    Daly, M J; Ouyang, L; Fuchs, P.; Minton, K W

    1994-01-01

    Deinococcus radiodurans R1 and other members of this genus share extraordinary resistance to the lethal and mutagenic effects of ionizing radiation. We have recently identified a RecA homolog in strain R1 and have shown that mutation of the corresponding gene causes marked radiosensitivity. We show here that following high-level exposure to gamma irradiation (1.75 megarads, the dose required to yield 37% of CFU for plateau-phase wild-type R1), the wild-type strain repairs > 150 double-strand ...

  20. Protein kinase a dependent phosphorylation of apical membrane antigen 1 plays an important role in erythrocyte invasion by the malaria parasite.

    Directory of Open Access Journals (Sweden)

    Kerstin Leykauf

    Full Text Available Apicomplexan parasites are obligate intracellular parasites that infect a variety of hosts, causing significant diseases in livestock and humans. The invasive forms of the parasites invade their host cells by gliding motility, an active process driven by parasite adhesion proteins and molecular motors. A crucial point during host cell invasion is the formation of a ring-shaped area of intimate contact between the parasite and the host known as a tight junction. As the invasive zoite propels itself into the host-cell, the junction moves down the length of the parasite. This process must be tightly regulated and signalling is likely to play a role in this event. One crucial protein for tight-junction formation is the apical membrane antigen 1 (AMA1. Here we have investigated the phosphorylation status of this key player in the invasion process in the human malaria parasite Plasmodium falciparum. We show that the cytoplasmic tail of P. falciparum AMA1 is phosphorylated at serine 610. We provide evidence that the enzyme responsible for serine 610 phosphorylation is the cAMP regulated protein kinase A (PfPKA. Importantly, mutation of AMA1 serine 610 to alanine abrogates phosphorylation of AMA1 in vivo and dramatically impedes invasion. In addition to shedding unexpected new light on AMA1 function, this work represents the first time PKA has been implicated in merozoite invasion.

  1. DNA binding of the cell cycle transcriptional regulator GcrA depends on N6-adenosine methylation in Caulobacter crescentus and other Alphaproteobacteria.

    Science.gov (United States)

    Fioravanti, Antonella; Fumeaux, Coralie; Mohapatra, Saswat S; Bompard, Coralie; Brilli, Matteo; Frandi, Antonio; Castric, Vincent; Villeret, Vincent; Viollier, Patrick H; Biondi, Emanuele G

    2013-05-01

    Several regulators are involved in the control of cell cycle progression in the bacterial model system Caulobacter crescentus, which divides asymmetrically into a vegetative G1-phase (swarmer) cell and a replicative S-phase (stalked) cell. Here we report a novel functional interaction between the enigmatic cell cycle regulator GcrA and the N6-adenosine methyltransferase CcrM, both highly conserved proteins among Alphaproteobacteria, that are activated early and at the end of S-phase, respectively. As no direct biochemical and regulatory relationship between GcrA and CcrM were known, we used a combination of ChIP (chromatin-immunoprecipitation), biochemical and biophysical experimentation, and genetics to show that GcrA is a dimeric DNA-binding protein that preferentially targets promoters harbouring CcrM methylation sites. After tracing CcrM-dependent N6-methyl-adenosine promoter marks at a genome-wide scale, we show that these marks recruit GcrA in vitro and in vivo. Moreover, we found that, in the presence of a methylated target, GcrA recruits the RNA polymerase to the promoter, consistent with its role in transcriptional activation. Since methylation-dependent DNA binding is also observed with GcrA orthologs from other Alphaproteobacteria, we conclude that GcrA is the founding member of a new and conserved class of transcriptional regulators that function as molecular effectors of a methylation-dependent (non-heritable) epigenetic switch that regulates gene expression during the cell cycle.

  2. DNA binding of the cell cycle transcriptional regulator GcrA depends on N6-adenosine methylation in Caulobacter crescentus and other Alphaproteobacteria.

    Directory of Open Access Journals (Sweden)

    Antonella Fioravanti

    2013-05-01

    Full Text Available Several regulators are involved in the control of cell cycle progression in the bacterial model system Caulobacter crescentus, which divides asymmetrically into a vegetative G1-phase (swarmer cell and a replicative S-phase (stalked cell. Here we report a novel functional interaction between the enigmatic cell cycle regulator GcrA and the N6-adenosine methyltransferase CcrM, both highly conserved proteins among Alphaproteobacteria, that are activated early and at the end of S-phase, respectively. As no direct biochemical and regulatory relationship between GcrA and CcrM were known, we used a combination of ChIP (chromatin-immunoprecipitation, biochemical and biophysical experimentation, and genetics to show that GcrA is a dimeric DNA-binding protein that preferentially targets promoters harbouring CcrM methylation sites. After tracing CcrM-dependent N6-methyl-adenosine promoter marks at a genome-wide scale, we show that these marks recruit GcrA in vitro and in vivo. Moreover, we found that, in the presence of a methylated target, GcrA recruits the RNA polymerase to the promoter, consistent with its role in transcriptional activation. Since methylation-dependent DNA binding is also observed with GcrA orthologs from other Alphaproteobacteria, we conclude that GcrA is the founding member of a new and conserved class of transcriptional regulators that function as molecular effectors of a methylation-dependent (non-heritable epigenetic switch that regulates gene expression during the cell cycle.

  3. Functional characterization of a veA-dependent polyketide synthase gene in Aspergillus flavus necessary for the synthesis of asparasone, a sclerotium-specific pigment.

    Science.gov (United States)

    Cary, Jeffrey W; Harris-Coward, Pamela Y; Ehrlich, Kenneth C; Di Mavungu, José Diana; Malysheva, Svetlana V; De Saeger, Sarah; Dowd, Patrick F; Shantappa, Sourabha; Martens, Stacey L; Calvo, Ana M

    2014-03-01

    The filamentous fungus, Aspergillus flavus, produces the toxic and carcinogenic, polyketide synthase (PKS)-derived family of secondary metabolites termed aflatoxins. While analysis of the A. flavus genome has identified many other PKSs capable of producing secondary metabolites, to date, only a few other metabolites have been identified. In the process of studying how the developmental regulator, VeA, affects A. flavus secondary metabolism we discovered that mutation of veA caused a dramatic down-regulation of transcription of a polyketide synthase gene belonging to cluster 27 and the loss of the ability of the fungi to produce sclerotia. Inactivation of the cluster 27 pks (pks27) resulted in formation of greyish-yellow sclerotia rather than the dark brown sclerotia normally produced by A. flavus while conidial pigmentation was unaffected. One metabolite produced by Pks27 was identified by thin layer chromatography and mass spectral analysis as the known anthraquinone, asparasone A. Sclerotia produced by pks27 mutants were significantly less resistant to insect predation than were the sclerotia produced by the wild-type and more susceptible to the deleterious effects of ultraviolet light and heat. Normal sclerotia were previously thought to be resistant to damage because of a process of melanization similar to that known for pigmentation of conidia. Our results show that the dark brown pigments in sclerotia derive from anthraquinones produced by Pks27 rather than from the typical tetrahydronapthalene melanin production pathway. To our knowledge this is the first report on the genes involved in the biosynthesis of pigments important for sclerotial survival. PMID:24412484

  4. NPM-ALK up-regulates iNOS expression through a STAT3/microRNA-26a-dependent mechanism.

    Science.gov (United States)

    Zhu, Haifeng; Vishwamitra, Deeksha; Curry, Choladda V; Manshouri, Roxsan; Diao, Lixia; Khan, Aarish; Amin, Hesham M

    2013-05-01

    NPM-ALK chimeric oncogene is aberrantly expressed in an aggressive subset of T-cell lymphomas that frequently occurs in children and young adults. The mechanisms underlying the oncogenic effects of NPM-ALK are not completely elucidated. Inducible nitric oxide synthase (iNOS) promotes the survival and maintains the malignant phenotype of cancer cells by generating NO, a highly active free radical. We tested the hypothesis that iNOS is deregulated in NPM-ALK(+) T-cell lymphoma and promotes the survival of this lymphoma. In line with this possibility, an iNOS inhibitor and NO scavenger decreased the viability, adhesion, and migration of NPM-ALK(+) T-cell lymphoma cells, and an NO donor reversed these effects. Moreover, the NO donor salvaged the viability of lymphoma cells treated with ALK inhibitors. In further support of an important role of iNOS, we found iNOS protein to be highly expressed in NPM-ALK(+) T-cell lymphoma cell lines and in 79% of primary tumours but not in human T lymphocytes. Although expression of iNOS mRNA was identified in NPM-ALK(+) T-cell lymphoma cell lines and tumours, iNOS mRNA was remarkably elevated in T lymphocytes, suggesting post-transcriptional regulation. Consistently, we found that miR-26a contains potential binding sites and interacts with the 3'-UTR of iNOS. In addition, miR-26a was significantly decreased in NPM-ALK(+) T-cell lymphoma cell lines and tumours compared with T lymphocytes and reactive lymph nodes. Restoration of miR-26a in lymphoma cells abrogated iNOS protein expression and decreased NO production and cell viability, adhesion, and migration. Importantly, the effects of miR-26a were substantially attenuated when the NO donor was simultaneously used to treat lymphoma cells. Our investigation of the mechanisms underlying the decrease in miR-26a in this lymphoma revealed novel evidence that STAT3, a major downstream substrate of NPM-ALK tyrosine kinase activity, suppresses MIR26A1 gene expression.

  5. Characterization of the gacA-dependent surface and coral mucus colonization by an opportunistic coral pathogen Serratia marcescens PDL100.

    Science.gov (United States)

    Krediet, Cory J; Carpinone, Emily M; Ritchie, Kim B; Teplitski, Max

    2013-05-01

    Opportunistic pathogens rely on global regulatory systems to assess the environment and to control virulence and metabolism to overcome host defenses and outcompete host-associated microbiota. In Gammaproteobacteria, GacS/GacA is one such regulatory system. GacA orthologs direct the expression of the csr (rsm) small regulatory RNAs, which through their interaction with the RNA-binding protein CsrA (RsmA), control genes with functions in carbon metabolism, motility, biofilm formation, and virulence. The csrB gene was controlled by gacA in Serratia marcescens PDL100. A disruption of the S. marcescens gacA gene resulted in an increased fitness of the mutant on mucus of the host coral Acropora palmata and its high molecular weight fraction, whereas the mutant was as competitive as the wild type on the low molecular weight fraction of the mucus. Swarming motility and biofilm formation were reduced in the gacA mutant. This indicates a critical role for gacA in the efficient utilization of specific components of coral mucus and establishment within the surface mucopolysaccharide layer. While significantly affecting early colonization behaviors (coral mucus utilization, swarming motility, and biofilm formation), gacA was not required for virulence of S. marcescens PDL100 in either a model polyp Aiptasia pallida or in brine shrimp Artemia nauplii. PMID:23278392

  6. Accumulation of cyclin B1 requires E2F and cyclin-A-dependent rearrangement of the anaphase-promoting complex

    DEFF Research Database (Denmark)

    Lukas, C; Sørensen, Claus Storgaard; Kramer, E;

    1999-01-01

    genes beyond the G1/S transition is required for coordinating S-phase progression with cell division, a process driven by cyclin-B-dependent kinase and anaphase-promoting complex (APC)-mediated proteolysis. How E2F-dependent events at G1/S transition are orchestrated with cyclin B and APC activity...... in the timely accumulation of cyclin B1 and the coordination of cell-cycle progression during the post-restriction point period....

  7. A degenerate tri-partite DNA binding site required for activation of ComA-dependent quorum response gene expression in Bacillus subtilis

    OpenAIRE

    Griffith, Kevin L.; Grossman, Alan D.

    2008-01-01

    In Bacillus subtilis, the transcription factor ComA activates several biological processes in response to increasing population density. Extracellular peptide signaling is used to coordinate the activity of ComA with population density. At low culture densities, when the concentration of signaling peptides is lowest, ComA is largely inactive. At higher densities, when the concentration of signaling peptides is higher, ComA is active and activates transcription of at least 9 operons involved i...

  8. Characterization of the gacA-dependent surface and coral mucus colonization by an opportunistic coral pathogen Serratia marcescens PDL100.

    Science.gov (United States)

    Krediet, Cory J; Carpinone, Emily M; Ritchie, Kim B; Teplitski, Max

    2013-05-01

    Opportunistic pathogens rely on global regulatory systems to assess the environment and to control virulence and metabolism to overcome host defenses and outcompete host-associated microbiota. In Gammaproteobacteria, GacS/GacA is one such regulatory system. GacA orthologs direct the expression of the csr (rsm) small regulatory RNAs, which through their interaction with the RNA-binding protein CsrA (RsmA), control genes with functions in carbon metabolism, motility, biofilm formation, and virulence. The csrB gene was controlled by gacA in Serratia marcescens PDL100. A disruption of the S. marcescens gacA gene resulted in an increased fitness of the mutant on mucus of the host coral Acropora palmata and its high molecular weight fraction, whereas the mutant was as competitive as the wild type on the low molecular weight fraction of the mucus. Swarming motility and biofilm formation were reduced in the gacA mutant. This indicates a critical role for gacA in the efficient utilization of specific components of coral mucus and establishment within the surface mucopolysaccharide layer. While significantly affecting early colonization behaviors (coral mucus utilization, swarming motility, and biofilm formation), gacA was not required for virulence of S. marcescens PDL100 in either a model polyp Aiptasia pallida or in brine shrimp Artemia nauplii.

  9. Tauroursodeoxycholate Protects Rat Hepatocytes from Bile Acid-Induced Apoptosis via β1-Integrin- and Protein Kinase A-Dependent Mechanisms

    Directory of Open Access Journals (Sweden)

    Annika Sommerfeld

    2015-05-01

    Full Text Available Background/Aims: Ursodeoxycholic acid, which in vivo is rapidly converted into its taurine conjugate, is frequently used for the treatment of cholestatic liver disease. Apart from its choleretic effects, tauroursodeoxycholate (TUDC can protect hepatocytes from bile acid-induced apoptosis, but the mechanisms underlying its anti-apoptotic effects are poorly understood. Methods: These mechanisms were investigated in perfused rat liver and isolated rat hepatocytes. Results: It was found that TUDC inhibited the glycochenodeoxycholate (GCDC-induced activation of the CD95 death receptor at the level of association between CD95 and the epidermal growth factor receptor. This was due to a rapid TUDC-induced β1-integrin-dependent cyclic AMP (cAMP signal with induction of the dual specificity mitogen-activated protein (MAP kinase phosphatase 1 (MKP-1, which prevented GCDC-induced phosphorylation of mitogen-activated protein kinase kinase 4 (MKK4 and c-jun-NH2-terminal kinase (JNK activation. Furthermore, TUDC induced a protein kinase A (PKA-mediated serine/threonine phosphorylation of the CD95, which was recently identified as an internalization signal for CD95. Furthermore, TUDC inhibited GCDC-induced CD95 targeting to the plasma membrane in a β1-integrin-and PKA-dependent manner. In line with this, the β1-integrin siRNA knockdown in sodium taurocholate cotransporting polypeptide (Ntcp-transfected HepG2 cells abolished the protective effect of TUDC against GCDC-induced apoptosis. Conclusion: TUDC exerts its anti-apoptotic effect via a β1-integrin-mediated formation of cAMP, which prevents CD95 activation by hydrophobic bile acids at the levels of JNK activation and CD95 serine/threonine phosphorylation.

  10. Melanoma dormancy in a mouse model is linked to GILZ/FOXO3A-dependent quiescence of disseminated stem-like cells.

    Science.gov (United States)

    Touil, Yasmine; Segard, Pascaline; Ostyn, Pauline; Begard, Severine; Aspord, Caroline; El Machhour, Raja; Masselot, Bernadette; Vandomme, Jerome; Flamenco, Pilar; Idziorek, Thierry; Figeac, Martin; Formstecher, Pierre; Quesnel, Bruno; Polakowska, Renata

    2016-01-01

    Metastatic cancer relapses following the reactivation of dormant, disseminated tumour cells; however, the cells and factors involved in this reactivation are just beginning to be identified. Using an immunotherapy-based syngeneic model of melanoma dormancy and GFP-labelled dormant cell-derived cell lines, we determined that vaccination against melanoma prevented tumour growth but did not prevent tumour cell dissemination or eliminate all tumour cells. The persistent disseminated melanoma tumour cells were quiescent and asymptomatic for one year. The quiescence/activation of these cells in vitro and the dormancy of melanoma in vivo appeared to be regulated by glucocorticoid-induced leucine zipper (GILZ)-mediated immunosuppression. GILZ expression was low in dormant cell-derived cultures, and re-expression of GILZ inactivated FOXO3A and its downstream target, p21CIP1. The ability of dormancy-competent cells to re-enter the cell cycle increased after a second round of cellular dormancy in vivo in association with shortened tumour dormancy period and faster and more aggressive melanoma relapse. Our data indicate that future cancer treatments should be adjusted according to the stage of disease progression. PMID:27465291

  11. Can weighting compensate for nonresponse bias in a dependent variable? An evaluation of weighting methods to correct for substantive bias in a mail survey among Dutch municipalities

    NARCIS (Netherlands)

    van Goor, H; Stuiver, B

    1998-01-01

    Due to a lack of pertinent data, little is known about nonresponse in substantive, generally "dependent" variables and its consequences. However, in a study on policy performance of Dutch municipalities, we were fortunately able to gather performance data fur respondents and nonrespondents from inde

  12. UvrD303, a Hyperhelicase Mutant That Antagonizes RecA-Dependent SOS Expression by a Mechanism That Depends on Its C Terminus▿ †

    OpenAIRE

    Centore, Richard C.; Leeson, Michael C.; Sandler, Steven J.

    2008-01-01

    Genomic integrity is critical for an organism's survival and ability to reproduce. In Escherichia coli, the UvrD helicase has roles in nucleotide excision repair and methyl-directed mismatch repair and can limit reactions by RecA under certain circumstances. UvrD303 (D403A D404A) is a hyperhelicase mutant, and when expressed from a multicopy plasmid, it results in UV sensitivity (UVs), recombination deficiency, and antimutability. In order to understand the molecular mechanism underlying the ...

  13. UvrD303, a hyperhelicase mutant that antagonizes RecA-dependent SOS expression by a mechanism that depends on its C terminus.

    Science.gov (United States)

    Centore, Richard C; Leeson, Michael C; Sandler, Steven J

    2009-03-01

    Genomic integrity is critical for an organism's survival and ability to reproduce. In Escherichia coli, the UvrD helicase has roles in nucleotide excision repair and methyl-directed mismatch repair and can limit reactions by RecA under certain circumstances. UvrD303 (D403A D404A) is a hyperhelicase mutant, and when expressed from a multicopy plasmid, it results in UV sensitivity (UV(s)), recombination deficiency, and antimutability. In order to understand the molecular mechanism underlying the UV(s) phenotype of uvrD303 cells, this mutation was transferred to the E. coli chromosome and studied in single copy. It is shown here that uvrD303 mutants are UV sensitive, recombination deficient, and antimutable and additionally have a moderate defect in inducing the SOS response after UV treatment. The UV-sensitive phenotype is epistatic with recA and additive with uvrA and is partially suppressed by removing the LexA repressor. Furthermore, uvrD303 is able to inhibit constitutive SOS expression caused by the recA730 mutation. The ability of UvrD303 to antagonize SOS expression was dependent on its 40 C-terminal amino acids. It is proposed that UvrD303, via its C terminus, can decrease the levels of RecA activity in the cell. PMID:19074381

  14. Group psycho-education in patients with bipolar disorder associated with a dependency of toxic substances in patients who are in abstinence

    Directory of Open Access Journals (Sweden)

    Patricia González Alegre

    2009-01-01

    Full Text Available The high comorbility that exists among psychiatric disorders and addictive is important. In the latest years it is produced an increase of the sensibility related to this problem. A great deal it is due to the demand of Mental Health Services and also due to drug dependency, as a consequence of the lack of an integral approach. Because of this fact and because of the mentioned demand, we though it should be pertinent developing a research project in order to check if the carrying the psycho-educative preventive group project out in patients with a diagnose of bipolar disorder with an abuse of drugs history and/or dependency of toxic substance in abstinence at the moment influents in a positive way in the course of the number of relapses in the toxic consumption during at least six months subsequent to the intervention. And at this way, these patients will purchase a greater consciousness of the important of healthy habits in the bipolar disorder and the recovery in the toxic substance abuse. The program will be developed in an experimental research where the patients will be randomly assigned in group control/ experimentally, the intervention will last twenty sessions, each session will be an hour and a half long and will be held weekly. In these sessions we will deal with topics related to the psychiatric disorder and the toxic consume. At the same time we will bank on the development of practical relaxation workshops on in some of the sessions with the object of providing a resource in view of stress situations.

  15. Protein crystallographic studies of CoA-dependent proteins: new insight into the binding mode and exchange mechanism of acyl-CoA

    OpenAIRE

    Taskinen, J. (Jukka)

    2006-01-01

    Abstract Multifunctional enzyme type 1 (MFE-1) is a monomeric member of the hydratase/isomerase superfamily (H/I) involved in the β-oxidation of fatty acids. MFE-1 has 2-enoyl-CoA hydratase-1, Δ3-Δ2-enoyl-CoA isomerase, and several other enoyl-CoA isomerase activities at the N-terminus. The C-terminus has (3S)-hydroxyacyl-CoA dehydrogenase activity. MFE-1 can also convert certain hydroxylated C27 bile acid synthesis intermediates. In these studies, a domain assignment of MFE-1 by sequ...

  16. Protein kinase A-dependent Neuronal Nitric Oxide Synthase Activation Mediates the Enhancement of Baroreflex Response by Adrenomedullin in the Nucleus Tractus Solitarii of Rats

    Directory of Open Access Journals (Sweden)

    Ho I-Chun

    2011-05-01

    Full Text Available Abstract Background Adrenomedullin (ADM exerts its biological functions through the receptor-mediated enzymatic mechanisms that involve protein kinase A (PKA, or neuronal nitric oxide synthase (nNOS. We previously demonstrated that the receptor-mediated cAMP/PKA pathway involves in ADM-enhanced baroreceptor reflex (BRR response. It remains unclear whether ADM may enhance BRR response via activation of nNOS-dependent mechanism in the nucleus tractus solitarii (NTS. Methods Intravenous injection of phenylephrine was administered to evoke the BRR before and at 10, 30, and 60 min after microinjection of the test agents into NTS of Sprague-Dawley rats. Western blotting analysis was used to measure the level and phosphorylation of proteins that involved in BRR-enhancing effects of ADM (0.2 pmol in NTS. The colocalization of PKA and nNOS was examined by immunohistochemical staining and observed with a laser confocal microscope. Results We found that ADM-induced enhancement of BRR response was blunted by microinjection of NPLA or Rp-8-Br-cGMP, a selective inhibitor of nNOS or protein kinase G (PKG respectively, into NTS. Western blot analysis further revealed that ADM induced an increase in the protein level of PKG-I which could be attenuated by co-microinjection with the ADM receptor antagonist ADM22-52 or NPLA. Moreover, we observed an increase in phosphorylation at Ser1416 of nNOS at 10, 30, and 60 min after intra-NTS administration of ADM. As such, nNOS/PKG signaling may also account for the enhancing effect of ADM on BRR response. Interestingly, biochemical evidence further showed that ADM-induced increase of nNOS phosphorylation was prevented by co-microinjection with Rp-8-Br-cAMP, a PKA inhibitor. The possibility of PKA-dependent nNOS activation was substantiated by immunohistochemical demonstration of co-localization of PKA and nNOS in putative NTS neurons. Conclusions The novel finding of this study is that the signal transduction cascade that underlies the enhancement of BRR response by ADM in NTS is composed sequentially of cAMP/PKA and nNOS/PKG pathways.

  17. Measurements of observables in the pion-nucleon system, nuclear a- dependence of heavy quark production and rare decays of D and B mesons

    International Nuclear Information System (INIS)

    This report discusses research on the following topics: pion-nucleon interactions; detector tomography facility; nuclear dependence of charm and beauty quark production and a study of two-prong decays of neutral D and B mesons; N* collaboration at CEBAF; and pilac experiments

  18. Identification of genes responsible for RelA-dependent proliferation arrest in human mammary epithelial cells conditionally expressing RelA

    OpenAIRE

    Bose S Kochupurakkal; J Dirk Iglehart

    2015-01-01

    The molecular mechanisms responsible for opposing oncogenic and tumor-suppressor activities of NF-kB are obscure. Semi-quantitative immunohistochemistry of primary breast tumors using antibodies to RelA, the pleiotropic NF-kB factor, and Ki67 revealed a negative correlation between RelA levels and Ki67-index among ER +/HER2 − tumors [1]. Similarly, expression of AURKA, a marker for proliferation, negatively correlates with expression of NFKBIA, a surrogate for RelA expression and activity, in...

  19. A RelA-dependent two-tiered regulated proteolysis cascade controls synthesis of a contact-dependent intercellular signal in Myxococcus xanthus.

    Science.gov (United States)

    Konovalova, Anna; Löbach, Stephanie; Søgaard-Andersen, Lotte

    2012-04-01

    Proteolytic cleavage of precursor proteins to generate intercellular signals is a common mechanism in all cells. In Myxococcus xanthus the contact-dependent intercellular C-signal is a 17 kDa protein (p17) generated by proteolytic cleavage of the 25 kDa csgA protein (p25), and is essential for starvation-induced fruiting body formation. p25 accumulates in the outer membrane and PopC, the protease that cleaves p25, in the cytoplasm of vegetative cells. PopC is secreted in response to starvation, therefore, restricting p25 cleavage to starving cells. We focused on identifying proteins critical for PopC secretion in response to starvation. PopC secretion depends on the (p)ppGpp synthase RelA and the stringent response, and is regulated post-translationally. PopD, which is encoded in an operon with PopC, forms a soluble complex with PopC and inhibits PopC secretion whereas the integral membrane AAA+ protease FtsH(D) is required for PopC secretion. Biochemical and genetic evidence suggest that in response to starvation, RelA is activated and induces the degradation of PopD thereby releasing pre-formed PopC for secretion and that FtsH(D) is important for PopD degradation. Hence, regulated PopC secretion depends on regulated proteolysis. Accordingly, p17 synthesis depends on a proteolytic cascade including FtsH(D) -dependent degradation of PopD and PopC-dependent cleavage of p25. PMID:22404381

  20. HipA-triggered growth arrest and β-lactam tolerance in Escherichia coli are mediated by RelA-dependent ppGpp synthesis.

    Science.gov (United States)

    Bokinsky, Gregory; Baidoo, Edward E K; Akella, Swetha; Burd, Helcio; Weaver, Daniel; Alonso-Gutierrez, Jorge; García-Martín, Héctor; Lee, Taek Soon; Keasling, Jay D

    2013-07-01

    Persistence is a phenomenon whereby a subpopulation of bacterial cells enters a transient growth-arrested state that confers antibiotic tolerance. While entrance into persistence has been linked to the activities of toxin proteins, the molecular mechanisms by which toxins induce growth arrest and the persistent state remain unclear. Here, we show that overexpression of the protein kinase HipA in Escherichia coli triggers growth arrest by activating synthesis of the alarmone guanosine tetraphosphate (ppGpp) by the enzyme RelA, a signal typically associated with amino acid starvation. We further demonstrate that chemically suppressing ppGpp synthesis with chloramphenicol relieves inhibition of DNA replication initiation and RNA synthesis in HipA-arrested cells and restores vulnerability to β-lactam antibiotics. HipA-arrested cells maintain glucose uptake and oxygen consumption and accumulate amino acids as a consequence of translational inhibition. We harness the active metabolism of HipA-arrested cells to provide a bacteriophage-resistant platform for the production of biotechnologically relevant compounds, which may represent an innovative solution to the costly problem of phage contamination in industrial fermentations.

  1. Synthesis of Asymmetric Propanetriol Analogues

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    From natural tartaric acid, (R)-2-benzyloxy-3-(2-tetrahydropyranyloxy) propanol 3 was designed and synthesized, and (R)-2-benzyloxy-3-(4-methoxybenzyloxy) propanol 7 was prepared in a new method. They can be used as chiral synthons of lysophosphatidic acid and other compounds with asymmetric propanetriol backbone.

  2. Rac controls PIP5K localisation and PtdIns(4,5)P-2 synthesis, which modulates vinculin localisation and neurite dynamics

    NARCIS (Netherlands)

    J.R. Halstead; N.E. Savaskan; I. van den Bout; F. van Horck; A. Hajdo-Milasinovic; M. Snell; W.J. Keune; J.P.T. Klooster; P.L. Hordijk; N. Divecha

    2010-01-01

    In N1E-115 cells, neurite retraction induced by neurite remodelling factors such as lysophosphatidic acid, sphingosine 1-phosphate and semaphorin 3A require the activity of phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks). PIP5Ks synthesise the phosphoinositide lipid second messenger phosphatidyl

  3. Production of LPA by autotaxin and toxic phospholipases D : biochemical characterization and physiological implications

    NARCIS (Netherlands)

    Meeteren, Laurens Arend van

    2007-01-01

    Lysophosphatidic acid (LPA) is a lipid mediator with a wide variety of biological actions, particularly as an inducer of cell proliferation, migration and survival. LPA binds to at least five distinct G protein-coupled receptors and thereby activates multiple signal transduction pathways, including

  4. Skeletal Muscle Magnetic Resonance Imaging of the Lower Limbs in Late-onset Lipid Storage Myopathy with Electron Transfer Flavoprotein Dehydrogenase Gene Mutations

    Institute of Scientific and Technical Information of China (English)

    Xin-Yi Liu; Ming Jin; Zhi-Qiang Wang; Dan-Ni Wang; Jun-Jie He; Min-Ting Lin; Hong-Xia Fu

    2016-01-01

    Background:Lipid storage myopathy (LSM) is a genetically heterogeneous group with variable clinical phenotypes.Late-onset multiple acyl-coenzyme A dehydrogenation deficiency (MADD) is a rather common form of LSM in China.Diagnosis and clinical management of it remain challenging,especially without robust muscle biopsy result and genetic detection.As the noninvasion and convenience,muscle magnetic resonance imaging (MRI) is a helpful assistant,diagnostic tool for neuromuscular disorders.However,the disease-specific MRI patterns of muscle involved and its diagnostic value in late-onset MADD have not been systematic analyzed.Methods:We assessed the MRI pattern and fat infiltration degree of the lower limb muscles in 28 late-onset MADD patients,combined with detailed clinical features and gene spectrum.Fat infiltration degree of the thigh muscle was scored while that ofgluteus was described as obvious or not.Associated muscular atrophy was defined as obvious muscle bulk reduction.Results:The mean scores were significantly different among the anterior,medial,and posterior thigh muscle groups.The mean of fat infiltration scores on posterior thigh muscle group was significantly higher than either anterior or medial thigh muscle group (P < 0.001).Moreover,the mean score on medial thigh muscle group was significantly higher than that of anterior thigh muscle group (P < 0.01).About half of the patients displayed fat infiltration and atrophy in gluteus muscles.Of 28 patients,12 exhibited atrophy in medial and/or posterior thigh muscle groups,especially in posterior thigh muscle group.Muscle edema pattern was not found in all the patients.Conclusions:Late-onset MADD patients show a typical muscular imaging pattern of fat infiltration and atrophy on anterior,posterior,and medial thigh muscle groups,with major involvement of posterior thigh muscle group and gluteus muscles and a sparing involvement of anterior thigh compartment.Our findings also suggest that muscle MRI of

  5. Cell Suspension Culture of Eriobotrya japonica Regulates the Diabetic and Hyperlipidemic Signs of High-Fat-Fed Mice

    Directory of Open Access Journals (Sweden)

    Jin-Bin Wu

    2013-03-01

    Full Text Available The present study investigates the anti-hyperlipidemic and antihyperglycemic effects and mechanism in high-fat (HF-fed mice of cell suspension culture of Eriobotrya japonica (TA, which contains a great number of pentacyclic terpenoids. Firstly, C57BL/6J mice were randomly divided into two groups: the control (CON group was fed with a low-fat diet (n = 9, whereas the experimental group was fed a 45% HF diet for 8 weeks. Afterwards, the CON group was treated with vehicle, whereas the HF group was subdivided into five groups and was orally given TA or rosiglitazone or not for 4 weeks. Blood and visceral adipose tissue, liver tissue and skeletal muscle were examined. Treatment with TA reduced body weight gain, weights of white adipose tissue (WAT (including epididymal, perirenal, mesenteric WAT and visceral fat, and hepatic triacylglycerol content significantly without affecting food intake in diet-induced diabetic mice. TA effectively prevented HF diet-induced increases in the levels of blood glucose, insulin, leptin and HOMA-IR index (p < 0.001, p < 0.05, p < 0.05, p < 0.01, respectively and attenuated insulin resistance. Treatment with TA, adipocytes in the visceral depots showed a reduction in size. TA effectively significantly increased the protein contents of phosphorylation of AMPK-α (Thr172 both in liver and adipose tissue. It is shown that TA exhibits hypolipidemic effect in HF-fed mice by decreasing gene expressions of fatty acid synthesis, including acyl-coenzyme A: diacylglycerol acyltransferase (DGAT 2, which catalyzes the final step in the synthesis of triglycerides, and antidiabetic properties occurred as a result of decreased hepatic glucose production via phosphenolpyruvate carboxykinase (PEPCK down- regulation, improved insulin sensitization and TA (at 1.0 g/kg dose decreased expression of hepatic and adipose 11-β-hydroxysteroid dehydroxygenase (11β-HSD1 gene, which contributed in attenuating diabetic state. Futhermore, TA at

  6. Validation of the Antidiabetic and Hypolipidemic Effects of Clitocybe nuda by Assessment of Glucose Transporter 4 and Gluconeogenesis and AMPK Phosphorylation in Streptozotocin-Induced Mice

    Directory of Open Access Journals (Sweden)

    Chun-Ching Shih

    2014-01-01

    Full Text Available The study was designed to investigate the effects of extract of Clitocybe nuda (CNE on type 1 diabetes mellitus and dyslipidemia in streptozotocin- (STZ- induced diabetic mice. Diabetes was induced by injection of STZ. Diabetic mice were randomly divided into five groups and given orally CNE (C1: 0.2, C2: 0.5, and C3: 1.0 g/kg body weight or metformin (Metf or vehicle for 4 weeks. STZ induction decreased in the levels of insulin, body weight, and the weight of skeletal muscle, whereas the levels of blood glucose, hemoglobin nonenzymatically (percent HbA1c, and circulating triglyceride (P < 0.001, P < 0.001, and P < 0.01, resp. were increased. CNE decreased the levels of blood glucose, HbA1c, and triglyceride levels, whereas it increased the levels of insulin and leptin compared with the vehicle-treated STZ group. STZ induction caused a decrease in the protein contents of skeletal muscular and hepatic phosphorylation of AMP-activated protein kinase (phospho-AMPK and muscular glucose transporter 4 (GLUT4. Muscular phospho-AMPK contents were increased in C2-, C3-, and Metf-treated groups. CNE and Metf significantly increased the muscular proteins of GLUT4. Liver phospho-AMPK showed an increase in all CNE- and Metf-treated groups combined with the decreased hepatic glucose production by decreasing phosphenolpyruvate carboxykinase (PEPCK, glucose-6-phosphatase (G6Pase, and 11beta hydroxysteroid dehydroxygenase (11β-HSD1 gene, which contributed to attenuating diabetic state. The study indicated that the hypoglycemic properties of CNE were related to both the increased muscular glucose uptake and the reduction in hepatic gluconeogenesis. CNE exerts hypolipidemic effect by increasing gene expressions of peroxisome proliferator-activated receptor α (PPARα and decreasing expressions of fatty acid synthesis, including acyl-coenzyme A: diacylglycerol acyltransferase (DGAT 2. Therefore, amelioration of diabetic and dyslipidemic state by CNE in STZ

  7. Mechanisms of toxicity of di(2-ethylhexyl) phthalate on the reproductive health of male zebrafish

    Energy Technology Data Exchange (ETDEWEB)

    Uren-Webster, Tamsyn M.; Lewis, Ceri; Filby, Amy L.; Paull, Gregory C. [Hatherly Laboratories, School of Biosciences, University of Exeter, Prince of Wales Road, Exeter, Devon EX4 4PS (United Kingdom); Santos, Eduarda M., E-mail: e.santos@exeter.ac.uk [Hatherly Laboratories, School of Biosciences, University of Exeter, Prince of Wales Road, Exeter, Devon EX4 4PS (United Kingdom)

    2010-09-01

    Phthalates are ubiquitous in the aquatic environment and are known to adversely affect male reproductive health in mammals through interactions with multiple receptor systems. However, little is known about the risks they pose to fish. This project investigated the effects of di(2-ethylhexyl) phthalate (DEHP), the most commonly used phthalate, on the reproductive health of male zebrafish (Danio rerio). Males were treated with 0.5, 50 and 5000 mg DEHP kg{sup -1} (body weight) for a period of 10 days via intraperitoneal injection. The effects of the exposure were assessed by analysing fertilisation success, testis histology, sperm DNA integrity and transcript profiles of the liver and testis. A significant increase in the hepatosomatic index and levels of hepatic vitellogenin transcript were observed following exposure to 5000 mg DEHP kg{sup -1}. Exposure to 5000 mg DEHP kg{sup -1} also resulted in a reduction in fertilisation success of oocytes spawned by untreated females. However, survival and development of the resulting embryos were unaffected by all treatments, and no evidence of DEHP-induced sperm DNA damage was observed. Exposure to 50 and 5000 mg DEHP kg{sup -1} caused alterations in the proportion of germ cells at specific stages of spermatogenesis in the testis, including a reduction in the proportion of spermatozoa and an increase in the proportion of spermatocytes, suggesting that DEHP may inhibit the progression of meiosis. In parallel, exposure to 5000 mg DEHP kg{sup -1} increased the levels of two peroxisome proliferator-activated receptor (PPAR) responsive genes (acyl-coenzyme A oxidase 1 (acox1) and enoyl-coenzyme A, hydratase/3-hydroxyacyl coenzyme A dehydrogenase (ehhadh). These data demonstrated that exposure to high concentrations of DEHP disrupts spermatogenesis in adult zebrafish with a consequent decrease in their ability to fertilise oocytes spawned by untreated females. Furthermore, our data suggest that the adverse effects caused by

  8. mRNA expression of genes regulating lipid metabolism in ringed seals (Pusa hispida) from differently polluted areas

    Energy Technology Data Exchange (ETDEWEB)

    Castelli, Martina Galatea [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway); University of Bergen, Department of Biology, 5020 Bergen (Norway); Rusten, Marte; Goksøyr, Anders [University of Bergen, Department of Biology, 5020 Bergen (Norway); Routti, Heli, E-mail: heli.routti@npolar.no [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway)

    2014-01-15

    Highlights: •Genes regulating lipid metabolism were studied in ringed seals. •We compared highly contaminated Baltic seals and less contaminated Svalbard seals. •mRNA expression of hepatic PPARγ was higher in the Baltic seals. •mRNA expression of adipose PPARγ target genes was higher in the Baltic seals. •Contaminant exposure may affect lipid metabolism in the Baltic ringed seals. -- Abstract: There is a growing concern about the ability of persistent organic pollutants (POPs) to influence lipid metabolism. Although POPs are found at high concentrations in some populations of marine mammals, for example in the ringed seal (Pusa hispida) from the Baltic Sea, little is known about the effects of POPs on their lipid metabolism. An optimal regulation of lipid metabolism is crucial for ringed seals during the fasting/molting season. This is a physiologically stressful period, during which they rely on the energy stored in their fat reserves. The mRNA expression levels for seven genes involved in lipid metabolism were analyzed in liver and/or blubber tissue from molting ringed seals from the polluted Baltic Sea and a less polluted reference location, Svalbard (Norway). mRNA expression of genes encoding peroxisome proliferator-activated receptors (PPAR) α and γ and their target genes acyl-coenzyme A oxidase 1 (ACOX1) and cluster of differentiation 36 (CD36) were analyzed in liver. mRNA expression level of genes encoding PPARβ, PPARγ and their target genes encoding fatty acid binding protein 4 (FABP4) and adiponectin (ADIPOQ) were measured in inner and middle blubber layers. In addition, we evaluated the influence of molting status on hepatic mRNA expression of genes encoding PPARs and their target genes in ringed seals from Svalbard. Our results show higher mRNA expression of genes encoding hepatic PPARγ and adipose PPARβ, FABP4, and ADIPOQ in the Baltic seals compared to the Svalbard seals. A positive relationship between mRNA expressions of genes

  9. Male Sterile2 Encodes a Plastid-Localized Fatty Acyl Carrier Protein Reductase Required for Pollen Exine Development in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Chen, W.; Shanklin, J.; Yu, X.-H.; Zhang, K.; Shi, J.; De Oliveira, S.; Schreiber, L.; Zhang, D.

