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1.
Aust Dent J ; 65(1): 96-99, 2020 03.
Article in English | MEDLINE | ID: mdl-31659747

ABSTRACT

Intracranial abscess is a rare but life-threatening disease. There have been no reports on intracranial abscess induced by the residual primary tooth and the impacted successive permanent tooth with infection. We report on an interesting case of a 29-year-old man suffering from an epidural abscess, potentially caused by an infection of the residual primary maxillary right canine and the impacted permanent maxillary right canine. The patient recovered completely after prolonged antibiotic treatment and extraction of both of the suspected teeth. Fusobacterium sp. was isolated from the culture of a peripheral blood sample. This case alerts us to realize that the lack of suitable and timely intervention in oral conditions might produce a harmful effect on general health.


Subject(s)
Abscess , Tooth, Impacted , Adult , Cuspid/diagnostic imaging , Humans , Male , Maxilla , Tooth, Deciduous
2.
Mol Oral Microbiol ; 32(2): 131-141, 2017 04.
Article in English | MEDLINE | ID: mdl-27004566

ABSTRACT

Streptococcus mutans, the primary etiologic agent of dental caries, can gain access to the bloodstream and has been associated with cardiovascular disease. However, the roles of S. mutans in inflammation in cardiovascular disease remain unclear. The aim of this study was to examine cytokine production induced by S. mutans in human aortic endothelial cells (HAECs) and to evaluate the participation of toll-like receptors (TLRs) and cytoplasmic nucleotide-binding oligomerization domain (NOD) -like receptors in HAECs. Cytokine production by HAECs was determined using enzyme-linked immunosorbent assays, and the expression of TLRs and NOD-like receptors was evaluated by real-time polymerase chain reaction, flow cytometry and immunocytochemistry. The involvement of TLR2 and NOD2 in cytokine production by invaded HAECs was examined using RNA interference. The invasion efficiencies of S. mutans strains were evaluated by means of antibiotic protection assays. Five of six strains of S. mutans of various serotypes induced interleukin-6, interleukin-8 and monocyte chemoattractant protein-1 production by HAECs. All S. mutans strains upregulated TLR2 and NOD2 mRNA levels in HAECs. Streptococcus mutans Xc upregulated the intracellular TLR2 and NOD2 protein levels in HAECs. Silencing of the TLR2 and NOD2 genes in HAECs invaded by S. mutans Xc led to a reduction in interleukin-6, interleukin-8 and monocyte chemoattractant protein-1 production. Cytokine production induced by invasive S. mutans via intracellular TLR2 and NOD2 in HAECs may be associated with inflammation in cardiovascular disease.


Subject(s)
Aorta/microbiology , Cytokines/biosynthesis , Endothelial Cells/microbiology , Endothelium, Vascular/microbiology , Inflammation Mediators/immunology , Nod2 Signaling Adaptor Protein/immunology , Streptococcus mutans/immunology , Toll-Like Receptor 2/immunology , Aorta/cytology , Aorta/immunology , Cytokines/immunology , Endothelial Cells/immunology , Endothelium, Vascular/cytology , Humans , Mouth/microbiology , Nod2 Signaling Adaptor Protein/biosynthesis , Signal Transduction , Streptococcus mutans/pathogenicity , Toll-Like Receptor 2/biosynthesis , Up-Regulation
3.
Mol Oral Microbiol ; 27(4): 295-307, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22759314

ABSTRACT

Streptococcus oralis, belonging to the oral viridans group streptococci, has been detected in human cardiovascular lesions including infective endocarditis and atheromatous plaques. The organism has coaggregation receptor polysaccharides (RPS) on the cell wall, which function as receptors for surface adhesins on other members of the oral biofilm community. The present study examined the capacity of S. oralis RPS to induce inflammatory responses in human aortic endothelial cells (HAECs). Purified RPS was used to stimulate HAECs, and the induction of cytokines, adhesion molecules and Toll-like receptors (TLRs) was examined. Involvement of RPS in HAEC invasion by S. oralis was also examined. RPS-stimulated HAECs produced more cytokines (interleukin-6, interleukin-8 and monocyte chemoattractant protein-1) and intercellular adhesion molecule-1 than non-stimulated HAECs. The messenger RNA (mRNA) expression of cytokines and adhesion molecules in RPS-stimulated HAECs increased markedly compared with that in non-stimulated HAECs. Upregulation of TLR-2 mRNA expression was demonstrated in RPS-stimulated HAECs. Moreover, TLR-2 mRNA expression and cytokine production were reduced by the incubation of HAECs with inhibitors against p38 mitogen-activated protein kinase and nuclear factor-κB. An RPS-defective mutant of S. oralis showed greater invasion into HAECs than an RPS-possessing strain. However, HAECs invaded by the RPS-defective mutant produced less cytokines than HAECs invaded by the RPS-possessing strain, indicating that RPS can stimulate HAECs intracellularly. These results suggest that S. oralis RPS may be an important contributor to the pathogenesis of cardiovascular diseases such as infective endocarditis and atherosclerosis.


