Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0243026 (sepsis)
 52,417 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the present study, rat cardiac sarcoplasmic reticulum (SR) phospholamban (PLB) phosphatase was partially purified by chromatography on DEAE-Sephacel. This PLB phosphatase was indentical to phosphatase-1. It was shown on electrophoresis of SDS-PAGE autoradiography that the PLB phosphatase in rats during early sepsis (ES) depressed dephosphorylation of substrates (32P-phosphorylase a and 32P-SR). However, dephosphorylation of the substrates by the partially purified phosphatase during late sepsis (LS) was same as that in control rats. The partially purified PLB phosphatase activity in ES rats was significantly decreased, but showed no change in LS rats. The results above were confirmed by a studing of the substrate concentration (enzyme concentration, time)--enzyme reaction velocity curve in showing that both affinity and maximum initial velocity (Vmax) of the phosphatase in the ES rats were decreased, but had no change in those in the LS rats.
 ...
PMID:[Alteration of phospholamban phosphatase activity associated with cardiac sarcoplasmic reticulum during sepsis in rats]. 748 77

Homopolymeric alpha-2,8-linked sialic acid (PSA) has been found as a capsular component of sepsis- and meningitis-causing bacterial pathogens, and on eukaryotic cells as a post-translational modification of the neural cell adhesion molecule (NCAM). The polysaccharide is specifically recognized and degraded by a phage-encoded enzyme, the endo-N-acetylneuraminidase E (Endo NE). Endo NE therefore has become a valuable tool in the study of bacterial pathogenesis and eukaryotic morphogenesis. In this report we describe the molecular cloning of Endo NE and the expression of a functionally active recombinant enzyme. The cloned DNA sequence (2436 bp) encodes a polypeptide of 811 amino acids, which at the 5' end contains a totally conserved neuraminidase motif. Expressed in Escherichia coli, the enzyme migrates as a single band of approximately 74 kDa in SDS-PAGE. A central domain of 669 amino acid residues is about 90% homologous to the recently cloned Endo NF. Both phage-induced lysis of bacteria and the catalysis of PSA degradation by the recombinant enzyme are efficiently inhibited by a polyclonal antiserum raised against the intact phage particle. The C-terminal region seems to be essential to enzymatic functions, as truncation of 32 amino acids outside the homology domain completely abolishes Endo NE activity. Our data also indicate that the 38 kDa protein, previously assumed to be a subunit of the Endo NE holoenzyme, is the product of a separate gene locus and is not necessary for in vitro depolymerase activity.
 ...
PMID:Molecular cloning and functional expression of bacteriophage PK1E-encoded endoneuraminidase Endo NE. 756 5

Radiolabeled antithrombin III (ATIII) was incubated at 37 degrees C with purified vitronectin (VN) or fibrinogen-deficient plasma before thrombin was added to initiate complex formation. Incorporation of radiolabeled ATIII was detected using polyacrylamide gel electrophoresis (PAGE) and autoradiography. The PAGE conditions appeared to be crucial for the detection of VN.TAT complexes. In the absence of SDS, ternary complexes formed instantaneously, whereas in the presence of SDS, only 50% of the TAT was associated with VN after a 60-min incubation. Formation of ternary complexes could be confirmed by gel filtration of the plasma to which thrombin was added. Furthermore, TAT in patient plasmas (disseminated intravascular coagulation and sepsis) was found to bind to heparin-Sepharose, indicating that this endogenously formed TAT was also associated with VN. The amino-terminal region of VN and the thrombin moiety of the TAT complex were found to be responsible for their interaction, which was stabilized by disulfide bridges. These results indicate that in normal plasma all TAT is complexed with VN. This association alters the conformational state of plasma VN, which appears to be responsible for the clearance of thrombin complexes from the circulation.
 ...
PMID:Ternary vitronectin-thrombin-antithrombin III complexes in human plasma. Detection and mode of association. 767 52