    2011-10-01

    Male Sterile2 (MS2) is predicted to encode a fatty acid reductase required for pollen wall development in Arabidopsis (Arabidopsis thaliana). Transient expression of MS2 in tobacco (Nicotiana benthamiana) leaves resulted in the accumulation of significant levels of C16 and C18 fatty alcohols. Expression of MS2 fused with green fluorescent protein revealed that an amino-terminal transit peptide targets the MS2 to plastids. The plastidial localization of MS2 is biologically important because genetic complementation of MS2 in ms2 homozygous plants was dependent on the presence of its amino-terminal transit peptide or that of the Rubisco small subunit protein amino-terminal transit peptide. In addition, two domains, NAD(P)H-binding domain and sterile domain, conserved in MS2 and its homologs were also shown to be essential for MS2 function in pollen exine development by genetic complementation testing. Direct biochemical analysis revealed that purified recombinant MS2 enzyme is able to convert palmitoyl-Acyl Carrier Protein to the corresponding C16:0 alcohol with NAD(P)H as the preferred electron donor. Using optimized reaction conditions (i.e. at pH 6.0 and 30 C), MS2 exhibits a K{sub m} for 16:0-Acyl Carrier Protein of 23.3 {+-} 4.0 {mu}m, a V{sub max} of 38.3 {+-} 4.5 nmol mg{sup -1} min{sup -1}, and a catalytic efficiency/K{sub m} of 1,873 m{sup -1} s{sup -1}. Based on the high homology of MS2 to other characterized fatty acid reductases, it was surprising that MS2 showed no activity against palmitoyl- or other acyl-coenzyme A; however, this is consistent with its plastidial localization. In summary, genetic and biochemical evidence demonstrate an MS2-mediated conserved plastidial pathway for the production of fatty alcohols that are essential for pollen wall biosynthesis in Arabidopsis.

  10. Homozygous and heterozygous GH transgenesis alters fatty acid composition and content in the liver of Amago salmon (Oncorhynchus masou ishikawae

    Directory of Open Access Journals (Sweden)

    Manabu Sugiyama

    2012-08-01

    Growth hormone (GH transgenic Amago (Oncorhynchus masou ishikawae, containing the sockeye GH1 gene fused with metallothionein-B promoter from the same species, were generated and the physiological condition through lipid metabolism compared among homozygous (Tg/Tg and heterozygous GH transgenic (Tg/+ Amago and the wild type control (+/+. Previously, we have reported that the adipose tissue was generally smaller in GH transgenic fish compared to the control, and that the Δ-6 fatty acyl desaturase gene was down-regulated in the Tg/+ fish. However, fatty acid (FA compositions have not been measured previously in these fish. In this study we compared the FAs composition and content in the liver using gas chromatography. Eleven kinds of FA were detected. The composition of saturated and monounsaturated fatty acids (SFA and MUFA such as myristic acid (14:0, palmitoleic acid (16:1n-7, and cis-vaccenic acid (cis-18:1n-7 was significantly (P<0.05 decreased in GH transgenic Amago. On the other hand, the composition of polyunsaturated fatty acids (PUFAs such as linoleic acid (18:2n-6, arachidonic acid (20:4n-6, and docosapentaenoic acid (22:5n-3 was significantly (P<0.05 increased. Levels of serum glucose and triacylglycerol were significantly (P<0.05 decreased in the GH transgenics compared with +/+ fish. Furthermore, 3′-tag digital gene expression profiling was performed using liver tissues from Tg/Tg and +/+ fish, and showed that Mid1 interacting protein 1 (Mid1ip1, which is an important factor to activate Acetyl-CoA carboxylase (ACC, was down-regulated in Tg/Tg fish, while genes involved in FA catabolism were up-regulated, including long-chain-fatty-acid–CoA ligase 1 (ACSL1 and acyl-coenzyme A oxidase 3 (ACOX3. These data suggest that liver tissue from GH transgenic Amago showed starvation by alteration in glucose and lipid metabolism due to GH overexpression. The decrease of serum glucose suppressed Mid1ip1, and caused a decrease of de novo FA synthesis, resulting

  11. Acyl-CoA:cholesterol acyltransferases (ACATs/SOATs): Enzymes with multiple sterols as substrates and as activators.

    Science.gov (United States)

    Rogers, Maximillian A; Liu, Jay; Song, Bao-Liang; Li, Bo-Liang; Chang, Catherine C Y; Chang, Ta-Yuan

    2015-07-01

    Cholesterol is essential to the growth and viability of cells. The metabolites of cholesterol include: steroids, oxysterols, and bile acids, all of which play important physiological functions. Cholesterol and its metabolites have been implicated in the pathogenesis of multiple human diseases, including: atherosclerosis, cancer, neurodegenerative diseases, and diabetes. Thus, understanding how cells maintain the homeostasis of cholesterol and its metabolites is an important area of study. Acyl-coenzyme A:cholesterol acyltransferases (ACATs, also abbreviated as SOATs) converts cholesterol to cholesteryl esters and play key roles in the regulation of cellular cholesterol homeostasis. ACATs are most unusual enzymes because (i) they metabolize diverse substrates including both sterols and certain steroids; (ii) they contain two different binding sites for steroidal molecules. In mammals, there are two ACAT genes that encode two different enzymes, ACAT1 and ACAT2. Both are allosteric enzymes that can be activated by a variety of sterols. In addition to cholesterol, other sterols that possess the 3-beta OH at C-3, including PREG, oxysterols (such as 24(S)-hydroxycholesterol and 27-hydroxycholesterol, etc.), and various plant sterols, could all be ACAT substrates. All sterols that possess the iso-octyl side chain including cholesterol, oxysterols, various plant sterols could all be activators of ACAT. PREG can only be an ACAT substrate because it lacks the iso-octyl side chain required to be an ACAT activator. The unnatural cholesterol analogs epi-cholesterol (with 3-alpha OH in steroid ring B) and ent-cholesterol (the mirror image of cholesterol) contain the iso-octyl side chain but do not have the 3-beta OH at C-3. Thus, they can only serve as activators and cannot serve as substrates. Thus, within the ACAT holoenzyme, there are site(s) that bind sterol as substrate and site(s) that bind sterol as activator; these sites are distinct from each other. These features form

  12. Serum Autotaxin/ENPP2 Correlates with Insulin Resistance in Older Humans with Obesity

    OpenAIRE

    Reeves, Valerie L.; Trybula, Joy S.; Wills, Rachel C.; Goodpaster, Bret H.; Dubé, John J.; Kienesberger, Petra C; Kershaw, Erin E.

    2015-01-01

    Objective Autotaxin (ATX) is an adipocyte-derived lysophospholipase D that generates the lipid signaling molecule lysophosphatidic acid (LPA). The ATX/LPA pathway in adipose tissue has recently been implicated in obesity and insulin resistance in animal models, but the role of circulating ATX in humans remains unclear. The aim of the present study was to determine the relationship between serum ATX and insulin resistance. Methods In this retrospective study, older (60–75 years), non-diabetic ...

  13. Biosynthesis of phosphatidylcholine by human lysophosphatidylcholine acyltransferase 11

    OpenAIRE

    Harayama, Takeshi; Shindou, Hideo; Shimizu, Takao

    2009-01-01

    Pulmonary surfactant is a complex of phospholipids and proteins lining the alveolar walls of the lung. It reduces surface tension in the alveoli, and is critical for normal respiration. Pulmonary surfactant phospholipids consist mainly of phosphatidylcholine (PC) and phosphatidylglycerol (PG). Although the phospholipid composition of pulmonary surfactant is well known, the enzyme(s) involved in its biosynthesis have remained obscure. We previously reported the cloning of murine lysophosphatid...

  14. LPA receptor signaling: pharmacology, physiology, and pathophysiology

    OpenAIRE

    Yung, Yun C.; Stoddard, Nicole C.; Chun, Jerold

    2014-01-01

    Lysophosphatidic acid (LPA) is a small ubiquitous lipid found in vertebrate and nonvertebrate organisms that mediates diverse biological actions and demonstrates medicinal relevance. LPA’s functional roles are driven by extracellular signaling through at least six 7-transmembrane G protein-coupled receptors. These receptors are named LPA1–6 and signal through numerous effector pathways activated by heterotrimeric G proteins, including Gi/o, G12/13, Gq, and Gs. LPA receptor-mediated effects ha...

  15. Non-Invasive Imaging of Tumors by Monitoring Autotaxin Activity Using an Enzyme-Activated Near-Infrared Fluorogenic Substrate

    OpenAIRE

    Madan, Damian; Ferguson, Colin G.; Lee, Won Yong; Prestwich, Glenn D.; Testa, Charles A.

    2013-01-01

    Autotaxin (ATX), an autocrine motility factor that is highly upregulated in metastatic cancer, is a lysophospholipase D enzyme that produces the lipid second messenger lysophosphatidic acid (LPA) from lysophosphatidylcholine (LPC). Dysregulation of the lysolipid signaling pathway is central to the pathophysiology of numerous cancers, idiopathic pulmonary fibrosis, rheumatoid arthritis, and other inflammatory diseases. Consequently, the ATX/LPA pathway has emerged as an important source of bio...

  16. Novel point mutations attenuate autotaxin activity

    OpenAIRE

    Stracke Mary L; Roberts David D; Bandle Russell W; Koh Eunjin; Clair Timothy

    2009-01-01

    Abstract Background The secreted enzyme autotaxin (ATX) stimulates tumor cell migration, tumorigenesis, angiogenesis, and metastasis. ATX hydrolyzes nucleotides, but its hydrolysis of lysophospholipids to produce lysophosphatidic acid (LPA) accounts for its biological activities. ATX has been identified only as a constitutively active enzyme, and regulation of its activity is largely unexplored. In spite of its presence in plasma along with abundant putative substrate LPC, the product LPA is ...

  17. Autotaxin: Its Role in Biology of Melanoma Cells and as a Pharmacological Target

    Directory of Open Access Journals (Sweden)

    Maciej Jankowski

    2011-01-01

    Full Text Available Autotaxin (ATX is an extracellular lysophospholipase D (lysoPLD released from normal cells and cancer cells. Activity of ATX is detected in various biological fluids. The lysophosphatidic acid (LPA is the main product of ATX. LPA acting through specific G protein-coupled receptors (LPA1-LPA6 affects immunological response, normal development, and malignant tumors' formation and progression. In this review, the impact of autotoxin on biology of melanoma cells and potential treatment is discussed.

  18. The Effects of EPA, DHA, and Aspirin Ingestion on Plasma Lysophospholipids and Autotaxin

    OpenAIRE

    Block, RC; Duff, R; Lawrence, P.; Kakinami, L.; Brenna, JT; Shearer, GC; Meednu, N; Mousa, S; Friedman, A.; Harris, WS; Larson, Mark; Georas, S

    2010-01-01

    Lysophophatidylcholine (LPC) and lysophosphatidic acid (LPA) are potent lysolipid mediators increasingly linked with atherosclerosis and inflammation. A current model proposing that plasma LPA is produced when LPC is hydrolyzed by the enzyme autotaxin has not been rigorously investigated in human subjects. We conducted a clinical trial of eicosapentaenoic acid/docosahexaenoic acid (EPA/DHA) and aspirin ingestion in normal volunteers. Fasting blood samples were drawn at baseline and after 4-we...

  19. A role for 1-acylglycerol-3-phosphate-O-acyltransferase-1 in myoblast differentiation

    OpenAIRE

    Subauste, Angela R; Elliott, Brandon; Das, Arun K.; Burant, Charles F.

    2010-01-01

    AGPAT isoforms catalyze the acylation of lysophosphatidic acid (LPA) to form phosphatidic acid (PA). AGPAT2 mutations are associated with defective adipogenesis. Muscle and adipose tissue share common precursor cells. We investigated the role of AGPAT isoforms in skeletal muscle development. We demonstrate that small interference RNA-mediated knockdown of AGPAT1 expression prevents the induction of myogenin, a key transcriptional activator of the myogenic program, and inhibits the expression ...

  20. A dependência pela prática de exercícios físicos e o uso de recursos ergogênicos = Physical exercise dependence and the use of ergogenic resources

    Directory of Open Access Journals (Sweden)

    José Luiz Lopes Vieira

    2010-01-01

    Full Text Available Objetivou-se Investigar a ocorrência de dependência por exercícios físicos quanto às características de praticantes de musculação e ginástica em academias, como uso de recursos ergogênicos, sexo e índice de massa corporal. Participaram do estudo 80 sujeitos (27,12 ± 6,60 anos, praticantes de ginástica e/ou musculação em academias, de ambos os sexos. Utilizou-se a Escala de Dependência por Exercícios Físicos, a listagem do tipo de suplemento alimentar utilizado como recurso ergogênico e o Índice de Massa Corporal – IMC (Kg/cm². A análise estatística foi realizada por meio da correlação de Spearman e o teste de Wilcoxon (p The aim of this study is to investigate the occurrence of physical exercise dependence in regards to the characteristics of participants in weight training and exercises at gyms, such as the use ofergogenic resources, gender and body mass index. Eighty subjects (27.12 ± 6.60 years from both genders took part in the study, all of whom practiced gymnastics and/or weight training in gyms. The study utilized the Exercise Dependence Scale, a check list of the kinds of nutritional supplementation used as ergogenic resources, and the Body Mass Index – BMI (Kg cm-². Statistical analysis was performed using Spearman's correlation and the Wilcoxon test (p < 0.05. The results showed that there was no statistically significant difference between physical exercise dependence in men (5.14 ± 1.28 and women (5.60 ± 1.45. The body mass index did not show statistically significant correlation with the scores of dependency, either. However, dependentmen showed high prevalence of use of ergogenic resources (63.63%, p = 0.01, while for dependent women there were no statistically significant results. The body mass index does not relate to the scores of exercise dependence. However, even with a normal BMI, the use of ergogenic resources presents high prevalence among dependent men. As a result, there is evidence that physical exercise dependence is a risk factor for the development of emotional disturbances related to exercise, such as muscle dysmorphia and overtraining.

  1. Measurements of observables in the pion-nucleon system, nuclear a- dependence of heavy quark production and rare decays of D and B mesons. Progress report, 1 December, 1990--15 February, 1992

    Energy Technology Data Exchange (ETDEWEB)

    Sadler, M.E.; Isenhower, L.D.

    1992-02-15

    This report discusses research on the following topics: pion-nucleon interactions; detector tomography facility; nuclear dependence of charm and beauty quark production and a study of two-prong decays of neutral D and B mesons; N* collaboration at CEBAF; and pilac experiments. (LSP)

  2. Gli2a protein localization reveals a role for Iguana/DZIP1 in primary ciliogenesis and a dependence of Hedgehog signal transduction on primary cilia in the zebrafish

    Directory of Open Access Journals (Sweden)

    van Eeden Freek

    2010-04-01

    Full Text Available Abstract Background In mammalian cells, the integrity of the primary cilium is critical for proper regulation of the Hedgehog (Hh signal transduction pathway. Whether or not this dependence on the primary cilium is a universal feature of vertebrate Hedgehog signalling has remained contentious due, in part, to the apparent divergence of the intracellular transduction pathway between mammals and teleost fish. Results Here, using a functional Gli2-GFP fusion protein, we show that, as in mammals, the Gli2 transcription factor localizes to the primary cilia of cells in the zebrafish embryo and that this localization is modulated by the activity of the Hh pathway. Moreover, we show that the Igu/DZIP1protein, previously implicated in the modulation of Gli activity in zebrafish, also localizes to the primary cilium and is required for its proper formation. Conclusion Our findings demonstrate a conserved role of the primary cilium in mediating Hedgehog signalling activity across the vertebrate phylum and validate the use of the zebrafish as a representative model for the in vivo analysis of vertebrate Hedgehog signalling.

  3. China y Argentina: "Oportunidades y desafíos", o cristalización de una asociación dependiente | China and Argentina: opportunities and challenges or the result of a dependent association

    Directory of Open Access Journals (Sweden)

    Pablo Alejandro NACHT

    2012-06-01

    Full Text Available El presente artículo tiene como objetivo central realizar un análisis del proceso de vinculación y articulación de los diferentes actores argentinos con la República Popular China. Se busca entender el rol de las fracciones del bloque en el poder (burguesía rural, burguesía industrial nacional relevantes en la vinculación con China en el período planteado. El trabajo abarca desde 2002 hasta 2007 y da cuenta del cambio del régimen de acumulación capitalista, evidenciando una nueva configuración de fuerzas dentro del bloque y que encuentra, en la relación argentina con el gigante asiático, un socio destacado. Los sectores agroexportadores y las petroleras se erigen como las más beneficiadas en esta vinculación, al tiempo que la burguesía mercado internista menos competitiva presiona para frenar la entrada de manufacturas chinas. El estado argentino, lejos de ser un actor neutral, recibe importantes recursos por lo recaudado como derechos de exportación y es utilizado por el Gobierno; plasmándose de esta manera un escenario de tensiones y pujas entre los sectores, al tiempo que se cristaliza un modelo neo-extractivista con el monocultivo de soja transgénica como commoditie de exportación.This research article focus on the process of vinculation and coordination between different argentinian actors and China. It seeks to understand the role of the fractions of the power bloc (rural bourgeoisie, national industrial bourgeoisie, etc. relevant to the relationship with China in the period stated. The work ranges from 2002 to 2007 and shows the capitalist accumulation regime´s change, showing a new configuration of forces within the bloc and the relationship with the asian giant as leading partner. The agro-exporters and oil sectors stand as the main beneficiaries in this relationship, while the national industrial bourgeoisie pressed to stem the flow of Chinese products. The argentine state far from being a neutral actor, receives substantial resources from the proceeds as export duties and is used by the Government, shaping itself in this way scenario tensions and struggles between sectors, crystallizes as a neo extractive model with transgenic soybean monoculture and export commodities.

  4. China y Argentina: "Oportunidades y desafíos", o cristalización de una asociación dependiente | China and Argentina: opportunities and challenges or the result of a dependent association

    OpenAIRE

    Pablo Alejandro NACHT

    2012-01-01

    El presente artículo tiene como objetivo central realizar un análisis del proceso de vinculación y articulación de los diferentes actores argentinos con la República Popular China. Se busca entender el rol de las fracciones del bloque en el poder (burguesía rural, burguesía industrial nacional) relevantes en la vinculación con China en el período planteado. El trabajo abarca desde 2002 hasta 2007 y da cuenta del cambio del régimen de acumulación capitalista, evidenciando una nueva configuraci...

  5. A dependência pela prática de exercícios físicos e o uso de recursos ergogênicos = Physical exercise dependence and the use of ergogenic resources

    OpenAIRE

    José Luiz Lopes Vieira; Priscila Garcia Marques da Rocha; Ricardo Aparecido Ferrarezzi

    2010-01-01

    Objetivou-se Investigar a ocorrência de dependência por exercícios físicos quanto às características de praticantes de musculação e ginástica em academias, como uso de recursos ergogênicos, sexo e índice de massa corporal. Participaram do estudo 80 sujeitos (27,12 ± 6,60 anos), praticantes de ginástica e/ou musculação em academias, de ambos os sexos. Utilizou-se a Escala de Dependência por Exercícios Físicos, a listagem do tipo de suplemento alimentar utilizado como recurso ergogênico e o Índ...

  6. The T Allele of the ACAT-2 734 C/T Polymorphism is Associated with the Changes of TG/HDLC, log (TG/HDLC) and LDLC/HDLC Induced by High-carbohydrate/ Low-Fat Diet in Healthy Young Women%酰基辅酶A:胆固醇酰基转移酶2基因734位点T等位基因与高糖低脂膳食诱导的健康青年女性TG/HDLC、log( TG/HDLC)及LDLC/HDLC改变相关联

    Institute of Scientific and Technical Information of China (English)

    姜喆; 龚仁蓉; 李元昊; 樊梅; 方定志

    2012-01-01

    Aim To investigate the role of the acyl-coenzyme A-' cholesterol acyltransferase-2 (ACAT-2) 734 C/T polymorphism on ratios of serum lipids and its interactions on the ratios with a high-carbohydrate/low-fat ( HC/LF) diet in a young healthy Chinese population. Methods After a washout diet for seven days, fifty six young healthy subjects were given the HC/LF diet for six days. The serum lipid profiles were analyzed using the twelve-hour fasting venous blood samples collected in the mornings of the first, the eighth and the fourteenth days. The ratios of TG/HDLC, log (TG/HDLC) , TC/HDLC and LDLC/HDLC were calculated. The ACAT-2 734 C/T polymorphism was analyzed by restriction fragments patterns after amplified by polymerase chain reactions. Results At baseline, the female carriers of the T allele had significantly lower log (TG/HDLC) than the female subjects with the CC genotype. The male subjects%目的 探讨酰基辅酶A:胆固醇酰基转移酶2(ACAT-2)基因734位点C/T多态性对健康青年血脂比值的影响及在高糖低脂膳食诱导的血脂比值变化中的作用.方法 给予56名健康青年志愿者7天平衡膳食和6天高糖低脂膳食,于第1天、第8天以及第14天清晨抽取12 h空腹静脉血,测定血脂,计算甘油三酯/高密度脂蛋白胆固醇(TG/HDLC)、log(TrG/HDLC)、总胆固醇/高密度脂蛋白胆固醇(TC/HDLC)和低密度脂蛋白胆固醇/高密度脂蛋白胆固醇(LDLC/HDLC)比值.提取基因组DNA,聚合酶链反应限制性酶切法分析ACAT-2基因734位点多态性.结果 女性T等位基因携带者log( TG/HDLC)显著低于CC纯合子受试者,男性CC纯合子受试者TC/HDLC显著高于女性CC纯合子受试者,男性T等位基因携带者log(TG/HDLC)显著高于女性.高糖低脂膳食前,男性T等位基因携带者TG/HDLC、log(TG/HDLC)显著高于女性.与高糖低脂膳食前相比,高糖低脂膳食后男性TC/HDLC、LDLC/HDLC显著降低,不受基因型影响.女性CC纯合子受试者

  7. Action and Signaling of Lysophosphatidylethanolamine in MDA-MB-231 Breast Cancer Cells

    OpenAIRE

    Park, Soo-Jin; Lee, Kyoung-Pil; Im, Dong-Soon

    2014-01-01

    Previously, we reported that lysophosphatidylethanolamine (LPE), a lyso-type metabolite of phosphatidylethanolamine, can increase intracellular Ca2+ ([Ca2+]i) via type 1 lysophosphatidic acid (LPA) receptor (LPA1) and CD97, an adhesion G-protein-coupled receptor (GPCR), in MDA-MB-231 breast cancer cells. Furthermore, LPE signaling was suggested as like LPA1/CD97-Gi/o proteins-phospholipase C-IP3-Ca2+ increase in these cells. In the present study, we further investigated actions of LPE not onl...

  8. Autotaxin: A protein with two faces

    International Nuclear Information System (INIS)

    Research highlights: → Autotaxin (ATX) has lysophospholipase D activity. → ATX catalyzes the formation of lysophosphatidic acid (LPA). → LPA is a mitogen, and thus is responsible for cancer. → ATX also catalyzes the formation of anti-cancerous cyclic phosphatidic acid. → Autotaxin is a novel target of cancer therapy research. -- Abstract: Autotaxin (ATX) is a catalytic protein, which possesses lysophospholipase D activity, and thus involved in cellular membrane lipid metabolism and remodeling. Primarily, ATX was thought as a culprit protein for cancer, which potently stimulates cancer cell proliferation and tumor cell motility, augments the tumorigenicity and induces angiogenic responses. The product of ATX catalyzed reaction, lysophosphatidic acid (LPA) is a potent mitogen, which facilitates cell proliferation and migration, neurite retraction, platelet aggregation, smooth muscle contraction, actin stress formation and cytokine and chemokine secretion. In addition to LPA formation, later ATX has been found to catalyze the formation of cyclic phosphatidic acid (cPA), which have antitumor role by antimitogenic regulation of cell cycle, inhibition of cancer invasion and metastasis. Furthermore, the very attractive information to the scientists is that the LPA/cPA formation can be altered at different physiological conditions. Thus the dual role of ATX with the scope of product manipulation has made ATX a novel target for cancer treatment.

  9. Autotaxin: A protein with two faces

    Energy Technology Data Exchange (ETDEWEB)

    Tania, Mousumi; Khan, Md. Asaduzzaman; Zhang, Huaiyuan; Li, Jinhua [Department of Biochemistry, School of Biological Science and Technology, Central South University, Changsha, Hunan 410013 (China); Song, Yuanda, E-mail: yuanda_song@hotmail.com [Department of Biochemistry, School of Biological Science and Technology, Central South University, Changsha, Hunan 410013 (China)

    2010-10-29

    Research highlights: {yields} Autotaxin (ATX) has lysophospholipase D activity. {yields} ATX catalyzes the formation of lysophosphatidic acid (LPA). {yields} LPA is a mitogen, and thus is responsible for cancer. {yields} ATX also catalyzes the formation of anti-cancerous cyclic phosphatidic acid. {yields} Autotaxin is a novel target of cancer therapy research. -- Abstract: Autotaxin (ATX) is a catalytic protein, which possesses lysophospholipase D activity, and thus involved in cellular membrane lipid metabolism and remodeling. Primarily, ATX was thought as a culprit protein for cancer, which potently stimulates cancer cell proliferation and tumor cell motility, augments the tumorigenicity and induces angiogenic responses. The product of ATX catalyzed reaction, lysophosphatidic acid (LPA) is a potent mitogen, which facilitates cell proliferation and migration, neurite retraction, platelet aggregation, smooth muscle contraction, actin stress formation and cytokine and chemokine secretion. In addition to LPA formation, later ATX has been found to catalyze the formation of cyclic phosphatidic acid (cPA), which have antitumor role by antimitogenic regulation of cell cycle, inhibition of cancer invasion and metastasis. Furthermore, the very attractive information to the scientists is that the LPA/cPA formation can be altered at different physiological conditions. Thus the dual role of ATX with the scope of product manipulation has made ATX a novel target for cancer treatment.

  10. A dependência pela prática de exercícios físicos e o uso de recursos ergogênicos - DOI: 10.4025/actascihealthsci.v32i1.4475 Physical exercise dependence and the use of ergogenic resources - DOI: 10.4025/actascihealthsci.v32i1.4475

    Directory of Open Access Journals (Sweden)

    Ricardo Aparecido Ferrarezi

    2009-12-01

    Full Text Available Objetivou-se Investigar a ocorrência de dependência por exercícios físicos quanto às características de praticantes de musculação e ginástica em academias, como uso de recursos ergogênicos, sexo e índice de massa corporal. Participaram do estudo 80 sujeitos (27,12 ± 6,60 anos, praticantes de ginástica e/ou musculação em academias, de ambos os sexos. Utilizou-se a Escala de Dependência por Exercícios Físicos, a listagem do tipo de suplemento alimentar utilizado como recurso ergogênico e o Índice de Massa Corporal – IMC (Kg/cm². A análise estatística foi realizada por meio da correlação de Spearman e o teste de Wilcoxon (p The aim of this study is to investigate the occurrence of physical exercise dependence in regards to the characteristics of participants in weight training and exercises at gyms, such as the use of ergogenic resources, gender and body mass index. Eighty subjects (27.12 ± 6.60 years from both genders took part in the study, all of whom practiced gymnastics and/or weight training in gyms. The study utilized the Exercise Dependence Scale, a check list of the kinds of nutritional supplementation used as ergogenic resources, and the Body Mass Index – BMI (Kg cm-². Statistical analysis was performed using Spearman's correlation and the Wilcoxon test (p < 0.05. The results showed that there was no statistically significant difference between physical exercise dependence in men (5.14 ± 1.28 and women (5.60 ± 1.45. The body mass index did not show statistically significant correlation with the scores of dependency, either. However, dependent men showed high prevalence of use of ergogenic resources (63.63%, p = 0.01, while for dependent women there were no statistically significant results. The body mass index does not relate to the scores of exercise dependence. However, even with a normal BMI, the use of ergogenic resources presents high prevalence among dependent men. As a result, there is evidence that physical exercise dependence is a risk factor for the development of emotional disturbances related to exercise, such as muscle dysmorphia and overtraining.

  11. Regulation of insulin-like growth factor I transcription by cyclic adenosine 3',5'-monophosphate (cAMP) in fetal rat bone cells through an element within exon 1: protein kinase A-dependent control without a consensus AMP response element

    Science.gov (United States)

    McCarthy, T. L.; Thomas, M. J.; Centrella, M.; Rotwein, P.

    1995-01-01

    Insulin-like growth factor I (IGF-I) is a locally synthesized anabolic growth factor for bone. IGF-I synthesis by primary fetal rat osteoblasts (Ob) is stimulated by agents that increase the intracellular cAMP concentration, including prostaglandin E2 (PGE2). Previous studies with Ob cultures demonstrated that PGE2 enhanced IGF-I transcription through selective use of IGF-I promoter 1, with little effect on IGF-I messenger RNA half-life. Transient transfection of Ob cultures with an array of promoter 1-luciferase reporter fusion constructs has now allowed localization of a potential cis-acting promoter element(s) responsible for cAMP-stimulated gene expression to the 5'-untranslated region (5'-UTR) of IGF-I exon 1, within a segment lacking a consensus cAMP response element. Our evidence derives from three principal observations: 1) a transfection construct containing only 122 nucleotides (nt) of promoter 1 and 328 nt of the 5'-UTR retained full PGE2-stimulated reporter expression; 2) maximal PGE2-driven reporter expression required the presence of nt 196 to 328 of exon 1 when tested within the context of IGF-I promoter 1; 3) cotransfection of IGF-I promoter-luciferase-reporter constructs with a plasmid encoding the alpha-isoform of the catalytic subunit of murine cAMP-dependent protein kinase (PKA) produced results comparable to those seen with PGE2 treatment, whereas cotransfection with a plasmid encoding a mutant regulatory subunit of PKA that cannot bind cAMP blocked PGE2-induced reporter expression. Deoxyribonuclease I footprinting of the 5'-UTR of exon 1 demonstrated protected sequences at HS3A, HS3B, and HS3D, three of six DNA-protein binding sites previously characterized with rat liver nuclear extracts. Of these three regions, only the HS3D binding site is located within the functionally identified hormonally responsive segment of IGF-I exon 1. These results directly implicate PKA in the control of IGF-I gene transcription by PGE2 and identify a segment of IGF-I exon 1 as being essential for this hormonal regulation.

  12. O estresse físico e a dependência de exercício físico El estrés físico y la dependencia de ejercicio físico Physical stress and physical exercise dependence

    Directory of Open Access Journals (Sweden)

    Hanna K.M. Antunes

    2006-10-01

    Full Text Available O objetivo do presente estudo foi verificar os escores referentes à escala de dependência de exercício, qualidade de vida, bem como os escores indicativos de humor em atletas de corrida de aventura (CA. Participaram deste estudo 17 atletas de ambos os gêneros com histórico de prática da modalidade de pelo menos três anos, com experiência em provas nacionais e internacionais e que figuram nas primeiras posições do ranking brasileiro. A média (± desvio-padrão da idade, altura, massa corporal, índice de massa corpórea (IMC e consumo de oxigênio foram: 31,11 ± 6,30 anos; 1,73 ± 0,07cm; 70,75 ± 7,96kg; 23,48 ± 1,48kg/m² e 58,70 ± 6,63ml.min¹.kg¹, respectivamente. Os voluntários responderam aos seguintes questionários: Escala de Dependência de Exercício (EDE, Idate Traço e Estado, Profile of Mood States (POMS, SF-36 Pesquisa em Saúde e Questionário de Padrão Social. Os resultados revelaram que os escores observados na EDE foram indicativos de dependência de exercício; já os questionários de humor revelaram ansiedade moderada, enquanto o POMS não detectou escores indicativos de distúrbios de humor. Quanto à qualidade de vida, a média das oito dimensões referentes ao questionário SF-36 se mostrou acima de 85%, sugerindo que, apesar de haver dependência de exercício, parece que esse fato não foi capaz de promover alterações significativas no estado de humor e na qualidade de vida. Esses dados sugerem que atletas de CA apresentam dependência de exercício não associada aos distúrbios de humor.El objetivo del presente estudio ha sido el de verificar los escores referentes a la escala de dependencia de ejercicio, calidad de vida, así como los escores indicativos de humor en atletas de Carrera de Aventura (CA. Participaron de este estudio 17 atletas de ambos géneros con histórico de práctica de la modalidad de por lo menos tres años, con experiencia en pruebas nacionales e internacionales y que figuran en las primeras posiciones del ranking brasileño. La media (± desvío padrón de edad, altura, masa corporal, índice de masa corpórea (IMC y consumo de oxígeno fueron: 31,11 ± 6,30 años; 1,73 ± 0,07 cm; 70,75 ± 7,96 kg; 23,48 ± 1,48 kg/m² y 58,70 ± 6,63 ml.min-1.kg-1, respectivamente. Los voluntarios respondieron a los siguientes cuestionarios: Escala de Dependencia de Ejercicio (EDE, "Idate Traço" y Estado, Profile of Mood States (POMS, SF-36 Investigación e Salud y Cuestionario de Padrón Social. Los resultados revelaron que los escores observados en la EDE fueron indicativos de dependencia de ejercicio, mientras que los cuestionarios de humor revelaron ansiedad moderada, entretanto el POMS no detectó escores indicativos de disturbios de humor. En relación a la calidad de vida, la media de las 8 dimensiones referentes al cuestionario SF-36 se mostró por encima de 85%, lo que sugiere que a pesar de haber dependencia de ejercicio, parece que ese hecho no fue capaz de promover alteraciones significativas en el estado de humor y en la calidad de vida. Nuestros datos sugieren que atletas de CA presentan dependencia de ejercicio no asociada a los disturbios de humor.The aim of this study was to verify the referring scores of exercise dependence, quality of life as well as the mood indicators in adventure race (AR athletes. 17 athletes of both sexes participated in the study and all had a history of three years in this modality, with national and international experience, and figured in the first positions in the Brazilian ranking. The age, height, weight, body mass index (BMI and oxygen uptake averages ± standard deviations were: 31.11 ± 6.30 years; 1.73 ± 0.07 cm; 70.75 ± 7.96 kg; 23.48 ± 1.48 wt/ht² and 58.70 ± 6.63 ml.min-1.kg-1, respectively. The volunteers were given the following questionnaires: Exercise Dependency Scale (EDE, Idate Trait and State, Profile of Mood States (POMS, SF-36 Health Research and Social Patterns Questionnaire. The results showed that scores in EDE indicated exercise dependence, and the mood questionnaires revealed moderate anxiety, while the POMS did not detect any indicative scores of mood disorders. Concerning the quality of life, the average of 8 dimensions of the SF-36 was higher than 85%, suggesting that although there was exercise dependence, this fact alone did not promote significant alterations in mood and quality of life. Thus, our data suggested that athletes of AR have exercise dependence not associated to mood disorders.

