Isolation of a Helicobacter strain from the human liver.
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Adult↗
Biomedical subjects
A Santos
Publications and source records attributed to A Santos.
Production and characteristics of Debaryomyces hansenii killer toxin.
The optimal conditions for the production of the killer toxin of Debaryomyces hansenii CYC 1021 have been studied. The lethal activity of the killer toxin increased with the presence of NaCl in the medium used for testing the killing action. Production of the killer toxin was stimulated in the presence of proteins of complex culture media. Addition of nonionic detergents and other additives, such as dimethylsulfoxide enhanced killer toxin production significantly. Killer toxin secretion pattern followed the growth curve and reached its maximum activity at the early stationary phase. Optimal stability was observed at pH 4.5 and temperatures up to 20 degrees C. Above pH 4.5 a steep decrease of the stability was noted. The activity was hardly detectable at pH 5.1.
Cell Division↗
The potential for CYP2D6 inhibition screening using a novel scintillation proximity assay-based approach.
High throughput inhibition screens for human cytochrome P450s (CYPs) are being used in preclinical drug metabolism to support drug discovery programs. The versatility of scintillation proximity assay (SPA) technology has enabled the development of a homogeneous high throughput assay for cytochrome P450 2D6 (CYP2D6) inhibition screen using [O-methyl-(14)C]dextromethorphan as substrate. The basis of the assay was the trapping of the O-demethylation product, [(14)C]HCHO, on SPA beads. Enzyme kinetics parameters V(max) and apparent K(m), determined using pooled human liver microsomes and microsomes from baculovirus cells coexpressing human CYP2D6 and NADPH-cytochrome P450 reductase, were 245 pmol [(14)C]HCHO/min/mg protein and 11 microM, and 27 pmol [(14)C]HCHO/min/pmol and 1.6 microM, respectively. In incubations containing either pooled microsomes or recombinant CYP2D6, [(14)C]dextromethorphan O-demethylase activity was inhibited in the presence of quinidine (IC(50) = 1.0 microM and 20 nM, respectively). By comparison, inhibitors selective for other CYP isoforms were relatively weak (IC(50) > 25 microM). In agreement, a selective CYP2D6 inhibitory monoclonal antibody caused greater than 90% inhibition of [(14)C]dextromethorphan O-demethylase activity in human liver microsomes, whereas CYP2C9/19- and CYP3A4/5-selective antibodies elicited a minimal inhibitory effect. SPA-based [(14)C]dextromethorphan O-demethylase activity was also shown to correlate (r(2) = 0.6) with dextromethorphan O-demethylase measured by high-performance liquid chromatography in a bank of human liver microsomes (N = 15 different organ donors). In a series of known CYP2D6 inhibitors/substrates, the SPA-based assay resolved potent inhibitors (IC(50) < 2 microM) from weak inhibitors (IC(50) >or= 20 microM). It is concluded that the SPA-based assay described herein is suitable for CYP2D6 inhibition screening using either native human liver microsomes or cDNA-expressed CYP2D6.
Antibodies, Monoclonal↗
The peroxisome proliferator-activated receptor gamma is an inhibitor of ErbBs activity in human breast cancer cells.
One of the most interesting recent developments in the nuclear receptor field has been the identification of natural and synthetic agonists of the peroxisome proliferator-activated receptor (PPAR) family, coupled with a growing recognition that the gamma isoform (PPARgamma) affects pathways important in a variety of human diseases. Here we show that the activation of PPARgamma through the 15-deoxy-Delta-12,14-prostaglandin J(2) (PG-J(2)) ligand causes a dramatic inhibition of ErbB-2 and ErbB-3 tyrosine phosphorylation caused by neuregulin 1 (NRG1) and neuregulin 2 (NRG2) in MCF-7 cells. This effect is accompanied by a very efficient blocking of ErbBs effects upon proliferation, differentiation and cell death in these cells. Preincubation of MCF-7 cells with PG-J(2) before addition of NRG1 and NRG2 had a dramatic growth-suppressive effect accompanied by accumulation of cells in the G0/G1 compartment of the cell cycle, and a marked increase in apoptosis. NRG1 and NRG2 induce G1 progression, which was associated with stimulation of the phosphatidylinositol-3 kinase (PI 3-K) pathway, whereas survival was dependent on ERK1/ERK2 activation. Both pathways were inhibited by PG-J(2). Furthermore, PG-J(2) can abolish the NRG1 and NRG2-induced increase in anchorage-independent growth of these cells. PG-J(2) also blocks phosphorylation of other receptor tyrosine kinases, such as IGF-IR, in MCF-7 cells, and suppress proliferation of other breast cancer cell lines. In summary, our data show a specific inhibitory action of PG-J(2) on the activity of the ErbB receptors in breast cancer cells.