Subject(s)
Cell Adhesion Molecules/biosynthesis , Endocarditis, Bacterial/microbiology , Inflammation Mediators/metabolism , Plaque, Atherosclerotic/microbiology , Polysaccharides, Bacterial/metabolism , Streptococcus oralis/metabolism , Aortitis/microbiology , Bacterial Adhesion , Cells, Cultured , Cytokines/biosynthesis , Endothelial Cells/microbiology , Endothelium, Vascular/cytology , Endothelium, Vascular/microbiology , Epithelial Cells/microbiology , Humans , Streptococcus oralis/immunology , Toll-Like Receptor 2/biosynthesis
4.
J Fish Dis ; 35(7): 471-80, 2012 Jul.
Article in English | MEDLINE | ID: mdl-22536999

ABSTRACT

We developed a simple genotyping method for Flavobacterium psychrophilum for analysing two single nucleotide polymorphisms (SNPs) in the gyrA gene and to distinguish between isolates that are virulent and avirulent to ayu, Plecoglossus altivelis altivelis (Temminck & Schlegel). The genotyping method is an on/off switch assay and is based on the polymerase chain reaction technique with phosphorothioated primers. We classified 232 isolates from four families of fish (i.e. Plecoglossidae, Osmeridae, Cyprinidae and Salmonidae) into four genotypes (G-C, A-T, A-C and G-T). The G-C type isolates exhibited strong pathogenicity to ayu, whereas the A-T and G-T types did not show any pathogenicity to this species. The A-C type exhibited no or weak pathogenicity to ayu. These results indicate that genotyping F. psychrophilum isolates with two SNPs from gyrA can clearly distinguish between isolates potentially harmful to ayu (G-C type) and those that are potentially not harmful or less harmful (A-C, A-T and G-T type). The on/off switch assay provides a quick, simple, and very powerful DNA genotyping technique for F. psychrophilum isolates.


Subject(s)
Fish Diseases/microbiology , Flavobacteriaceae Infections/veterinary , Flavobacterium/genetics , Flavobacterium/pathogenicity , Genotyping Techniques/veterinary , Osmeriformes , Animals , Base Sequence , DNA Gyrase/genetics , Fish Diseases/mortality , Flavobacteriaceae Infections/microbiology , Flavobacteriaceae Infections/mortality , Flavobacterium/classification , Japan , Molecular Sequence Data , Polymorphism, Single Nucleotide , Survival Analysis , Virulence/genetics
5.
Horm Metab Res ; 44(1): 15-20, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22205568

ABSTRACT

Glucocorticoid (GC) excess promotes adipose tissue accumulation, and 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) plays an important role in the local amplification of GC. Therefore, in this study, we investigated the effects of carbenoxolone (CBX), an 11ß-HSD1 inhibitor, on morphological changes in visceral fat, and the expression of genes involved in adipogenesis and lipid metabolism in high-fat (HF) diet-fed mice. Mice were fed a HF diet from 5 weeks of age. At 10 weeks of age, the mice received an intraperitoneal injection of CBX or vehicle every day for 2 weeks. CBX decreased body weight and visceral fat mass, and improved insulin sensitivity in HF-fed mice. This was accompanied by reduced adipocyte size and a decrease in large-sized adipocytes in visceral fat. The expression of adipogenesis (PPARγ and C/EBPα), glucose transport (GLUT4) and lipid metabolism (LPL, ATGL, and HSL)-related genes were suppressed in CBX mice. CBX treatment induced beneficial morphological changes in visceral fat and decreased the expression of adipogenesis, glucose transport and lipid metabolism-related genes. These findings reveal a potential mechanism underling the effects of CBX on reduced fat accumulation and improved insulin sensitivity.