  13. A dinâmica da área, do rendimento e dos preços sobre o valor da produção do feijão e da soja no Rio Grande do Sul e a dependência temporal entre esses componentes

    Directory of Open Access Journals (Sweden)

    Dienice Ana Bini

    2015-06-01

    Full Text Available Objetiva-se neste artigo avaliar o crescimento do valor da produção (VP do feijão e da soja no Rio Grande do Sul (RS, baseado no comportamento de seus determinantes área, rendimento e preço. Também se pretende determinar como choques nestes componentes transmitem-se de uma safra para outra no VP. Utilizaram-se dados de área, rendimento e preço do feijão preto e da soja no RS de 1977 até 2010. A metodologia shift-share permitiu decompor a importância de cada item sobre a variação do valor da produção. O ferramental de séries temporais permitiu obter o efeito de um choque em uma variável, ceteris paribus, sobre as demais. No curto prazo, o rendimento é o principal responsável pela variação do VP. Entretanto, no longo prazo, a queda dos preços reais de ambos os produtos é o fator determinante na redução dos seus valores da produção. O valor da produção reage positivamente a choques positivos na área, rendimento e preço no ano corrente ao choque. Em geral, os ajustes são mais dinâmicos na soja do que no feijão

  14. A dependência pela prática de exercícios físicos e o uso de recursos ergogênicos - DOI: 10.4025/actascihealthsci.v32i1.4475 Physical exercise dependence and the use of ergogenic resources - DOI: 10.4025/actascihealthsci.v32i1.4475

    OpenAIRE

    Ricardo Aparecido Ferrarezi; Priscila Garcia Marques Rocha; José Luiz Lopes Vieira

    2009-01-01

    Objetivou-se Investigar a ocorrência de dependência por exercícios físicos quanto às características de praticantes de musculação e ginástica em academias, como uso de recursos ergogênicos, sexo e índice de massa corporal. Participaram do estudo 80 sujeitos (27,12 ± 6,60 anos), praticantes de ginástica e/ou musculação em academias, de ambos os sexos. Utilizou-se a Escala de Dependência por Exercícios Físicos, a listagem do tipo de suplemento alimentar utilizado como recurso ergogênico e o Índ...

  15. Optimal Investment Strategy for an Insurer under Mean-Variance in a Dependent Risk Model%保险公司在风险相依模型中均值-方差准则下的最优投资策略*

    Institute of Scientific and Technical Information of China (English)

    谷爱玲; 李仲飞; 申曙光

    2013-01-01

    研究了具有两个业务部门的保险公司的最优投资问题,其中每个业务部门的盈余过程由二维的Lévy过程描述。保险公司可将其盈余投资于金融市场,其中金融市场由一个无风险资产和两个具有风险相关性的风险资产组成,而且风险资产的价格过程由二维的Lévy过程所驱动。文中讨论了两个优化问题。一个是基准问题,即选择适当的投资策略使保险公司的终端财富与一个基准值之差的平方期望最小;另一个是均值-方差(M-V)问题,即在保险公司终端财富给定的情形下,选择适当的投资策略使终端财富的方差最小。利用动态规划的方法,得到第一个优化问题的最优投资策略和最优值函数的解析式。结合第一个优化问题的结果,利用对偶定理得到第二个优化问题的最优投资策略和有效前沿。%Two optimal investment problems for an insurer with two business lines are considered,where each business line's risk process is modeled by two-dimensional Lévy process.It is assumed that the in-surer can invest its surplus in a risk-free asset and two risky assets,where the risky assets'price processes are described by a two-dimensional Lévy process.A benchmark problem and a mean-variance problem are discussed.The first problem is to choose the optimal investment strategy to minimize the expected quadratic distance of the risk reserve to a given benchmark;the second problem is to minimize the vari-ance of the terminal wealth when the expected terminal reserve is given.By employing stochastic dynamic programming approach,the explicit expressions of the optimal investment strategy and the optimal value function are derived for the first problem;with the results of the first problem and the duality theory,the optimal investment strategy and the efficient frontier for the second problem are derived.

  16. Clinical study on lysophospholipids acid, D_dimers detection in the prediction of progress and application effect of cerebral apoplexy%溶血磷脂酸、D-二聚体检测预测进展性脑卒中的临床应用研究

    Institute of Scientific and Technical Information of China (English)

    胡红晓

    2014-01-01

    Objective:To study the lysophospholipids acid , D_dimers detection in the prediction of progress and application effect of cerebral apoplexy .Methods:Our hospital in April 2012 to April 2013 were 35 patients with progressive stroke ( research group ) , physical examination, 35 cases (control group) as the research object, research on two groups of objects of D -dimer, lysophosphatide acid level testing and comparison .Results:The group D -dimer level of patients >75 mu g/L, positive;Control the research object of D -dimer level <75 mu g/L, negative;Group patients were lysophospholipids , D -dimer acid average levels were significantly higher than that of control group, P <0.05).Conclusion:D -dimer, lysophosphatide acid was an important molecular markers of early thrombosis in vivo, by detecting D -dimer, lysophosphatide acid levels , could provide effective basis for clinical prediction progress sex stroke .%目的:探讨溶血磷脂酸、D-二聚体检测在预测进展性脑卒中的应用效果。方法:选取我院在2012年4月~2013年4月收治的进展性脑卒中患者35例(研究组)、健康体检者35例(对照组)作为研究对象,对2组研究对象的D-二聚体、溶血磷脂酸水平进行检测、比较。结果:研究组患者的D-二聚体水平均>75μg/L,呈阳性;对照组研究对象的D-二聚体水平均<75μg/L,呈阴性;研究组患者的溶血磷脂酸、D-二聚体平均水平均显著高于对照组,P<0.05。结论:D-二聚体、溶血磷脂酸是体内早期血栓形成的重要分子标志物,通过检测D-二聚体、溶血磷脂酸水平,可为临床预测进展性脑卒中提供有效依据。

  17. Differential phospholipid-labeling suggests two subtypes of phospholipase D in rat Leydig cells

    DEFF Research Database (Denmark)

    Lauritzen, L.; Hansen, Harald S.

    1995-01-01

    The aim of the present study was to compare the transphosphatidylation activity of phospholipase D (PLD) under different substrate labeling conditions, in order to investigate whether PLD in rat Leydig cells exhibited any substrate preferences for the alkyl- or acyl-form of phosphatidylcholine (Ptd......Cho). The [H] phosphatidylethanol formation in response to 4ß-phorbol 12-myristate 13-acetate (PMA), sphingosine, or Ca-ionophore A23187, was lower when Leydig cells were labeled with 1-O-[H]alkyl lysoPtdCho compared with the responses when [H]myristic acid was employed. In contrast, the results...... for the receptor agonists (vasopressin, bradykinin, and lysophosphatidic acid), using the two labels, showed mole consistency. Thus, the PLD-activity induced by PMA, sphingosine, or A23187 has a more selective substrate range (i.e. mainly acyl-linked PtdCho) than the PLD-activity stimulated via a receptor. Our...

  18. Non-cell autonomous and non-catalytic activities of ATX in the developing brain

    Directory of Open Access Journals (Sweden)

    Raanan eGreenman

    2015-03-01

    Full Text Available The intricate formation of the cerebral cortex requires a well-coordinated series of events, which are regulated at the level of cell-autonomous and non-cell autonomous mechanisms. Whereas cell-autonomous mechanisms that regulate cortical development are well-studied, the non cell-autonomous mechanisms remain poorly understood. A non-biased screen allowed us to identify Autotaxin (ATX as a non cell-autonomous regulator of neural stem cell proliferation. ATX (also known as ENPP2 is best known to catalyze lysophosphatidic acid (LPA production. Our results demonstrate that ATX affects the localization and adhesion of neuronal progenitors in a cell autonomous and non-cell autonomous manner, and strikingly, this activity is independent from its catalytic activity in producing LPA.

  19. Structure-Driven Pharmacology of Transient Receptor Potential Channel Vanilloid 1.

    Science.gov (United States)

    Díaz-Franulic, Ignacio; Caceres-Molina, Javier; Sepulveda, Romina V; Gonzalez-Nilo, Fernando; Latorre, Ramon

    2016-09-01

    The transient receptor potential vanilloid 1 (TRPV1) ion channel is a polymodal receptor that mediates the flux of cations across the membrane in response to several stimuli, including heat, voltage, and ligands. The best known agonist of TRPV1 channels is capsaicin, the pungent component of "hot" chili peppers. In addition, peptides found in the venom of poisonous animals, along with the lipids phosphatidylinositol 4,5-biphosphate, lysophosphatidic acid, and cholesterol, bind to TRPV1 with high affinity to modulate channel gating. Here, we discuss the functional evidence regarding ligand-dependent activation of TRPV1 channels in light of structural data recently obtained by cryoelectron microscopy. This review focuses on the mechanistic insights into ligand binding and allosteric gating of TRPV1 channels and the relevance of accurate polymodal receptor biophysical characterization for drug design in novel pain therapies. PMID:27335334

  20. DIFFERENTIAL EXPRESSION OF GENES INVOLVED IN METABOLISM BETWEEN TUMORIGENITIC HUMAN LEUKEMIA CELL LINES K562 AND K562-n

    Institute of Scientific and Technical Information of China (English)

    吕书晴; 许小平; 夏放; 居小萍; 李瑶; 应康; 毛裕民

    2003-01-01

    Objective: To study the molecular mechanism of different tumorigenicity in nude mice of human leukemia cell lines K562-n and K562. Methods: To analyze the genes differently expressed between K562 and K562-n cells by using cDNA microarray technique. Results: Among the 12800 genes detected, some genes involved in material metabolism and material transport were differently expressed between K562-n and K562 cells. These genes include homo sapiens placenta-specific ATP-binding cassette transporter gene, dihydrodiol dehydrogenase gene, hepatic dihydrodiol dehydrogenase gene, NAD-dependent methylene tetrahydrofolate dehydrogenase cyclohydrolase, lysophosphatidic acid acyltransferase, alpha gene, argininosuccinate lyase gene, mitochondrial isocitrtate dehydrogenase, adhesion protein SQM1 gene, dimethylarginine dimethylamino-hydrolase gene, M1 subunit of ribonucleotide reductase and farnesyl pyrophosphate synthetase gene. Conclusion: The high tumorigenicity of K562-n cells is related to the different expression of some genes concerned with cell metabolism and material transpoert.

  1. Migratory properties of cultured olfactory ensheathing cells by single-cell migration assay

    Institute of Scientific and Technical Information of China (English)

    Zhi-hui Huang; Ying Wang; Li Cao; Zhi-da Su; Yan-ling Zhu; Yi-zhang Chen; Xiao-bing Yuan; Cheng He

    2008-01-01

    Olfactory ensheathing cells (OECs) are a unique type of glial cells that have axonal growth-promoting properties. OEC transplantation has emerged as a promising experimental therapy of axonal injuries and demyelinating diseases. However, some fundamental cellular properties of OECs remain unclear. In this study, we found that the distinct OEC subpopulations exhibited different migratory properties based on time-lapse imaging of single isolated cells, possibly due to their different cytoskeletal organizations. Moreover, OEC subpopulations displayed different attractive migratory responses to a gradient of lysophosphatidic acid (LPA) in single-cell migration assays. Finally, we found that OEC subpopulations transformed into each other spontaneously. Together, these results demonstrate, for the first time to our knowledge, that distinct OEC subpopulations display different migratory properties in vitro and provide new evidence to support the notion of OECs as a single cell type with malleable functional phenotypes.

  2. PPARγ Networks in Cell Signaling: Update and Impact of Cyclic Phosphatidic Acid

    Directory of Open Access Journals (Sweden)

    Tamotsu Tsukahara

    2013-01-01

    Full Text Available Lysophospholipid (LPL has long been recognized as a membrane phospholipid metabolite. Recently, however, the LPL has emerged as a candidate for diagnostic and pharmacological interest. LPLs include lysophosphatidic acid (LPA, alkyl glycerol phosphate (AGP, cyclic phosphatidic acid (cPA, and sphingosine-1-phosphate (S1P. These biologically active lipid mediators serve to promote a variety of responses that include cell proliferation, migration, and survival. These LPL-related responses are mediated by cell surface G-protein-coupled receptors and also intracellular receptor peroxisome proliferator-activated receptor gamma (PPARγ. In this paper, we focus mainly on the most recent findings regarding the biological function of nuclear receptor-mediated lysophospholipid signaling in mammalian systems, specifically as they relate to health and diseases. Also, we will briefly review the biology of PPARγ and then provide an update of lysophospholipids PPARγ ligands that are under investigation as a therapeutic compound and which are targets of PPARγ relevant to diseases.

  3. Analysis of the ligand binding properties of recombinant bovine liver-type fatty acid binding protein

    DEFF Research Database (Denmark)

    Rolf, B; Oudenampsen-Krüger, E; Börchers, T;

    1995-01-01

    The coding part of the cDNA for bovine liver-type fatty acid binding protein (L-FABP) has been amplified by RT-PCR, cloned and used for the construction of an Escherichia coli (E. coli) expression system. The recombinant protein made up to 25% of the soluble E. coli proteins and could be isolated...... by a simple two step protocol combining ion exchange chromatography and gel filtration. Dissociation constants for binding of oleic acid, arachidonic acid, oleoyl-CoA, lysophosphatidic acid and the peroxisomal proliferator bezafibrate to L-FABP have been determined by titration calorimetry. All ligands were...... bound in a 2:1 stoichiometry, the dissociation constants for the first ligand bound were all in the micro molar range. Oleic acid was bound with the highest affinity and a Kd of 0.26 microM. Furthermore, binding of cholesterol to L-FABP was investigated with the Lipidex assay, a liposome binding assay...

  4. Structure-Driven Pharmacology of Transient Receptor Potential Channel Vanilloid 1.

    Science.gov (United States)

    Díaz-Franulic, Ignacio; Caceres-Molina, Javier; Sepulveda, Romina V; Gonzalez-Nilo, Fernando; Latorre, Ramon

    2016-09-01

    The transient receptor potential vanilloid 1 (TRPV1) ion channel is a polymodal receptor that mediates the flux of cations across the membrane in response to several stimuli, including heat, voltage, and ligands. The best known agonist of TRPV1 channels is capsaicin, the pungent component of "hot" chili peppers. In addition, peptides found in the venom of poisonous animals, along with the lipids phosphatidylinositol 4,5-biphosphate, lysophosphatidic acid, and cholesterol, bind to TRPV1 with high affinity to modulate channel gating. Here, we discuss the functional evidence regarding ligand-dependent activation of TRPV1 channels in light of structural data recently obtained by cryoelectron microscopy. This review focuses on the mechanistic insights into ligand binding and allosteric gating of TRPV1 channels and the relevance of accurate polymodal receptor biophysical characterization for drug design in novel pain therapies.

  5. Antiphospholipid Antibody and Antiphospholipid Syndrome

    Institute of Scientific and Technical Information of China (English)

    吴竞生

    2008-01-01

    @@ Antiphospholipid antibodies (APA) APA is a big category for all kinds of negative charge phospholipid or lecithin - a protein complex autoantibodies or the same antibody, through its recognition of antigen (target protein) different, and phospholipids or lecithin - protein complex combination of various rely on the interference Phospholipid clotting and anti-coagulation factor, and promote endothelial cells, platelets, complement activation and play a role. APA including lupus anticoagulant(LA) and anticardiolipin antibody (ACA), In addition, there are anti-β2 glycoprotein-I (β2-GPI) antibody, anti-prothrombin (a- PT) antibody, anti-lysophosphatidic acid antibody and anti-phosphatidylserine antibody, and so on. APA as the main target of phospholipid-binding protein, including β2-GPI, prothrombin, annexin, protein C (PC) and protein S (PS), plasminogen, and so on.

  6. Alterations of red cell membrane properties in neuroacanthocytosis.

    Directory of Open Access Journals (Sweden)

    Claudia Siegl

    Full Text Available Neuroacanthocytosis (NA refers to a group of heterogenous, rare genetic disorders, namely chorea acanthocytosis (ChAc, McLeod syndrome (MLS, Huntington's disease-like 2 (HDL2 and pantothenate kinase associated neurodegeneration (PKAN, that mainly affect the basal ganglia and are associated with similar neurological symptoms. PKAN is also assigned to a group of rare neurodegenerative diseases, known as NBIA (neurodegeneration with brain iron accumulation, associated with iron accumulation in the basal ganglia and progressive movement disorder. Acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions, is frequently observed in ChAc and MLS patients but less prevalent in PKAN (about 10% and HDL2 patients. The pathological factors that lead to the formation of the acanthocytic red blood cell shape are currently unknown. The aim of this study was to determine whether NA/NBIA acanthocytes differ in their functionality from normal erythrocytes. Several flow-cytometry-based assays were applied to test the physiological responses of the plasma membrane, namely drug-induced endocytosis, phosphatidylserine exposure and calcium uptake upon treatment with lysophosphatidic acid. ChAc red cell samples clearly showed a reduced response in drug-induced endovesiculation, lysophosphatidic acid-induced phosphatidylserine exposure, and calcium uptake. Impaired responses were also observed in acanthocyte-positive NBIA (PKAN red cells but not in patient cells without shape abnormalities. These data suggest an "acanthocytic state" of the red cell where alterations in functional and interdependent membrane properties arise together with an acanthocytic cell shape. Further elucidation of the aberrant molecular mechanisms that cause this acanthocytic state may possibly help to evaluate the pathological pathways leading to neurodegeneration.

  7. The Peanut (Arachis hypogaea L. Gene AhLPAT2 Increases the Lipid Content of Transgenic Arabidopsis Seeds.

    Directory of Open Access Journals (Sweden)

    Silong Chen

    Full Text Available Lysophosphatidic acid acyltransferase (LPAT, which converts lysophosphatidic acid (LPA to phosphatidic acid (PA, catalyzes the addition of fatty acyl moieties to the sn-2 position of the LPA glycerol backbone in triacylglycerol (TAG biosynthesis. We recently reported the cloning and temporal-spatial expression of a peanut (Arachis hypogaea AhLPAT2gene, showing that an increase in AhLPAT2 transcript levels was closely correlated with an increase in seed oil levels. However, the function of the enzyme encoded by the AhLPAT2 gene remains unclear. Here, we report that AhLPAT2 transcript levels were consistently higher in the seeds of a high-oil cultivar than in those of a low-oil cultivar across different seed developmental stages. Seed-specific overexpression of AhLPAT2 in Arabidopsis results in a higher percentage of oil in the seeds and greater-than-average seed weight in the transgenic plants compared with the wild-type plants, leading to a significant increase in total oil yield per plant. The total fatty acid (FA content and the proportion of unsaturated FAs also increased. In the developing siliques of AhLPAT2-overexpressing plants, the expression levels of genes encoding crucial enzymes involved in de novo FA synthesis, acetyl-CoA subunit (AtBCCP2 and acyl carrier protein 1 (AtACP1 were elevated. AhLPAT2 overexpression also promoted the expression of several key genes related to TAG assembly, sucrose metabolism, and glycolysis. These results demonstrate that the expression of AhLPAT2 plays an important role in glycerolipid production in peanuts.

  8. Structure and function of sterol carrier proteins in insects%昆虫固醇转运蛋白的结构与功能

    Institute of Scientific and Technical Information of China (English)

    张丽丽; 郭兴荣; 冯启理; 郑思春

    2011-01-01

    , cholesterol derivatives, fatty acids,acyl-coenzyme A and phospholipids. Over-expression of SlSCP-x and SlSCP-2 genes can increase the uptake of cholesterol into cells and RNAi inhibits the expression of SlSCP-x and SlSCP-2 genes in S. litura larvae,resulting in a decrease in cholesterol level in the hemolymph and a delay in larval growth and pupation.%在昆虫中,胆固醇不仅是细胞膜的重要成分之一,也是昆虫蜕皮激素生物合成的前体.由于昆虫体内缺少两种合成胆固醇所必需的关键性酶,所以昆虫不能自主地从简单的前体化合物从头合成胆同醇,而必须通过吸收食物中的甾醇转化为胆固醇来满足生长、发育和繁殖的需要.胆固醇在组织和细胞内的运输主要由固醇转运蛋白(sterol carrier proteins,SCPs)执行.因此,对同醇转运蛋白结构与功能的研究对于阐明昆虫中固醇运输具有重要的意义.本文对同醇转运蛋白的基因和蛋白结构、细胞内表达和定位、翻译后修饰、蛋白三维结构、底物特异性和可能的运输途径等方面的研究进展进行了综述,并对其作为害虫防治分子靶标的可能性进行了初步的讨论.研究发现,不同物种的SCP蛋白的基因编码形式和蛋白剪切形式不同;双翅目昆虫埃及伊蚊Aedes aegypti和黑腹果蝇Drosophila melanogaster除了SCP-x基因可编码SCP-x和SCP-2蛋白外,还有另外的SCP-2和类SCP-2(SCP-2L)基因编码SCP-2和类SCP-2蛋白;而鳞翅目昆虫棉贪夜蛾Spodoptera littoralis、斜纹夜蛾Spodoptera litura和家蚕Bombyx mori中SCP-x 基因的表达和转录方式与脊椎动物的SCP-x基因类似,通过转录和翻译后剪切形成SCP-2蛋白.SCP-x和SCP-2蛋白定位于过氧化物酶体.SCP-2蛋白由5个α-螺旋和5个β-折叠组成,其中α5-螺旋可影响蛋白与底物的结合.SCP-2蛋白以不同的亲和力与固醇、胆同醇衍生物、脂肪酸、脂酰辅酶A和磷脂等化合物结合.超表达斜纹夜蛾SlSCP-x和SlSCP-2

  9. História da enfermagem psiquiátrica e a dependência química no Brasil: atravessando a história para reflexão Historia de la enfermería psiquiátrica y la dependencia química en el Brasil: atravesando la historia para la reflexión History of the psychiatric nursing and chemical dependency in Brazil: crossing the history for reflection

    OpenAIRE

    Amanda Márcia dos Santos Reinaldo; Sandra Cristina Pillon

    2007-01-01

    A formação do enfermeiro em Enfermagem Psiquiátrica e na área de dependências químicas norteia a discussão desse artigo, tendo em vista a complexidade dos problemas relacionados ao ensino de enfermagem, saúde mental, psiquiatria e álcool e drogas. Trata-se de um artigo de revisão de literatura onde as autoras compilaram fontes primárias e secundárias sobre o tema e, a partir da leitura do material bibliográfico, fizeram análises e reflexões acerca dos atravessamentos históricos que permeiam a...

  10. 鹅脱氧胆酸对高果糖饮食致大鼠肾损伤的保护作用及其机制%Protective effect of chenodeoxycholic acid against lipid kidney injury induced by high-fructose-feeding in rats and the underlying mechanism

    Institute of Scientific and Technical Information of China (English)

    胡志娟; 任路平; 王超; 李芳; 董春霞; 李刚; 刘冰; 宋光耀

    2013-01-01

    Objective To study the intervention of chenodeoxycholic acid (CDCA) on kidney of high-fructose-fed rats,and investigate the mechanism of CDCA on lipid kidney injury.Methods Forty-eight healthy male Wistar rats were randomly divided into three groups:normal control group (n =16),high fructose group (n =16) and CDCA group (n =16).Eight rats were sacrificed at the end of 8 and 16 weeks in each group.BUN,Scr,uric acid (UA),fast glucose,serum lipid concentration,urinary albumin were measured.The triglyceride content of renal cortices was detected.The change of renal histopathology was observed by Periodic acid Schiff staining and electron microscopy.The mRNA expressions of farnesoid X receptor (FXR),small heterodimer partner (SHP),sterol regulatory elementbinding protein 1c (SREBP-1c),stearoyl-CoA carboxylase (SCD-1),peroxisome proliferator-activated receptor α (PPARα),acyl coenzyme A oxidase (ACO),transforming growth factor β1 (TGF-β1),type 1 plasminogen activator inhibitor (PAI-1),tumor necrosis factor α (TNF-α),interleukin 6 (IL-6) and NADPH oxidase 2 (Nox2) were measured by real-time PCR,and the protein expressions of which were analyzed by Western blotting.Results Left kidney weight/body weight,triglyceride,very-low-density-lipoprotein and UA in blood were significantly increased in high fructose group (all P < 0.05).Renal function and fast glucose did not change much (P > 0.05).The urinary albumin significantly increased in high fructose group (P < 0.01).The triglyceride content in renal cortex was much more abundant than that in control group (P<0.01).Renal injuries including mesangial expansion,glomerular basement membrane thickening and podocyte foot process effacement were found in fructosefed Wistar rats.The gene and protein expressions of FXR and SHP in kidneys of rats fed with high fructose were significantly down-regulated (all P < 0.01).The gene and protein expressions of SREBP-1c and SCD-1 in kidneys of rats fed with high fructose were

  11. Comparison of glycerolipid biosynthesis in non-green plastids from sycamore (Acer pseudoplatanus) cells and cauliflower (Brassica oleracea) buds.

    Science.gov (United States)

    Alban, C; Joyard, J; Douce, R

    1989-05-01

    The availability of methods to fractionate non-green plastids and to prepare their limiting envelope membranes [Alban, Joyard & Douce (1988) Plant Physiol. 88, 709-717] allowed a detailed analysis of the biosynthesis of lysophosphatidic acid, phosphatidic acid, diacylglycerol and monogalactosyl-diacylglycerol (MGDG) in two different types of non-green starch-containing plastids: plastids isolated from cauliflower buds and amyloplasts isolated from sycamore cells. An enzyme [acyl-ACP (acyl carrier protein):sn-glycerol 3-phosphate acyltransferase) recovered in the soluble fraction of non-green plastids transfers oleic acid from oleoyl-ACP to the sn-1 position of sn-glycerol 3-phosphate to form lysophosphatidic acid. Then a membrane-bound enzyme (acyl-ACP:monoacyl-sn-glycerol 3-phosphate acyltransferase), localized in the envelope membrane, catalyses the acylation of the available sn-2 position of 1-oleoyl-sn-glycerol 3-phosphate by palmitic acid from palmitoyl-ACP. Therefore both the soluble phase and the envelope membranes are necessary for acylation of sn-glycerol 3-phosphate. The major difference between cauliflower (Brassica oleracea) and sycamore (Acer pseudoplatanus) membranes is the very low level of phosphatidate phosphatase activity in sycamore envelope membrane. Therefore, very little diacylglycerol is available for MGDG synthesis in sycamore, compared with cauliflower. These findings are consistent with the similarities and differences described in lipid metabolism of mature chloroplasts from 'C18:3' and 'C16:3' plants (those with MGDG containing C18:3 and C16:3 fatty acids). Sycamore contains only C18 fatty acids in MGDG, and the envelope membranes from sycamore amyloplasts have a low phosphatidate phosphatase activity and therefore the enzymes of the Kornberg-Pricer pathway have a low efficiency of incorporation of sn-glycerol 3-phosphate into MGDG. By contrast, cauliflower contains MGDG with C16:3 fatty acid, and the incorporation of sn-glycerol 3

  12. Structural basis of substrate discrimination and integrin binding by autotaxin

    Energy Technology Data Exchange (ETDEWEB)

    Hausmann, Jens; Kamtekar, Satwik; Christodoulou, Evangelos; Day, Jacqueline E.; Wu, Tao; Fulkerson, Zachary; Albers, Harald M.H.G.; van Meeteren, Laurens A.; Houben, Anna J.S.; van Zeijl, Leonie; Jansen, Silvia; Andries, Maria; Hall, Troii; Pegg, Lyle E.; Benson, Timothy E.; Kasiem, Mobien; Harlos, Karl; Vander Kooi, Craig W.; Smyth, Susan S.; Ovaa, Huib; Bollen, Mathieu; Morris, Andrew J.; Moolenaar, Wouter H.; Perrakis, Anastassis (Pfizer); (Leuven); (Oxford); (NCI-Netherlands); (Kentucky)

    2013-09-25

    Autotaxin (ATX, also known as ectonucleotide pyrophosphatase/phosphodiesterase-2, ENPP2) is a secreted lysophospholipase D that generates the lipid mediator lysophosphatidic acid (LPA), a mitogen and chemoattractant for many cell types. ATX-LPA signaling is involved in various pathologies including tumor progression and inflammation. However, the molecular basis of substrate recognition and catalysis by ATX and the mechanism by which it interacts with target cells are unclear. Here, we present the crystal structure of ATX, alone and in complex with a small-molecule inhibitor. We have identified a hydrophobic lipid-binding pocket and mapped key residues for catalysis and selection between nucleotide and phospholipid substrates. We have shown that ATX interacts with cell-surface integrins through its N-terminal somatomedin B-like domains, using an atypical mechanism. Our results define determinants of substrate discrimination by the ENPP family, suggest how ATX promotes localized LPA signaling and suggest new approaches for targeting ATX with small-molecule therapeutic agents.

  13. A role for 1-acylglycerol-3-phosphate-O-acyltransferase-1 in myoblast differentiation.

    Science.gov (United States)

    Subauste, Angela R; Elliott, Brandon; Das, Arun K; Burant, Charles F

    2010-01-01

    AGPAT isoforms catalyze the acylation of lysophosphatidic acid (LPA) to form phosphatidic acid (PA). AGPAT2 mutations are associated with defective adipogenesis. Muscle and adipose tissue share common precursor cells. We investigated the role of AGPAT isoforms in skeletal muscle development. We demonstrate that small interference RNA-mediated knockdown of AGPAT1 expression prevents the induction of myogenin, a key transcriptional activator of the myogenic program, and inhibits the expression of myosin heavy chain. This effect is rescued by transfection with AGPAT1 but not AGPAT2. Knockdown of AGPAT2 has no effect. The regulation of myogenesis by AGPAT1 is associated with alterations on actin cytoskeleton. The role of AGPAT1 on actin cytoskeleton is further supported by colocalization of AGPAT1 to areas of active actin polymerization. AGPAT1 overexpression was not associated with an increase in PA levels. Our observations strongly implicate AGPAT1 in the development of skeletal muscle, specifically to terminal differentiation. These findings are linked to the regulation of actin cytoskeleton. PMID:20561744

  14. Autotaxin: a protein with two faces.

    Science.gov (United States)

    Tania, Mousumi; Khan, Md Asaduzzaman; Zhang, Huaiyuan; Li, Jinhua; Song, Yuanda

    2010-10-29

    Autotaxin (ATX) is a catalytic protein, which possesses lysophospholipase D activity, and thus involved in cellular membrane lipid metabolism and remodeling. Primarily, ATX was thought as a culprit protein for cancer, which potently stimulates cancer cell proliferation and tumor cell motility, augments the tumorigenicity and induces angiogenic responses. The product of ATX catalyzed reaction, lysophosphatidic acid (LPA) is a potent mitogen, which facilitates cell proliferation and migration, neurite retraction, platelet aggregation, smooth muscle contraction, actin stress formation and cytokine and chemokine secretion. In addition to LPA formation, later ATX has been found to catalyze the formation of cyclic phosphatidic acid (cPA), which have antitumor role by antimitogenic regulation of cell cycle, inhibition of cancer invasion and metastasis. Furthermore, the very attractive information to the scientists is that the LPA/cPA formation can be altered at different physiological conditions. Thus the dual role of ATX with the scope of product manipulation has made ATX a novel target for cancer treatment. PMID:20888793

  15. Role of mitochondrial lipids in guiding fission and fusion.

    Science.gov (United States)

    Frohman, Michael A

    2015-03-01

    Clinically important links have been established between mitochondrial function and cardiac physiology and disease in the context of signaling mechanisms, energy production, and muscle cell development. The proteins and processes that drive mitochondrial fusion and fission are now known to have emergent functions in intracellular calcium homeostasis, apoptosis, vascular smooth muscle cell proliferation, myofibril organization, and Notch-driven cell differentiation, all key issues in cardiac disease. Moreover, decreasing fission may confer protection against ischemic heart disease, particularly in the setting of obesity, diabetes, and heart failure. The importance of lipids in controlling mitochondrial fission and fusion is increasingly becoming appreciated. Roles for the bulk and signaling lipids cardiolipin, phosphatidylethanolamine, phosphatidic acid, diacylglycerol, and lysophosphatidic acid and the enzymes that synthesize or metabolize them in the control of mitochondrial shape and function are reviewed here. A number of diseases have been linked to loss-of-function alleles for a subset of the enzymes, emphasizing the importance of the lipid environment in this context. PMID:25471483

  16. Fatty acid composition of muscle fat and enzymes of storage lipid synthesis in whole muscle from beef cattle.

    Science.gov (United States)

    Kazala, E Chris; Lozeman, Fred J; Mir, Priya S; Aalhus, Jennifer L; Schmutz, Sheila M; Weselake, Randall J

    2006-11-01

    Enhanced intramuscular fat content (i.e., marbling) in beef is a desirable trait, which can result in increased product value. This study was undertaken with the aim of revealing biochemical factors associated with the marbling trait in beef cattle. Samples of longissimus lumborum (LL) and pars costalis diaphragmatis (PCD) were taken from a group of intact crossbred males and females at slaughter, lipids extracted, and the resulting FAME examined for relationships with marbling fat deposition. For LL, significant associations were found between degree of marbling and myristic (14:0, r = 0.55, P muscle were assayed for diacylglycerol acyltransferase (DGAT), lysophosphatidic acid acyltransferase (LPAAT), and phosphatidic acid phosphatase-1 (PAP-1) activity, and the results examined for relationships with degree of intramuscular fat deposition. None of the enzyme activities from PCD displayed an association with marbling fat content, but DGAT specific activity showed significant positive associations with LPAAT (r = 0.54, P muscle tissues provide insight into possible enzyme action associated with the production of specific FA. The increased proportion of oleic acid associated with enhanced lipid content of whole muscle is noteworthy given the known health benefits of this FA. PMID:17263304

  17. Alterations in lipid signaling underlie lipodystrophy secondary to AGPAT2 mutations.

    Science.gov (United States)

    Subauste, Angela R; Das, Arun K; Li, Xiangquan; Elliott, Brandon G; Elliot, Brandon; Evans, Charles; El Azzouny, Mahmoud; Treutelaar, Mary; Oral, Elif; Leff, Todd; Burant, Charles F

    2012-11-01

    Congenital generalized lipodystrophy (CGL), secondary to AGPAT2 mutation is characterized by the absence of adipocytes and development of severe insulin resistance. In the current study, we investigated the adipogenic defect associated with AGPAT2 mutations. Adipogenesis was studied in muscle-derived multipotent cells (MDMCs) isolated from vastus lateralis biopsies obtained from controls and subjects harboring AGPAT2 mutations and in 3T3-L1 preadipocytes after knockdown or overexpression of AGPAT2. We demonstrate an adipogenic defect using MDMCs from control and CGL human subjects with mutated AGPAT2. This defect was rescued in CGL MDMCs with a retrovirus expressing AGPAT2. Both CGL-derived MDMCs and 3T3-L1 cells with knockdown of AGPAT2 demonstrated an increase in cell death after induction of adipogenesis. Lack of AGPAT2 activity reduces Akt activation, and overexpression of constitutively active Akt can partially restore lipogenesis. AGPAT2 modulated the levels of phosphatidic acid, lysophosphatidic acid, phosphatidylinositol species, as well as the peroxisome proliferator-activated receptor γ (PPARγ) inhibitor cyclic phosphatidic acid. The PPARγ agonist pioglitazone partially rescued the adipogenic defect in CGL cells. We conclude that AGPAT2 regulates adipogenesis through the modulation of the lipome, altering normal activation of phosphatidylinositol 3-kinase (PI3K)/Akt and PPARγ pathways in the early stages of adipogenesis. PMID:22872237

  18. Horse chestnut extract induces contraction force generation in fibroblasts through activation of Rho/Rho kinase.

    Science.gov (United States)

    Fujimura, Tsutomu; Moriwaki, Shigeru; Hotta, Mitsuyuki; Kitahara, Takashi; Takema, Yoshinori

    2006-06-01

    Contraction forces generated by non-muscle cells such as fibroblasts play important roles in determining cell morphology, vasoconstriction, and/or wound healing. However, few factors that induce cell contraction forces are known, such as lysophosphatidic acid and thrombin. Our study analyzed various plant extracts for ingredients that induce generation of cell contraction forces in fibroblasts populating collagen gels. We found that an extract of Horse chestnut (Aesculus hippocastanum) is able to induce such contraction forces in fibroblasts. The involvement of actin polymerization and stress fiber formation in the force generation was suggested by inhibition of this effect by cytochalasin D and by Rhodamine phalloidin. Rho kinase inhibitors (Y27632 and HA1077) and a Rho inhibitor (exoenzyme C3) significantly inhibited the force generation induced by the Horse chestnut extract. H7, which inhibits Rho kinase as well as other protein kinases, also significantly inhibited induction of force generation. However, inhibitors of other protein kinases such as myosin light chain kinase (ML-9), protein kinase C (Calphostin), protein kinase A (KT5720), and tyrosine kinase (Genistein, Herbimycin A) had no effect on force generation induced by Horse chestnut extract. These results suggest that the Horse chestnut extract induces generation of contraction forces in fibroblasts through stress fiber formation followed by activation of Rho protein and Rho kinase but not myosin light chain kinase or other protein kinases. PMID:16754996

  19. Fatty acid composition of muscle fat and enzymes of storage lipid synthesis in whole muscle from beef cattle.

    Science.gov (United States)

    Kazala, E Chris; Lozeman, Fred J; Mir, Priya S; Aalhus, Jennifer L; Schmutz, Sheila M; Weselake, Randall J

    2006-11-01

    Enhanced intramuscular fat content (i.e., marbling) in beef is a desirable trait, which can result in increased product value. This study was undertaken with the aim of revealing biochemical factors associated with the marbling trait in beef cattle. Samples of longissimus lumborum (LL) and pars costalis diaphragmatis (PCD) were taken from a group of intact crossbred males and females at slaughter, lipids extracted, and the resulting FAME examined for relationships with marbling fat deposition. For LL, significant associations were found between degree of marbling and myristic (14:0, r = 0.55, P muscle were assayed for diacylglycerol acyltransferase (DGAT), lysophosphatidic acid acyltransferase (LPAAT), and phosphatidic acid phosphatase-1 (PAP-1) activity, and the results examined for relationships with degree of intramuscular fat deposition. None of the enzyme activities from PCD displayed an association with marbling fat content, but DGAT specific activity showed significant positive associations with LPAAT (r = 0.54, P muscle tissues provide insight into possible enzyme action associated with the production of specific FA. The increased proportion of oleic acid associated with enhanced lipid content of whole muscle is noteworthy given the known health benefits of this FA.