Apoptosis↗
Report of Didymocystis wedli Ariola, 1902 (Digenea; Didymozoidae) from Thunnus albacares in Brazil.
Didymocystis wedli a parasite from the gills of Thunnus albacares from the coast of the State of Rio de Janeiro, is described by use of light and scanning electron microscopy. This is the first report of this species in Brazil and South America. New data are presented on the surface topography as demonstrated by scanning electron microscopy.
Animals↗
[Surgery of atrial fibrillation with intraoperative radiofrequency].
Radiofrequency ablation is successful in most supraventricular tachyarrhythmias and the first results in percutaneous applications in atrial fibrillation have been published. At present, all the surgical techniques for the treatment of atrial fibrillation use extensive atrial incisions. A case report is presented of a 67 year old woman with chronic atrial fibrillation and rheumatic mitral and tricuspid valve disease. At surgery, pulmonary vein isolation was performed using a continuous linear ablation around the pulmonary veins, instead of using extensive atrial incisions. Concomitantly, a mitral valve replacement and tricuspid valve annuloplasty were performed. Five months after surgery the patient remains with a stable sinus rhythm surgery and in functional class I (NYHA).
Aged↗
[Comparative study of the eradication of Helicobacter pylori: ranitidine bismuth citrate versus omeprazole plus two antibiotics for seven days].
AIMS: To compare the efficacy of ranitidine bismuth citrate (RB) combined with two antibiotics during seven days in comparison with OCA-7 for treatment of Helicobacter pylori. METHODS: In a randomized clinical trial 200, consecutive patients infected by Helicobacter pylori were studied prospectively. One of two regimens was given for 7 days; OCA-7 (omeprazole 20 mg b.i.d., amoxycillin 1 gr b.i.d., clarithromycin 500 mg b.i.d.) or RBCA (RB 400 b.i.d., amoxicillin 1 gr b.i.d., clarithromycin 500 mg b.i.d.). Eradication was defined as a negative breath test one month after therapy. RESULTS: 112 male and 88 female with a mean age of 48 years (range 18-82). 100 patients were randomized to OCA-7 and 100 patients to RBCA-7. Overall 186 patients completed the treatment. Intention to-treat eradication was in the group OCA-7 of 72% and 73% in group RBCA-7. Per-protocol eradication was in the group OCA-7 of 78% and 77% in group RBCA-7 (p = 0.3). Five patients in the group OCA-7 and nine in the group RBCA-7, had side effects, three of these patients did not complete therapy by diarrhea. CONCLUSIONS: RB with two antibiotics is as effective and safe as OCA-7 for the eradication of Helicobacter pylori.
Adult↗
Kinetic modeling of lactose hydrolysis with an immobilized beta-galactosidase from Kluyveromyces fragilis.