Subject(s)
Adipogenesis/genetics , Adipose Tissue/anatomy & histology , Adipose Tissue/metabolism , Carbenoxolone/pharmacology , Down-Regulation/drug effects , Glucose/metabolism , Lipid Metabolism/genetics , Adipocytes/cytology , Adipocytes/drug effects , Adipocytes/metabolism , Adipogenesis/drug effects , Adipose Tissue/drug effects , Animals , Biological Transport/drug effects , Body Weight/drug effects , Diet, High-Fat , Down-Regulation/genetics , Feeding Behavior/drug effects , Glucose Tolerance Test , Insulin/metabolism , Intra-Abdominal Fat/drug effects , Intra-Abdominal Fat/metabolism , Lipid Metabolism/drug effects , Lipogenesis/drug effects , Lipogenesis/genetics , Male , Mice , Mice, Inbred BALB C , Organ Size/drug effects , RNA, Messenger/genetics , RNA, Messenger/metabolism
6.
Mol Oral Microbiol ; 26(3): 221-7, 2011 Jun.
Article in English | MEDLINE | ID: mdl-21545699

ABSTRACT

Streptococcus anginosus, an anginosus group bacterium, is frequently isolated from odontogenic abscesses, and is the oral bacterium that is primarily responsible for producing hydrogen sulfide from l-cysteine through the action of its l-cysteine desulfhydrase (ßC-S lyase) enzyme. However, the relationship between its production of hydrogen sulfide and abscess formation has not been investigated. To elucidate the etiological role of hydrogen sulfide in abscess formation, we initially measured, using specific primers, expression of the lcd gene, which encodes ßC-S lyase, in the pus of abscesses that formed in BALB/c mice following subcutaneous injection of S. anginosus into the dorsa. Expression of lcd was >15-fold higher when l-cysteine was present than when it was absent. A mouse virulence assay revealed that the mean diameter of abscesses caused by S. anginosus FW73 plus l-cysteine was greater than that of abscesses caused by S. anginosus FW73 in the absence of l-cysteine. These findings demonstrate that the lcd gene of S. anginosus is upregulated in mouse abscesses and that hydrogen sulfide, the product of a reaction catalyzed by ßC-S lyase, plays an etiological role in odontogenic abscess formation.


Subject(s)
Abscess/enzymology , Cystathionine gamma-Lyase/metabolism , Streptococcal Infections/enzymology , Streptococcus anginosus/enzymology , Abscess/etiology , Animals , Bacterial Proteins/genetics , Bacteriological Techniques , Cystathionine gamma-Lyase/genetics , Cysteine/metabolism , DNA Gyrase/genetics , Gene Expression Regulation, Enzymologic/genetics , Humans , Hydrogen Sulfide/metabolism , Mice , Mice, Inbred BALB C , Reverse Transcriptase Polymerase Chain Reaction , Skin Diseases, Bacterial/microbiology , Streptococcus anginosus/pathogenicity , Suppuration , Tongue/microbiology , Up-Regulation , Virulence
7.
Horm Metab Res ; 43(3): 178-82, 2011 Mar.
Article in English | MEDLINE | ID: mdl-21264794

ABSTRACT

Intrauterine growth restriction (IUGR) is associated with a substantially greater incidence of metabolic syndrome in adulthood. Animal studies have shown that IUGR offspring are hyperphagic during the early postnatal period and therefore exhibit obesity. The molecular mechanisms underlying food intake regulation in the gastrointestinal tract have not been clarified in IUGR. In the present study, we utilized a rat model of IUGR by restricting the food intake of the mother (50% of the normal intake, ad libitum; FR group) from day 7 of gestation until delivery. Pups from undernourished mothers were fostered by control mothers. We examined the food intake and assessed the gene expressions of ghrelin, peptide YY (PYY), and cholecystokinin (CCK) in the alimentary tract of male newborns (postnatal day1) and adult offspring (age, 7 months). Compared to the offspring whose mothers received the standard diet ad libitum (CON offspring), FR offspring were hyperphagic from the weaning time until the end of the experiment, and resulted in a heavier final weight. Both newborn and adult FR offspring had higher ghrelin gene expression in the stomach and higher ghrelin plasma levels than did the controls. Although the gastrointestinal gene expressions and plasma levels of the anorexic peptides, PYY and CCK, were elevated in the FR newborns, they decreased in the FR adults. Our findings suggest that the altered gene expressions of orexigenic and anorexigenic gut peptides in the gastrointestinal tract in the maternal undernutrition-induced IUGR offspring provide a potential mechanism to explain hyperphagia and obesity seen in these offspring.