  20. Alterations in lipid signaling underlie lipodystrophy secondary to AGPAT2 mutations.

    Science.gov (United States)

    Subauste, Angela R; Das, Arun K; Li, Xiangquan; Elliott, Brandon G; Elliot, Brandon; Evans, Charles; El Azzouny, Mahmoud; Treutelaar, Mary; Oral, Elif; Leff, Todd; Burant, Charles F

    2012-11-01

    Congenital generalized lipodystrophy (CGL), secondary to AGPAT2 mutation is characterized by the absence of adipocytes and development of severe insulin resistance. In the current study, we investigated the adipogenic defect associated with AGPAT2 mutations. Adipogenesis was studied in muscle-derived multipotent cells (MDMCs) isolated from vastus lateralis biopsies obtained from controls and subjects harboring AGPAT2 mutations and in 3T3-L1 preadipocytes after knockdown or overexpression of AGPAT2. We demonstrate an adipogenic defect using MDMCs from control and CGL human subjects with mutated AGPAT2. This defect was rescued in CGL MDMCs with a retrovirus expressing AGPAT2. Both CGL-derived MDMCs and 3T3-L1 cells with knockdown of AGPAT2 demonstrated an increase in cell death after induction of adipogenesis. Lack of AGPAT2 activity reduces Akt activation, and overexpression of constitutively active Akt can partially restore lipogenesis. AGPAT2 modulated the levels of phosphatidic acid, lysophosphatidic acid, phosphatidylinositol species, as well as the peroxisome proliferator-activated receptor γ (PPARγ) inhibitor cyclic phosphatidic acid. The PPARγ agonist pioglitazone partially rescued the adipogenic defect in CGL cells. We conclude that AGPAT2 regulates adipogenesis through the modulation of the lipome, altering normal activation of phosphatidylinositol 3-kinase (PI3K)/Akt and PPARγ pathways in the early stages of adipogenesis.

  1. Serum Autotaxin is a Marker of the Severity of Liver Injury and Overall Survival in Patients with Cholestatic Liver Diseases

    Science.gov (United States)

    Wunsch, Ewa; Krawczyk, Marcin; Milkiewicz, Malgorzata; Trottier, Jocelyn; Barbier, Olivier; Neurath, Markus F.; Lammert, Frank; Kremer, Andreas E.; Milkiewicz, Piotr

    2016-01-01

    Autotaxin (ATX) is involved in the synthesis of lysophosphatidic acid. Both have recently been linked to cholestatic pruritus and liver injury. We aimed to investigate whether ATX is an indicator of cholestatic liver injury, health-related quality of life (HRQoL) and prognosis based on a group of 233 patients, 118 with primary biliary cholangitis (PBC) and 115 with primary sclerosing cholangitis (PSC). Patients were followed for 1–60 months, cumulative survival rates were calculated. ATX activity was significantly higher in both groups than in the 103 controls, particularly in patients with cirrhosis and in patients with longer disease duration. Ursodeoxycholic acid (UDCA) non-responders with PBC exhibited increased ATX activity. ATX activity was correlated with liver biochemistry, MELD, Mayo Risk scores and was associated with worse disease-specific HRQoL aspects. In both groups, Cox model analysis indicated that ATX was a negative predictor of survival. Increased ATX levels were associated with a 4-fold higher risk of death/liver transplantation in patients with PBC and a 2.6-fold higher risk in patients with PSC. We conclude that in patients with cholestatic conditions, ATX is not only associated with pruritus but also indicates impairment of other HRQoL aspects, liver dysfunction, and can serve as a predictor of survival. PMID:27506882

  2. Enhancement of migration and invasion of hepatoma cells via a Rho GTPase signaling pathway

    Institute of Scientific and Technical Information of China (English)

    De-Sheng Wang; Ke-Feng Dou; Kai-Zong Li; Zhen-Shun Song

    2004-01-01

    AIM: Intrahepatic extension is the main cause of liver failure and death in hepatocellular carcinoma patients. The small GTPase Rho and one of its effector molecules ROCK regulate cytoskeleton and actomyosin contractility, and play a crucial role in cell adhesion and motility. We investigated the role of small GTPase Rho in biological behaviors of hepatocellular carcinoma to demonstrate the importance of Rho in cancer invasion and metastasis.METHODS: Using Western blotting, we quantitated Rho protein expression in SMMC-7721 cells induced by Lysophosphatidic acid (LPA). Furthermore, we examined the role of Rho signaling in regulating the motile and invasiveproperties of tumor cells.RESULTS: Rho protein expression was stimulated by LPA.Using the Rhotekin binding assay to assess Rho activation,we observed that the level of GTP-bound Rho was elevated transiently after the addition of LPA, and Y-27632 decreased the level of active Rho. LPA enhanced the motility of tumor cells and facilitated their invasion. Rho played an essential role in the migratory process, as evidenced by the inhibition of migration and motility of cancer cells by a specific inhibitor of ROCK, Y-27632.CONCLUSION: The finding that invasiveness of hepatocellular carcinoma is facilitated by the Rho/Rho-kinase pathway is likely to be relevant to tumor progression and Y-27632 may be a new potential effective agent for the prevention of intrahepatic extension of human liver cancer.

  3. Protein kinase D is increased and activated in lung epithelial cells and macrophages in idiopathic pulmonary fibrosis.

    Directory of Open Access Journals (Sweden)

    Huachen Gan

    Full Text Available Idiopathic pulmonary fibrosis (IPF is a relentlessly progressive and usually fatal lung disease of unknown etiology for which no effective treatments currently exist. Hence, there is a profound need for the identification of novel drugable targets to develop more specific and efficacious therapeutic intervention in IPF. In this study, we performed immunohistochemical analyses to assess the cell type-specific expression and activation of protein kinase D (PKD family kinases in normal and IPF lung tissue sections. We also analyzed PKD activation and function in human lung epithelial cells. We found that PKD family kinases (PKD1, PKD2 and PKD3 were increased and activated in the hyperplastic and regenerative alveolar epithelial cells lining remodeled fibrotic alveolar septa and/or fibroblast foci in IPF lungs compared with normal controls. We also found that PKD family kinases were increased and activated in alveolar macrophages, bronchiolar epithelium, and honeycomb cysts in IPF lungs. Interestingly, PKD1 was highly expressed and activated in the cilia of IPF bronchiolar epithelial cells, while PKD2 and PKD3 were expressed in the cell cytoplasm and nuclei. In contrast, PKD family kinases were not apparently increased and activated in IPF fibroblasts or myofibroblasts. We lastly found that PKD was predominantly activated by poly-L-arginine, lysophosphatidic acid and thrombin in human lung epithelial cells and that PKD promoted epithelial barrier dysfunction. These findings suggest that PKD may participate in the pathogenesis of IPF and may be a novel target for therapeutic intervention in this disease.

  4. The effect of charge reversal mutations in the alpha-helical region of liver fatty acid binding protein on the binding of fatty-acyl CoAs, lysophospholipids and bile acids.

    Science.gov (United States)

    Hagan, Robert M; Davies, Joanna K; Wilton, David C

    2002-10-01

    Liver fatty acid binding protein (LFABP) is unique among the various types of FABPs in that it can bind a variety of ligands in addition to fatty acids. LFABP is able to bind long chain fatty acids with a 2:1 stoichiometry and the crystal structure has identified two fatty acid binding sites in the binding cavity. The presumed primary site (site 1) involves the fatty acid binding with the carboxylate group buried in the cavity whereas the fatty acid at site 2 has the carboxylate group solvent-exposed within the ligand portal region and in the vicinity of alpha-helix II. The alpha-helical region contains three cationic residues, K20, K31, K33 and modelling studies suggest that K31 on alpha-helix II could make an electrostatic contribution to anionic ligands binding to site 2. The preparation of three charge reversal mutants of LFABP, K20E, K31E and K33E has allowed an investigation of the role of site 2 in ligand binding, particularly those ligands with a bulky anionic head group. The binding of oleoyl CoA, lysophosphatidic acid, lysophosphatidylcholine, lithocholic acid and taurolithocholate 3-sulphate to LFABP has been studied using the alpha-helical mutants. The results support the concept that such ligands bind at site 2 of LFABP where solvent exposure allows the accommodation of their bulky anionic group. PMID:12479568

  5. Stimulation of human red blood cells leads to Ca2+-mediated intercellular adhesion

    CERN Document Server

    Steffen, Patrick; Nguyen, Duc Bach; Müller, Torsten; Bernhardt, Ingolf; Kaestner, Lars; Wagner, Christian

    2011-01-01

    Red blood cells (RBCs) are a major component of blood clots, which form physiologically as a response to injury or pathologically in thrombosis. The active participation of RBCs in thrombus solidification has been previously proposed but not yet experimentally proven. Holographic optical tweezers and single-cell force spectroscopy were used to study potential cell-cell adhesion between RBCs. Irreversible intercellular adhesion of RBCs could be induced by stimulation with lysophosphatidic acid (LPA), a compound known to be released by activated platelets. We identified Ca2+ as an essential player in the signaling cascade by directly inducing Ca2+ influx using A23187. Elevation of the internal Ca2+ concentration leads to an intercellular adhesion of RBCs similar to that induced by LPA stimulation. Using single-cell force spectroscopy, the adhesion of the RBCs was identified to be approximately 100 pN, a value large enough to be of significance inside a blood clot or in pathological situations like the vasco-occ...

  6. Identification and expression analysis of castor bean (Ricinus communis) genes encoding enzymes from the triacylglycerol biosynthesis pathway.

    Science.gov (United States)

    Cagliari, Alexandro; Margis-Pinheiro, Márcia; Loss, Guilherme; Mastroberti, Alexandra Antunes; de Araujo Mariath, Jorge Ernesto; Margis, Rogério

    2010-11-01

    Castor bean (Ricinus communis) oil contains ricinoleic acid-rich triacylglycerols (TAGs). As a result of its physical and chemical properties, castor oil and its derivatives are used for numerous bio-based products. In this study, we survey the Castor Bean Genome Database to report the identification of TAG biosynthesis genes. A set of 26 genes encoding six distinct classes of enzymes involved in TAGs biosynthesis were identified. In silico characterization and sequence analysis allowed the identification of plastidic isoforms of glycerol-3-phosphate acyltransferase and lysophosphatidate acyltransferase enzyme families, involved in the prokaryotic lipid biosynthesis pathway, that form a cluster apart from the cytoplasmic isoforms, involved in the eukaryotic pathway. In addition, two distinct membrane bound diacylglycerol acyltransferase enzymes were identified. Quantitative expression pattern analyses demonstrated variations in gene expressions during castor seed development. A tendency of maximum expression level at the middle of seed development was observed. Our results represent snapshots of global transcriptional activities of genes encompassing six enzyme families involved in castor bean TAG biosynthesis that are present during seed development. These genes represent potential targets for biotechnological approaches to produce nutritionally and industrially desirable oils.

  7. Non-invasive imaging of tumors by monitoring autotaxin activity using an enzyme-activated near-infrared fluorogenic substrate.

    Directory of Open Access Journals (Sweden)

    Damian Madan

    Full Text Available Autotaxin (ATX, an autocrine motility factor that is highly upregulated in metastatic cancer, is a lysophospholipase D enzyme that produces the lipid second messenger lysophosphatidic acid (LPA from lysophosphatidylcholine (LPC. Dysregulation of the lysolipid signaling pathway is central to the pathophysiology of numerous cancers, idiopathic pulmonary fibrosis, rheumatoid arthritis, and other inflammatory diseases. Consequently, the ATX/LPA pathway has emerged as an important source of biomarkers and therapeutic targets. Herein we describe development and validation of a fluorogenic analog of LPC (AR-2 that enables visualization of ATX activity in vivo. AR-2 exhibits minimal fluorescence until it is activated by ATX, which substantially increases fluorescence in the near-infrared (NIR region, the optimal spectral window for in vivo imaging. In mice with orthotopic ATX-expressing breast cancer tumors, ATX activated AR-2 fluorescence. Administration of AR-2 to tumor-bearing mice showed high fluorescence in the tumor and low fluorescence in most healthy tissues with tumor fluorescence correlated with ATX levels. Pretreatment of mice with an ATX inhibitor selectively decreased fluorescence in the tumor. Together these data suggest that fluorescence directly correlates with ATX activity and its tissue expression. The data show that AR-2 is a non-invasive and selective tool that enables visualization and quantitation of ATX-expressing tumors and monitoring ATX activity in vivo.

  8. Expression of the lysophospholipid receptor family and investigation of lysophospholipid-mediated responses in human macrophages.

    Science.gov (United States)

    Duong, Chinh Quoc; Bared, Salim Maa; Abu-Khader, Ahmad; Buechler, Christa; Schmitz, Anna; Schmitz, Gerd

    2004-06-01

    Some of the biological effects of lipoproteins have been attributed to their association with lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), sphingosine-1-phosphate (S1P) and sphingosylphosphorylcholine (SPC). These lysophospholipids mediate multiple biological responses via several G protein-coupled receptors (GPR). The expression of these receptors, however, has not been systematically investigated in primary human monocytes and macrophages as major cells involved in atherosclerosis. The mRNAs for all 15 receptors described so far were detected in monocytes, macrophages, foam cells and high density lipoprotein (HDL(3))-treated cells using real time RT-PCR. Immunoblots revealed that S1P(1), S1P(2), S1P(4), LPA(1), LPA(2) and GPR65 are expressed in monocytes and macrophages, while S1P(5) and LPA(3) have not been detected. S1P(3) was induced during differentiation but down-regulated by lipid-loading and HDL(3), whereas LPA(1) was down-regulated in differentiated macrophages. The influence of S1P on macrophages was investigated and the induction of CD32 indicates an enhanced phagocytic activity. Altogether, these data give insights into the expression and regulation of lysophospholipid receptors in primary human monocytes, macrophages and foam cells. PMID:15158762

  9. Anatomical Location of LPA1 Activation and LPA Phospholipid Precursors in Rodent and Human Brain

    Science.gov (United States)

    González de San Román, E; Manuel, I; Giralt, MT; Chun, J; Estivill-Torrús, G; Rodriguez de Fonseca, F; Santín, LJ; Ferrer, I; Rodriguez-Puertas, R

    2016-01-01

    Lysophosphatidic acid (LPA) is a signaling molecule that binds to six known G protein-coupled receptors (GPCRs): LPA1–LPA6. LPA evokes several responses in the CNS including cortical development and folding, growth of the axonal cone and its retraction process. Those cell processes involve survival, migration, adhesion proliferation, differentiation and myelination. The anatomical localization of LPA1 is incompletely understood, particularly with regard to LPA binding. Therefore, we have used functional [35S]GTPγS autoradiography to verify the anatomical distribution of LPA1 binding sites in adult rodent and human brain. The greatest activity was observed in myelinated areas of the white matter such as corpus callosum, internal capsule and cerebellum. MaLPA1-null mice (a variant of LPA1-null) lack [35S]GTPγS basal binding in white matter areas, where the LPA1 receptor is expressed at high levels, suggesting a relevant role of the activity of this receptor in the most myelinated brain areas. In addition, phospholipid precursors of LPA were localized by MALDI-IMS in both rodent and human brain slices identifying numerous species of phosphatides (PA) and phosphatidylcholines (PC). Both PA and PC species represent potential LPA precursors. The anatomical distribution of these precursors in rodent and human brain may indicate a metabolic relationship between LPA and LPA1 receptors. PMID:25857358

  10. Autotaxin, Pruritus and Primary Biliary Cholangitis (PBC).

    Science.gov (United States)

    Sun, Ying; Zhang, Weici; Evans, Jilly F; Floreani, Annarosa; Zou, Zhengsheng; Nishio, Yukiko; Qi, Ruizhao; Leung, Patrick S C; Bowlus, Christopher L; Gershwin, M Eric

    2016-08-01

    Autotaxin (ATX) is a 125-kD type II ectonucleotide pyrophosphatase/phosphodiesterase (ENPP2 or NPP2) originally discovered as an unknown "autocrine motility factor" in human melanoma cells. In addition to its pyrophosphatase/phosphodiesterase activities ATX has lysophospholipase D (lysoPLD) activity, catalyzing the conversion of lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA). ATX is the only ENPP family member with lysoPLD activity and it produces most of the LPA in circulation. In support of this, ATX heterozygous mice have 50% of normal LPA plasma levels. The ATX-LPA signaling axis plays an important role in both normal physiology and disease pathogenesis and recently has been linked to pruritus in chronic cholestatic liver diseases, including primary biliary cholangitis (PBC). Several lines of evidence have suggested that a circulating puritogen is responsible, but the identification of the molecule has yet to be definitively identified. In contrast, plasma ATX activity is strongly associated with pruritus in PBC, suggesting a targetable molecule for treatment. We review herein the biochemistry of ATX and the rationale for its role in pruritus.

  11. [Regulatory role of mechanical stress response in cellular function: development of new drugs and tissue engineering].

    Science.gov (United States)

    Momose, Kazutaka; Matsuda, Takehisa; Oike, Masahiro; Obara, Kazuo; Laher, Ismail; Sugiura, Seiryo; Ohata, Hisayuki; Nakayama, Koichi

    2003-02-01

    The investigation of mechanotransduction in the cardiovascular system is essentially important for elucidating the cellular and molecular mechanisms involved in not only the maintenance of hemodynamic homeostasis but also etiology of cardiovascular diseases including arteriosclerosis. The present review summarizes the latest research performed by six academic groups, and presented at the 75th Annual Meeting of the Japanese Pharmacological Society. Technology of cellular biomechanics is also required for research and clinical application of a vascular hybrid tissue responding to pulsatile stress. 1) Vascular tissue engineering: Design of pulsatile stress-responsive scaffold and in vivo vascular wall reconstruction (T. Matsuda); 2) Cellular mechanisms of mechanosensitive calcium transients in vascular endothelium (M. Oike et al.); 3) Cross-talk of stimulation with fluid flow and lysophosphatidic acid in vascular endothelial cells (K. Momose et al.); 4) Mechanotransduction of vascular smooth muscles: Rate-dependent stretch-induced protein phosphorylations and contractile activation (K. Obara et al.); 5) Lipid mediators in vascular myogenic tone (I. Laher et al.); and 6) Caldiomyocyte regulates its mechanical output in response to mechanical load (S. Sugiura et al.).

  12. Drosophila homologue of Diaphanous 1 (DIAPH1) controls the metastatic potential of colon cancer cells by regulating microtubule-dependent adhesion.

    Science.gov (United States)

    Lin, Yuan-Na; Bhuwania, Ridhirama; Gromova, Kira; Failla, Antonio Virgilio; Lange, Tobias; Riecken, Kristoffer; Linder, Stefan; Kneussel, Matthias; Izbicki, Jakob R; Windhorst, Sabine

    2015-07-30

    Drosophila homologue of Diaphanous 1 (DIAPH1) regulates actin polymerization and microtubule (MT) stabilization upon stimulation with lysophosphatidic acid (LPA). Recently, we showed strongly reduced lung metastasis of DIAPH1-depleted colon cancer cells but we found accumulations of DIAPH1-depleted cells in bone marrow. Here, we analyzed possible organ- or tissue-specific metastasis of DIAPH1-depleted HCT-116 cells. Our data confirmed that depletion of DIAPH1 strongly inhibited lung metastasis and revealed that, in contrast to control cells, DIAPH1-depleted cells did not form metastases in further organs. Detailed mechanistic analysis on cells that were not stimulated with LPA to activate the cytoskeleton-modulating activity of DIAPH1, revealed that even under basal conditions DIAPH1 was essential for cellular adhesion to collagen. In non-stimulated cells DIAPH1 did not control actin dynamics but, interestingly, was essential for stabilization of microtubules (MTs). Additionally, DIAPH1 controlled directed vesicle trafficking and with this, local clustering of the adhesion protein integrin-β1 at the plasma membrane. Therefore, we conclude that under non-stimulating conditions DIAPH1 controls cellular adhesion by stabilizing MTs required for local clustering of integrin-β1 at the plasma membrane. Thus, blockade of DIAPH1-tubulin interaction may be a promising approach to inhibit one of the earliest steps in the metastatic cascade of colon cancer. PMID:26124177

  13. PRG-1 Regulates Synaptic Plasticity via Intracellular PP2A/β1-Integrin Signaling.

    Science.gov (United States)

    Liu, Xingfeng; Huai, Jisen; Endle, Heiko; Schlüter, Leslie; Fan, Wei; Li, Yunbo; Richers, Sebastian; Yurugi, Hajime; Rajalingam, Krishnaraj; Ji, Haichao; Cheng, Hong; Rister, Benjamin; Horta, Guilherme; Baumgart, Jan; Berger, Hendrik; Laube, Gregor; Schmitt, Ulrich; Schmeisser, Michael J; Boeckers, Tobias M; Tenzer, Stefan; Vlachos, Andreas; Deller, Thomas; Nitsch, Robert; Vogt, Johannes

    2016-08-01

    Alterations in dendritic spine numbers are linked to deficits in learning and memory. While we previously revealed that postsynaptic plasticity-related gene 1 (PRG-1) controls lysophosphatidic acid (LPA) signaling at glutamatergic synapses via presynaptic LPA receptors, we now show that PRG-1 also affects spine density and synaptic plasticity in a cell-autonomous fashion via protein phosphatase 2A (PP2A)/β1-integrin activation. PRG-1 deficiency reduces spine numbers and β1-integrin activation, alters long-term potentiation (LTP), and impairs spatial memory. The intracellular PRG-1 C terminus interacts in an LPA-dependent fashion with PP2A, thus modulating its phosphatase activity at the postsynaptic density. This results in recruitment of adhesome components src, paxillin, and talin to lipid rafts and ultimately in activation of β1-integrins. Consistent with these findings, activation of PP2A with FTY720 rescues defects in spine density and LTP of PRG-1-deficient animals. These results disclose a mechanism by which bioactive lipid signaling via PRG-1 could affect synaptic plasticity and memory formation. PMID:27453502

  14. Simulated microgravity inhibits osteogenic differentiation of mesenchymal stem cells via depolymerizing F-actin to impede TAZ nuclear translocation

    Science.gov (United States)

    Chen, Zhe; Luo, Qing; Lin, Chuanchuan; Kuang, Dongdong; Song, Guanbin

    2016-01-01

    Microgravity induces observed bone loss in space flight, and reduced osteogenesis of bone mesenchymal stem cells (BMSCs) partly contributes to this phenomenon. Abnormal regulation or functioning of the actin cytoskeleton induced by microgravity may cause the inhibited osteogenesis of BMSCs, but the underlying mechanism remains obscure. In this study, we demonstrated that actin cytoskeletal changes regulate nuclear aggregation of the transcriptional coactivator with PDZ-binding motif (TAZ), which is indispensable for osteogenesis of bone mesenchymal stem cells (BMSCs). Moreover, we utilized a clinostat to model simulated microgravity (SMG) and demonstrated that SMG obviously depolymerized F-actin and hindered TAZ nuclear translocation. Interestingly, stabilizing the actin cytoskeleton induced by Jasplakinolide (Jasp) significantly rescued TAZ nuclear translocation and recovered the osteogenic differentiation of BMSCs in SMG, independently of large tumor suppressor 1(LATS1, an upstream kinase of TAZ). Furthermore, lysophosphatidic acid (LPA) also significantly recovered the osteogenic differentiation of BMSCs in SMG through the F-actin-TAZ pathway. Taken together, we propose that the depolymerized actin cytoskeleton inhibits osteogenic differentiation of BMSCs through impeding nuclear aggregation of TAZ, which provides a novel connection between F-actin cytoskeleton and osteogenesis of BMSCs and has important implications in bone loss caused by microgravity. PMID:27444891

  15. Regulation of T cell motility in vitro and in vivo by LPA and LPA2.

    Directory of Open Access Journals (Sweden)

    Sara A Knowlden

    Full Text Available Lysophosphatidic acid (LPA and the LPA-generating enzyme autotaxin (ATX have been implicated in lymphocyte trafficking and the regulation of lymphocyte entry into lymph nodes. High local concentrations of LPA are thought to be present in lymph node high endothelial venules, suggesting a direct influence of LPA on cell migration. However, little is known about the mechanism of action of LPA, and more work is needed to define the expression and function of the six known G protein-coupled receptors (LPA 1-6 in T cells. We studied the effects of 18∶1 and 16∶0 LPA on naïve CD4+ T cell migration and show that LPA induces CD4+ T cell chemorepulsion in a Transwell system, and also improves the quality of non-directed migration on ICAM-1 and CCL21 coated plates. Using intravital two-photon microscopy, lpa2-/- CD4+ T cells display a striking defect in early migratory behavior at HEVs and in lymph nodes. However, later homeostatic recirculation and LPA-directed migration in vitro were unaffected by loss of lpa2. Taken together, these data highlight a previously unsuspected and non-redundant role for LPA2 in intranodal T cell motility, and suggest that specific functions of LPA may be manipulated by targeting T cell LPA receptors.

  16. Ecdysone and insulin signaling play essential roles in readjusting the altered body size caused by the dGPAT4 mutation in Drosophila.

    Science.gov (United States)

    Yan, Yan; Wang, Hao; Chen, Hanqing; Lindström-Battle, Anya; Jiao, Renjie

    2015-09-20

    Body size is one of the features that distinguish one species from another in the biological world. Animals have developed mechanisms to control their body size during normal development. However, how animals cope with genetic alterations and/or environmental stresses to develop into normal-sized adults remain poorly understood. The ability of the animals to develop into a normal-sized adult after the challenges of genetic alterations and/or environmental stresses reveals a robustness of body size control. Here we show that the mutation of dGPAT4, a de novo synthase of lysophosphatidic acid, is a genetic alteration that triggers such a robust response of the animals to body size challenges in Drosophila. Loss of dGPAT4 leads to a severe delay of development, slow growth and resultant small-sized animals during the larval stages, but results in normal-sized adult flies. The robust body size adjustment of the dGPAT4 mutant is likely achieved by corresponding changes in ecdysone and insulin signaling, which is also manifested by compromised food intake. Thus, we propose that a strategy has been evolved by the animals to reach final body size when challenged by genetic alterations, which requires the coordinated ecdysone and insulin signaling.

  17. LPA signaling initiates schizophrenia-like brain and behavioral changes in a mouse model of prenatal brain hemorrhage.

    Science.gov (United States)

    Mirendil, H; Thomas, E A; De Loera, C; Okada, K; Inomata, Y; Chun, J

    2015-04-07

    Genetic, environmental and neurodevelopmental factors are thought to underlie the onset of neuropsychiatric disorders such as schizophrenia. How these risk factors collectively contribute to pathology is unclear. Here, we present a mouse model of prenatal intracerebral hemorrhage--an identified risk factor for schizophrenia--using a serum-exposure paradigm. This model exhibits behavioral, neurochemical and schizophrenia-related gene expression alterations in adult females. Behavioral alterations in amphetamine-induced locomotion, prepulse inhibition, thigmotaxis and social interaction--in addition to increases in tyrosine hydroxylase-positive dopaminergic cells in the substantia nigra and ventral tegmental area and decreases in parvalbumin-positive cells in the prefrontal cortex--were induced upon prenatal serum exposure. Lysophosphatidic acid (LPA), a lipid component of serum, was identified as a key molecular initiator of schizophrenia-like sequelae induced by serum. Prenatal exposure to LPA alone phenocopied many of the schizophrenia-like alterations seen in the serum model, whereas pretreatment with an antagonist against the LPA receptor subtype LPA1 prevented many of the behavioral and neurochemical alterations. In addition, both prenatal serum and LPA exposure altered the expression of many genes and pathways related to schizophrenia, including the expression of Grin2b, Slc17a7 and Grid1. These findings demonstrate that aberrant LPA receptor signaling associated with fetal brain hemorrhage may contribute to the development of some neuropsychiatric disorders.

  18. YAP Induces Human Naive Pluripotency

    Directory of Open Access Journals (Sweden)

    Han Qin

    2016-03-01

    Full Text Available The human naive pluripotent stem cell (PSC state, corresponding to a pre-implantation stage of development, has been difficult to capture and sustain in vitro. We report that the Hippo pathway effector YAP is nuclearly localized in the inner cell mass of human blastocysts. Overexpression of YAP in human embryonic stem cells (ESCs and induced PSCs (iPSCs promotes the generation of naive PSCs. Lysophosphatidic acid (LPA can partially substitute for YAP to generate transgene-free human naive PSCs. YAP- or LPA-induced naive PSCs have a rapid clonal growth rate, a normal karyotype, the ability to form teratomas, transcriptional similarities to human pre-implantation embryos, reduced heterochromatin levels, and other hallmarks of the naive state. YAP/LPA act in part by suppressing differentiation-inducing effects of GSK3 inhibition. CRISPR/Cas9-generated YAP−/− cells have an impaired ability to form colonies in naive but not primed conditions. These results uncover an unexpected role for YAP in the human naive state, with implications for early human embryology.

  19. Proteomic screening of human targets of viral microRNAs reveals functions associated with immune evasion and angiogenesis.

    Directory of Open Access Journals (Sweden)

    Amelia M Gallaher

    Full Text Available Kaposi's sarcoma (KS is caused by infection with Kaposi's sarcoma-associated herpesvirus (KSHV. The virus expresses unique microRNAs (miRNAs, but the targets and functions of these miRNAs are not completely understood. In order to identify human targets of viral miRNAs, we measured protein expression changes caused by multiple KSHV miRNAs using pulsed stable labeling with amino acids in cell culture (pSILAC in primary endothelial cells. This led to the identification of multiple human genes that are repressed at the protein level, but not at the miRNA level. Further analysis also identified that KSHV miRNAs can modulate activity or expression of upstream regulatory factors, resulting in suppressed activation of a protein involved in leukocyte recruitment (ICAM1 following lysophosphatidic acid treatment, as well as up-regulation of a pro-angiogenic protein (HIF1α, and up-regulation of a protein involved in stimulating angiogenesis (HMOX1. This study aids in our understanding of miRNA mechanisms of repression and miRNA contributions to viral pathogenesis.

  20. LPA is a chemorepellent for B16 melanoma cells: action through the cAMP-elevating LPA5 receptor.

    Directory of Open Access Journals (Sweden)

    Maikel Jongsma

    Full Text Available Lysophosphatidic acid (LPA, a lipid mediator enriched in serum, stimulates cell migration, proliferation and other functions in many cell types. LPA acts on six known G protein-coupled receptors, termed LPA(1-6, showing both overlapping and distinct signaling properties. Here we show that, unexpectedly, LPA and serum almost completely inhibit the transwell migration of B16 melanoma cells, with alkyl-LPA(18:1 being 10-fold more potent than acyl-LPA(18:1. The anti-migratory response to LPA is highly polarized and dependent on protein kinase A (PKA but not Rho kinase activity; it is associated with a rapid increase in intracellular cAMP levels and PIP3 depletion from the plasma membrane. B16 cells express LPA(2, LPA(5 and LPA(6 receptors. We show that LPA-induced chemorepulsion is mediated specifically by the alkyl-LPA-preferring LPA(5 receptor (GPR92, which raises intracellular cAMP via a noncanonical pathway. Our results define LPA(5 as an anti-migratory receptor and they implicate the cAMP-PKA pathway, along with reduced PIP3 signaling, as an effector of chemorepulsion in B16 melanoma cells.

  1. Prostaglandinsvis-à-vis bovine embryonic mortality:a review

    Institute of Scientific and Technical Information of China (English)

    Jerome A; Srivastava N

    2012-01-01

    Decline in fertility in bovines is attributed to various reproductive problems viz. anoestrus, repeat breeding, abortions and post parturient disorders.Among these, repeat breeding has been an important cause for reducing the animals’ fertility and life-time productivity.Many researchers have reported embryonic mortality as a major cause of repeat breeding arising due to premature corpus luteumlysis.ProstaglandinF2α released from the uterus causes alterations in luteal blood flow, induces luteal lysis, and hence reduces progesterone secretion from the bovine corpus luteum.Therefore various strategies have been tried to modulate prostaglandinF2α synthesis and secretion in order to prolong the lifespan ofCL.Administration of cyclooxygenase inhibitors which include non-steroidal anti-inflammatory drugs acting by competitive inhibition of key enzymes of prostaglandin synthesis is one such method.Feeding of diet rich in polyunsaturated fatty acids during critical period significantly reduces prostaglandin synthesis.Other drugs, which are potential candidates for reducing prostaglandin synthesis, include oxytocin receptor antagonist, recombinant bovine somatotropin, lysophosphatidic acid and prostaglandinF synthase inhibitors. To conclude, there is much scope of using various compounds to reduce prostaglandins synthesis during the critical period of pregnancy for improving the embryo survival rate.

  2. Pharmacological Characterization of a Potent Inhibitor of Autotaxin in Animal Models of Inflammatory Bowel Disease and Multiple Sclerosis.