The kinetic model of the hydrolysis of lactose with a beta-galactosidase from Kluyveromyces fragilis immobilized on a commercial silica-alumina (KA-3, from Südchemie) has been determined. A wide experimental range of the main variables has been employed: temperature, concentrations of substrate, and products and concentration of enzyme. The runs were performed in a complex buffer with the salt composition of milk. The effect of pH and temperature on the stability and the activity of the enzyme have been studied. The optimum pH for the enzyme activity was, approximately, seven. The immobilized enzyme was more stable than the free one at acidic pH, but more instable at basic pH. The maximum temperature used for the hydrolysis runs performed to select the kinetic model was 40 degrees C, so inactivation of the enzyme during the kinetic runs has been avoided. Agitation, concentration of enzyme in the solid and particle size were selected to ensure that the overall rate was that of the chemical reaction. Eleven kinetic models were proposed to fit experimental data, from first order to more complex ones, such as those taking into account inhibition by one of the compounds involved in the hydrolysis reaction. Applying statistical and physical criteria, a Michaelis-Menten model with a competitive inhibition by galactose has been selected. The model is able to fit the experimental data correctly in the wide experimental range studied. Finally, the model obtained is compared to the one selected in a previous work for the hydrolysis of lactose with the free enzyme.
Journal Article↗
Activation of the human p27(Kip1) promoter by IFNalpha 2b.
p27(Kip1) is one of the key regulatory proteins in cell cycle through inhibition of pRB phosphorylation by suppression of the activity of several cyclin/Cdk complexes. The expression of p27(Kip1) has been shown to be controlled by a posttranslational mechanism, although vitamin D(3) and neuronal differentiation can also induce its mRNA. Recently, the p27(Kip1) promoter was isolated and sequenced from a human leukocyte genomic library. In this report, we demonstrate that IFNalpha 2b, activates the human p27(Kip1) promoter-driven luciferase reporter gene in transient expression assays in H82 cells. This induction might involve two IRF 1-like binding sites present in the p27(Kip1) promoter. To our knowledge this is the first report on the direct activation of the human p27(Kip1) promoter by IFNalpha 2b.
Base Sequence↗
False autoregulation (pseudoautoregulation) in patients with severe head injury. Its importance in CPP management.
False autoregulation has been described as an alteration of autoregulation in which the apparent maintenance of a constant cerebral blood flow (CBF) when increasing cerebral perfusion pressure (CPP) is due to an increase in brain tissue pressure. The objective of our study was to investigate how often false autoregulation occurred in patients with a severe head injury. In forty-six patients with a moderate or severe head injury autoregulation was studied using arteriojugular differences of oxygen (AVDO2) to estimate changes in CBF after inducing arterial hypertension with phenylephrine. Changes in mean arterial blood pressure (MABP), intracranial pressure (ICP), cerebral perfusion pressure (CPP) and AVDO2 were calculated before and after inducing hypertension. Ninety-five episodes of provoked hypertension were studied in 46 patients. In 28 tests (29.5%) a constant or even reduced CBF was detected simultaneously with a median increase in parenchymal ICP of 8.5 mm Hg (false autoregulation). In this group the median of the induced increase in MABP was 20.6 mm Hg with a median increase in CPP of 11.5 mm Hg. From our data we can conclude that false autoregulation is frequently found in patients after a severe head injury. Increasing MABP to obtain a better CPP in these patients is not beneficial because CBF is not modified or may even be reduced.
Adolescent↗
Does an increase in cerebral perfusion pressure always mean a better oxygenated brain? A study in head-injured patients.