Subject(s)
Cholecystokinin/genetics , Fetal Growth Retardation/genetics , Gastrointestinal Tract/metabolism , Ghrelin/genetics , Hyperphagia/genetics , Peptide YY/genetics , Up-Regulation , Adult , Animals , Animals, Newborn , Body Weight , Cholecystokinin/blood , Disease Models, Animal , Eating , Female , Fetal Growth Retardation/blood , Fetal Growth Retardation/physiopathology , Gastrointestinal Tract/growth & development , Gene Expression , Gene Expression Regulation, Developmental , Ghrelin/blood , Humans , Hyperphagia/blood , Hyperphagia/physiopathology , Male , Peptide YY/blood , Rats , Rats, Sprague-Dawley
8.
Mol Oral Microbiol ; 26(1): 78-88, 2011 Feb.
Article in English | MEDLINE | ID: mdl-21214874

ABSTRACT

Oral viridans group streptococci are the major commensal bacteria of the supragingival oral biofilm and have been detected in human atheromatous plaque. Atherosclerosis involves an ongoing inflammatory response, reportedly involving chronic infection caused by multiple pathogens. The aim of this study was to examine the invasion of human aortic endothelial cells (HAECs) by oral viridans group streptococci and the subsequent cytokine production by viable invaded HAECs. The invasion of HAECs by bacteria was examined using antibiotic protection assays and was visualized by confocal scanning laser microscopy. The inhibitory effects of catalase and cytochalasin D on the invasion of HAECs were also examined. The production of cytokines by invaded or infected HAECs was determined using enzyme-linked immunosorbent assays, and a real-time polymerase chain reaction method was used to evaluate the expression of cytokine messenger RNA. The oral streptococci tested were capable of invading HAECs. The number of invasive bacteria increased with the length of the co-culture period. After a certain co-culture period, some organisms were cytotoxic to the HAECs. Catalase and cytochalasin D inhibited the invasion of HAECs by the organism. HAECs invaded by Streptococcus mutans Xc, Streptococcus gordonii DL1 (Challis), Streptococcus gordonii ATCC 10558 and Streptococcus salivarius ATCC 13419 produced more cytokine(s) (interleukin-6, interleukin-8, monocyte chemoattractant protein-1) than non-invaded HAECs. The HAECs invaded by S. mutans Xc produced the largest amounts of cytokines, and the messenger RNA expression of cytokines by invaded HAECs increased markedly compared with that by non-invaded HAECs. These results suggest that oral streptococci may participate in the pathogenesis of atherosclerosis.


Subject(s)
Aorta/microbiology , Cytokines/biosynthesis , Endothelial Cells/microbiology , Endothelium, Vascular/microbiology , Inflammation Mediators/metabolism , Mouth/microbiology , Viridans Streptococci/physiology , Aorta/cytology , Atherosclerosis/microbiology , Catalase/pharmacology , Cells, Cultured , Chemokine CCL2/biosynthesis , Coculture Techniques , Cytochalasin D/pharmacology , Endothelial Cells/immunology , Endothelium, Vascular/cytology , Humans , Interleukin-6/biosynthesis , Interleukin-8/biosynthesis , Microscopy, Confocal , Streptococcus/physiology , Streptococcus anginosus/physiology , Streptococcus gordonii/physiology , Streptococcus intermedius/physiology , Streptococcus mitis/physiology , Streptococcus mutans/physiology , Streptococcus oralis/physiology , Viridans Streptococci/drug effects , Viridans Streptococci/immunology , Virulence
9.
Horm Metab Res ; 42(11): 821-5, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20711951

ABSTRACT

Peroxisomal proliferator-activated receptors (PPARs) play an important role in the regulation of lipid metabolism. The aim of this study was to investigate the effects of a maternal high-fat (HF) diet on serum lipid concentration and PPAR gene expression in liver and adipose tissue in the early life of the rat offspring. Female Sprague-Dawley rats were fed either an HF or control (CON) diet 6 weeks before mating and throughout gestation and lactation. Blood and tissue samplings of male offspring were carried out at birth or weaning. Birth weights were similar and serum triglyceride (TG) and nonesterified fatty acid (NEFA) levels showed no significant difference between HF and CON newborns, despite greatly increased hepatic PPARα mRNA expression in the HF newborns (p<0.05). Both HF newborns and weanlings revealed significantly decreased hepatic PPARγ expression compared with controls (p<0.0001). Hepatic PPARα expression in the HF weanlings was reduced markedly compared with CON weanlings (p<0.0001) and showed a negative correlation with serum TG levels (r=-0.743, p<0.05). However, epididymal expression of PPARγ in the HF weanlings was upregulated significantly compared with controls (p<0.05) and demonstrated a positive correlation with epididymal fat mass (r=0.733, p<0.05). These were accompanied by obesity as well as a rise in serum TG by 79% (p<0.05) and NEFA concentration by 36% (p<0.05) in these HF weanlings. Our findings suggest that maternal HF diet leads to alterations in PPAR gene expression in the weanling offspring, which is associated with the disturbed lipid homeostasis.