    Science.gov (United States)

    Thirunavukkarasu, Kannan; Tan, Bailin; Swearingen, Craig A; Rocha, Guilherme; Bui, Hai H; McCann, Denis J; Jones, Spencer B; Norman, Bryan H; Pfeifer, Lance A; Saha, Joy K

    2016-10-01

    Autotaxin is a secreted enzyme that catalyzes the conversion of lysophosphatidyl choline into the bioactive lipid mediator lysophosphatidic acid (LPA). It is the primary enzyme responsible for LPA production in plasma. It is upregulated in inflammatory conditions and inhibition of autotaxin may have anti-inflammatory activity in a variety of inflammatory diseases. To determine the role of autotaxin and LPA in the pathophysiology of inflammatory disease states, we used a potent and orally bioavailable inhibitor of autotaxin that we have recently identified, and characterized it in mouse models of inflammation, inflammatory bowel disease (IBD), multiple sclerosis (MS), and visceral pain. Compound-1, a potent inhibitor of autotaxin with an IC50 of ∼2 nM, has good oral pharmacokinetic properties in mice and results in a substantial inhibition of plasma LPA that correlates with drug exposure levels. Treatment with the inhibitor resulted in significant anti-inflammatory and analgesic effects in the carrageenan-induced paw inflammation and acetic acid-induced visceral pain tests, respectively. Compound-1 also significantly inhibited disease activity score in the dextran sodium sulfate-induced model of IBD, and in the experimental autoimmune encephalomyelitis model of MS. In conclusion, the present study demonstrates the anti-inflammatory and analgesic properties of a novel inhibitor of autotaxin that may serve as a therapeutic option for IBD, MS, and pain associated with inflammatory states.

  3. Age-related changes in retinoic, docosahexaenoic and arachidonic acid modulation in nuclear lipid metabolism.

    Science.gov (United States)

    Gaveglio, Virginia L; Pascual, Ana C; Giusto, Norma M; Pasquaré, Susana J

    2016-08-15

    The aim of this work was to study how age-related changes could modify several enzymatic activities that regulate lipid mediator levels in nuclei from rat cerebellum and how these changes are modulated by all-trans retinoic acid (RA), docosahexaenoic acid (DHA) and arachidonic acid (AA). The higher phosphatidate phosphohydrolase activity and lower diacylglycerol lipase (DAGL) activity observed in aged animals compared with adults could augment diacylglycerol (DAG) availability in the former. Additionally, monoacylglycerol (MAG) availability could be high due to an increase in lysophosphatidate phosphohydrolase (LPAPase) activity and a decrease in monocylglycerol lipase activity. Interestingly, RA, DHA and AA were observed to modulate these enzymatic activities and this modulation was found to change in aged rats. In adult nuclei, whereas RA led to high DAG and MAG production through inhibition of their hydrolytic enzymes, DHA and AA promoted high MAG production by LPAPase and DAGL stimulation. In contrast, in aged nuclei RA caused high MAG generation whereas DHA and AA diminished it through LPAPase activity modulation. These results demonstrate that aging promotes a different nuclear lipid metabolism as well as a different type of non-genomic regulation by RA, DHA and AA, which could be involved in nuclear signaling events. PMID:27355428

  4. PPAR γ Networks in Cell Signaling: Update and Impact of Cyclic Phosphatidic Acid.

    Science.gov (United States)

    Tsukahara, Tamotsu

    2013-01-01

    Lysophospholipid (LPL) has long been recognized as a membrane phospholipid metabolite. Recently, however, the LPL has emerged as a candidate for diagnostic and pharmacological interest. LPLs include lysophosphatidic acid (LPA), alkyl glycerol phosphate (AGP), cyclic phosphatidic acid (cPA), and sphingosine-1-phosphate (S1P). These biologically active lipid mediators serve to promote a variety of responses that include cell proliferation, migration, and survival. These LPL-related responses are mediated by cell surface G-protein-coupled receptors and also intracellular receptor peroxisome proliferator-activated receptor gamma (PPAR γ ). In this paper, we focus mainly on the most recent findings regarding the biological function of nuclear receptor-mediated lysophospholipid signaling in mammalian systems, specifically as they relate to health and diseases. Also, we will briefly review the biology of PPAR γ and then provide an update of lysophospholipids PPAR γ ligands that are under investigation as a therapeutic compound and which are targets of PPAR γ relevant to diseases. PMID:23476786

  5. Discovery and synthetic optimization of a novel scaffold for hydrophobic tunnel-targeted autotaxin inhibition.

    Science.gov (United States)

    Ragle, Lauren E; Palanisamy, Dilip J; Joe, Margaux J; Stein, Rachel S; Norman, Derek D; Tigyi, Gabor; Baker, Daniel L; Parrill, Abby L

    2016-10-01

    Autotaxin (ATX) is a ubiquitous ectoenzyme that hydrolyzes lysophosphatidylcholine (LPC) to form the bioactive lipid mediator lysophosphatidic acid (LPA). LPA activates specific G-protein coupled receptors to elicit downstream effects leading to cellular motility, survival, and invasion. Through these pathways, upregulation of ATX is linked to diseases such as cancer and cardiovascular disease. Recent crystal structures confirm that the catalytic domain of ATX contains multiple binding regions including a polar active site, hydrophobic tunnel, and a hydrophobic pocket. This finding is consistent with the promiscuous nature of ATX hydrolysis of multiple and diverse substrates and prior investigations of inhibitor impacts on ATX enzyme kinetics. The current study used virtual screening methods to guide experimental identification and characterization of inhibitors targeting the hydrophobic region of ATX. An initially discovered inhibitor, GRI392104 (IC50 4μM) was used as a lead for synthetic optimization. In total twelve newly synthesized inhibitors of ATX were more potent than GRI392104 and were selective for ATX as they had no effect on other LPC-specific NPP family members or on LPA1-5 GPCR. PMID:27544588

  6. Sphingolipids in human synovial fluid--a lipidomic study.

    Directory of Open Access Journals (Sweden)

    Marta Krystyna Kosinska

    Full Text Available Articular synovial fluid (SF is a complex mixture of components that regulate nutrition, communication, shock absorption, and lubrication. Alterations in its composition can be pathogenic. This lipidomic investigation aims to quantify the composition of sphingolipids (sphingomyelins, ceramides, and hexosyl- and dihexosylceramides and minor glycerophospholipid species, including (lysophosphatidic acid, (lysophosphatidylglycerol, and bis(monoacylglycerophosphate species, in the SF of knee joints from unaffected controls and from patients with early (eOA and late (lOA stages of osteoarthritis (OA, and rheumatoid arthritis (RA. SF without cells and cellular debris from 9 postmortem donors (control, 18 RA, 17 eOA, and 13 lOA patients were extracted to measure lipid species using electrospray ionization tandem mass spectrometry--directly or coupled with hydrophilic interaction liquid chromatography. We provide a novel, detailed overview of sphingolipid and minor glycerophospholipid species in human SF. A total of 41, 48, and 50 lipid species were significantly increased in eOA, lOA, and RA SF, respectively when compared with normal SF. The level of 21 lipid species differed in eOA SF versus SF from lOA, an observation that can be used to develop biomarkers. Sphingolipids can alter synovial inflammation and the repair responses of damaged joints. Thus, our lipidomic study provides the foundation for studying the biosynthesis and function of lipid species in health and most prevalent joint diseases.

  7. Phosphatidate Kinase, A Novel Enzyme in Phospholipid Metabolism (Characterization of the Enzyme from Suspension-Cultured Catharanthus roseus Cells).

    Science.gov (United States)

    Wissing, J. B.; Kornak, B.; Funke, A.; Riedel, B.

    1994-01-01

    Phosphatidate kinase (adenosine 5[prime]-triphosphate:phosphatidic acid phosphotransferase), a novel enzyme of phospholipid metabolism, was detected recently in the plasma membranes of suspension-cultured Catharanthus roseus cells and purified (J.B. Wissing, H. Behrbohm [1993] Plant Physiol 102: 1243-1249). In the present work the properties of phosphatidate kinase are described. The enzyme showed a pH optimum of 6.1 and an isoelectric point of 4.8, and was rather stable in the presence of its substrates. Although the kinase accepted both ATP and GTP, with Km values of about 12 and 18 [mu]M, respectively, the only lipid substrate was phosphatidic acid; neither lysophosphatidic acid nor any other lipid tested was phosphorylated. With 32P- and 14C-labeled diacylglycerol pyrophosphate, the product of the enzyme, it was shown that the kinase catalyzes a reversible reaction. The activity of the extracted enzyme depended on the presence of surfactants such as Triton X-100 or [beta]-octylglucoside, whereas deoxycholate was strongly inhibitory. Kinetic analysis with Triton X-100/phosphatidate mixed micelles performed according to the "surface dilution" kinetic model showed saturation kinetics with respect to both bulk and surface concentration of phosphatidate. The interfacial Michaelis constant for phosphatidate was determined as 0.6 mol %. PMID:12232252

  8. Glycerol-3-phosphate acyltransferase 4 gene is involved in mouse spermatogenesis

    Institute of Scientific and Technical Information of China (English)

    Qingming Qiu; Gang Liu; Weina Li; Qiuwen Shi; Fuxi Zhu; Guangxiu Lu

    2009-01-01

    Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the first committed step of de novo triacylglycerol syn-thesis by converting glycerol-3-phosphate to lysopho-sphatidic acid (LPA). LPA is a mitogen that mediates multiple cellular processes including cell proliferation. Four GPAT isoforms have been cloned to date. GPAT4 is strongly expressed in the mouse testis. Reverse tran-scription-polymerase chain reaction (PCR), real-time PCR, and in situ hybridization (ISH) were used to analyze the GPAT4 expression and to localize the expressing cell types in the mouse testis during post-natal development. GPAT4 cDNA was inserted into pcDNA4/His to construct a recombinant vector, which was transfected into a mouse spermatogonial cell line (GC-lspg). GPAT4 was first expressed in mice at 2 weeks postnatally. Expression was abundant from the third week, plateaued at week 5-6 and then maintained at a high level in the adult. ISH revealed that GPAT4 gene was expressed abundantly in spermatocytes and around spermatids during meiosis but not in elongated spermatids during later spermiogenesis. GC-1spg cells showed a marked increase in proliferation after trans-fection with GPAT4; cell cycle analysis showed a decrease in the percentage of cells in the Go/G1 phase and an increase in the S phase. Thus, GPAT4 might play an important role in spermatogenesis, especially in mid-meiosis.

  9. Characterization of a small acyl-CoA-binding protein (ACBP) from Helianthus annuus L. and its binding affinities.

    Science.gov (United States)

    Aznar-Moreno, Jose A; Venegas-Calerón, Mónica; Du, Zhi-Yan; Garcés, Rafael; Tanner, Julian A; Chye, Mee-Len; Martínez-Force, Enrique; Salas, Joaquín J

    2016-05-01

    Acyl-CoA-binding proteins (ACBPs) bind to acyl-CoA esters and promote their interaction with other proteins, lipids and cell structures. Small class I ACBPs have been identified in different plants, such as Arabidopsis thaliana (AtACBP6), Brassica napus (BnACBP) and Oryza sativa (OsACBP1, OsACBP2, OsACBP3), and they are capable of binding to different acyl-CoA esters and phospholipids. Here we characterize HaACBP6, a class I ACBP expressed in sunflower (Helianthus annuus) tissues, studying the specificity of its corresponding recombinant HaACBP6 protein towards various acyl-CoA esters and phospholipids in vitro, particularly using isothermal titration calorimetry and protein phospholipid binding assays. This protein binds with high affinity to de novo synthetized derivatives palmitoly-CoA, stearoyl-CoA and oleoyl-CoA (Kd 0.29, 0.14 and 0.15 μM respectively). On the contrary, it showed lower affinity towards linoleoyl-CoA (Kd 5.6 μM). Moreover, rHaACBP6 binds to different phosphatidylcholine species (dipalmitoyl-PC, dioleoyl-PC and dilinoleoyl-PC), yet it displays no affinity towards other phospholipids like lyso-PC, phosphatidic acid and lysophosphatidic acid derivatives. In the light of these results, the possible involvement of this protein in sunflower oil synthesis is considered. PMID:26938582

  10. Lysophospholipid receptors are differentially expressed in rat terminal Schwann cells, as revealed by a single cell rt-PCR and in situ hybridization.

    Science.gov (United States)

    Kobashi, Hiroaki; Yaoi, Takeshi; Oda, Ryo; Okajima, Seiichiro; Fujiwara, Hiroyoshi; Kubo, Toshikazu; Fushiki, Shinji

    2006-04-22

    Terminal Schwann cells (TSCs) that cover motor neuron terminals, are known to play an important role in maintaining neuromuscular junctions, as well as in the repair process after nerve injury. However, the molecular characteristics of TSCs remain unknown, because of the difficulties in analyzing them due to their paucity. By using our previously reported method of selectively and efficiently collecting TSCs, we have analyzed the difference in expression patterns of lysophospholipid (LPL) receptor genes (LPA1, LPA2, LPA3, S1P1, S1P2, S1P3, S1P4, and S1P5) between TSCs and myelinating Schwann cells (MSCs). LPL, which includes lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P), is the bioactive lipid that induces a myriad of cellular responses through specific members of G-protein coupled receptors for LPA. It turned out that LPA3 was expressed only in TSCs, whereas S1P1 was expressed in TSCs and skeletal muscle, but not in MSCs. Other types of LPL receptor genes, including LPA1, S1P2, S1P3, S1P4, were expressed in both types of Schwann cells. None of the LPL receptor gene family showed MSCs-specific expression. PMID:17375210

  11. Role of mitochondrial lipids in guiding fission and fusion.

    Science.gov (United States)

    Frohman, Michael A

    2015-03-01

    Clinically important links have been established between mitochondrial function and cardiac physiology and disease in the context of signaling mechanisms, energy production, and muscle cell development. The proteins and processes that drive mitochondrial fusion and fission are now known to have emergent functions in intracellular calcium homeostasis, apoptosis, vascular smooth muscle cell proliferation, myofibril organization, and Notch-driven cell differentiation, all key issues in cardiac disease. Moreover, decreasing fission may confer protection against ischemic heart disease, particularly in the setting of obesity, diabetes, and heart failure. The importance of lipids in controlling mitochondrial fission and fusion is increasingly becoming appreciated. Roles for the bulk and signaling lipids cardiolipin, phosphatidylethanolamine, phosphatidic acid, diacylglycerol, and lysophosphatidic acid and the enzymes that synthesize or metabolize them in the control of mitochondrial shape and function are reviewed here. A number of diseases have been linked to loss-of-function alleles for a subset of the enzymes, emphasizing the importance of the lipid environment in this context.

  12. Compartmentation of hepatic fatty-acid-binding protein in liver cells and its effect on microsomal phosphatidic acid biosynthesis.

    Science.gov (United States)

    Bordewick, U; Heese, M; Börchers, T; Robenek, H; Spener, F

    1989-03-01

    Fatty-acid-binding proteins are known to occur in the cytosol of mammalian cells and to bind fatty acids and their CoA-esters. Application of the postembedding protein A-gold labeling method with antibody against the hepatic type fatty-acid-binding protein (hFABP) to cross-sections of liver cells and a newly developed gel-chromatographic immunofluorescence assay established qualitatively (1) that hFABP in mitochondria was confined to outer mitochondrial membranes, (2) the presence of this protein in microsomes and (3) that nuclei were also filled with hFABP. Quantitative data elaborated with a non-competitive ELISA confirmed these results. A significant difference to the distribution of cardiac FABP in heart muscle cells, where this type of protein was found in cytosol, matrix and nuclei, was observed (Börchers et al. (1989) Biochim. Biophys. Acta, in the press). hFABP-containing rat liver microsomes were incubated with long-chain acyl-CoAs in the presence of hFABP (isolated from rat liver cytosol) in a study on the acylation of sn-glycerol-3-phosphate and lysophosphatidic acid. Both acyltransferases were stimulated by addition of hFABP to the incubation medium. The morphological, immunochemical as well as kinetic data infer a direct interaction of hFABP with microsomal membranes in liver cells.

  13. Segregation of Incomplete Achromatopsia and Alopecia Due to PDE6H and LPAR6 Variants in a Consanguineous Family from Pakistan

    Science.gov (United States)

    Pedurupillay, Christeen Ramane J.; Landsend, Erlend Christoffer Sommer; Vigeland, Magnus Dehli; Ansar, Muhammad; Frengen, Eirik; Misceo, Doriana; Strømme, Petter

    2016-01-01

    We report on two brothers with visual impairment, and non-syndromic alopecia in the elder proband. The parents were first-degree Pakistani cousins. Whole exome sequencing of the elder brother and parents, followed by Sanger sequencing of all four family members, led to the identification of the variants responsible for the two phenotypes. One variant was a homozygous nonsense variant in the inhibitory subunit of the cone-specific cGMP phosphodiesterase gene, PDE6H:c.35C>G (p.Ser12*). PDE6H is expressed in the cones of the retina, which are involved in perception of color vision. This is the second report of a homozygous PDE6H:c.35C>G variant causing incomplete achromatopsia (OMIM 610024), thus strongly supporting the hypothesis that loss-of-function variants in PDE6H cause this visual deficiency phenotype. The second variant was a homozygous missense substitution in the lysophosphatidic acid receptor 6, LPAR6:c.188A>T (p.Asp63Val). LPAR6 acts as a G-protein-coupled receptor involved in hair growth. Biallelic loss-of-function variants in LPAR6 cause hypotrichosis type 8 (OMIM 278150), with or without woolly hair, a form of non-syndromic alopecia. Biallelic LPAR6:c.188A>T was previously described in five families from Pakistan. PMID:27472364

  14. Arabidopsis non-specific phospholipase C1: Characterisation and its involvement in response to heat stress

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    Zuzana eKrčková

    2015-11-01

    Full Text Available The Arabidopsis non-specific phospholipase C (NPC protein family is encoded by the genes NPC1 – NPC6. It has been shown that NPC4 and NPC5 possess phospholipase C activity; NPC3 has lysophosphatidic acid phosphatase activity. NPC3, 4 and 5 play roles in the responses to hormones and abiotic stresses. NPC1, 2 and 6 has not been studied functionally yet.We found that Arabidopsis NPC1 expressed in E. coli possesses phospholipase C activity in vitro. This protein was able to hydrolyse phosphatidylcholine to diacylglycerol. NPC1-green fluorescent protein was localized to secretory pathway compartments in Arabidopsis roots. In the knock out T-DNA insertion line NPC1 (npc1 basal thermotolerance was impaired compared with wild-type; npc1 exhibited significant decreases in survival rate and chlorophyll content at the seventh day after heat stress. Conversely, plants overexpressing NPC1 (NPC1-OE were more resistant to heat stress compared with wild-type. These findings suggest that NPC1 is involved in the plant response to heat

  15. HIV-1 Tat Inhibits Autotaxin Lysophospholipase D Activity and Modulates Oligodendrocyte Differentiation

    Science.gov (United States)

    Wheeler, Natalie A.; Fuss, Babette; Knapp, Pamela E.

    2016-01-01

    White matter injury has been frequently reported in HIV+ patients. Previous studies showed that HIV-1 Tat (transactivator of transcription), a viral protein that is produced and secreted by HIV-infected cells, is toxic to young, immature oligodendrocytes (OLGs). Adding Tat to the culture medium reduced the viability of immature OLGs, and the surviving OLGs exhibited reduced process networks. OLGs produce and secrete autotaxin (ATX), an ecto-enzyme containing a lysophospholipase D (lysoPLD) activity that converts lysophosphatidylcholine (LPC) to lysophosphatidic acid (LPA), a lipid signaling molecule that stimulates OLG differentiation. We hypothesized that Tat affects OLG development by interfering with the ATX-LPA signaling pathway. Our data show that Tat treatment leads to changes in the expression of OLG differentiation genes and the area of OLG process networks, both of which can be rescued by LPA. Tat-treated OLGs showed no change in LPA receptor expression but significantly decreased extracellular ATX levels and lysoPLD activity. In Tat transgenic mice, expression of Tat in vivo leads to decreased OLG ATX secretion. Furthermore, co-immunoprecipitation experiments revealed a potential physical interaction between Tat and ATX. Together, these data strongly suggest two functional implications of Tat blocking ATX’s lysoPLD activity. On one hand, it attenuates OLG differentiation, and on the other hand it interferes with the protective effects of LPA on OLG process morphology. PMID:27659560

  16. Requirement of Osteopontin in the migration and protection against Taxol-induced apoptosis via the ATX-LPA axis in SGC7901 cells

    Directory of Open Access Journals (Sweden)

    Huang Zuhu

    2011-03-01

    Full Text Available Abstract Background Autotaxin (ATX possesses lysophospholipase D (lyso PLD activity, which converts lysophosphatidylcholine (LPC into lysophosphatidic acid (LPA. The ATX-LPA signaling axis has been implicated in angiogenesis, chronic inflammation and tumor progression. Osteopontin (OPN is an important chemokine involved in the survival, proliferation, migration, invasion and metastasis of gastric cancer cells. The focus of the present study was to investigate the relationship between the ATX-LPA axis and OPN. Results In comparison with non-treated cells, we found that the ATX-LPA axis up-regulated OPN expression by 1.92-fold in protein levels and 1.3-fold in mRNA levels. The ATX-LPA axis activates LPA2, Akt, ERK and ELK-1 and also protects SGC7901 cells from apoptosis induced by Taxol treatment. Conclusions This study provides the first evidence that expression of OPN induced by ATX-LPA axis is mediated by the activation of Akt and MAPK/ERK pathways through the LPA2 receptor. In addition, OPN is required for the protective effects of ATX-LPA against Taxol-induced apoptosis and ATX-LPA-induced migration of SGC7901 cells.

  17. Targeting the myofibroblast genetic switch: inhibitors of myocardin-related transcription factor/serum response factor-regulated gene transcription prevent fibrosis in a murine model of skin injury.

    Science.gov (United States)

    Haak, Andrew J; Tsou, Pei-Suen; Amin, Mohammad A; Ruth, Jeffrey H; Campbell, Phillip; Fox, David A; Khanna, Dinesh; Larsen, Scott D; Neubig, Richard R

    2014-06-01

    Systemic sclerosis (SSc), or scleroderma, similar to many fibrotic disorders, lacks effective therapies. Current trials focus on anti-inflammatory drugs or targeted approaches aimed at one of the many receptor mechanisms initiating fibrosis. In light of evidence that a myocardin-related transcription factor (MRTF)-and serum response factor (SRF)-regulated gene transcriptional program induced by Rho GTPases is essential for myofibroblast activation, we explored the hypothesis that inhibitors of this pathway may represent novel antifibrotics. MRTF/SRF-regulated genes show spontaneously increased expression in primary dermal fibroblasts from patients with diffuse cutaneous SSc. A novel small-molecule inhibitor of MRTF/SRF-regulated transcription (CCG-203971) inhibits expression of connective tissue growth factor (CTGF), α-smooth muscle actin (α-SMA), and collagen 1 (COL1A2) in both SSc fibroblasts and in lysophosphatidic acid (LPA)-and transforming growth factor β (TGFβ)-stimulated fibroblasts. In vivo treatment with CCG-203971 also prevented bleomycin-induced skin thickening and collagen deposition. Thus, targeting the MRTF/SRF gene transcription pathway could provide an efficacious new approach to therapy for SSc and other fibrotic disorders.

  18. Molecular cloning, sequence characterization, and gene expression profiling of a novel water buffalo (Bubalus bubalis) gene, AGPAT6.

    Science.gov (United States)

    Song, S; Huo, J L; Li, D L; Yuan, Y Y; Yuan, F; Miao, Y W

    2013-01-01

    Several 1-acylglycerol-3-phosphate-O-acyltransferases (AGPATs) can acylate lysophosphatidic acid to produce phosphatidic acid. Of the eight AGPAT isoforms, AGPAT6 is a crucial enzyme for glycerolipids and triacylglycerol biosynthesis in some mammalian tissues. We amplified and identified the complete coding sequence (CDS) of the water buffalo AGPAT6 gene by using the reverse transcription-polymerase chain reaction, based on the conversed sequence information of the cattle or expressed sequence tags of other Bovidae species. This novel gene was deposited in the NCBI database (accession No. JX518941). Sequence analysis revealed that the CDS of this AGPAT6 encodes a 456-amino acid enzyme (molecular mass = 52 kDa; pI = 9.34). Water buffalo AGPAT6 contains three hydrophobic transmembrane regions and a signal 37-amino acid peptide, localized in the cytoplasm. The deduced amino acid sequences share 99, 98, 98, 97, 98, 98, 97 and 95% identity with their homologous sequences from cattle, horse, human, mouse, orangutan, pig, rat, and chicken, respectively. The phylogenetic tree analysis based on the AGPAT6 CDS showed that water buffalo has a closer genetic relationship with cattle than with other species. Tissue expression profile analysis shows that this gene is highly expressed in the mammary gland, moderately expressed in the heart, muscle, liver, and brain; weakly expressed in the pituitary gland, spleen, and lung; and almost silently expressed in the small intestine, skin, kidney, and adipose tissues. Four predicted microRNA target sites are found in the water buffalo AGPAT6 CDS. These results will establish a foundation for further insights into this novel water buffalo gene. PMID:24114207

  19. Mitochondrial citrate synthase crystals: novel finding in Sengers syndrome caused by acylglycerol kinase (AGK) mutations.

    Science.gov (United States)

    Siriwardena, Komudi; Mackay, Nevena; Levandovskiy, Valeriy; Blaser, Susan; Raiman, Julian; Kantor, Paul F; Ackerley, Cameron; Robinson, Brian H; Schulze, Andreas; Cameron, Jessie M

    2013-01-01

    We report on two families with Sengers syndrome and mutations in the acylglycerol kinase gene (AGK). In the first family, two brothers presented with vascular strokes, lactic acidosis, cardiomyopathy and cataracts, abnormal muscle cell histopathology and mitochondrial function. One proband had very abnormal mitochondria with citrate synthase crystals visible in electron micrographs, associated with markedly high citrate synthase activity. Exome sequencing was used to identify mutations in the AGK gene in the index patient. Targeted sequencing confirmed the same homozygous mutation (c.3G>A, p.M1I) in the brother. The second family had four affected members, of which we examined two. They also presented with similar clinical symptoms, but no strokes. Postmortem heart and skeletal muscle tissues showed low complex I, III and IV activities in the heart, but normal in the muscle. Skin fibroblasts showed elevated lactate/pyruvate ratios and low complex I+III activity. Targeted sequencing led to identification of a homozygous c.979A>T, p.K327* mutation. AGK is located in the mitochondria and phosphorylates monoacylglycerol and diacylglycerol to lysophosphatidic acid and phosphatidic acid. Disruption of these signaling molecules affects the mitochondria's response to superoxide radicals, resulting in oxidative damage to mitochondrial DNA, lipids and proteins, and stimulation of cellular detoxification pathways. High levels of manganese superoxide dismutase protein were detected in all four affected individuals, consistent with increased free radical damage. Phosphatidic acid is also involved in the synthesis of phospholipids and its loss will result in changes to the lipid composition of the inner mitochondrial membrane. These effects manifest as cataract formation in the eye, respiratory chain dysfunction and cardiac hypertrophy in heart tissue. These two pedigrees confirm that mutation of AGK is responsible for the severe neonatal presentation of Sengers syndrome. The

  20. High specificity of human secretory class II phospholipase A2 for phosphatidic acid.

    Science.gov (United States)

    Snitko, Y; Yoon, E T; Cho, W

    1997-02-01

    Lysophosphatidic acid (LPA) is a potent lipid second messenger which stimulates platelet aggregation, cell proliferation and smooth-muscle contraction. The phospholipase A2 (PLA2)-catalysed hydrolysis of phosphatidic acid (PA) is thought to be a primary synthetic route for LPA. Of the multiple forms of PLA2 present in human tissues, human secretory class-II PLA2 (hs-PLA2) has been implicated in the production of LPA from platelets and whole blood cells challenged with inflammatory stimuli. To explore further the possibility that hs-PLA2 is involved in the production of LPA, we rigorously measured the phospholipid head group specificity of hs-PLA2 by a novel PLA2 kinetic system using polymerized mixed liposomes. Kinetic analysis of recombinant hs-PLA2 demonstrates that hs-PLA2 strongly prefers PA as substrate over other phospholipids found in the mammalian plasma membrane including phosphatidylserine (PS), phosphatidylcholine (PC) and phosphatidylethanolamine (PE). The order of preference is PA > PE approximately PS > PC. To identify amino acid residues of hs-PLA2 that are involved in its unique substrate specificity, we mutated two residues, Glu-56 and Lys-69, which were shown to interact with the phospholipid head group in the X-ray-crystallographic structure of the hs-PLA2-transition-state-analogue complex. The K69Y mutant showed selective inactivation toward PA whereas the E56K mutant displayed a most pronounced inactivation to PE. Thus it appears that Lys-69 is at least partially involved in the PA specificity of hs-PLA2 and Glu-56 in the distinction between PE and PC. In conjunction with a recent cell study [Fourcade, Simon, Viode, Rugani, Leballe, Ragab, Fournie, Sarda and Chap (1995) Cell 80, 919-927], these studies suggest that hs-PLA2 can rapidly hydrolyse PA molecules exposed to the outer layer of cell-derived microvesicles and thereby produce LPA.

  1. Molecular cloning of magnesium-independent type 2 phosphatidic acid phosphatases from airway smooth muscle.

    Science.gov (United States)

    Tate, R J; Tolan, D; Pyne, S

    1999-07-01

    Members of the type 2 phosphatidic acid phosphatase (PAP2) family catalyse the dephosphorylation of phosphatidic acid (PA), lysophosphatidate and sphingosine 1-phosphate. Here, we demonstrate the presence of a Mg(2+)-independent and N-ethymaleimide-insensitive PAP2 activity in cultured guinea-pig airway smooth muscle (ASM) cells. Two PAP2 cDNAs of 923 and 926 base pairs were identified and subsequently cloned from these cells. The ORF of the 923 base pair cDNA encoded a protein of 285 amino acids (Mr = 32.1 kDa), which had 94% homology with human PAP2a (hPAP2a) and which probably represents a guinea-pig specific PAP2a (gpPAP2a1). The ORF of the 926 base pair cDNA encoded a protein of 286 amino acids (Mr = 32.1 kDa) which had 84% and 91% homology with hPAP2a and gpPAP2a1, respectively. This protein, termed gpPAP2a2, has two regions (aa 21-33 and 51-74) of marked divergence and altered hydrophobicity compared with hPAP2a and gpPAP2a1. This occurs in the predicted first and second transmembrane domains and at the extremes of the first outer loop. Other significant differences between gpPAP2a1/2 and hPAP2a, hPAP2b and hPAP2c occur at the cytoplasmic C-terminal. Transient expression of gpPAP2a2 in Cos-7 cells resulted in an approx. 4-fold increase in Mg(2+)-independent PAP activity, thereby confirming that gpPAP2a2 is another catalytically active member of an extended PAP2 family.

  2. Pinoresinol-4,4'-di-O-beta-D-glucoside from Valeriana officinalis root stimulates calcium mobilization and chemotactic migration of mouse embryo fibroblasts.

    Science.gov (United States)

    Do, Kee Hun; Choi, Young Whan; Kim, Eun Kyoung; Yun, Sung Ji; Kim, Min Sung; Lee, Sun Young; Ha, Jung Min; Kim, Jae Ho; Kim, Chi Dae; Son, Beung Gu; Kang, Jum Soon; Khan, Ikhlas A; Bae, Sun Sik

    2009-06-01

    Lignans are major constituents of plant extracts and have important pharmacological effects on mammalian cells. Here we showed that pinoresinol-4,4'-di-O-beta-D-glucoside (PDG) from Valeriana officinalis induced calcium mobilization and cell migration through the activation of lysophosphatidic acid (LPA) receptor subtypes. Stimulation of mouse embryo fibroblast (MEF) cells with 10 microM PDG resulted in strong stimulation of MEF cell migration and the EC(50) was about 2 microM. Pretreatment with pertussis toxin (PTX), an inhibitor of G(i) protein, completely blocked PDG-induced cell migration demonstrating that PDG evokes MEF cell migration through the activation of the G(i)-coupled receptor. Furthermore, pretreatment of MEF cells with Ki16425 (10 microM), which is a selective antagonist for LPA(1) and LPA(3) receptors, completely blocked PDG-induced cell migration. Likewise, PDG strongly induced calcium mobilization, which was also blocked by Ki16425 in a dose-dependent manner. Prior occupation of the LPA receptor with LPA itself completely blocked PDG-induced calcium mobilization. Finally, PDG-induced MEF cell migration was attenuated by pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor such as LY294002. Cells lacking downstream mediator of PI3K such as Akt1 and Akt2 (DKO cells) showed loss of PDG-induced migration. Re-expression of Akt1 (but not Akt2) completely restored PDG-induced DKO cell migration. Given these results, we conclude that PDG is a strong inducer of cell migration. We suggest that the pharmacological action of PDG may occur through the activation of an LPA receptor whereby activation of PI3K/Akt signaling pathway mediates PDG-induced MEF cell migration.

  3. Kaempferol suppresses lipid accumulation by inhibiting early adipogenesis in 3T3-L1 cells and zebrafish.

    Science.gov (United States)

    Lee, Yeon-Joo; Choi, Hyeon-Son; Seo, Min-Jung; Jeon, Hui-Jeon; Kim, Kui-Jin; Lee, Boo-Yong

    2015-08-01

    Kaempferol is a flavonoid present in Kaempferia galanga and Opuntia ficus indica var. saboten. Recent studies have suggested that it has anti-oxidant, anti-inflammatory, anti-cancer, and anti-obesity effects. In this study, we focused on the anti-adipogenic effects of kaempferol during adipocyte differentiation. The results showed that kaempferol inhibits lipid accumulation in adipocytes and zebrafish. Oil Red O and Nile Red staining showed that the number of intracellular lipid droplets decreased in adipocytes and zebrafish treated with kaempferol. LPAATθ (lysophosphatidic acid acyltransferase), lipin1, and DGAT1 (triglyceride synthetic enzymes) and FASN and SREBP-1C (fatty acid synthetic proteins) showed decreased expression levels in the presence of kaempferol. In addition, treatment of kaempferol showed an inhibitory activity on cell cycle progression. Kaempferol delayed cell cycle progression from the S to G2/M phase through the regulation of cyclins in a dose-dependent manner. Kaempferol blocked the phosphorylation of AKT (protein kinase B) and mammalian target of rapamycin (mTOR) signaling pathway during the early stages of adipogenesis. In addition, kaempferol down-regulated pro-early adipogenic factors such as CCAAT-enhancer binding proteins β (C/EBPβ), and Krüppel-like factors (KLFs) 4 and 5, while anti-early adipogenic factors, such as KLF2 and pref-1(preadipocyte factor-1), were upregulated. These kaempferol-mediated regulations of early adipogenic factors resulted in the attenuation of late adipogenic factors such as C/EBPα and peroxisome proliferator-activated receptor γ (PPARγ). These results were supported in zebrafish based on the decrease in lipid accumulation and expression of adipogenic factors. Our results indicated that kaempferol might have an anti-obesity effect by regulating lipid metabolism. PMID:26174858

  4. NT-3 protein levels are enhanced in the hippocampus of PRG1-deficient mice but remain unchanged in PRG1/LPA2 double mutants.

    Science.gov (United States)

    Petzold, Sandra; Sommer, Babette; Kröber, Andrea; Nitsch, Robert; Schwegler, Herbert; Vogt, Johannes; Roskoden, Thomas

    2016-01-26

    The plasticity-related gene 1 (PRG1) modulates bioactive lipids at the postsynaptic density and is a novel player in neuronal plasticity and regulation of glutamatergic transmission at principal neurons. PRG1, a neuronal molecule, is highly expressed during development and regeneration processes at the postsynaptic density, modulates synaptic lysophosphatidic acid (LPA) levels and is related to epilepsy and brain injury. In the present study, we analyzed the interaction between the synaptic molecules PRG1 and LPA2R with other plasticity-related molecules the neurotrophins. The protein levels of NGF, BDNF and NT-3 were measured using ELISA in hippocampal tissue of homozygous (PRG(-/-)) and heterozygous (PRG(+/-)) PRG1 deficient mice and compared to their wild type (PRG(+/+)/WT) littermates. In the hippocampus, protein levels of NT-3 were significantly increased in PRG(-/-) mice (compared to WT-litters) while protein levels of NGF and BDNF were not affected. Since PRG1 deficiency leads to increased neuronal excitability and higher hippocampal network activity, which may well influence neurotrophin levels, we further assessed PRG1 deficient mice on an LPA2-receptor (LPA2R) deficient background, reported to normalize hippocampal over-excitability in PRG1(-/-) mice. However, on an LPA2R deficient background, protein levels of NT-3 in PRG1(-/-) mice (PRG1(-/-)/LPA2R(-/-)) were not significantly different when compared to WT animals. Since PRG1 deficient mice showed over-excitability in glutamatergic neurons. This was normalized by additional LPA2R deletion, and we conclude the increased NT3-levels were directly or indirectly attributable to increased hippocampal network activity, possibly exerting a protective effect against over-excitability.