Phospholipase A2 (PLA2) activity was purified 12,544-fold with a 13% yield from the plasma of patients diagnosed of septic shock by the sequential use of heparin-agarose affinity chromatography, gel filtration, and reverse-phase f.p.l.c. Gel-filtration chromatography of plasma omitting high-ionic-strength buffer revealed a molecular mass different from that of purified PLA2 and co-elution with apolipoprotein A-I peaks, which suggests its association with high-density lipoproteins (HDL). N-terminal analysis of the enzyme activity protein band, electroblotted from a SDS-acrylamide gel and with an assessed molecular mass of 19 kDa, showed an identical sequence to that of alpha-chain of human C3 complement component, suggesting the presence in this band of a complex formed by a complement C3-derived anaphylatoxin (C3a)-related fragment and the PLA2 linked side-by-side. Because the preparation of plasma enzyme showed lower activity than the enzyme obtained from fibroblasts transfected with the coding sequence of human group-II PLA2, and because the addition of C3-derived anaphylatoxins from human serum inhibited the activity of this recombinant PLA2, it was considered that C3a-related peptides behave as inhibitors of group-II PLA2. The enzyme showed optimal activity on [14C]oleate-labelled autoclaved E. coli, on synthetic phosphatidylethanolamine, and on [3H]arachidonate-labelled membranes of the monoblast cell line U937, but it did not show any activity on the release of [3H]arachidonate from pre-labelled human polymorphonuclear leukocytes (PMNs). In short, PLA2 from plasma of sepsis patients shows unique associations with other plasma proteins which may influence its functional properties. The association with C3-related peptides shows an inhibitory effect on the enzyme activity, whereas the association with HDL might influence its environment and/or its interaction with cells. The study of the catalytic properties shows a prominent effect on bacterial phospholipids, synthetic phosphatidylethanolamine, and membranes from U937 monoblasts, but not on synthetic phosphatidylcholine or on PMNs, even when these cells were maintained in culture to allow spontaneous apoptosis and became a good substrate for pancreatic type PLA2.
 ...
PMID:Phospholipase A2 from plasma of patients with septic shock is associated with high-density lipoproteins and C3 anaphylatoxin: some implications for its functional role. 786 6

A secreted form of phospholipase A2 (PLA2) has been implicated in inflammatory disorders such as rheumatoid arthritis and sepsis. To determine if PLA2 may also play a role in allergic rhinitis, we have measured enzymic activity in nasal lavage from allergic subjects. Enhanced activity of PLA2 in the lavage was observed following nasal challenge with antigen or histamine. The PLA2 in the nasal lavage was partially purified by acid extraction, size exclusion chromatography, and ion exchange chromatography. The partially purified enzyme from nasal lavage was subsequently compared to a recombinant form of human PLA2 identified in synovial fluid from arthritic patients. The two enzymes showed similar molecular weights (15 to 16 kD) on SDS-PAGE, and both reacted with a rabbit polyclonal antiserum raised to a galactokinase-PLA2 fusion protein. The enzymatic activities of the two PLA2s were indistinguishable when compared for ionic dependence, substrate selectivity, and sensitivity to inhibitors. These results suggest that the PLA2 induced in nasal lavage in response to challenge by antigen is very similar to the extracellular PLA2 found in synovial fluid from subjects with rheumatoid arthritis and may play a role in the inflammatory processes associated with allergic rhinitis.
 ...
PMID:Characterization of phospholipase A2 from human nasal lavage. 801 33

The effect of supernatant from phorbol myristate acetate (PMA) stimulated human polymorphonuclear granulocytes (PMN) on human factor VII was studied in vitro. The supernatant caused a rapid loss in factor VII coagulant activity by the action of human leukocyte elastase (HLE) and cathepsin G in the supernatant, as demonstrated by the use of specific inhibitors of the two serine proteases, respectively. Preincubation of the supernatant with the elastase inhibitor and the cathepsin G inhibitor preserved 80% and 25% of the clotting activity, respectively. Calcium protected factor VII completely from the supernatant mediated inactivation. Cathepsin G and HLE purified from PMN each destroyed the coagulant activity of factor VII when added to a non-plasma system. There were, however, no effect on factor VII activity when cathepsin G was added to plasma. Polyacrylamide gel electrophoresis in the presence of SDS indicated that HLE and cathepsin G cleaved the zymogen in the same manner, producing (a) peptide(s) of low molecular mass and a single large product of 48 kDa. Preincubation of factor VII with calcium ions inhibited the proteolytic action of HLE and cathepsin G. It is suggested that HLE and cathepsin G from activated granulocytes may be partly responsible for the loss in factor VII activity that is observed during sepsis.
 ...
PMID:Human leukocyte elastase and cathepsin G inactivate factor VII by limited proteolysis. 805 78

The ability of cefotaxime, ciprofloxacin, piperacillin and tobramycin to cause release of endotoxin was examined in vitro with cultures of Enterobacter cloacae and Escherichia coli. Endotoxin was measured by a quantitative limulus amoebocyte lysate assay and its presence was confirmed by silver staining of the lipopolysaccharide moiety following SDS-PAGE. The morphology of the bacteria during antibiotic exposure was examined by scanning electronmicroscopy. Cefotaxime, ciprofloxacin and piperacillin caused significant endotoxin release, correlating with their ability to affect cell-wall morphology, causing filamentation, wall breakage and cell lysis. In contrast, little endotoxin was released when bacteria were exposed to tobramycin and no morphological changes were observed when bacteria were exposed to bactericidal concentrations of this aminoglycoside. Its antimicrobial spectrum and bactericidal activity make tobramycin an appropriate agent for treatment of sepsis caused by gram-negative bacteria and its lack of propensity to elicit excessive release of endotoxin may avoid exacerbation of endotoxin-related shock in sepsis.
 ...
PMID:Antibiotic-induced release of endotoxin from bacteria in vitro. 828 10