Subject(s)
Dietary Fats/pharmacology , Lipids/blood , Peroxisome Proliferator-Activated Receptors/metabolism , Animals , Animals, Newborn , Body Weight/drug effects , Female , Rats , Rats, Sprague-Dawley
10.
J Fish Dis ; 32(10): 873-81, 2009 Oct.
Article in English | MEDLINE | ID: mdl-19500209

ABSTRACT

Flavobacterium psychrophilum is the causative agent of bacterial cold-water disease and rainbow trout fry syndrome of salmonids. The pathogen has been reported from all regions in the world involved in salmonid aquaculture, but also from natural fresh-water environments. We established a quantitative loop-mediated isothermal amplification of DNA (LAMP) method to estimate quantities of F. psychrophilum. LAMP primers were designed based on the sequence of the DNA topoisomerase IV subunit B gene, parE, of F. psychrophilum. parE LAMP exhibited a high specificity for the parE gene of F. psychrophilum but not for other related species. parE LAMP detected the gene in a wide range of concentrations from 2.0 x 10(1) to 2.0 x 10(9) copies/reaction within 70 min and revealed a good correlation between threshold times and gene copy number.


Subject(s)
Fish Diseases/microbiology , Flavobacteriaceae Infections/veterinary , Flavobacterium/isolation & purification , Polymerase Chain Reaction/veterinary , Salmonidae , Animals , DNA Topoisomerase IV/chemistry , DNA Topoisomerase IV/genetics , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Fish Diseases/diagnosis , Flavobacteriaceae Infections/diagnosis , Flavobacteriaceae Infections/microbiology , Flavobacterium/genetics , Polymerase Chain Reaction/methods , RNA, Ribosomal, 16S/chemistry , RNA, Ribosomal, 16S/genetics , Sensitivity and Specificity
11.
Eur J Neurol ; 14(11): 1288-91, 2007 Nov.
Article in English | MEDLINE | ID: mdl-17868276

ABSTRACT

Miyoshi myopathy (MM) is an autosomal recessive distal muscular dystrophy characterized by mutations of the dysferlin gene. Although several pairs of homozygous/heterozygous mutations have been reported, few effective treatments of MM are available. We had observed the decreased serum creatine kinase (CK) before and after administration of dantrolene in the elder brother and the increased serum CK before and after discontinuance of the drug on suspicion of drug-induced hepatopathy in the younger sister. We report a novel pair of heterozygous mutations in the 3'-splicing site of exon 26 and the translation site of exon 28 of the dysferlin gene in two siblings, and effective treatment of their MM with dantrolene.


Subject(s)
Dantrolene/therapeutic use , Heterozygote , Membrane Proteins/genetics , Muscle Proteins/genetics , Muscular Dystrophies/genetics , Mutation , Siblings , Adolescent , Asian People/genetics , Dysferlin , Female , Humans , Male , Muscular Diseases/diagnosis , Muscular Diseases/drug therapy , Muscular Diseases/genetics , Muscular Dystrophies/diagnosis , Muscular Dystrophies/drug therapy , Pedigree
12.
Eur J Neurol ; 14(4): 464-6, 2007 Apr.
Article in English | MEDLINE | ID: mdl-17389000

ABSTRACT

We report a novel missense mutation of the Notch3 gene in a Japanese family with CADASIL. The Cys49Gly mutation in this family is located in exon 2 of the Notch3 gene. Most of the documented Notch3 gene mutations occur in exons 3 or 4. On the other hand, there are few reports around the world of mutations in exon 2 of the Notch3 gene, and this is the first report of a mutation in exon 2 of the gene in a Japanese family. In general, CADASIL mutations involve a cysteine residue. Such mutations may influence the tertiary structure of the Notch3 protein, resulting in protein dysfunction. Thus, the CADASIL in the present case may be a consequence of the mutation in exon 2 causing a structural change in the Notch3 protein.