  5. Pathogenesis and Management of Pruritus in PBC and PSC.

    Science.gov (United States)

    Kremer, Andreas E; Namer, Barbara; Bolier, Ruth; Fischer, Michael J; Oude Elferink, Ronald P; Beuers, Ulrich

    2015-01-01

    Pruritus is a preeminent symptom in patients with chronic cholestatic liver disorders such as primary biliary cirrhosis and primary sclerosing cholangitis. More than two-thirds of these patients experience itching during the course of their disease. This symptom is also frequently observed in patients with other causes of cholestasis such as cholangiocarcinoma, inherited forms of cholestasis and intrahepatic cholestasis of pregnancy, but may accompany almost any other liver disease. The pathogenesis of pruritus of cholestasis remains largely elusive. Increased concentrations of bile salts, histamine, serotonin, progesterone metabolites and endogenous opioids have been controversially discussed as potential pruritogens. However, for these molecules, neither a correlation with itch intensity nor a causative link could be established. The G protein-coupled receptor for bile salts, TGR5, has been shown to be expressed in dorsal root ganglia and give rise to itch in rodents, albeit upon stimuli with suprapathological concentrations of bile salts. The potent neuronal activator lysophosphatidic acid (LPA) and its forming enzyme, autotaxin (ATX), could be identified in the serum of patients with cholestatic pruritus. ATX activity correlated with itch severity and effectiveness of several anti-pruritic therapeutic interventions in cholestatic patients. Thus, the ATX-LPA-axis may represent a key element in the pathogenesis of this agonizing symptom. Treatment options for pruritus of cholestasis remain limited to a few evidence-based and several experimental medical and interventional therapies. The current guideline-based recommendations include the anion exchange resins colestyramine, the pregnane X receptor-agonist and enzyme inducer rifampicin, the μ-opioid antagonist naltrexone, and the selective serotonin reuptake inhibitors sertraline. Still, a considerable part of patients is unresponsive to these drugs and requires experimental approaches including phototherapy

  6. Enhancement of endothelial cell migration by constitutively active LPA{sub 1}-expressing tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Kitayoshi, Misaho; Kato, Kohei; Tanabe, Eriko; Yoshikawa, Kyohei; Fukui, Rie [Division of Cancer Biology and Bioinformatics, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Fukushima, Nobuyuki [Division of Molecular Neurobiology, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan); Tsujiuchi, Toshifumi, E-mail: ttujiuch@life.kindai.ac.jp [Division of Cancer Biology and Bioinformatics, Department of Life Science, Faculty of Science and Engineering, Kinki University, 3-4-1, Kowakae, Higashiosaka, Osaka 577-8502 (Japan)

    2012-06-01

    Highlights: Black-Right-Pointing-Pointer Mutated LPA{sub 1} stimulates cell migration of endothelial cells. Black-Right-Pointing-Pointer VEGF expressions are increased by mutated LPA{sub 1}. Black-Right-Pointing-Pointer LPA signaling via mutated LPA{sub 1} is involved in angiogenesis. Black-Right-Pointing-Pointer Mutated LPA{sub 1} promotes cancer cell progression. -- Abstract: Lysophosphatidic acid (LPA) receptors belong to G protein-coupled transmembrane receptors (LPA receptors; LPA{sub 1} to LPA{sub 6}). They indicate a variety of cellular response by the interaction with LPA, including cell proliferation, migration and differentiation. Recently, we have reported that constitutive active mutated LPA{sub 1} induced the strong biological effects of rat neuroblastoma B103 cells. In the present study, we examined the effects of mutated LPA{sub 1} on the interaction between B103 cells and endothelial F-2 cells. Each LPA receptor expressing B103 cells were maintained in serum-free DMEM and cell motility assay was performed with a Cell Culture Insert. When F-2 cells were cultured with conditioned medium from Lpar1 and Lpar3-expressing cells, the cell motility of F-2 cells was significantly higher than control cells. Interestingly, the motile activity of F-2 cells was strongly induced by mutated LPA{sub 1} than other cells, correlating with the expression levels of vascular endothelial growth factor (Vegf)-A and Vegf-C. Pretreatment of LPA signaling inhibitors inhibited F-2 cell motility stimulated by mutated LPA{sub 1}. These results suggest that activation of LPA signaling via mutated LPA{sub 1} may play an important role in the promotion of angiogenesis in rat neuroblastoma cells.

  7. Understanding the local actions of lipids in bone physiology.

    Science.gov (United States)

    During, Alexandrine; Penel, Guillaume; Hardouin, Pierre

    2015-07-01

    The adult skeleton is a metabolically active organ system that undergoes continuous remodeling to remove old and/or stressed bone (resorption) and replace it with new bone (formation) in order to maintain a constant bone mass and preserve bone strength from micro-damage accumulation. In that remodeling process, cellular balances--adipocytogenesis/osteoblastogenesis and osteoblastogenesis/osteoclastogenesis--are critical and tightly controlled by many factors, including lipids as discussed in the present review. Interest in the bone lipid area has increased as a result of in vivo evidences indicating a reciprocal relationship between bone mass and marrow adiposity. Lipids in bones are usually assumed to be present only in the bone marrow. However, the mineralized bone tissue itself also contains small amounts of lipids which might play an important role in bone physiology. Fatty acids, cholesterol, phospholipids and several endogenous metabolites (i.e., prostaglandins, oxysterols) have been purported to act on bone cell survival and functions, the bone mineralization process, and critical signaling pathways. Thus, they can be regarded as regulatory molecules important in bone health. Recently, several specific lipids derived from membrane phospholipids (i.e., sphingosine-1-phosphate, lysophosphatidic acid and different fatty acid amides) have emerged as important mediators in bone physiology and the number of such molecules will probably increase in the near future. The present paper reviews the current knowledge about: (1°) bone lipid composition in both bone marrow and mineralized tissue compartments, and (2°) local actions of lipids on bone physiology in relation to their metabolism. Understanding the roles of lipids in bone is essential to knowing how an imbalance in their signaling pathways might contribute to bone pathologies, such as osteoporosis. PMID:26118851

  8. Targeting melanoma growth and viability reveals dualistic functionality of the phosphonothionate analogue of carba cyclic phosphatidic acid

    Directory of Open Access Journals (Sweden)

    Prestwich Glenn D

    2010-06-01

    Full Text Available Abstract Background Although the incidence of melanoma in the U.S. is rising faster than any other cancer, the FDA-approved chemotherapies lack efficacy for advanced disease, which results in poor overall survival. Lysophosphatidic acid (LPA, autotaxin (ATX, the enzyme that produces LPA, and the LPA receptors represent an emerging group of therapeutic targets in cancer, although it is not known which of these is most effective. Results Herein we demonstrate that thio-ccPA 18:1, a stabilized phosphonothionate analogue of carba cyclic phosphatidic acid, ATX inhibitor and LPA1/3 receptor antagonist, induced a marked reduction in the viability of B16F10 metastatic melanoma cells compared with PBS-treated control by 80-100%. Exogenous LPA 18:1 or D-sn-1-O-oleoyl-2-O-methylglyceryl-3-phosphothioate did not reverse the effect of thio-ccPA 18:1. The reduction in viability mediated by thio-ccPA 18:1 was also observed in A375 and MeWo melanoma cell lines, suggesting that the effects are generalizable. Interestingly, siRNA to LPA3 (siLPA3 but not other LPA receptors recapitulated the effects of thio-ccPA 18:1 on viability, suggesting that inhibition of the LPA3 receptor is an important dualistic function of the compound. In addition, siLPA3 reduced proliferation, plasma membrane integrity and altered morphology of A375 cells. Another experimental compound designed to antagonize the LPA1/3 receptors significantly reduced viability in MeWo cells, which predominantly express the LPA3 receptor. Conclusions Thus the ability of thio-ccPA 18:1 to inhibit the LPA3 receptor and ATX are key to its molecular mechanism, particularly in melanoma cells that predominantly express the LPA3 receptor. These observations necessitate further exploration and exploitation of these targets in melanoma.

  9. To the Root of the Curl: A Signature of a Recent Selective Sweep Identifies a Mutation That Defines the Cornish Rex Cat Breed.

    Directory of Open Access Journals (Sweden)

    Barbara Gandolfi

    Full Text Available The cat (Felis silvestris catus shows significant variation in pelage, morphological, and behavioral phenotypes amongst its over 40 domesticated breeds. The majority of the breed specific phenotypic presentations originated through artificial selection, especially on desired novel phenotypic characteristics that arose only a few hundred years ago. Variations in coat texture and color of hair often delineate breeds amongst domestic animals. Although the genetic basis of several feline coat colors and hair lengths are characterized, less is known about the genes influencing variation in coat growth and texture, especially rexoid - curly coated types. Cornish Rex is a cat breed defined by a fixed recessive curly coat trait. Genome-wide analyses for selection (di, Tajima's D and nucleotide diversity were performed in the Cornish Rex breed and in 11 phenotypically diverse breeds and two random bred populations. Approximately 63K SNPs were used in the analysis that aimed to localize the locus controlling the rexoid hair texture. A region with a strong signature of recent selective sweep was identified in the Cornish Rex breed on chromosome A1, as well as a consensus block of homozygosity that spans approximately 3 Mb. Inspection of the region for candidate genes led to the identification of the lysophosphatidic acid receptor 6 (LPAR6. A 4 bp deletion in exon 5, c.250_253_delTTTG, which induces a premature stop codon in the receptor, was identified via Sanger sequencing. The mutation is fixed in Cornish Rex, absent in all straight haired cats analyzed, and is also segregating in the German Rex breed. LPAR6 encodes a G protein-coupled receptor essential for maintaining the structural integrity of the hair shaft; and has mutations resulting in a wooly hair phenotype in humans.

  10. Localization and translocation of RhoA protein in the human gastric cancer cell line SGC-7901

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To elucidate the localization of RhoA in gastric SGC-7901 cancer cells and its translocation by lysophosphatidic acid (LPA) and/or 8-chlorophenylthio cAMP (CPT-cAMP). METHODS: Immunofluorescence microscopy was used to determine the localization of RhoA. Western blotting was used to detect both endogenous and exogenous RhoA in different cellular compartments (membrane, cytosol, nucleus) and the translocation of RhoA following treatment with LPA, CPT-cAMP, or CPT-cAMP+LPA. RESULTS: Immunofluorescence staining revealed endogenous RhoA to be localized in the membrane, the cytosol, and the nucleus, and its precise localization within the nucleus to be the nucleolus. Western blotting identified both endogenous and exogenous RhoA within different cellular compartments (membrane, cytosol, nucleus, nucleolus). After stimulation with LPA, the amount of RhoA within membrane and nuclear extracts increased, while it decreased in the cytosol fractions. After treatment with CPT-cAMP the amount of RhoA within the membrane and the nuclear extracts decreased, while it increased within the cytosol fraction. Treatment with a combination of both substances led to a decrease in RhoA in the membrane and the nucleus but to an increase in the cytosol. CONCLUSION: In SGC-7901 cells RhoA was found to be localized within the membrane, the cytosol, and the nucleus. Within the nucleus its precise localization could be demonstrated to be the nucleolus. Stimulation with LPA caused a translocation of RhoA from the cytosol towards the membrane and the nucleus; treatment with CPT-cAMP caused the opposite effect. Furthermore, pre-treatment with CPT-cAMP was found to block the effect of LPA.

  11. Human neural progenitors express functional lysophospholipid receptors that regulate cell growth and morphology

    Directory of Open Access Journals (Sweden)

    Callihan Phillip

    2008-12-01

    Full Text Available Abstract Background Lysophospholipids regulate the morphology and growth of neurons, neural cell lines, and neural progenitors. A stable human neural progenitor cell line is not currently available in which to study the role of lysophospholipids in human neural development. We recently established a stable, adherent human embryonic stem cell-derived neuroepithelial (hES-NEP cell line which recapitulates morphological and phenotypic features of neural progenitor cells isolated from fetal tissue. The goal of this study was to determine if hES-NEP cells express functional lysophospholipid receptors, and if activation of these receptors mediates cellular responses critical for neural development. Results Our results demonstrate that Lysophosphatidic Acid (LPA and Sphingosine-1-phosphate (S1P receptors are functionally expressed in hES-NEP cells and are coupled to multiple cellular signaling pathways. We have shown that transcript levels for S1P1 receptor increased significantly in the transition from embryonic stem cell to hES-NEP. hES-NEP cells express LPA and S1P receptors coupled to Gi/o G-proteins that inhibit adenylyl cyclase and to Gq-like phospholipase C activity. LPA and S1P also induce p44/42 ERK MAP kinase phosphorylation in these cells and stimulate cell proliferation via Gi/o coupled receptors in an Epidermal Growth Factor Receptor (EGFR- and ERK-dependent pathway. In contrast, LPA and S1P stimulate transient cell rounding and aggregation that is independent of EGFR and ERK, but dependent on the Rho effector p160 ROCK. Conclusion Thus, lysophospholipids regulate neural progenitor growth and morphology through distinct mechanisms. These findings establish human ES cell-derived NEP cells as a model system for studying the role of lysophospholipids in neural progenitors.

  12. Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea

    Indian Academy of Sciences (India)

    Si-Long Chen; Jia-Quan Huang; Lei Yong; Yue-Ting Zhang; Xiao-Ping Ren; Yu-Ning Chen; Hui-Fang Jiang; Li-Ying Yan; Yu-Rong Li; Bo-Shou Liao

    2012-12-01

    Lysophosphatidyl acyltransferase (LPAT) is the important enzyme responsible for the acylation of lysophosphatidic acid (LPA), leading to the generation of phosphatidic acid (PA) in plant. Its encoding gene is an essential candidate for oil crops to improve oil composition and increase seed oil content through genetic engineering. In this study, a full-length AhLPAT4 gene was isolated via cDNA library screening and rapid amplification of cDNA ends (RACE); our data demonstrated that AhLPAT4 had 1631 nucleotides, encoding a putative 43.8 kDa protein with 383 amino acid residues. The deduced protein included a conserved acyltransferase domain and four motifs (I–IV) with putative LPA and acyl-CoA catalytic and binding sites. Bioinformatic analysis indicated that AhLPAT4 contained four transmembrane domains (TMDs), localized to the endoplasmic reticulum (ER) membrane; detailed analysis indicated that motif I and motifs II–III in AhLPAT4 were separated by the third TMD, which located on cytosolic and ER luminal side respectively, and hydrophobic residues on the surface of AhLPAT4 protein fold to form a hydrophobic tunnel to accommodate the acyl chain. Subcellular localization analysis confirmed that AhLPAT4 was a cytoplasm protein. Phylogenetic analysis revealed that AhLPAT4 had a high homology (63.7–78.3%) with putative LPAT4 proteins from Glycine max, Arabidopsis thaliana and Ricinus communis. AhLPAT4 was ubiquitously expressed in diverse tissues except in flower, which is almost undetectable. The expression analysis in different developmental stages in peanut seeds indicated that AhLPAT4 did not coincide with oil accumulation.

  13. Lysophospholipid Growth Factors and Their G Protein-Coupled Receptors in Immunity, Coronary Artery Disease, and Cancer

    Directory of Open Access Journals (Sweden)

    Edward J. Goetzl

    2002-01-01

    Full Text Available The physiological lysophospholipids (LPLs, exemplified by lysophosphatidic acid (LPA and sphingosine 1-phosphate (S1P, are omnific mediators of normal cellular proliferation, survival, and functions. Although both LPA and S1P attain micromolar concentrations in many biological fluids, numerous aspects of their biosynthesis, transport, and metabolic degradation are unknown. Eight members of a new subfamily of G protein-coupled LPA/S1P receptors, originally termed Edg Rs, bind either LPA or S1P with high affinity and transduce a series of growth-related and/or cytoskeleton-based functional responses. The most critical areas of LPL biology and pathobiology are neural development and neurodegeneration, immunity, atherosclerosis and myocardial injury, and cancer. Data from analyses of T cells established two basic points: (1 the plasticity and adaptability of expression of LPA/S1P Rs by some cells as a function of activation, and (2 the role of opposing signals from two different receptors for the same ligand as a mechanism for fine control of effects of LPLs. In the heart, LPLs may promote coronary atherosclerosis, but are effectively cytoprotective for hypoxic cardiac myocytes and those exposed to oxygen free radicals. The findings of production of LPA by some types of tumor cells, overexpression of selected sets of LPA receptors by the same tumor cells, and augmentation of the effects of protein growth factors by LPA have suggested pathogenetic roles for the LPLs in cancer. The breadth of physiologic and pathologic activities of LPLs emphasizes the importance of developing bioavailable nonlipid agonists and antagonists of the LPA/S1P receptors for diverse therapeutic applications.

  14. Phospholipase Cϵ Activates Nuclear Factor-κB Signaling by Causing Cytoplasmic Localization of Ribosomal S6 Kinase and Facilitating Its Phosphorylation of Inhibitor κB in Colon Epithelial Cells.

    Science.gov (United States)

    Wakita, Masahiro; Edamatsu, Hironori; Li, Mingzhen; Emi, Aki; Kitazawa, Sohei; Kataoka, Tohru

    2016-06-10

    Phospholipase Cϵ (PLCϵ), an effector of Ras and Rap small GTPases, plays a crucial role in inflammation by augmenting proinflammatory cytokine expression. This proinflammatory function of PLCϵ is implicated in its facilitative role in tumor promotion and progression during skin and colorectal carcinogenesis, although their direct link remains to be established. Moreover, the molecular mechanism underlying these functions of PLCϵ remains unknown except that PKD works downstream of PLCϵ. Here we show by employing the colitis-induced colorectal carcinogenesis model, where Apc(Min) (/+) mice are administered with dextran sulfate sodium, that PLCϵ knock-out alleviates the colitis and suppresses the following tumorigenesis concomitant with marked attenuation of proinflammatory cytokine expression. In human colon epithelial Caco2 cells, TNF-α induces sustained expression of proinflammatory molecules and sustained activation of nuclear factor-κB (NF-κB) and PKD, the late phases of which are suppressed by not only siRNA-mediated PLCϵ knockdown but also treatment with a lysophosphatidic acid (LPA) receptor antagonist. Also, LPA stimulation induces these events in an early time course, suggesting that LPA mediates TNF-α signaling in an autocrine manner. Moreover, PLCϵ knockdown results in inhibition of phosphorylation of IκB by ribosomal S6 kinase (RSK) but not by IκB kinases. Subcellular fractionation suggests that enhanced phosphorylation of a scaffolding protein, PEA15 (phosphoprotein enriched in astrocytes 15), downstream of the PLCϵ-PKD axis causes sustained cytoplasmic localization of phosphorylated RSK, thereby facilitating IκB phosphorylation in the cytoplasm. These results suggest the crucial role of the TNF-α-LPA-LPA receptor-PLCϵ-PKD-PEA15-RSK-IκB-NF-κB pathway in facilitating inflammation and inflammation-associated carcinogenesis in the colon. PMID:27053111

  15. Exploring the role of sphingolipid machinery during the epithelial to mesenchymal transition program using an integrative approach

    Science.gov (United States)

    Meshcheryakova, Anastasia; Köfeler, Harald C.; Triebl, Alexander; Mungenast, Felicitas; Heinze, Georg; Gerner, Christopher; Zimmermann, Philip; Jaritz, Markus; Mechtcheriakova, Diana

    2016-01-01

    The epithelial to mesenchymal transition (EMT) program is activated in epithelial cancer cells and facilitates their ability to metastasize based on enhanced migratory, proliferative, anti-apoptotic, and pluripotent capacities. Given the fundamental impact of sphingolipid machinery to each individual process, the sphingolipid-related mechanisms might be considered among the most prominent drivers/players of EMT; yet, there is still limited knowledge. Given the complexity of the interconnected sphingolipid system, which includes distinct sphingolipid mediators, their synthesizing enzymes, receptors and transporters, we herein apply an integrative approach for assessment of the sphingolipid-associated mechanisms underlying EMT program. We created the sphingolipid-/EMT-relevant 41-gene/23-gene signatures which were applied to denote transcriptional events in a lung cancer cell-based EMT model. Based on defined 35-gene sphingolipid/EMT-attributed signature of regulated genes, we show close associations between EMT markers, genes comprising the sphingolipid network at multiple levels and encoding sphingosine 1-phosphate (S1P)-/ceramide-metabolizing enzymes, S1P and lysophosphatidic acid (LPA) receptors and S1P transporters, pluripotency genes and inflammation-related molecules, and demonstrate the underlying biological pathways and regulators. Mass spectrometry-based sphingolipid analysis revealed an EMT-attributed shift towards increased S1P and LPA accompanied by reduced ceramide levels. Notably, using transcriptomics data across various cell-based perturbations and neoplastic tissues (24193 arrays), we identified the sphingolipid/EMT signature primarily in lung adenocarcinoma tissues; besides, bladder, colorectal and prostate cancers were among the top-ranked. The findings also highlight novel regulatory associations between influenza virus and the sphingolipid/EMT-associated mechanisms. In sum, data propose the multidimensional contribution of sphingolipid machinery

  16. Fear extinction and acute stress reactivity reveal a role of LPA(1) receptor in regulating emotional-like behaviors.

    Science.gov (United States)

    Pedraza, C; Sánchez-López, J; Castilla-Ortega, E; Rosell-Valle, C; Zambrana-Infantes, E; García-Fernández, M; Rodriguez de Fonseca, F; Chun, J; Santín, L J; Estivill-Torrús, G

    2014-09-01

    LPA1 receptor is one of the six characterized G protein-coupled receptors (LPA1-6) through which lysophosphatidic acid acts as an intercellular signaling molecule. It has been proposed that this receptor has a role in controlling anxiety-like behaviors and in the detrimental consequences of stress. Here, we sought to establish the involvement of the LPA1 receptor in emotional regulation. To this end, we examined fear extinction in LPA1-null mice, wild-type and LPA1 antagonist-treated animals. In LPA1-null mice we also characterized the morphology and GABAergic properties of the amygdala and the medial prefrontal cortex. Furthermore, the expression of c-Fos protein in the amygdala and the medial prefrontal cortex, and the corticosterone response following acute stress were examined in both genotypes. Our data indicated that the absence of the LPA1 receptor significantly inhibited fear extinction. Treatment of wild-type mice with the LPA1 antagonist Ki16425 mimicked the behavioral phenotype of LPA1-null mice, revealing that the LPA1 receptor was involved in extinction. Immunohistochemistry studies revealed a reduction in the number of neurons, GABA+ cells, calcium-binding proteins and the volume of the amygdala in LPA1-null mice. Following acute stress, LPA1-null mice showed increased corticosterone and c-Fos expression in the amygdala. In conclusion, LPA1 receptor is involved in emotional behaviors and in the anatomical integrity of the corticolimbic circuit, the deregulation of which may be a susceptibility factor for anxiety disorders and a potential therapeutic target for the treatment of these diseases.

  17. Phospholipase C-η1 is activated by intracellular Ca(2+) mobilization and enhances GPCRs/PLC/Ca(2+) signaling.

    Science.gov (United States)

    Kim, Jung Kuk; Choi, Jung Woong; Lim, Seyoung; Kwon, Ohman; Seo, Jeong Kon; Ryu, Sung Ho; Suh, Pann-Ghill

    2011-06-01

    Phospholipase C-η1 (PLC-η1) is the most recently identified PLC isotype and is primarily expressed in nerve tissue. However, its functional role is unclear. In the present study, we report for the first time that PLC-η1 acts as a signal amplifier in G protein-coupled receptor (GPCR)-mediated PLC and Ca(2+) signaling. Short-hairpin RNA (shRNA)-mediated knockdown of endogenous PLC-η1 reduced lysophosphatidic acid (LPA)-, bradykinin (BK)-, and PACAP-induced PLC activity in mouse neuroblastoma Neuro2A (N2A) cells, indicating that PLC-η1 participates in GPCR-mediated PLC activation. Interestingly, ionomycin-induced PLC activity was significantly decreased by PLC-η1, but not PLC-η2, knockdown. In addition, we found that intracellular Ca(2+) source is enough for PLC-η1 activation. Furthermore, the IP(3) receptor inhibitor, 2-APB, inhibited LPA-induced PLC activity in control N2A cells, whereas this effect was not observed in PLC-η1 knockdown N2A cells, suggesting a pivotal role of intracellular Ca(2+) mobilization in PLC-η1 activation. Finally, we found that LPA-induced ERK1/2 phosphorylation and expression of the downstream target gene, krox-24, were significantly decreased by PLC-η1 knockdown, and these knockdown effects were abolished by 2-APB. Taken together, our results strongly suggest that PLC-η1 is activated via intracellular Ca(2+) mobilization from the ER, and therefore amplifies GPCR-mediated signaling.

  18. Role and regulation of EGFR in actin remodeling in sperm capacitation and the acrosome reaction

    Institute of Scientific and Technical Information of China (English)

    Haim Breitbart; Nir Etkovitz

    2011-01-01

    To bind and fertilize the egg,the spermatozoon should undergo few biochemical and motility changes in the female reproductive tract collectively called capacitation.The capacitated spermatozoon binds to the egg zona pellucida,and then undergoes the acrosome reaction(AR),which allows its penetration into the egg.The mechanisms regulating sperm capacitation and the AR are not completely understood.In the present review,we summarize some data regarding the role and regulation of the epidermal growth factor receptor(EGFR)in these processes.In the capacitation process,the EGFR is partially activated by protein kinase A(PKA),resulting in phospholipase D(PLD)activation and actin polymerization.Protein kinase C alpha(PKCα),which is already activated at the beginning of the capacitation,also participates in PLD activation.Further activation of the EGFR at the end of the capacitation enhances intracellular Ca2+concentration leading to F-actin breakdown and allows the AR to take place.Under in vivo conditions,the EGFR can be directly activated by its known ligand epidermal growth factor(EGF),and indirectly by activating PKA or by transactivation mediated by G protein-coupled receptors(GPCRs)activation or by ouabain.Under physiological conditions,sperm PKA is activated mainly by bicarbonate,which activates the soluble adenylyl cyclase to produce cyclic adenosine monophosphate(cAMP),the activator of PKA.The GPCR activators angiotensin ll or lysophosphatidic acid,as well as ouabain and EGF are physiological components present in the female reproductive tract.

  19. The pathological effects of Heminecrolysin, a dermonecrotic toxin from Hemiscorpius lepturus scorpion venom are mediated through its lysophospholipase D activity.

    Science.gov (United States)

    Borchani, Lamia; Sassi, Atfa; Ben Gharsa, Haifa; Safra, Ines; Shahbazzadeh, Delavar; Ben Lasfar, Zakaria; El Ayeb, Mohamed

    2013-06-01

    We have previously identified Heminecrolysin, a sphingomyelinase D (SMaseD), as the major protein responsible for the main pathological effects observed following Hemiscorpius (H.) lepturus scorpion envenomation. We aimed herein to further investigate the kinetics and molecular mechanisms triggered by Heminecrolysin to initiate hematological disorders and inflammatory reaction. We show that Heminecrolysin highly hydrolyzes lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA) and choline, with a Vmax = 1481 ± 51 μmol/min/mg and a Km = 97 ± 16.78 μM, at a much lesser extend sphingomyelin but not phosphatidylcholine substrates. Its lysophospholipase D (lysoPLD) catalytic efficiency, up to three orders of magnitude higher, comparatively to spider's SMaseDs (newly referred as phospholipases D; PLDs), could explain its strong hemolytic capacity. Chelating agents such as EDTA, EGTA, and 1, 10-phenantroline blocked Heminecrolysin-induced LPC hydrolysis at 98, 48, and 70% respectively. Hemolysis blockade occurs only when the toxin is added to erythrocytes in the presence of serum, source of LPC and complement, indicating that the production of LPA and the presence of complement are mandatory for hemolysis. Moreover, we show that Heminecrolysin efficiently binds to erythrocyte's membrane and provokes phosphatidylserine (PS) translocation without cleavage of glycophorin A, suggesting that, unlike spider's PLDs, complement was activated only via the classical pathway. Interestingly, Heminecrolysin was found to induce PS exposure on human nucleated Jurkat T cells, to stimulate secretion of the pro-inflammatory (TNF-α, IL-6), and anti-inflammatory (IL-10) cytokines by human monocytes, and to provoke a disseminated intravascular coagulation on chick embryo chorioallantoic membrane model system. Taken together, our results indicate that Heminecrolysin evokes the major characteristic clinical features of H. lepturus envenomation by using mainly its lyso

  20. Platelet lipidomic.

    Science.gov (United States)

    Dolegowska, B; Lubkowska, A; De Girolamo, L

    2012-01-01

    Lipids account for 16-19 percent dry platelet matter and includes 65 percent phospholipids, 25 percent neutral lipids and about 8 percent glycosphingolipids. The cell membrane that surrounds platelets is a bilayer that contains different types phospholipids symmetrically distributed in resting platelets, such as phosphatidylserine (PS), phosphatidylethanolamine (PE), phosphatidylcholine, and sphingomyelin. The collapse of lipid asymmetry is exposure of phosphatidylserine in the external leaflet of the plasma bilayer, where it is known to serve at least two major functions: providing a platform for development of the blood coagulation cascade and presenting the signal that induces phagocytosis of apoptotic cells. During activation, this asymmetrical distribution becomes disrupted, and PS and PE become exposed on the cell surface. The transbilayer movement of phosphatidylserine is responsible for the platelet procoagulant activity. Exposure of phosphatidylserine is a flag for macrophage recognition and clearance from the circulation. Platelets, stored at room temperature for transfusion for more than 5 days, undergo changes collectively known as platelet storage lesions. Thus, the platelet lipid composition and its possible modifications over time are crucial for efficacy of platelet rich plasma therapy. Moreover, a number of substances derived from lipids are contained into platelets. Eicosanoids are lipid signaling mediators generated by the action of lipoxygenase and include prostaglandins, thromboxane A2, 12-hydroxyeicosatetraenoic acid. Isoprostanes have a chemical structure similar to this of prostanoids, but are differently produced into the particle, and are ligands for prostaglandins receptors, exhibiting biological activity like thromboxane A2. Endocannabinoids are derivatives from arachidonic acid which could reduce local pain. Phospholipids growth factors (sphingolipids, lysophosphatidic acid, platelet-activating factor) are involved in tissue

  1. Design of serum-free medium for suspension culture of CHO cells on the basis of general commercial media.

    Science.gov (United States)

    Miki, Hideo; Takagi, Mutsumi

    2015-08-01

    The design of serum-free media for suspension culture of genetically engineered Chinese hamster ovary (CHO) cells using general commercial media as a basis was investigated. Subcultivation using a commercial serum-free medium containing insulin-like growth factor (IGF)-1 with or without FCS necessitated additives other than IGF-1 to compensate for the lack of FCS and improve cell growth. Suspension culture with media containing several combinations of growth factors suggested the effectiveness of addition of both IGF-1 and the lipid signaling molecule lysophosphatidic acid (LPA) for promoting cell growth. Subcultivation of CHO cells in suspension culture using the commercial serum-free medium EX-CELL™302, which contained an IGF-1 analog, supplemented with LPA resulted in gradually increasing specific growth rate comparable to the serum-containing medium and in almost the same high antibody production regardless of the number of generations. The culture with EX-CELL™302 supplemented with LPA in a jar fermentor with pH control at 6.9 showed an apparently higher cell growth rate than the cultures without pH control and with pH control at 6.8. The cell growth in the medium supplemented with aurintricarboxylic acid (ATA), which was much cheaper than IGF-1, in combination with LPA was synergistically promoted similarly to that in the medium supplemented with IGF-1 and LPA. In conclusion, the serum-free medium designed on the basis of general commercial media could support the growth of CHO cells and antibody production comparable to serum-containing medium in suspension culture. Moreover, the possibility of cost reduction by the substitution of IGF-1 with ATA was also shown.

  2. Reassessing the Potential Activities of Plant CGI-58 Protein.

    Science.gov (United States)

    Khatib, Abdallah; Arhab, Yani; Bentebibel, Assia; Abousalham, Abdelkarim; Noiriel, Alexandre

    2016-01-01

    Comparative Gene Identification-58 (CGI-58) is a widespread protein found in animals and plants. This protein has been shown to participate in lipolysis in mice and humans by activating Adipose triglyceride lipase (ATGL), the initial enzyme responsible for the triacylglycerol (TAG) catabolism cascade. Human mutation of CGI-58 is the cause of Chanarin-Dorfman syndrome, an orphan disease characterized by a systemic accumulation of TAG which engenders tissue disorders. The CGI-58 protein has also been shown to participate in neutral lipid metabolism in plants and, in this case, a mutation again provokes TAG accumulation. Although its roles as an ATGL coactivator and in lipid metabolism are quite clear, the catalytic activity of CGI-58 is still in question. The acyltransferase activities of CGI-58 have been speculated about, reported or even dismissed and experimental evidence that CGI-58 expressed in E. coli possesses an unambiguous catalytic activity is still lacking. To address this problem, we developed a new set of plasmids and site-directed mutants to elucidate the in vivo effects of CGI-58 expression on lipid metabolism in E. coli. By analyzing the lipid composition in selected E. coli strains expressing CGI-58 proteins, and by reinvestigating enzymatic tests with adequate controls, we show here that recombinant plant CGI-58 has none of the proposed activities previously described. Recombinant plant and mouse CGI-58 both lack acyltransferase activity towards either lysophosphatidylglycerol or lysophosphatidic acid to form phosphatidylglycerol or phosphatidic acid and recombinant plant CGI-58 does not catalyze TAG or phospholipid hydrolysis. However, expression of recombinant plant CGI-58, but not mouse CGI-58, led to a decrease in phosphatidylglycerol in all strains of E. coli tested, and a mutation of the putative catalytic residues restored a wild-type phenotype. The potential activities of plant CGI-58 are subsequently discussed. PMID:26745266

  3. 卵巢过度刺激综合征发病机制的研究进展%Research Progress of Ovarian Hyperstimulation Syndrome Pathogenesis

    Institute of Scientific and Technical Information of China (English)

    叶尔登切切克

    2012-01-01

    卵巢过度刺激综合征(OHSS)是促排卵过程中严重的医源性并发症,人绒毛膜促性腺激素诱导排卵使过多卵泡受刺激有关.其发病机制不明,病理生理特征主要为毛细血管通透性增加以及富含蛋白的液体停留在血管外间隙,导致血液浓缩,第三间隙水肿.现从血管内皮生长因子、血小板活化因子、溶血磷脂酸诱导炎性因子、囊性纤维化跨膜传导调节因子、肾素-血管紧张素系统对OHSS发病机制进行探讨分析,为临床更好地治疗和预防OHSS 的发生提供理论依据.%The ovarian hyperstimulation syndrome( OHSS )is a serious iatrogenic syndrome of controlled ovarian hyperstimulation. related tohuman chorionic gonadotropin( HCG ) induced ovulation caused follicles stimulation. Its pathogenesis is unknown, the main pathophysiological characteristics are increased capillary permeahility and protein-rich liquid stasis in the extravascular space, resulting in hlood concentration, third interstitial edema. Here is to discuss the possible pathogenesis of OHSS from the perspective of blood vessels endothelial growth factor, platelet-activating factor, lysophosphatidic acid-induced inflammatory factor, cystic fibrosis transmembrane conductance regulator,renin-angiotensin system,to provide a theoretical basis for better prevention and treatment of OHSS in clinic.

  4. Association of genetic loci with sleep apnea in European Americans and African-Americans: the Candidate Gene Association Resource (CARe.

    Directory of Open Access Journals (Sweden)

    Sanjay R Patel

    Full Text Available Although obstructive sleep apnea (OSA is known to have a strong familial basis, no genetic polymorphisms influencing apnea risk have been identified in cross-cohort analyses. We utilized the National Heart, Lung, and Blood Institute (NHLBI Candidate Gene Association Resource (CARe to identify sleep apnea susceptibility loci. Using a panel of 46,449 polymorphisms from roughly 2,100 candidate genes on a customized Illumina iSelect chip, we tested for association with the apnea hypopnea index (AHI as well as moderate to severe OSA (AHI≥15 in 3,551 participants of the Cleveland Family Study and two cohorts participating in the Sleep Heart Health Study.Among 647 African-Americans, rs11126184 in the pleckstrin (PLEK gene was associated with OSA while rs7030789 in the lysophosphatidic acid receptor 1 (LPAR1 gene was associated with AHI using a chip-wide significance threshold of p-value<2×10(-6. Among 2,904 individuals of European ancestry, rs1409986 in the prostaglandin E2 receptor (PTGER3 gene was significantly associated with OSA. Consistency of effects between rs7030789 and rs1409986 in LPAR1 and PTGER3 and apnea phenotypes were observed in independent clinic-based cohorts.Novel genetic loci for apnea phenotypes were identified through the use of customized gene chips and meta-analyses of cohort data with replication in clinic-based samples. The identified SNPs all lie in genes associated with inflammation suggesting inflammation may play a role in OSA pathogenesis.