Plasmin-alpha2-antiplasmin complexes (PAP) are considered good markers of fibrinolytic activation in vivo. The presence of neoantigens in these complexes offers the possibility to develop specific immunoassays to determine PAP levels. We have developed a sensitive PAP purification method in vitro by adding urokinase to fresh plasma followed by affinity chromatography to lysine-sepharose and elution with epsilon-aminocaproic acid. This material, characterized by SDS-PAGE and Western blotting, was used to raise monoclonal antibodies (MoAbs). We describe a new enzyme linked immunosorbent assay (ELISA) to quantify PAP complexes in plasma. The assay follows the sandwich principle and is based on two MoAbs, CPL12 and CPL15, that bind to the modified alpha2-antiplasmin moiety and the plasmin moiety of the complex respectively. The calibration curve was constructed with definite concentrations of purified PAP. The lower limit of the assay is 75 ng/ml and the variation coefficients are 3.5% (intra-assay) and 10-6% (interassay). A mean value of 573.5+/-131.4 ng/ml was obtained from PAP concentration in a healthy population (n = 30). Significantly higher PAP levels were observed under diverse clinical conditions in which fibrinolysis is activated: clinical sepsis, acute myocardial infarction (AMI), malignancy, diabetes, pregnancy, elderly people and thrombolytic therapy. From our results we conclude that this ELISA is suitable to measure in vivo plasma PAP levels.
 ...
PMID:Development and clinical application of a new ELISA assay to determine plasmin-alpha2-antiplasmin complexes in plasma. 861 97

The present study includes 178 Haemophilus influenzae strains isolated in different pediatric hospitals from Havana, Cuba, during 1991-1994, associated to divers infections (meningitis, respiratory sepsis, primary bacteremia). A combination of various typing and subtyping methods was used as epidemiological markers: serotyping (slide agglutination with diagnostical serum a-f and latex agglutination), biotyping according to Killian's procedures (by determination of indole production, urease and ornithine decarboxylase activity), subtyping by fermentative profiles according to Roberts' methods (glucose, maltose, xylose and fructose) and outer membrane protein profile subtyping (vesicles extraction by a modified Barenkamp's method, analysis by lineal and gradient SDS-PAGE and assessment according to our own classification system). Serotype b was identified in 89.3%, biotype I was the most frequent (79.1%), other biotypes (II, III, IV and V) were also identified. Fermentative profile D (glucose, maltose, xylose and fructose positive) was the most frequent (52.8%) while profile G (glucose, maltose, xylose positive and fructose negative) represented 20.2%. Other known profiles were present. PA2 (33.7%) was the most frequent OMP subtype. Even though 11 different protein subtypes were found, the 77.5% of the strains were located in only three OMP electrophoretic subtypes (PA2, PC1, LA2).
 ...
PMID:[Utilization of different microbiological markers in the study of Haemophilus influenzae]. 902 20

Fifteen Bacteroides forsythus strains freshly isolated from patients with periodontitis were used together with three collection strains and one type strain for characterization of growth on various media; determination of enzymatic profiles, antibiotic susceptibility profiles, 16S rRNA ribotypes, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) outer membrane protein profiles, and pathogenicity; and gas chromatography analysis by using a wound chamber model in rabbits. All strains were stimulated by N-acetylmuramic acid, while one strain needed a further supplement such as yeast extract for optimal growth. All strains showed trypsin-like activity. While 10 different ribotypes were found, the SDS-PAGE profiles revealed similar patterns for all strains. All strains were sensitive to penicillin G (MICs, <0.5 microg/ml), ampicillin (MICs, <1.0 microg/ml), amoxicillin (MICs, <0.38 microg/ml), metronidazole (MICs, <0.005 microg/ml), tetracycline (MICs, <0.19 microg/ml), doxycycline (MICs, 0.05 microg/ml), erythromycin (MICs, <0.4 microg/ml), and clindamycin (MICs, <0.016 microg/ml), while they were less sensitive to ciprofloxacin (MICs, <4 microg/ml). B. forsythus did not cause abscess formation by monoinoculation. B. forsythus coinoculated with Fusobacterium nucleatum ATCC 10953 caused abscess formation in 75% of rabbits, while it caused abscess formation in 100% of rabbits when it was coinoculated with Porphyromonas gingivalis FDC 381. In the case of the latter combination, four of six rabbits died of sepsis after 6 to 7 days, and P. gingivalis and B. forsythus were recovered from the heart blood at a proportion of 10:1. B. forsythus strains were highly virulent and invasive in combination with P. gingivalis.
 ...
PMID:Characterization of Bacteroides forsythus isolates. 916 47


<< Previous 1 2 3 4 5 Next >>