Subject(s)
CADASIL/genetics , Mutation, Missense , Receptors, Notch/genetics , Adult , Asian People , Brain/pathology , CADASIL/diagnosis , CADASIL/physiopathology , Diagnosis, Differential , Female , Humans , Magnetic Resonance Angiography , Magnetic Resonance Imaging , Male , Meniere Disease/diagnosis , Meniere Disease/genetics , Meniere Disease/pathology , Meniere Disease/physiopathology , Microscopy, Electron, Transmission , Pedigree , Receptor, Notch3 , Skin/pathology , Tinnitus/physiopathology , Vertigo/physiopathology
13.
Oral Microbiol Immunol ; 21(6): 420-3, 2006 Dec.
Article in English | MEDLINE | ID: mdl-17064403

ABSTRACT

Streptococcus mutans and other viridans streptococci have been implicated as major etiological agents of infective endocarditis. The serotype-specific rhamnose-glucose polysaccharide (RGP) of S. mutans has several biological functions that appear to be essential for the induction of infective endocarditis. The aim of this study was to examine the contribution of RGP to the infectivity of S. mutans in infective endocarditis using a rat model. The RGP-defective mutant of S. mutans showed reduced ability to induce infective endocarditis compared to the parental strain. The ability of S. mutans to induce infective endocarditis was not consistent with the binding capacity of the organism to extracellular matrix proteins. The results suggest that S. mutans containing whole RGP is more virulent than the RGP-defective mutant, and the RGP has an important role for the induction of infective endocarditis by S. mutans.


Subject(s)
Endocarditis, Bacterial/microbiology , Polysaccharides, Bacterial/physiology , Streptococcus mutans/pathogenicity , Animals , Bacterial Adhesion , Extracellular Matrix Proteins/metabolism , Male , Protein Binding , Rats , Rats, Wistar , Serotyping , Species Specificity , Virulence Factors
14.
J Biol Chem ; 276(42): 39179-85, 2001 Oct 19.
Article in English | MEDLINE | ID: mdl-11502751

ABSTRACT

The inositol pyrophosphate disphosphoinositol pentakisphosphate (PP-InsP(3)/InsP(7)) is formed in mammals by two recently cloned inositol hexakiphosphate kinases, InsP(6)K1 and InsP(6)K2 (Saiardi, A., Erdjument-Bromage, H., Snowman, A. M., Tempst, P., and Snyder, S. H. (1999) Curr. Biol. 9, 1323-1326). We now report the identification, cloning, and characterization of a third InsP(7) forming enzyme designated InsP(6)K3. InsP(6)K3 displays 50 and 45% sequence identity to InsP(6)K1 and InsP(6)K2, respectively, with a smaller mass (46 kDa) and a more basic character than the other two enzymes. InsP(6)K3 is most enriched in the brain where its localization resembles InsP(6)K1 and InsP(6)K2. Intracellular disposition discriminates the three enzymes with InsP(6)K2 being exclusively nuclear, InsP(6)K3 predominating in the cytoplasm, and InsP(6)K1 displaying comparable nuclear and cytosolic densities.


Subject(s)
Phosphotransferases (Phosphate Group Acceptor)/biosynthesis , Phosphotransferases (Phosphate Group Acceptor)/chemistry , Phosphotransferases (Phosphate Group Acceptor)/isolation & purification , Amino Acid Sequence , Animals , Base Sequence , Blotting, Northern , Brain/metabolism , Catalysis , Cell Line , Cell Nucleus/enzymology , Cloning, Molecular , Cytosol/enzymology , DNA, Complementary/metabolism , Humans , In Situ Hybridization , Kinetics , Male , Mice , Mice, Inbred C57BL , Molecular Sequence Data , Point Mutation , Protein Binding , Sequence Homology, Amino Acid , Time Factors , Tissue Distribution
15.
Neuron ; 31(3): 439-51, 2001 Aug 16.
Article in English | MEDLINE | ID: mdl-11516400

ABSTRACT

Diphosphoinositol-pentakisphosphate (InsP7) and bis-diphosphoinositol tetrakisphosphate (InsP8) possess pyrophosphate bonds. InsP7 is formed from inositol hexakisphosphate (InsP6) by recently identified InsP6 kinases designated InsP6K1 and InsP6K2. We now report the identification, cloning, and characterization of a novel protein, GRAB (guanine nucleotide exchange factor for Rab3A), which interacts with both InsP6K1 and Rab3A, a Ras-like GTPase that regulates synaptic vesicle exocytosis. GRAB is a physiologic GEF (guanine nucleotide exchange factor) for Rab3A. Consistent with a role of Rab3A in synaptic vesicle exocytosis, GRAB regulates depolarization-induced release of dopamine from PC12 cells and nicotinic agonist-induced hGH release from bovine adrenal chromaffin cells. The association of InsP6K1 with GRAB fits with a role for InsP7 in vesicle exocytosis.