  5. Non-transactivational, dual pathways for LPA-induced Erk1/2 activation in primary cultures of brown pre-adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Holmstroem, Therese E.; Mattsson, Charlotte L.; Wang, Yanling; Iakovleva, Irina; Petrovic, Natasa [Department of Physiology, The Wenner-Gren Institute, Stockholm University, SE-106 91 Stockholm (Sweden); Nedergaard, Jan, E-mail: jan@metabol.su.se [Department of Physiology, The Wenner-Gren Institute, Stockholm University, SE-106 91 Stockholm (Sweden)

    2010-10-01

    In many cell types, G-protein-coupled receptor (GPCR)-induced Erk1/2 MAP kinase activation is mediated via receptor tyrosine kinase (RTK) transactivation, in particular via the epidermal growth factor (EGF) receptor. Lysophosphatidic acid (LPA), acting via GPCRs, is a mitogen and MAP kinase activator in many systems, and LPA can regulate adipocyte proliferation. The mechanism by which LPA activates the Erk1/2 MAP kinase is generally accepted to be via EGF receptor transactivation. In primary cultures of brown pre-adipocytes, EGF can induce Erk1/2 activation, which is obligatory and determinant for EGF-induced proliferation of these cells. Therefore, we have here examined whether LPA, via EGF transactivation, can activate Erk1/2 in brown pre-adipocytes. We found that LPA could induce Erk1/2 activation. However, the LPA-induced Erk1/2 activation was independent of transactivation of EGF receptors (or PDGF receptors) in these cells (whereas in transformed HIB-1B brown adipocytes, the LPA-induced Erk1/2 activation indeed proceeded via EGF receptor transactivation). In the brown pre-adipocytes, LPA instead induced Erk1/2 activation via two distinct non-transactivational pathways, one G{sub i}-protein dependent, involving PKC and Src activation, the other, a PTX-insensitive pathway, involving PI3K (but not Akt) activation. Earlier studies showing LPA-induced Erk1/2 activation being fully dependent on RTK transactivation have all been performed in cell lines and transfected cells. The present study implies that in non-transformed systems, RTK transactivation may not be involved in the mediation of GPCR-induced Erk1/2 MAP kinase activation.

  6. Reassessing the Potential Activities of Plant CGI-58 Protein.

    Directory of Open Access Journals (Sweden)

    Abdallah Khatib

    Full Text Available Comparative Gene Identification-58 (CGI-58 is a widespread protein found in animals and plants. This protein has been shown to participate in lipolysis in mice and humans by activating Adipose triglyceride lipase (ATGL, the initial enzyme responsible for the triacylglycerol (TAG catabolism cascade. Human mutation of CGI-58 is the cause of Chanarin-Dorfman syndrome, an orphan disease characterized by a systemic accumulation of TAG which engenders tissue disorders. The CGI-58 protein has also been shown to participate in neutral lipid metabolism in plants and, in this case, a mutation again provokes TAG accumulation. Although its roles as an ATGL coactivator and in lipid metabolism are quite clear, the catalytic activity of CGI-58 is still in question. The acyltransferase activities of CGI-58 have been speculated about, reported or even dismissed and experimental evidence that CGI-58 expressed in E. coli possesses an unambiguous catalytic activity is still lacking. To address this problem, we developed a new set of plasmids and site-directed mutants to elucidate the in vivo effects of CGI-58 expression on lipid metabolism in E. coli. By analyzing the lipid composition in selected E. coli strains expressing CGI-58 proteins, and by reinvestigating enzymatic tests with adequate controls, we show here that recombinant plant CGI-58 has none of the proposed activities previously described. Recombinant plant and mouse CGI-58 both lack acyltransferase activity towards either lysophosphatidylglycerol or lysophosphatidic acid to form phosphatidylglycerol or phosphatidic acid and recombinant plant CGI-58 does not catalyze TAG or phospholipid hydrolysis. However, expression of recombinant plant CGI-58, but not mouse CGI-58, led to a decrease in phosphatidylglycerol in all strains of E. coli tested, and a mutation of the putative catalytic residues restored a wild-type phenotype. The potential activities of plant CGI-58 are subsequently discussed.

  7. Lysophospholipids and their receptors in the central nervous system.

    Science.gov (United States)

    Choi, Ji Woong; Chun, Jerold

    2013-01-01

    Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P), two of the best-studied lysophospholipids, are known to influence diverse biological events, including organismal development as well as function and pathogenesis within multiple organ systems. These functional roles are due to a family of at least 11 G protein-coupled receptors (GPCRs), named LPA(1-6) and S1P(1-5), which are widely distributed throughout the body and that activate multiple effector pathways initiated by a range of heterotrimeric G proteins including G(i/o), G(12/13), G(q) and G(s), with actual activation dependent on receptor subtypes. In the central nervous system (CNS), a major locus for these signaling pathways, LPA and S1P have been shown to influence myriad responses in neurons and glial cell types through their cognate receptors. These receptor-mediated activities can contribute to disease pathogenesis and have therapeutic relevance to human CNS disorders as demonstrated for multiple sclerosis (MS) and possibly others that include congenital hydrocephalus, ischemic stroke, neurotrauma, neuropsychiatric disorders, developmental disorders, seizures, hearing loss, and Sandhoff disease, based upon the experimental literature. In particular, FTY720 (fingolimod, Gilenya, Novartis Pharma, AG) that becomes an analog of S1P upon phosphorylation, was approved by the FDA in 2010 as a first oral treatment for MS, validating this class of receptors as medicinal targets. This review will provide an overview and update on the biological functions of LPA and S1P signaling in the CNS, with a focus on results from studies using genetic null mutants for LPA and S1P receptors. This article is part of a Special Issue entitled Advances in Lysophospholipid Research. PMID:22884303

  8. Inhibition of lipid phosphate phosphatase activity by VPC32183 suppresses the ability of diacylglycerol pyrophosphate to activate ERK(1/2) MAP kinases.

    Science.gov (United States)

    Violet, Pierre-Christian; Billon-Denis, Emmanuelle; Robin, Philippe

    2012-11-01

    The lipidic metabolite, diacylglycerol pyrophosphate (DGPP), in its dioctanoyl form (DGPP 8:0), has been described as an antagonist for mammalian lysophosphatidic acid (LPA) receptors LPA1 and LPA3. In this study we show that DGPP 8:0 does not antagonize LPA dependent activation of ERK(1/2) MAP kinases but strongly stimulated them in various mammalian cell lines. LPA and DGPP 8:0 stimulation of ERK(1/2) occurred through different pathways. The DGPP 8:0 effect appeared to be dependent on PKC, Raf and MEK but was insensitive to pertussis toxin and did not involve G protein activation. Finally we showed that DGPP 8:0 effect on ERK(1/2) was dependent on its dephosphorylation by a phosphatase activity sharing lipid phosphate phosphatase properties. The inhibition of this phosphatase activity by VPC32183, a previously characterized LPA receptor antagonist, blocked the DGPP 8:0 effect on ERK(1/2) activation. Moreover, down-regulation of lipid phosphate phosphatase 1 (LPP1) expression by RNA interference technique also reduced DGPP 8:0-induced ERK(1/2) activation. Consistently, over expression of LPP1 in HEK293 cells increases DGPP 8:0 hydrolysis and this increased activity was inhibited by VPC32183. In conclusion, DGPP 8:0 does not exert its effect by acting on a G protein coupled receptor, but through its dephosphorylation by LPP1, generating dioctanoyl phosphatidic acid which in turn activates PKC. These results suggest that LPP1 could have a positive regulatory function on cellular signaling processes such as ERK(1/2) activation.

  9. Rho signaling in Entamoeba histolytica modulates actomyosin-dependent activities stimulated during invasive behavior.

    Science.gov (United States)

    Franco-Barraza, Janusz; Zamudio-Meza, Horacio; Franco, Elizabeth; del Carmen Domínguez-Robles, M; Villegas-Sepúlveda, Nicolás; Meza, Isaura

    2006-03-01

    Interaction of Entamoeba histolytica trophozoites with target cells and substrates activates signaling pathways in the parasite. Phosphorylation cascades triggered by phospho-inositide and adenyl-cyclase-dependent pathways modulate reorganization of the actin cytoskeleton to form structures that facilitate adhesion. In contrast, little is known about participation of Rho proteins and Rho signaling in actin rearrangements. We report here the in vivo expression of at least one Rho protein in trophozoites, whose activation induced actin reorganization and actin-myosin interaction. Antibodies to EhRhoA1 recombinant protein mainly localized Rho in the cytosol of nonactivated amoebae, but it was translocated to vesicular membranes and to some extent to the plasma membrane after treatment with lysophosphatidic acid (LPA), a specific agonist of Rho activation. Activated Rho was identified in LPA-treated trophozoites. LPA induced striking polymerization of actin into distinct dynamic structures. Disorganization of these structures by inhibition of Rho effector, Rho-kinase (ROCK), and by ML-7, an inhibitor of myosin light chain kinase dependent phosphorylation of myosin light chain, suggested that the actin structures also contained myosin. LPA stimulated concanavalin-A-mediated formation of caps, chemotaxis, invasion of extracellular matrix substrates, and erythrophagocytosis, but not binding to fibronectin. ROCK inhibition impaired LPA-stimulated functions and to some extent adhesion to fibronectin. Similar results were obtained with ML-7. These data suggest the presence and operation of Rho-signaling pathways in E. histolytica, that together with other, already described, signaling routes modulate actomyosin-dependent motile processes, particularly stimulated during invasive behavior.

  10. Proline-rich tyrosine kinase 2 (Pyk2 promotes cell motility of hepatocellular carcinoma through induction of epithelial to mesenchymal transition.

    Directory of Open Access Journals (Sweden)

    Chris K Sun

    Full Text Available AIMS: Proline-rich tyrosine kinase 2 (Pyk2, a non-receptor tyrosine kinase of the focal adhesion kinase (FAK family, is up-regulated in more than 60% of the tumors of hepatocellular carcinoma (HCC patients. Forced overexpression of Pyk2 can promote the proliferation and invasion of HCC cells. In this study, we aimed to explore the underlying molecular mechanism of Pyk2-mediated cell migration of HCC cells. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrated that Pyk2 transformed the epithelial HCC cell line Hep3B into a mesenchymal phenotype via the induction of epithelial to mesenchymal transition (EMT, signified by the up-regulation of membrane ruffle formation, activation of Rac/Rho GTPases, down-regulation of epithelial genes E-cadherin and cytokeratin as well as promotion of cell motility in presence of lysophosphatidic acid (LPA. Suppression of Pyk2 by overexpression of dominant negative PRNK domain in the metastatic HCC cell line MHCC97L transformed its fibroblastoid phenotype to an epithelial phenotype with up-regulation of epithelial genes, down-regulation of mesenchymal genes N-cadherin and STAT5b, and reduction of LPA-induced membrane ruffle formation and cell motility. Moreover, overexpression of Pyk2 in Hep3B cells promoted the phosphorylation and localization of mesenchymal gene Hic-5 onto cell membrane while suppression of Pyk2 in MHCC97L cells attenuated its phosphorylation and localization. CONCLUSION: These data provided new evidence of the underlying mechanism of Pyk2 in controlling cell motility of HCC cells through regulation of genes associated with EMT.

  11. Reassessing the Potential Activities of Plant CGI-58 Protein.

    Science.gov (United States)

    Khatib, Abdallah; Arhab, Yani; Bentebibel, Assia; Abousalham, Abdelkarim; Noiriel, Alexandre

    2016-01-01

    Comparative Gene Identification-58 (CGI-58) is a widespread protein found in animals and plants. This protein has been shown to participate in lipolysis in mice and humans by activating Adipose triglyceride lipase (ATGL), the initial enzyme responsible for the triacylglycerol (TAG) catabolism cascade. Human mutation of CGI-58 is the cause of Chanarin-Dorfman syndrome, an orphan disease characterized by a systemic accumulation of TAG which engenders tissue disorders. The CGI-58 protein has also been shown to participate in neutral lipid metabolism in plants and, in this case, a mutation again provokes TAG accumulation. Although its roles as an ATGL coactivator and in lipid metabolism are quite clear, the catalytic activity of CGI-58 is still in question. The acyltransferase activities of CGI-58 have been speculated about, reported or even dismissed and experimental evidence that CGI-58 expressed in E. coli possesses an unambiguous catalytic activity is still lacking. To address this problem, we developed a new set of plasmids and site-directed mutants to elucidate the in vivo effects of CGI-58 expression on lipid metabolism in E. coli. By analyzing the lipid composition in selected E. coli strains expressing CGI-58 proteins, and by reinvestigating enzymatic tests with adequate controls, we show here that recombinant plant CGI-58 has none of the proposed activities previously described. Recombinant plant and mouse CGI-58 both lack acyltransferase activity towards either lysophosphatidylglycerol or lysophosphatidic acid to form phosphatidylglycerol or phosphatidic acid and recombinant plant CGI-58 does not catalyze TAG or phospholipid hydrolysis. However, expression of recombinant plant CGI-58, but not mouse CGI-58, led to a decrease in phosphatidylglycerol in all strains of E. coli tested, and a mutation of the putative catalytic residues restored a wild-type phenotype. The potential activities of plant CGI-58 are subsequently discussed.

  12. Ectodomain cleavage of the EGF ligands HB-EGF, neuregulin1-beta, and TGF-alpha is specifically triggered by different stimuli and involves different PKC isoenzymes.

    Science.gov (United States)

    Herrlich, Andreas; Klinman, Eva; Fu, Jonathan; Sadegh, Cameron; Lodish, Harvey

    2008-12-01

    Metalloproteinase cleavage of transmembrane proteins (ectodomain cleavage), including the epidermal growth factor (EGF) ligands heparin-binding EGF-like growth factor (HB-EGF), neuregulin (NRG), and transforming growth factor-alpha (TGF-alpha), is important in many cellular signaling pathways and is disregulated in many diseases. It is largely unknown how physiological stimuli of ectodomain cleavage--hypertonic stress, phorbol ester, or activation of G-protein-coupled receptors [e.g., by lysophosphatidic acid (LPA)]--are molecularly connected to metalloproteinase activation. To study this question, we developed a fluorescence-activated cell sorting (FACS)- based assay that measures cleavage of EGF ligands in single living cells. EGF ligands expressed in mouse lung epithelial cells are differentially and specifically cleaved depending on the stimulus. Inhibition of protein kinase C (PKC) isoenzymes or metalloproteinase inhibition by batimastat (BB94) showed that different regulatory signals are used by different stimuli and EGF substrates, suggesting differential effects that act on the substrate, the metalloproteinase, or both. For example, hypertonic stress led to strong cleavage of HB-EGF and NRG but only moderate cleavage of TGF-alpha. HB-EGF, NRG, and TGF-alpha cleavage was not dependent on PKC, and only HB-EGF and NRG cleavage were inhibited by BB94. In contrast, phorbol 12-myristate-13-acetate (TPA) -induced cleavage of HB-EGF, NRG, and TGF-alpha was dependent on PKC and sensitive to BB94 inhibition. LPA led to significant cleavage of only NRG and TGF-alpha and was inhibited by BB94; only LPA-induced NRG cleavage required PKC. Surprisingly, specific inhibition of atypical PKCs zeta and iota [not activated by diacylglycerol (DAG) and calcium] significantly enhanced TPA-induced NRG cleavage. Employed in a high-throughput cloning strategy, our cleavage assay should allow the identification of candidate proteins involved in signal transduction of different

  13. Epidermal growth factor (EGF) ligand release by substrate-specific a disintegrin and metalloproteases (ADAMs) involves different protein kinase C (PKC) isoenzymes depending on the stimulus.

    Science.gov (United States)

    Dang, Michelle; Dubbin, Karen; D'Aiello, Antonio; Hartmann, Monika; Lodish, Harvey; Herrlich, Andreas

    2011-05-20

    The dysregulation of EGF family ligand cleavage has severe consequences for the developing as well as the adult organism. Therefore, their production is highly regulated. The limiting step is the ectodomain cleavage of membrane-bound precursors by one of several a disintegrin and metalloprotease (ADAM) metalloproteases, and understanding the regulation of cleavage is an important goal of current research. We have previously reported that in mouse lung epithelial cells, the pro-EGF ligands TGFα, neuregulin 1β (NRG), and heparin-binding EGF are differentially cleaved depending on the cleavage stimulus (Herrlich, A., Klinman, E., Fu, J., Sadegh, C., and Lodish, H. (2008) FASEB J.). In this study in mouse embryonic fibroblasts that lack different ADAMs, we show that induced cleavage of EGF ligands can involve the same substrate-specific metalloprotease but does require different stimulus-dependent signaling pathways. Cleavage was stimulated by phorbol ester (12-O-tetradecanoylphorbol-13-acetate (TPA), a mimic of diacylglycerol and PKC activator), hypertonic stress, lysophosphatidic acid (LPA)-induced G protein-coupled receptor activation, or by ionomycin-induced intracellular calcium release. Although ADAMs showed substrate preference (ADAM17, TGFα and heparin-binding EGF; and ADAM9, NRG), substrate cleavage differed substantially with the stimulus, and cleavage of the same substrate depended on the presence of different, sometimes multiple, PKC isoforms. For instance, classical PKC was required for TPA-induced but not hypertonic stress-induced cleavage of all EGF family ligands. Inhibition of PKCζ enhanced NRG release upon TPA stimulation, but it blocked NRG release in response to hypertonic stress. Our results suggest a model in which substantial regulation of ectodomain cleavage occurs not only on the metalloprotease level but also on the level of the substrate or of a third protein.

  14. Autotaxin is induced by TSA through HDAC3 and HDAC7 inhibition and antagonizes the TSA-induced cell apoptosis

    Directory of Open Access Journals (Sweden)

    Zhang Junjie

    2011-02-01

    Full Text Available Abstract Background Autotaxin (ATX is a secreted glycoprotein with the lysophospholipase D (lysoPLD activity to convert lysophosphatidylcholine (LPC into lysophosphatidic acid (LPA, a bioactive lysophospholipid involved in diverse biological actions. ATX is highly expressed in some cancer cells and contributes to their tumorigenesis, invasion, and metastases, while in other cancer cells ATX is silenced or expressed at low level. The mechanism of ATX expression regulation in cancer cells remains largely unknown. Results In the present study, we demonstrated that trichostatin A (TSA, a well-known HDAC inhibitor (HDACi, significantly induced ATX expression in SW480 and several other cancer cells with low or undetectable endogenous ATX expression. ATX induction could be observed when HDAC3 and HDAC7 were down-regulated by their siRNAs. It was found that HDAC7 expression levels were low in the cancer cells with high endogenous ATX expression. Exogenous over-expression of HDAC7 inhibited ATX expression in these cells in a HDAC3-dependent manner. These data indicate that HDAC3 and HDAC7 collaboratively suppress ATX expression in cancer cells, and suggest that TSA induce ATX expression by inhibiting HDAC3 and HDAC7. The biological significance of this regulation mechanism was revealed by demonstrating that TSA-induced ATX protected cancer cells against TSA-induced apoptosis by producing LPA through its lysoPLD activity, which could be reversed by BrP-LPA and S32826, the inhibitors of the ATX-LPA axis. Conclusions We have demonstrated that ATX expression is repressed by HDAC3 and HDAC7 in cancer cells. During TSA treatment, ATX is induced due to the HDAC3 and HDAC7 inhibition and functionally antagonizes the TSA-induced apoptosis. These results reveal an internal HDACi-resistant mechanism in cancer cells, and suggest that the inhibition of ATX-LPA axis would be helpful to improve the efficacy of HDACi-based therapeutics against cancer.

  15. The focal adhesion: a regulated component of aortic stiffness.

    Directory of Open Access Journals (Sweden)

    Robert J Saphirstein

    Full Text Available Increased aortic stiffness is an acknowledged predictor and cause of cardiovascular disease. The sources and mechanisms of vascular stiffness are not well understood, although the extracellular matrix (ECM has been assumed to be a major component. We tested here the hypothesis that the focal adhesions (FAs connecting the cortical cytoskeleton of vascular smooth muscle cells (VSMCs to the matrix in the aortic wall are a component of aortic stiffness and that this component is dynamically regulated. First, we examined a model system in which magnetic tweezers could be used to monitor cellular cortical stiffness, serum-starved A7r5 aortic smooth muscle cells. Lysophosphatidic acid (LPA, an activator of myosin that increases cell contractility, increased cortical stiffness. A small molecule inhibitor of Src-dependent FA recycling, PP2, was found to significantly inhibit LPA-induced increases in cortical stiffness, as well as tension-induced increases in FA size. To directly test the applicability of these results to force and stiffness development at the level of vascular tissue, we monitored mouse aorta ring stiffness with small sinusoidal length oscillations during agonist-induced contraction. The alpha-agonist phenylephrine, which also increases myosin activation and contractility, increased tissue stress and stiffness in a PP2- and FAK inhibitor 14-attenuated manner. Subsequent phosphotyrosine screening and follow-up with phosphosite-specific antibodies confirmed that the effects of PP2 and FAK inhibitor 14 in vascular tissue involve FA proteins, including FAK, CAS, and paxillin. Thus, in the present study we identify, for the first time, the FA of the VSMC, in particular the FAK-Src signaling complex, as a significant subcellular regulator of aortic stiffness and stress.

  16. First International Conference on Lysophospholipids and Related Bioactive Lipids in Biology and Disease Sponsored by the Federation of American Societies of Experimental Biology

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    Edward J. Goetzl

    2001-01-01

    Full Text Available The First International Conference on “Lysophospholipids and Related Bioactive Lipids in Biology and Diseases” was held in Tucson, AZ on June 10�14, 2001, under the sponsorship of the Federation of American Societies of Experimental Biology (FASEB. More than 100 scientists from 11 countries discussed the recent results of basic and clinical research in the broad biology of this emerging field. Immense progress was reported in defining the biochemistry of generation and biology of cellular effects of the bioactive lysophospholipids (LPLs. These aspects of LPLs described at the conference parallel in many ways those of the eicosanoid mediators, such as prostaglandins and leukotrienes. As for eicosanoids, the LPLs termed lysophosphatidic acid (LPA and sphingosine 1-phosphate (S1P are produced enzymatically from phospholipid precursors in cell membranes and act on cells at nanomolar concentrations through subfamilies of receptors of the G protein–coupled superfamily. The rate-limiting steps in production of LPLs were reported to be controlled by specific phospholipases for LPA and sphingosine kinases for S1P. The receptor subfamilies formerly were designated endothelial differentiation gene-encoded receptors or Edg Rs for their original discovery in endothelial cells. A currently active nomenclature committee at this conference suggested the ligand-based names: S1P1 = Edg-1, S1P2 = Edg-5, S1P3 = Edg-3, S1P4 = Edg-6, and S1P5 = Edg-8; LPA1 = Edg-2, LPA2 = Edg-4, and LPA3 = Edg-7 receptors. Several families of lysophospholipid phosphatases (LPPs have been characterized, which biodegrade LPA, whereas S1P is inactivated with similar rapidity by both a lyase and S1P phosphatases.

  17. A functional and transcriptomic analysis of NET1 bioactivity in gastric cancer

    LENUS (Irish Health Repository)

    Bennett, Gayle

    2011-02-01

    Abstract Background NET1, a RhoA guanine exchange factor, is up-regulated in gastric cancer (GC) tissue and drives the invasive phenotype of this disease. In this study, we aimed to determine the role of NET1 in GC by monitoring the proliferation, motility and invasion of GC cells in which NET1 has been stably knocked down. Additionally, we aimed to determine NET1-dependent transcriptomic events that occur in GC. Methods An in vitro model of stable knockdown of NET1 was achieved in AGS human gastric adenocarcinoma cells via lentiviral mediated transduction of short-hairpin (sh) RNA targeting NET1. Knockdown was assessed using quantitative PCR. Cell proliferation was assessed using an MTS assay and cell migration was assessed using a wound healing scratch assay. Cell invasion was assessed using a transwell matrigel invasion assay. Gene expression profiles were examined using affymetrix oligonucleotide U133A expression arrays. A student\\'s t test was used to determine changes of statistical significance. Results GC cells were transduced with NET1 shRNA resulting in a 97% reduction in NET1 mRNA (p < 0.0001). NET1 knockdown significantly reduced the invasion and migration of GC cells by 94% (p < 0.05) and 24% (p < 0.001) respectively, while cell proliferation was not significantly altered following NET1 knockdown. Microarray analysis was performed on non-target and knockdown cell lines, treated with and without 10 μM lysophosphatidic acid (LPA) allowing us to identify NET1-dependent, LPA-dependent and NET1-mediated LPA-induced gene transcription. Differential gene expression was confirmed by quantitative PCR. Shortlisted NET1-dependent genes included STAT1, TSPAN1, TGFBi and CCL5 all of which were downregulatd upon NET1 downregulation. Shortlisted LPA-dependent genes included EGFR and PPARD where EGFR was upregulated and PPARD was downregulated upon LPA stimulation. Shortlisted NET1 and LPA dependent genes included IGFR1 and PIP5K3. These LPA induced genes were

  18. Characteristics of Gintonin-Mediated Membrane Depolarization of Pacemaker Activity in Cultured Interstitial Cells of Cajal

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    Byung Joo Kim

    2014-08-01

    Full Text Available Background/Aims: Ginseng regulates gastrointestinal (GI motor activity but the underlying components and molecular mechanisms are unknown. We investigated the effect of gintonin, a novel ginseng-derived G protein-coupled lysophosphatidic acid (LPA receptor ligand, on the pacemaker activity of the interstitial cells of Cajal (ICC in murine small intestine and GI motility. Materials and Methods: Enzymatic digestion was used to dissociate ICC from mouse small intestines. The whole-cell patch-clamp configuration was used to record pacemaker potentials and currents from cultured ICC in the absence or presence of gintonin. In vivo effects of gintonin on gastrointestinal (GI motility were investigated by measuring the intestinal transit rate (ITR of Evans blue in normal and streptozotocin (STZ-induced diabetic mice. Results: We investigated the effects of gintonin on pacemaker potentials and currents in cultured ICC from mouse small intestine. Gintonin caused membrane depolarization in current clamp mode but this action was blocked by Ki16425, an LPA1/3 receptor antagonist, and by the addition of GDPβS, a GTP-binding protein inhibitor, into the ICC. To study the gintonin signaling pathway, we examined the effects of U-73122, an active PLC inhibitor, and chelerythrine and calphostin, which inhibit PKC. All inhibitors blocked gintonin actions on pacemaker potentials, but not completely. Gintonin-mediated depolarization was lower in Ca2+-free than in Ca2+-containing external solutions and was blocked by thapsigargin. We found that, in ICC, gintonin also activated Ca2+-activated Cl- channels (TMEM16A, ANO1, but not TRPM7 channels. In vivo, gintonin (10-100 mg/kg, p.o. not only significantly increased the ITR in normal mice but also ameliorated STZ-induced diabetic GI motility retardation in a dose-dependent manner. Conclusions: Gintonin-mediated membrane depolarization of pacemaker activity and ANO1 activation are coupled to the stimulation of GI

  19. FGF23 gene regulation by 1,25-dihydroxyvitamin D: opposing effects in adipocytes and osteocytes.

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    Kaneko, Ichiro; Saini, Rimpi K; Griffin, Kristin P; Whitfield, G Kerr; Haussler, Mark R; Jurutka, Peter W

    2015-09-01

    In a closed endocrine loop, 1,25-dihydroxyvitamin D3 (1,25D) induces the expression of fibroblast growth factor 23 (FGF23) in bone, with the phosphaturic peptide in turn acting at kidney to feedback repress CYP27B1 and induce CYP24A1 to limit the levels of 1,25D. In 3T3-L1 differentiated adipocytes, 1,25D represses FGF23 and leptin expression and induces C/EBPβ, but does not affect leptin receptor transcription. Conversely, in UMR-106 osteoblast-like cells, FGF23 mRNA concentrations are upregulated by 1,25D, an effect that is blunted by lysophosphatidic acid, a cell-surface acting ligand. Progressive truncation of the mouse FGF23 proximal promoter linked in luciferase reporter constructs reveals a 1,25D-responsive region between -400 and -200  bp. A 0.6  kb fragment of the mouse FGF23 promoter, linked in a reporter construct, responds to 1,25D with a fourfold enhancement of transcription in transfected K562 cells. Mutation of either an ETS1 site at -346  bp, or an adjacent candidate vitamin D receptor (VDR)/Nurr1-element, in the 0.6  kb reporter construct reduces the transcriptional activity elicited by 1,25D to a level that is not significantly different from a minimal promoter. This composite ETS1-VDR/Nurr1 cis-element may function as a switch between induction (osteocytes) and repression (adipocytes) of FGF23, depending on the cellular setting of transcription factors. Moreover, experiments demonstrate that a 1 kb mouse FGF23 promoter-reporter construct, transfected into MC3T3-E1 osteoblast-like cells, responds to a high calcium challenge with a statistically significant 1.7- to 2.0-fold enhancement of transcription. Thus, the FGF23 proximal promoter harbors cis elements that drive responsiveness to 1,25D and calcium, agents that induce FGF23 to curtail the pathologic consequences of their excess. PMID:26148725

  20. Measurements of Intracellular Ca2+ Content and Phosphatidylserine Exposure in Human Red Blood Cells: Methodological Issues

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    Mauro C. Wesseling

    2016-06-01

    Full Text Available Background/Aims: The increase of the intracellular Ca2+ content as well as the exposure of phosphatidylserine (PS on the outer cell membrane surface after activation of red blood cells (RBCs by lysophosphatidic acid (LPA has been investigated by a variety of research groups. Carrying out experiments, which we described in several previous publications, we observed some discrepancies when comparing data obtained by different investigators within our research group and also between batches of LPA. In addition, we found differences comparing the results of double and single labelling experiments (for Ca2+ and PS. Furthermore, the results of PS exposure depended on the fluorescent dye used (annexin V-FITC versus annexin V alexa fluor® 647. Therefore, it seems necessary to investigate these methodological approaches in more detail to be able to quantify results and to compare data obtained by different research groups. Methods: The intracellular Ca2+ content and the PS exposure of RBCs separated from whole blood have been investigated after treatment with LPA (2.5 µM obtained from three different companies (Sigma-Aldrich, Cayman Chemical Company, and Santa Cruz Biotechnology Inc.. Fluo-4 and x-rhod-1 have been used to detect intracellular Ca2+ content, annexin V alexa fluor® 647 and annexin V-FITC have been used for PS exposure measurements. Both parameters (Ca2+ content, PS exposure were studied using flow cytometry and fluorescence microscopy. Results: The percentage of RBCs showing increased intracellular Ca2+ content as well as PS exposure changes significantly between different LPA manufacturers as well as on the condition of mixing of LPA with the RBC suspension. Furthermore, the percentage of RBCs showing PS exposure is reduced in double labelling compared to single labelling experiments and depends also on the fluorescent dye used. Finally, data on Ca2+ content are slightly affected whereas PS exposure data are not affected significantly

  1. Role of LPAR3, PKC and EGFR in LPA-induced cell migration in oral squamous carcinoma cells

    International Nuclear Information System (INIS)

    Oral squamous cell carcinoma is an aggressive neoplasm with serious morbidity and mortality, which typically spreads through local invasive growth. Lysophosphatidic acid (LPA) is involved in a number of biological processes, and may have a role in cancer cell migration and invasiveness. LPA is present in most tissues and can activate cells through six different LPA receptors (LPAR1-6). Although LPA is predominantly promigratory, some of the receptors may have antimigratory effects in certain cells. The signalling mechanisms of LPA are not fully understood, and in oral carcinoma cells the specific receptors and pathways involved in LPA-stimulated migration are unknown. The oral carcinoma cell lines E10, SCC-9, and D2 were investigated. Cell migration was studied in a scratch wound assay, and invasion was demonstrated in organotypic three dimensional co-cultures. Protein and mRNA expression of LPA receptors was studied with Western blotting and qRT-PCR. Activation of signalling proteins was examined with Western blotting and isoelectric focusing, and signalling mechanisms were further explored using pharmacological agents and siRNA directed at specific receptors and pathways. LPA stimulated cell migration in the two oral carcinoma cell lines E10 and SCC-9, but was slightly inhibitory in D2. The receptor expression profile and the effects of specific pharmacological antagonist and agonists indicated that LPA-stimulated cell migration was mediated through LPAR3 in E10 and SCC-9. Furthermore, in both these cell lines, the stimulation by LPA was dependent on PKC activity. However, while LPA induced transactivation of EGFR and the stimulated migration was blocked by EGFR inhibitors in E10 cells, LPA did not induce EGFR transactivation in SCC-9 cells. In D2 cells, LPA induced EGFR transactivation, but this was associated with slowing of a very high inherent migration rate in these cells. The results demonstrate LPA-stimulated migration in oral carcinoma cells through LPAR3

  2. microRNA-146a inhibits G protein-coupled receptor-mediated activation of NF-κB by targeting CARD10 and COPS8 in gastric cancer

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    Crone Stephanie

    2012-09-01

    Full Text Available Abstract Background Gastric cancer is the second most common cause of cancer-related death in the world. Inflammatory signals originating from gastric cancer cells are important for recruiting inflammatory cells and regulation of metastasis of gastric cancer. Several microRNAs (miRNA have been shown to be involved in development and progression of gastric cancer. miRNA-146a (miR-146a is a modulator of inflammatory signals, but little is known about its importance in gastric cancer. We therefore wanted to identify targets of miR-146a in gastric cancer and examine its biological roles. Results The expression of miR-146a was evaluated by quantitative PCR (qPCR and found up-regulated in the gastrin knockout mice, a mouse model of gastric cancer, and in 73% of investigated human gastric adenocarcinomas. Expression of miR-146a by gastric cancer cells was confirmed by in situ hybridization. Global analysis of changes in mRNA levels after miR-146a transfection identified two transcripts, caspase recruitment domain-containing protein 10 (CARD10 and COP9 signalosome complex subunit 8 (COPS8, as new miR-146a targets. qPCR, Western blotting and luciferase assays confirmed these transcripts as direct miR-146a targets. CARD10 and COPS8 were shown to be part of the G protein-coupled receptor (GPCR pathway of nuclear factor-kappaB (NF-kappaB activation. Lysophosphatidic acid (LPA induces NF-kappaB activation via this pathway and over-expression of miR-146a inhibited LPA-induced NF-kappaB activation, reduced LPA-induced expression of tumor-promoting cytokines and growth factors and inhibited monocyte attraction. Conclusions miR-146a expression is up-regulated in a majority of gastric cancers where it targets CARD10 and COPS8, inhibiting GPCR-mediated activation of NF-kappaB, thus reducing expression of NF-kappaB-regulated tumor-promoting cytokines and growth factors. By targeting components of several NF-kappaB-activating pathways, miR-146a is a key component in

  3. Regulators of G-Protein signaling RGS10 and RGS17 regulate chemoresistance in ovarian cancer cells

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    Ali Mourad W

    2010-11-01

    Full Text Available Abstract Background A critical therapeutic challenge in epithelial ovarian carcinoma is the development of chemoresistance among tumor cells following exposure to first line chemotherapeutics. The molecular and genetic changes that drive the development of chemoresistance are unknown, and this lack of mechanistic insight is a major obstacle in preventing and predicting the occurrence of refractory disease. We have recently shown that Regulators of G-protein Signaling (RGS proteins negatively regulate signaling by lysophosphatidic acid (LPA, a growth factor elevated in malignant ascites fluid that triggers oncogenic growth and survival signaling in ovarian cancer cells. The goal of this study was to determine the role of RGS protein expression in ovarian cancer chemoresistance. Results In this study, we find that RGS2, RGS5, RGS10 and RGS17 transcripts are expressed at significantly lower levels in cells resistant to chemotherapy compared with parental, chemo-sensitive cells in gene expression datasets of multiple models of chemoresistance. Further, exposure of SKOV-3 cells to cytotoxic chemotherapy causes acute, persistent downregulation of RGS10 and RGS17 transcript expression. Direct inhibition of RGS10 or RGS17 expression using siRNA knock-down significantly reduces chemotherapy-induced cell toxicity. The effects of cisplatin, vincristine, and docetaxel are inhibited following RGS10 and RGS17 knock-down in cell viability assays and phosphatidyl serine externalization assays in SKOV-3 cells and MDR-HeyA8 cells. We further show that AKT activation is higher following RGS10 knock-down and RGS 10 and RGS17 overexpression blocked LPA mediated activation of AKT, suggesting that RGS proteins may blunt AKT survival pathways. Conclusions Taken together, our data suggest that chemotherapy exposure triggers loss of RGS10 and RGS17 expression in ovarian cancer cells, and that loss of expression contributes to the development of chemoresistance, possibly

  4. Characterization of Microvesicles Released from Human Red Blood Cells

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    Duc Bach Nguyen

    2016-03-01

    Full Text Available Background/Aims: Extracellular vesicles (EVs are spherical fragments of cell membrane released from various cell types under physiological as well as pathological conditions. Based on their size and origin, EVs are classified as exosome, microvesicles (MVs and apoptotic bodies. Recently, the release of MVs from human red blood cells (RBCs under different conditions has been reported. MVs are released by outward budding and fission of the plasma membrane. However, the outward budding process itself, the release of MVs and the physical properties of these MVs have not been well investigated. The aim of this study is to investigate the formation process, isolation and characterization of MVs released from RBCs under conditions of stimulating Ca2+ uptake and activation of protein kinase C. Methods: Experiments were performed based on single cell fluorescence imaging, fluorescence activated cell sorter/flow cytometer (FACS, scanning electron microscopy (SEM, atomic force microscopy (AFM and dynamic light scattering (DLS. The released MVs were collected by differential centrifugation and characterized in both their size and zeta potential. Results: Treatment of RBCs with 4-bromo-A23187 (positive control, lysophosphatidic acid (LPA, or phorbol-12 myristate-13 acetate (PMA in the presence of 2 mM extracellular Ca2+ led to an alteration of cell volume and cell morphology. In stimulated RBCs, exposure of phosphatidylserine (PS and formation of MVs were observed by using annexin V-FITC. The shedding of MVs was also observed in the case of PMA treatment in the absence of Ca2+, especially under the transmitted bright field illumination. By using SEM, AFM and DLS the morphology and size of stimulated RBCs, MVs were characterized. The sizes of the two populations of MVs were 205.8 ± 51.4 nm and 125.6 ± 31.4 nm, respectively. Adhesion of stimulated RBCs and MVs was observed. The zeta potential of MVs was determined in the range from - 40 mV to - 10 m

  5. Novel point mutations attenuate autotaxin activity

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    Stracke Mary L

    2009-02-01

    Full Text Available Abstract Background The secreted enzyme autotaxin (ATX stimulates tumor cell migration, tumorigenesis, angiogenesis, and metastasis. ATX hydrolyzes nucleotides, but its hydrolysis of lysophospholipids to produce lysophosphatidic acid (LPA accounts for its biological activities. ATX has been identified only as a constitutively active enzyme, and regulation of its activity is largely unexplored. In spite of its presence in plasma along with abundant putative substrate LPC, the product LPA is found in plasma at unexpectedly low concentrations. It is plausible that the LPA-producing activity of ATX is regulated by its expression and by access to substrate(s. For this reason studying the interaction of enzyme with substrate is paramount to understanding the regulation of LPA production. Results In this study we determine ATX hydrolytic activities toward several artificial and natural substrates. Two novel point mutations near the enzyme active site (H226Q and H434Q confer attenuated activity toward all substrates tested. The Vmax for LPC compounds depends upon chain length and saturation; but this order does not differ among wild type and mutants. However the mutant forms show disproportionately low activity toward two artificial substrates, pNpTMP and FS-3. The mutant forms did not significantly stimulate migration responses at concentrations that produced a maximum response for WT-ATX, but this defect could be rescued by inclusion of exogenous LPC. Conclusion H226Q-ATX and H434Q-ATX are the first point mutations of ATX/NPP2 demonstrated to differentially impair substrate hydrolysis, with hydrolysis of artificial substrates being disproportionately lower than that of LPC. This implies that H226 and H434 are important for substrate interaction. Assays that rely on hydrolyses of artificial substrates (FS-3 and pNpTMP, or that rely on hydrolysis of cell-derived substrate, might fail to detect certain mutated forms of ATX that are nonetheless capable of

  6. Autotaxin and LPA receptors represent potential molecular targets for the radiosensitization of murine glioma through effects on tumor vasculature.