The adequate management of cerebral perfusion pressure (CPP) continues to be a controversial issue in head-injured patients. The purpose of our study was to test two hypotheses. The first was that in patients with a CPP below 70 mm Hg, oxygen delivery is compromised and that therefore signs of tissue hypoxia would be reflected in low PtiO2 measurements. The second hypothesis was that manipulating mean arterial blood pressure to increase CPP improves oxygen delivery, particularly in patients with a CPP below 70 mm Hg. Twenty-five moderately or severely head-injured patients were included in the study. In all of them PtiO2 was monitored in the non-injured hemisphere using the Licox system (GMS, Kiel-Mielkendorf, Germany). Arterial hypertension was induced with phenylephrine 29 times. To quantify the effect of increasing mean arterial blood pressure (MABP) on oxygen delivery to the brain, the PtiO2-BP index was calculated (PtiO2-BP index = delta PtiO2/delta MABP). In 16 tests (55%) baseline CPP was above or equal to 70 mm Hg and in the remaining 13 (45%) it was below 70 mm Hg. Mean increase in MABP after phenylephrine was 23.7 +/- 10.2 mm Hg. Mean PtiO2 was 29.5 +/- 14.7 mm Hg in patients with a basal CPP of below 70 mm Hg and 28.9 +/- 10.6 mm Hg in patients in the high CPP group. These differences being not statistically significant. The PtiO2-BP index was 0.29 +/- 0.23 in patients with a basal CPP of below 70 mm Hg and in patients with a CPP of above 70 mm Hg this index was 0.16 +/- 0.11 Hg. These differences were not statistically significant (Student's t-test, P = 0.09). In our study a low PtiO2 was not observed in patients with marginally low CPPs (48-70 mm Hg) and readings below 15 mm Hg were observed in cases with both normal or supranormal CPPs. We conclude that episodes of low PtiO2 could not be predicted on the basis of CPP alone. On the other hand, raising CPP did not increase oxygen availability in the majority of cases, even if the CPP was markedly improved.
Adolescent↗
Impact of using marginal cadaver donors and long cold ischemia time in renal transplant survival.
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Cadaver↗
Factors associated with Helicobacter pylori infection by a cagA-positive strain in children.
Although infection with a cagA-positive Helicobacter pylori strain is considered a risk factor for the development of duodenal peptic ulcer in adults, this association has not been demonstrated in children. The presence of cagA was investigated by polymerase chain reaction in H. pylori strains isolated from 27 children with duodenal ulcer and 53 without duodenal ulcer. All patients (100%) with duodenal ulcer and 33 (62.3%) without ulcer were colonized by a cagA-positive strain (P=.00007). A cagA-positive status was also associated with a more marked macroscopic gastritis, with a greater inflammatory infiltrate of both mononuclear and polymorphonuclear cells in the antral and oxyntic gastric mucosae and degenerative and regenerative changes of the gastric mucosa. Increased cagA positivity was also associated with increased age, but no association between cagA-positive status and sex was observed.
Adolescent↗
Arterial distensibility in subjects with white-coat hypertension with and without diabetes or dyslipidaemia: comparison with normotensives and sustained hypertensives.
BACKGROUND: Arterial distensibility can be assessed by measuring pulse-wave velocity (PWV). OBJECTIVE: To determine whether diabetes, smoking and dyslipidaemia were associated with greater than normal stiffness of aortic walls in subjects with white-coat hypertension. METHODS: Arterial distensibility was assessed by automatic measurement of carotid-femoral PWV in 35 healthy normotensives, 46 white-coat hypertensives (WCH, clinic blood pressures >140/90 mm Hg, daytime blood pressures <130/85 mm Hg) and 81 ambulatory hypertensives (clinic blood pressures >140/90 mmHg, daytime blood pressures > or =130 mm Hg systolic or > or =85 mm Hg diastolic, or both) all matched for age, sex and body mass index. Nineteen normotensives (subgroup A), 28 WCH (subgroup A) and 37 ambulatory hypertensives (subgroup A) had only one or no other major cardiovascular risk factor whereas 16 normotensives (subgroup B), 18 WCH (subgroup B) and 44 ambulatory hypertensives (subgroup B) had also some combination of non-insulin-dependent diabetes, a smoking habit and dyslipidaemia. RESULTS: Both for the WCH and for ambulatory hypertensives diabetes and dyslipidaemia (subgroups B) were associated with higher (P<0.04) PWV (11.6+/-0.3 and 12.8+/-0.3m/s, respectively) than for subgroups A (9.3+/-0.5 and 10.9+/-0.6 m/s, respectively). In contrast, PWV for WCH in subgroup A (9.3+/-0.5m/s) did not differ (P>0.35) from those for the normotensive subgroups A (9.2+/-0.3m/s) and B (9.6+/-0.4m/s). PWV was not correlated to levels of glycaemia, glycosylated haemoglobin and cholesterolaemia. CONCLUSIONS: These results suggest that, both for ambulatory hypertensives and for WCH, diabetes and dyslipidaemia are associated with an impairment of arterial distensibility that can entail a greater than normal cardiovascular risk, which might dictate a more than usually stringent treatment of concomitant risk factors and possibly of high blood pressure. In contrast, PWV in WCH of the subgroup A did not differ from those in normotensives, reinforcing the hypothesis that WCH is associated with a benign cardiovascular outcome in the absence of other cardiovascular risk factors.