Subject(s)
Guanine Nucleotide Exchange Factors/metabolism , Intracellular Signaling Peptides and Proteins , Phosphotransferases (Phosphate Group Acceptor)/metabolism , Synaptic Vesicles/physiology , rab3A GTP-Binding Protein/metabolism , Adrenal Medulla/cytology , Adrenal Medulla/physiology , Amino Acid Sequence , Animals , Carrier Proteins/chemistry , Cattle , Chromaffin Cells/cytology , Chromaffin Cells/physiology , Cloning, Molecular , Dopamine/metabolism , Exocytosis , Guanine Nucleotide Exchange Factors/chemistry , Guanine Nucleotide Exchange Factors/genetics , Guanosine 5'-O-(3-Thiotriphosphate)/metabolism , Guanosine Triphosphate/metabolism , Human Growth Hormone/genetics , Human Growth Hormone/metabolism , Humans , Molecular Sequence Data , Nerve Growth Factor/pharmacology , Nicotinic Agonists/pharmacology , PC12 Cells , Phosphates/metabolism , Protein Conformation , Rats , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Transfection
16.
Infect Immun ; 69(5): 2788-96, 2001 May.
Article in English | MEDLINE | ID: mdl-11292690

ABSTRACT

Toll-like receptors 2 and 4 (TLR2 and TLR4) have been found to transduce signals of peptidoglycan (PGN) and lipopolysaccharide (LPS), respectively, for NF-kappa B activation. However, little is known about the expression and regulation of the TLR2 gene in monocytes/macrophages in response to the two typical bacterial products. We show in the present study that both PGN and a high concentration of LPS increase TLR2 gene expression in macrophage-like cells, 1 alpha,25-dihydroxyvitamin D(3)-differentiated human HL60 and mouse RAW264.7 cells, and human monocytes in a dose- and time-dependent manner. Actinomycin D and pyrrolidine dithiocarbamate inhibition of gene transcription and NF-kappa B activation, respectively, blocks LPS- and PGN-elevated TLR2 mRNA in monocytic cells. The LPS-induced increase in TLR2 mRNA in monocytic cells is abolished by polymyxin B pretreatment and is observed in peripheral blood mononuclear cells from pigs subjected to endotoxic shock. Further, high concentrations of LPS and synthetic lipid A increase TLR2 mRNA expression in peritoneal macrophages from both TLR4-deficient C3H/HeJ mice and normal C3H/HeN mice, a process that constitutes induction of TLR4-independent TLR2 expression. These findings demonstrate that TLR2 gene expression is upregulated in macrophage responses to PGN and to high concentrations of LPS in vitro and in vivo and correlates with NF-kappa B activation.


Subject(s)
Drosophila Proteins , Gene Expression Regulation/drug effects , Lipopolysaccharides/pharmacology , Macrophages/drug effects , Membrane Glycoproteins/genetics , NF-kappa B/physiology , Peptidoglycan/pharmacology , Receptors, Cell Surface/genetics , Animals , HL-60 Cells , Humans , Lipid A/pharmacology , Macrophages/metabolism , Mice , Mice, Inbred C3H , Shock, Septic/blood , Swine , Toll-Like Receptor 2 , Toll-Like Receptor 4 , Toll-Like Receptors , Transcription, Genetic/drug effects , Up-Regulation
17.
Anesth Analg ; 92(3): 676-80, 2001 Mar.
Article in English | MEDLINE | ID: mdl-11226100

ABSTRACT

UNLABELLED: Decreased hepatic blood flow, and impaired hepatic oxygen delivery caused by endotoxin, result in hepatic metabolic deterioration followed by liver dysfunction and multiple organ failure. Among phosphodiesterase III inhibitors, only olprinone increases hepatosplanchnic blood flow. We evaluated the effects of olprinone on systemic hemodynamics, hepatic circulation, and hepatic oxygen delivery in a porcine model of endotoxemia. Fifteen pigs received a continuous infusion (1.7 microg. kg(-1). h(-1)) of endotoxin (lipopolysaccharide [LPS]) via the portal vein for 240 min. Seven of these pigs received olprinone infusion (0.3 microg. kg(-1). min(-1)) via a central vein from t = 150 min to t = 240 min, whereas the eight remaining pigs served as LPS controls. Continuous infusion of LPS caused significant reductions in hemodynamic variables and a significant increase in arterial lactate. After the administration of olprinone during the LPS infusion, portal venous flow and hepatic oxygen delivery were increased and were higher than in the LPS group. Furthermore, olprinone prevented any further increase in arterial lactate. We conclude that the administration of olprinone halted the disturbances in the hepatic circulation, especially in portal venous flow and hepatic oxygen delivery, in a porcine model of endotoxemia. IMPLICATIONS: Endotoxin is a causative factor in peripheral vascular failure, resulting in a hemodynamic depression that includes a reduction in liver blood flow. The administration of olprinone (phosphodiesterase III inhibitor) improves the liver blood flow circulation in a porcine model of endotoxemia.