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    Stephen M Schleicher

    Full Text Available Despite wide margins and high dose irradiation, unresectable malignant glioma (MG is less responsive to radiation and is uniformly fatal. We previously found that cytosolic phospholipase A2 (cPLA(2 is a molecular target for radiosensitizing cancer through the vascular endothelium. Autotaxin (ATX and lysophosphatidic acid (LPA receptors are downstream from cPLA(2 and highly expressed in MG. Using the ATX and LPA receptor inhibitor, α-bromomethylene phosphonate LPA (BrP-LPA, we studied ATX and LPA receptors as potential molecular targets for the radiosensitization of tumor vasculature in MG. Treatment of Human Umbilical Endothelial cells (HUVEC and mouse brain microvascular cells bEND.3 with 5 µmol/L BrP-LPA and 3 Gy irradiation showed decreased clonogenic survival, tubule formation, and migration. Exogenous addition of LPA showed radioprotection that was abrogated in the presence of BrP-LPA. In co-culture experiments using bEND.3 and mouse GL-261 glioma cells, treatment with BrP-LPA reduced Akt phosphorylation in both irradiated cell lines and decreased survival and migration of irradiated GL-261 cells. Using siRNA to knock down LPA receptors LPA1, LPA2 or LPA3 in HUVEC, we demonstrated that knockdown of LPA2 but neither LPA1 nor LPA3 led to increased viability and proliferation. However, knockdown of LPA1 and LPA3 but not LPA2 resulted in complete abrogation of tubule formation implying that LPA1 and LPA3 on endothelial cells are likely targets of BrP-LPA radiosensitizing effect. Using heterotopic tumor models of GL-261, mice treated with BrP-LPA and irradiation showed a tumor growth delay of 6.8 days compared to mice treated with irradiation alone indicating that inhibition of ATX and LPA receptors may significantly improve malignant glioma response to radiation therapy. These findings identify ATX and LPA receptors as molecular targets for the development of radiosensitizers for MG.

  7. Modulation of membrane phospholipids, the cytosolic calcium influx and cell proliferation following treatment of B16-F10 cells with recombinant phospholipase-D from Loxosceles intermedia (brown spider) venom.

    Science.gov (United States)

    Wille, Ana Carolina Martins; Chaves-Moreira, Daniele; Trevisan-Silva, Dilza; Magnoni, Mariana Gabriel; Boia-Ferreira, Marianna; Gremski, Luiza Helena; Gremski, Waldemiro; Chaim, Olga Meiri; Senff-Ribeiro, Andrea; Veiga, Silvio Sanches

    2013-06-01

    The mechanism through which brown spiders (Loxosceles genus) cause dermonecrosis, dysregulated inflammatory responses, hemolysis and platelet aggregation, which are effects reported following spider bites, is currently attributed to the presence of phospholipase-D in the venom. In the present investigation, through two-dimensional immunoblotting, we observed immunological cross-reactivity for at least 25 spots in crude Loxosceles intermedia venom, indicating high expression levels for different isoforms of phospholipase-D. Using a recombinant phospholipase-D from the venom gland of L. intermedia (LiRecDT1) in phospholipid-degrading kinetic experiments, we determined that this phospholipase-D mainly hydrolyzes synthetic sphingomyelin in a time-dependent manner, generating ceramide 1-phosphate plus choline, as well as lysophosphatidylcholine, generating lysophosphatidic acid plus choline, but exhibits little activity against phosphatidylcholine. Through immunofluorescence assays with antibodies against LiRecDT1 and using a recombinant GFP-LiRecDT1 fusion protein, we observed direct binding of LiRecDT1 to the membrane of B16-F10 cells. We determined that LiRecDT1 hydrolyzes phospholipids in detergent extracts and from ghosts of B16-F10 cells, generating choline, indicating that the enzyme can access and modulate and has activity against membrane phospholipids. Additionally, using Fluo-4, a calcium-sensitive fluorophore, it was shown that treatment of cells with phospholipase-D induced an increase in the calcium concentration in the cytoplasm, but without altering viability or causing damage to cells. Finally, based on the known endogenous activity of phospholipase-D as an inducer of cell proliferation and the fact that LiRecDT1 binds to the cell surface, hydrolyzing phospholipids to generate bioactive lipids, we employed LiRecDT1 as an exogenous source of phospholipase-D in B16-F10 cells. Treatment of the cells was effective in increasing their proliferation in a

  8. Downregulation of kinin B1 receptor function by B2 receptor heterodimerization and signaling.

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    Zhang, Xianming; Brovkovych, Viktor; Zhang, Yongkang; Tan, Fulong; Skidgel, Randal A

    2015-01-01

    Signaling through the G protein-coupled kinin receptors B1 (kB1R) and B2 (kB2R) plays a critical role in inflammatory responses mediated by activation of the kallikrein-kinin system. The kB2R is constitutively expressed and rapidly desensitized in response to agonist whereas kB1R expression is upregulated by inflammatory stimuli and it is resistant to internalization and desensitization. Here we show that the kB1R heterodimerizes with kB2Rs in co-transfected HEK293 cells and natively expressing endothelial cells, resulting in significant internalization and desensitization of the kB1R response in cells pre-treated with kB2R agonist. However, pre-treatment of cells with kB1R agonist did not affect subsequent kB2R responses. Agonists of other G protein-coupled receptors (thrombin, lysophosphatidic acid) had no effect on a subsequent kB1R response. The loss of kB1R response after pretreatment with kB2R agonist was partially reversed with kB2R mutant Y129S, which blocks kB2R signaling without affecting endocytosis, or T342A, which signals like wild type but is not endocytosed. Co-endocytosis of the kB1R with kB2R was dependent on β-arrestin and clathrin-coated pits but not caveolae. The sorting pathway of kB1R and kB2R after endocytosis differed as recycling of kB1R to the cell surface was much slower than that of kB2R. In cytokine-treated human lung microvascular endothelial cells, pre-treatment with kB2R agonist inhibited kB1R-mediated increase in transendothelial electrical resistance (TER) caused by kB1R stimulation (to generate nitric oxide) and blocked the profound drop in TER caused by kB1R activation in the presence of pyrogallol (a superoxide generator). Thus, kB1R function can be downregulated by kB2R co-endocytosis and signaling, suggesting new approaches to control kB1R signaling in pathological conditions. PMID:25289859

  9. Cannabinoid receptors in brain: pharmacogenetics, neuropharmacology, neurotoxicology, and potential therapeutic applications.

    Science.gov (United States)

    Onaivi, Emmanuel S

    2009-01-01

    Much progress has been achieved in cannabinoid research. A major breakthrough in marijuana-cannabinoid research has been the discovery of a previously unknown but elaborate endogenous endocannabinoid system (ECS), complete with endocannabinoids and enzymes for their biosynthesis and degradation with genes encoding two distinct cannabinoid (CB1 and CB2) receptors (CBRs) that are activated by endocannabinoids, cannabinoids, and marijuana use. Physical and genetic localization of the CBR genes CNR1 and CNR2 have been mapped to chromosome 6 and 1, respectively. A number of variations in CBR genes have been associated with human disorders including osteoporosis, attention deficit hyperactivity disorder (ADHD), posttraumatic stress disorder (PTSD), drug dependency, obesity, and depression. Other family of lipid receptors including vanilloid (VR1) and lysophosphatidic acid (LPA) receptors appear to be related to the CBRs at the phylogenetic level. The ubiquitous abundance and differential distribution of the ECS in the human body and brain along with the coupling to many signal transduction pathways may explain the effects in most biological system and the myriad behavioral effects associated with smoking marijuana. The neuropharmacological and neuroprotective features of phytocannabinoids and endocannabinoid associated neurogenesis have revealed roles for the use of cannabinoids in neurodegenerative pathologies with less neurotoxicity. The remarkable progress in understanding the biological actions of marijuana and cannabinoids have provided much richer results than previously appreciated cannabinoid genomics and raised a number of critical issues on the molecular mechanisms of cannabinoid induced behavioral and biochemical alterations. These advances will allow specific therapeutic targeting of the different components of the ECS in health and disease. This review focuses on these recent advances in cannabinoid genomics and the surprising new fundamental roles that the

  10. 溶血磷脂酸对小胶质细胞活化和吞噬功能的影响%Effect of LPA on activation and endocytosis of microglia

    Institute of Scientific and Technical Information of China (English)

    付佩彩; 喻志源; 刘淼; 唐荣华; 王伟; 骆翔

    2012-01-01

    目的:观察溶血磷脂酸(LPA)对小胶质细胞活化及其吞噬功能的影响.方法:小胶质细胞系BV2细胞复苏后传代培养,取对数生长期细胞分为对照组和LPA组,LPA干预30 min后各组加入FITC标记的葡聚糖,在干预后1h、2h后收取细胞,用流式细胞仪测定各孔FITC-葡聚糖微粒吞噬率及荧光强度.结果:LPA干预1h时,吞噬率较对照组明显增加(P<0.05),2h时两组细胞吞噬率达到最高值.LPA干预后1h及2h时,吞噬荧光强度较对照组明显增加(P<0.01),而且吞噬率与荧光强度的乘积较对照组亦明显增加(P<0.01).结论:LPA可提高BV2细胞活化及吞噬功能,提示LPA在小胶质细胞活化及吞噬过程中发挥了重要作用.%Objectives To observe the effect of lysophosphatidic acid (LPA) on activation and endocytosis of microglia. Methods;BV2 cells in exponential phase were randomly divided into two groups: control group and LPA group. At time points (1 h and 2 h), cells were harvested for later use. FCM was used to detect the phagocytic rate of FITC-dextran and the fluorescence intensity. Results: As compared with control group, the phagocytic rate of FITC-dextran in LPA group was significantly increased at 1 h (P<0.05). At 2 h, the phagocytic rate of FITC-dextran reached the peak in both two groups. At time points (1 h and 2 h), as compared with control group, the fluorescence intensity in LPA group was increased (P<0. 01) and the product of phagocytic rate and fluorescence intensity in LPA group was significantly higher than that of control group (P<0. 01). Conclusion: LPA can promote activation and endocytosis of microglia in vitro, suggesting the roles of LPA in activation and endocytosis of microglia.

  11. CARMA3: A novel scaffold protein in regulation of NF-κB activation and diseases

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    CARD recruited membrane associated protein 3 (CARMA3) is a novel scaffold protein. It belongs to the CARMA protein family, and is known to activate nuclear factor (NF)- κB. However, it is still unknown which receptor functions upstream of CARMA3 to trigger NF-κB activation. Recently, several studies have demonstrated that CARMA3 serves as an indispensable adaptor protein in NF-κB signaling under some G protein-coupled receptors (GP- CRs), such as lysophosphatidic acid (LPA) receptor and angiotensin (Ang) Ⅱ receptor. Mechanistically, CARMA3 recruits its essential downstream molecules Bcl10 and MALT1 to form the CBM (CARMA3-Bcl10-MALT1) signalosome whereby it triggers NF-κB activation. GPCRs and NF-κB play pivotal roles in the regulation of various cellular functions, therefore, aberrant regulation of the GPCR/NF-κB signaling axis leads to the development of many types of diseases, such as cancer and atherogenesis. Recently, the GPCR/CARMA3/NF-κB signaling axis has been confirmed in these specific diseases and it plays crucial roles in the pathogenesis of disease progression. In ovarian cancer cell lines, knockdown of CARMA3 abolishes LPA receptor-induced NF-κB activation, and reduces LPA-induced ovarian cancer invasion. In vascular smooth cells, downregulation of CARMA3 substantially impairs Ang-Ⅱ-receptor-induced NF-κB activation, and in vivo studies have confirmed that Bcl10- deficient mice are protected from developing Ang-Ⅱ-receptor-induced atherosclerosis and aortic aneurysms. In this review, we summarize the biology of CARMA3, describe the role of the GPCR/CARMA3/NF-κB signaling axis in ovarian cancer and atherogenesis, and speculate about the potential roles of this signaling axis in other types of cancer and diseases. With a significant increase in the identification of LPA- and Ang-Ⅱ-like ligands, such as endothelin-1, which also activates NF-κB via CARMA3 and contributes to the development of many diseases, CARMA3 is emerging as a novel

  12. S-Nitrosothiols modulate G protein-coupled receptor signaling in a reversible and highly receptor-specific manner

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    Mönkkönen Kati S

    2005-04-01

    Full Text Available Abstract Background Recent studies indicate that the G protein-coupled receptor (GPCR signaling machinery can serve as a direct target of reactive oxygen species, including nitric oxide (NO and S-nitrosothiols (RSNOs. To gain a broader view into the way that receptor-dependent G protein activation – an early step in signal transduction – might be affected by RSNOs, we have studied several receptors coupling to the Gi family of G proteins in their native cellular environment using the powerful functional approach of [35S]GTPγS autoradiography with brain cryostat sections in combination with classical G protein activation assays. Results We demonstrate that RSNOs, like S-nitrosoglutathione (GSNO and S-nitrosocysteine (CysNO, can modulate GPCR signaling via reversible, thiol-sensitive mechanisms probably involving S-nitrosylation. RSNOs are capable of very targeted regulation, as they potentiate the signaling of some receptors (exemplified by the M2/M4 muscarinic cholinergic receptors, inhibit others (P2Y12 purinergic, LPA1lysophosphatidic acid, and cannabinoid CB1 receptors, but may only marginally affect signaling of others, such as adenosine A1, μ-opioid, and opiate related receptors. Amplification of M2/M4 muscarinic responses is explained by an accelerated rate of guanine nucleotide exchange, as well as an increased number of high-affinity [35S]GTPγS binding sites available for the agonist-activated receptor. GSNO amplified human M4 receptor signaling also under heterologous expression in CHO cells, but the effect diminished with increasing constitutive receptor activity. RSNOs markedly inhibited P2Y12 receptor signaling in native tissues (rat brain and human platelets, but failed to affect human P2Y12 receptor signaling under heterologous expression in CHO cells, indicating that the native cellular signaling partners, rather than the P2Y12 receptor protein, act as a molecular target for this action. Conclusion These in vitro studies

  13. Intrahepatic expression of genes related to metabotropic receptors in chronic hepatitis

    Institute of Scientific and Technical Information of China (English)

    Andrzej Cie(s)la,; Maciej Ku(s)mider,; Agata Faron-Górecka; Marta Dziedzicka-Wasylewska; Monika Bociaga-Jasik; Danuta Owczarek; Irena Cie(c)ko-Michalska

    2012-01-01

    AIM:To screen for genes related to metabotropic receptors that might be involved in the development of chronic hepatitis.METHODS:Assessment of 20 genes associated with metabotropic receptors was performed in liver specimens obtained by punch biopsy from 12 patients with autoimmune and chronic hepatitis type B and C.For this purpose,a microarray with low integrity grade and with oligonucleotide DNA probes complementary to target transcripts was used.Evaluation of gene expression was performed in relation to transcript level,correlation between samples and grouping of clinical parameters used in chronic hepatitis assessment.Clinical markers of chronic hepatitis included alanine and aspartate aminotransferase,γ-glutamyltranspeptidase,alkaline phosphatase and cholinesterase activity,levels of iron ions,total cholesterol,triglycerides,albumin,glucose,hemoglobin,platelets,histological analysis of inflammatory and necrotic status,fibrosis according to METAVIR score,steatosis,as well as anthropometric body mass index,waist/hip index,percentage of adipose tissue and liver size in ultrasound examination.Gender,age,concomitant diseases and drugs were also taken into account.Validation of oligonucleotide microarray gene expression results was done with the use of quantitative real-time polymerase chain reaction (qRT-PCR).RESULTS:The highest (0.002 < P < 0.046) expression among genes encoding main components of metabotropic receptor pathways,such as the a subunit of G-coupled protein,phosphoinositol-dependent protein kinase or arrestin was comparable to that of angiotensinogen synthesized in the liver.Carcinogenesis suppressor genes,such as chemokine ligand 4,transcription factor early growth response protein 1 and lysophosphatidic acid receptor,were characterized by the lowest expression (0.002 < P < 0.046),while the factor potentially triggering hepatic cancer,transcription factor JUN-B,had a 20-fold higher expression.The correlation between expression of genes of

  14. A designated centre for people with disabilities operated by St Michael's House, Dublin 5

    LENUS (Irish Health Repository)

    Bennett, Gayle

    2011-02-01

    Abstract Background NET1, a RhoA guanine exchange factor, is up-regulated in gastric cancer (GC) tissue and drives the invasive phenotype of this disease. In this study, we aimed to determine the role of NET1 in GC by monitoring the proliferation, motility and invasion of GC cells in which NET1 has been stably knocked down. Additionally, we aimed to determine NET1-dependent transcriptomic events that occur in GC. Methods An in vitro model of stable knockdown of NET1 was achieved in AGS human gastric adenocarcinoma cells via lentiviral mediated transduction of short-hairpin (sh) RNA targeting NET1. Knockdown was assessed using quantitative PCR. Cell proliferation was assessed using an MTS assay and cell migration was assessed using a wound healing scratch assay. Cell invasion was assessed using a transwell matrigel invasion assay. Gene expression profiles were examined using affymetrix oligonucleotide U133A expression arrays. A student\\'s t test was used to determine changes of statistical significance. Results GC cells were transduced with NET1 shRNA resulting in a 97% reduction in NET1 mRNA (p < 0.0001). NET1 knockdown significantly reduced the invasion and migration of GC cells by 94% (p < 0.05) and 24% (p < 0.001) respectively, while cell proliferation was not significantly altered following NET1 knockdown. Microarray analysis was performed on non-target and knockdown cell lines, treated with and without 10 μM lysophosphatidic acid (LPA) allowing us to identify NET1-dependent, LPA-dependent and NET1-mediated LPA-induced gene transcription. Differential gene expression was confirmed by quantitative PCR. Shortlisted NET1-dependent genes included STAT1, TSPAN1, TGFBi and CCL5 all of which were downregulatd upon NET1 downregulation. Shortlisted LPA-dependent genes included EGFR and PPARD where EGFR was upregulated and PPARD was downregulated upon LPA stimulation. Shortlisted NET1 and LPA dependent genes included IGFR1 and PIP5K3. These LPA induced genes were

  15. ATX-LPA生物学功能及其在乳腺癌中的研究进展%Biological function of ATX-LPA axis and its progress in cancer

    Institute of Scientific and Technical Information of China (English)

    邵营波; 刘慧

    2015-01-01

    OBJECTIVE To summarize the recent advances in the biological function of ATX-LPA axis and its association with breast cancer.METHODS The PubMed and CNKI database from Jan.2006 to Apr.2014 were filtered by search teams " breast cancer,autotaxin,lysophosphatidic acid".Exclusion criteria:non breast cancer and non ATX-LPA.Inclusion criterion:Recent literature about the biological function of ATX-LPA axis.Latest advances regards to some of the key findings of ATX-LPA signalling axis in breast cancer.Totally 45 literatures were enrolled in the study according to the criterions.RESULTS The ATX-LPA signaling axis acts on a series of high-affinity cell surface G protein-coupled receptors (GPCRs),leading to diverse biological actions.Many subsequent studies have combined to establish an important function for the ATX-LPA axis in mammary tumourigenesis,metastasis and drug resistance,suggesting that the ATX-LPA signalling axis is a novel target in breast cancer.The focus of researches now turn to understanding the mechanisms by which ATX and LPA promote mammary tumourigenesis,metastasis and and drug resistance.Researches about the inhibitors and monoclonal antibodies in the above-mentioned mechanisms have achieved some improvements in laboratory and clinical studies.CONCLUSIONS The biological function of ATX-LPA signaling axis in mammary tumourigenesis,metastasis and drug resistance has been confirmed,more laboratory studies are needed to provide powerful evidence to clarify detailed mechanism between ATX-LPA signaling axis and breast cancer.Targeting the ATX-LPA signalling axis for drug development may further improve the outcomes in patients of breast cancer.%目的 总结近年来关于自分泌运动因子-溶血磷脂酸(autotaxin-lysophosphatidic acid,ATX-LPA)生物学功能及其在乳腺癌中的研究进展.方法 应用PubMed和CNKI期刊全文数据库检索系统,以“乳腺肿瘤、自分泌运动因子、溶血磷脂酸”为关键词,检索2006-01-2014-04

  16. Clinical significance of combined detection of three tumor markers in the diagnosis of non-small cell lung cancer%3种肿瘤标志物联合检测对非小细胞肺癌诊断的临床应用价值

    Institute of Scientific and Technical Information of China (English)

    张小南; 卢祥珍; 颜永乾

    2015-01-01

    目的:探讨溶血磷脂酸(LPA)、肿瘤特异性生长因子(TSGF)及糖类抗原19-9(CA19-9)联合检测对非小细胞肺癌(NSCLC)诊断的临床价值。方法对102例 NSCLC 患者(NSCLC 组)、63例肺良性疾病患者(肺良性疾病组)、50例体检健康者(健康对照组)的血清 LPA、TSGF 和 CA19-9水平进行测定,分析3种肿瘤标志物联合检测对 NSCLC 的诊断价值。结果NSCLC 组血清 LPA、TSGF、CA19-9水平高于肺良性疾病组与健康对照组,差异均有统计学意义(均 P <0.05)。LPA、TSGF 和CA19-9联合检测的灵敏度和特异度分别为68.63%、37.17%,其灵敏度高于单项检测和2项联合检测。结论肿瘤标志物联合检测可提高对 NSCLC 诊断的灵敏度,为 NSCLC 的诊断提供可靠的实验依据。%Objective To explore the clinical value of combined detection of lysophosphatidic acid(LPA),tumor-specific growth factor(TSGF)and carbohydrate antigen 1 9-9(CA1 9-9)in the diagnosis of non-small cell lung cancer(NSCLC).Methods The ser-um levels of LPA,TSGF and CA1 9-9 in 97 cases of patients with NSCLC(NSCLC group),43 cases of patients with benign lung disease(benign lung disease group)and 50 cases of healthy individuals(healthy control group)were detected,and diagnostic value of combined detection of these three tumor makers in the diagnosis of NSCLC was analysed.Results Serum levels of LPA,TSGF and CA1 9-9 in the NSCLC group were significantly higher than those in the benign lung disease group and healthy control group,had statistically significant differences(P <0.05).The sensitivity and specificity of combined detection of LPA,TSGF and CA1 9-9 was 68.63% and 37.1 7% respectively,and the sensitivity was higher than that of single detection and combined detection of any two of the three indicators.Conclusion Combined detection of tumor markers can improve the sensitivity in the diagnosis of NSCLC, which could provide reliable laboratory references for diagnosing NSCLC.

  17. Dynamic spatial panels : models, methods, and inferences

    NARCIS (Netherlands)

    Elhorst, J. Paul

    2012-01-01

    This paper provides a survey of the existing literature on the specification and estimation of dynamic spatial panel data models, a collection of models for spatial panels extended to include one or more of the following variables and/or error terms: a dependent variable lagged in time, a dependent

  18. Predicting Group-Level Outcome Variables from Variables Measured at the Individual Level: A Latent Variable Multilevel Model

    Science.gov (United States)

    Croon, Marcel A.; van Veldhoven, Marc J. P. M.

    2007-01-01

    In multilevel modeling, one often distinguishes between macro-micro and micro-macro situations. In a macro-micro multilevel situation, a dependent variable measured at the lower level is predicted or explained by variables measured at that lower or a higher level. In a micro-macro multilevel situation, a dependent variable defined at the higher…

  19. Summary of the parallel session on hadron dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Corcoran, M.D. [Rice Univ., Houston, TX (United States); Carroll, A.S. [Brookhaven National Lab., Upton, NY (United States)

    1994-12-31

    The hadron dynamics sessions focussed on A-dependent effects in hard interactions and heavy quark production, as well as color transparency effects. Other topics covered included exclusive processes and spin effects.

  20. Psychological Time and Sociology: A Research Agenda.

    Science.gov (United States)

    Hogan, H. Wayne

    1979-01-01

    Psychological time is hypothesized as potentially being both an independent and a dependent dimension associated with such sociological and psychological phenomena as social change, environmental design, personal space, esthetics, and color considerations. (Author)

  1. An exploration of the strategic implementation of marketing communication within social networking communication context

    OpenAIRE

    Cloete, Ewoudt

    2012-01-01

    Taking into consideration the dramatic changes ushered in by the exponential growth of social networking, marketers are left without a dependable framework on how to implement social networking strategically and in alignment with other modern as well as traditional marketing disciplines. In light of this, the study aims to explore the strategic implementation of social networking within the context of a dependable marketing theoretical model, known as the marketing communications mix. D...

  2. Anxiety, Affect, Self-Esteem, and Stress: Mediation and Moderation Effects on Depression

    OpenAIRE

    Nima, Ali Al; Rosenberg, Patricia; Archer, Trevor; Garcia, Danilo

    2013-01-01

    Background Mediation analysis investigates whether a variable (i.e., mediator) changes in regard to an independent variable, in turn, affecting a dependent variable. Moderation analysis, on the other hand, investigates whether the statistical interaction between independent variables predict a dependent variable. Although this difference between these two types of analysis is explicit in current literature, there is still confusion with regard to the mediating and moderating effects of differ...

  3. Anxiety, Affect, Self-Esteem, and Stress: Mediation and Moderation Effects on Depression

    OpenAIRE

    Ali Al Nima; Patricia Rosenberg; Trevor Archer; Danilo Garcia

    2013-01-01

    BACKGROUND: Mediation analysis investigates whether a variable (i.e., mediator) changes in regard to an independent variable, in turn, affecting a dependent variable. Moderation analysis, on the other hand, investigates whether the statistical interaction between independent variables predict a dependent variable. Although this difference between these two types of analysis is explicit in current literature, there is still confusion with regard to the mediating and moderating effects of diffe...

  4. Chemoprotective activity of boldine: modulation of drug-metabolizing enzymes.

    Science.gov (United States)

    Kubínová, R; Machala, M; Minksová, K; Neca, J; Suchý, V

    2001-03-01

    Possible chemoprotective effects of the naturally occurring alkaloid boldine, a major alkaloid of boldo (Peumus boldus Mol.) leaves and bark, including in vitro modulations of drug-metabolizing enzymes in mouse hepatoma Hepa-1 cell line and mouse hepatic microsomes, were investigated. Boldine manifested inhibition activity on hepatic microsomal CYP1A-dependent 7-ethoxyresorufin O-deethylase and CYP3A-dependent testosterone 6 beta-hydroxylase activities and stimulated glutathione S-transferase activity in Hepa-1 cells. In addition to the known antioxidant activity, boldine could decrease the metabolic activation of other xenobiotics including chemical mutagens. PMID:11265593

  5. Effect of temperature on nitrate removal from polluted groundwater by biofilm technology; Efecto de la temperatura en la eliminacion de nitrato de aguas subterraneas contaminadas mediante sistemas de biopeliculas

    Energy Technology Data Exchange (ETDEWEB)

    Rua Ruiz, A. de la; Gonzalez Lopez, J.; Gomez Nieto, M. A.

    2007-07-01

    The effect of temperature on biofilm formation and denitrification activity was evaluated, using a lab-scale submerged filter for the denitrification of polluted groundwater, inoculated with activated sludge and with a selected strain of Pseudomonas sp. Different temperatures were tested: 5, 10, 20 and 30 degree centigree. Our results showed that colonization of the support material of the filter and denitrification capacity have a dependency with temperature, although this factor influences the process in a different way depending on the inoculum used. Nitrite concentration also showed a dependency with temperature. (Author) 12 refs.

  6. Neutron spin precession and test of linearity of wave mechanics

    International Nuclear Information System (INIS)

    A new experimental approach consisting in the search for a dependence of the Larmor precession of a neutron on the direction of the polarization vector is proposed to test the linearity of wave mechanics. The sensitivity of the experiment suggested is approximately four orders of magnitude higher than that achieved presently. 5 refs

  7. PARAMETRIC AND NON PARAMETRIC (MARS: MULTIVARIATE ADDITIVE REGRESSION SPLINES) LOGISTIC REGRESSIONS FOR PREDICTION OF A DICHOTOMOUS RESPONSE VARIABLE WITH AN EXAMPLE FOR PRESENCE/ABSENCE OF AMPHIBIANS

    Science.gov (United States)

    The purpose of this report is to provide a reference manual that could be used by investigators for making informed use of logistic regression using two methods (standard logistic regression and MARS). The details for analyses of relationships between a dependent binary response ...

  8. The Paralinguistic Encoding Capability of Children. Report from the Project on Studies of Instructional Programming for the Individual Student. Technical Report No. 441.

    Science.gov (United States)

    Plazewski, Joseph G.; Allen, Vernon L.

    A study was conducted of the capacity of sixth-grade children to communicate accurately paralinguistic affect. A dependent measure indicating the accuracy of paralinguistic communication of affect was obtained by comparing the level of affect which children intended to encode with ratings of vocal inflections from adult judges. Four independent…

  9. Tax Information for Refugees and Their Sponsors: Questions and Answers. Revised.

    Science.gov (United States)

    Internal Revenue Service (Dept. of Treasury), Washington, DC.

    This guide provides Federal income tax information for refugees and their sponsors. Issues covered in a question and answer format include: (1) the tax status of refugees; (2) the criteria for declaring a refugee a dependent; (3) deductions for contributions to refugees or organizations that support them; (4) the distinction between resident and…

  10. Resilience Approach for Medical Residents

    NARCIS (Netherlands)

    Bezemer, R.A.; Bos, E.W.

    2014-01-01

    Medical residents are in a vulnerable position. While still in training, they are responsible for patient care. They have a dependent relation with their supervisor and low decision latitude. An intervention was developed to increase individual and system resilience, addressing burnout, patient safe

  11. Energy Transition Initiative: Islands Playbook (Book)

    Energy Technology Data Exchange (ETDEWEB)

    2015-01-01

    The Island Energy Playbook (the Playbook) provides an action-oriented guide to successfully initiating, planning, and completing a transition to an energy system that primarily relies on local resources to eliminate a dependence on one or two imported fuels. It is intended to serve as a readily available framework that any community can adapt to organize its own energy transition effort.

  12. Rethinking the dependent variable in voting behavior: On the measurement and analysis of electoral utilities

    NARCIS (Netherlands)

    Eijk, van der Cees; Brug, van der Wouter; Kroh, Martin; Franklin, Mark

    2006-01-01

    As a dependent variable, party choice did not lend itself to analysis by means of powerful multivariate methods until the coming of discrete-choice models, most notably conditional logit and multinomial logit. These methods involve estimating effects on party preferences (utilities) that are post ho

  13. Longitudinal and Transverse Nuclear Shadowing

    OpenAIRE

    Barone, V.; M.; Genovese

    1996-01-01

    Nuclear shadowing arises from multiple scattering of the hadronic fluctuations of the virtual photon in a nucleus. We predict different longitudinal and transverse shadowing and an A-dependence of R which can be up to a 50% effect. The possibility of detecting nuclear effects on R at HERA is discussed.

  14. Can a Siphon Work in Vacuo?

    Science.gov (United States)

    Boatwright, Adrian L.; Puttick, Simon; Licence, Peter

    2011-01-01

    Used since the time of the ancient Egyptians as a method for transferring liquids from one vessel to another, the siphon is a dependable tool. Although, the act of siphoning beer from a fermentation barrel or wine from a demijohn is a skill that has been passed down from generation to generation, do we really know how the siphon works? It is…

  15. Purified tomato spotted wilt virus particles support both genome replication and transcription

    NARCIS (Netherlands)

    Knippenberg, van I.; Goldbach, R.W.; Kormelink, R.

    2002-01-01

    Purified Tomato spotted wilt virus particles were shown to support either genome replication or transcription in vitro, depending on the conditions chosen. Transcriptional activity was observed only upon addition of rabbit reticulocyte lysate, indicating a dependence on translation. Under these cond

  16. Predamage threshold electron emission from insulator and semiconductor surfaces

    International Nuclear Information System (INIS)

    Predamage electron emission shows a dependence on fluence, bandgap and wavelength consistent with multiphoton excitation across the bandgap and inconsistent with avalanche ionization and thermionic emission models. The electron emission scales with pulselength as 1/√T. 6 references, 8 figures, 1 table

  17. Validity of a Residualized Dependent Variable after Pretest Covariance Adjustments: Still the Same Variable?

    Science.gov (United States)

    Nimon, Kim; Henson, Robin K.

    2015-01-01

    The authors empirically examined whether the validity of a residualized dependent variable after covariance adjustment is comparable to that of the original variable of interest. When variance of a dependent variable is removed as a result of one or more covariates, the residual variance may not reflect the same meaning. Using the pretest-posttest…

  18. Matérn thinned Cox processes

    DEFF Research Database (Denmark)

    Andersen, Ina Trolle; Hahn, Ute

    2016-01-01

    and hard core behaviour can be achieved by applying a dependent Matérn thinning to a Cox process. An exact formula for the intensity of a Matérn thinned shot noise Cox process is derived from the Palm distribution. For the more general class of Matérn thinned Cox processes, formulae for the intensity...

  19. Matérn thinned Cox processes

    DEFF Research Database (Denmark)

    Andersen, Ina Trolle; Hahn, Ute

    of clustering and hard core behaviour can be achieved by applying a dependent Matérn thinning to a Cox process. An exact formula for the intensity of a Matérn thinned shot noise Cox process is derived from the Palm distribution. For the more general class of Matérn thinned Cox processes, formulae...

  20. 45 CFR 264.1 - What restrictions apply to the length of time Federal TANF assistance may be provided?

    Science.gov (United States)

    2010-10-01

    ..., or attempts at, physical or sexual abuse; (vi) Mental abuse; or (vii) Neglect or deprivation of... subjected to: (i) Physical acts that resulted in, or threatened to result in, physical injury to the individual; (ii) Sexual abuse; (iii) Sexual activity involving a dependent child; (iv) Being forced as...