Subject(s)
Endotoxemia/drug therapy , Imidazoles/pharmacology , Liver Circulation/drug effects , Phosphodiesterase Inhibitors/pharmacology , Pyridones/pharmacology , Animals , Disease Models, Animal , Endotoxemia/physiopathology , Imidazoles/therapeutic use , Oxygen/metabolism , Pyridones/therapeutic use , Swine
18.
Proc Natl Acad Sci U S A ; 98(5): 2306-11, 2001 Feb 27.
Article in English | MEDLINE | ID: mdl-11226235

ABSTRACT

Using a consensus sequence in inositol phosphate kinase, we have identified and cloned a 44-kDa mammalian inositol phosphate kinase with broader catalytic capacities than any other member of the family and which we designate mammalian inositol phosphate multikinase (mIPMK). By phosphorylating inositol 4,5-bisphosphate, mIPMK provides an alternative biosynthesis for inositol 1,4,5-trisphosphate [Ins(1,4,5)P(3)]. mIPMK also can form the pyrophosphate disphosphoinositol tetrakisphosphate (PP-InsP(4)) from InsP(5). Additionally, mIPMK forms InsP(4) from Ins(1,4,5)P(3) and InsP(5) from Ins(1,3,4,5)P(4).


Subject(s)
Inositol 1,4,5-Trisphosphate/biosynthesis , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Amino Acid Sequence , Animals , Base Sequence , Cloning, Molecular , DNA Primers , In Situ Hybridization , Male , Molecular Sequence Data , Phosphotransferases (Alcohol Group Acceptor)/chemistry , Phosphotransferases (Alcohol Group Acceptor)/genetics , Rats , Rats, Sprague-Dawley , Sequence Homology, Amino Acid
20.
Brain Res ; 873(1): 83-93, 2000 Aug 04.
Article in English | MEDLINE | ID: mdl-10915813

ABSTRACT

NS-7 is a novel blocker of voltage-sensitive Ca(2+) and Na(+) channels, and it significantly reduces infarct size after occlusion of the middle cerebral artery. Persistent activation of cyclic AMP response element binding protein (CREB), which can be induced by increase in intracellular Ca(2+) concentrations or other second messengers, has recently been found to be closely associated with neuronal survival in cerebral ischemia. The present study was therefore undertaken to evaluate the neuroprotective effects of NS-7 by analyzing changes in CREB phosphorylation in a focal cerebral ischemia model. CREB phosphorylation in the brain of rats was investigated immunohistochemically at 3.5-48-h recirculation after 1. 5-h occlusion of the middle cerebral artery. NS-7 (1 mg/kg; NS-7 group) or saline (saline group) was intravenously injected 5 min after the start of recirculation. The NS-7 group showed significantly milder activation of CREB phosphorylation in various cortical regions after 3.5 h of recirculation than the saline group. The inner border zone of ischemia in the NS-7 group subsequently exhibited a moderate, but persistent, increase in number of phosphorylated CREB-positive neurons with no apparent histological damage. By contrast, the saline group displayed a marked, but only transient, increase in number of immunopositive neurons in this border zone after 3.5 h of recirculation, and this was followed by clear suppression of CREB phosphorylation and subsequent loss of normal neurons. These findings suggest that: (1) the marked enhancement of CREB phosphorylation in the acute post-ischemic phase may be triggered largely by an influx of calcium ions as a result of activation of the voltage-sensitive Ca(2+) and Na(+) channels; and that (2) the neuroprotective effects of NS-7 may be accompanied by persistent activation of CREB phosphorylation in the inner border zone of ischemia.


Subject(s)
Brain Ischemia/metabolism , Calcium Channel Blockers/pharmacology , Cyclic AMP Response Element-Binding Protein/metabolism , Neuroprotective Agents/pharmacology , Pyrimidines/pharmacology , Sodium Channel Blockers , Animals , Brain/metabolism , Calcium Channels/physiology , Immunohistochemistry , Male , Phosphorylation/drug effects , Rats , Rats, Sprague-Dawley , Sodium Channels/physiology
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