Adult↗
Nonlinear viscosity and velocity distribution function in a simple longitudinal flow
A compressible flow characterized by a velocity field u(x)(x, t)=ax/(1+at) is analyzed by means of the Boltzmann equation and the Bhatnagar-Gross-Krook kinetic model. The sign of the control parameter (the longitudinal deformation rate a) distinguishes between an expansion (a>0) and a condensation (a<0) phenomenon. The temperature is a decreasing function of time in the former case, while it is an increasing function in the latter. The non-Newtonian behavior of the gas is described by a dimensionless nonlinear viscosity eta(*)(a(*)), that depends on the dimensionless longitudinal rate a(*). The Chapman-Enskog expansion of eta(*) in powers of a(*) is seen to be only asymptotic (except in the case of Maxwell molecules). The velocity distribution function is also studied. At any value of a(*), it exhibits an algebraic high-velocity tail that is responsible for the divergence of velocity moments. For sufficiently negative a(*), moments of degree 4 and higher may diverge, while for positive a(*) the divergence occurs in moments of degree equal to or larger than 8.
Journal Article↗
(1-->6)-beta-D-glucan as cell wall receptor for Pichia membranifaciens killer toxin.
The killer toxin from Pichia membranifaciens CYC 1106, a yeast isolated from fermenting olive brines, binds primarily to the (1-->6)-beta-D-glucan of the cell wall of a sensitive yeast (Candida boidinii IGC 3430). The (1-->6)-beta-D-glucan was purified from cell walls of C. boidinii by alkali and hot-acetic acid extraction, a procedure which solubilizes glucans. The major fraction of receptor activity remained with the alkali-insoluble (1-->6)-beta- and (1-->3)-beta-D-glucans. The chemical (gas-liquid chromatography) and structural (periodate oxidation, infrared spectroscopy, and (1)H nuclear magnetic resonance) analyses of the fractions obtained showed that (1-->6)-beta-D-glucan was a receptor. Adsorption of most of the killer toxin to the (1-->6)-beta-D-glucan was complete within 2 min. Killer toxin adsorption to the linear (1-->6)-beta-D-glucan, pustulan, and a glucan from Penicillium allahabadense was observed. Other polysaccharides with different linkages failed to bind the killer toxin. The specificity of the killer toxin for its primary receptor provides an effective means to purify the killer toxin, which may have industrial applications for fermentations in which salt is present as an adjunct, such as olive brines. This toxin shows its maximum killer activity in the presence of NaCl. This report is the first to identify the (1-->6)-beta-D-glucan as a receptor for this novel toxin.
Adsorption↗
Detection of p56(lck) kinase activity using scintillation proximity assay in 384-well format and imaging proximity assay in 384- and 1536-well format.
p56(lck) is a lymphocyte-specific tyrosine kinase that plays an important role in both T-cell maturation and activation. We have developed a homogeneous assay in which p56(lck) catalyzes the transfer of the gamma-phosphate group from [gamma-(33)P]ATP to a biotinylated peptide substrate. The labeled peptide is then captured on a streptavidin-coated scintillation proximity assay (SPA) bead or imaging proximity bead. The SPA is counted in a microplate scintillation counter and the imaging proximity assay is counted in a charge-coupled device-based imaging system called LEADseekertrade mark, recently launched as a homogeneous imaging system by Amersham Pharmacia Biotech. We show, via time-dependence assays and inhibitor studies, that this assay can be performed in 1536-well microplate format using imaging proximity as the method of detection. The results compare favorably with the same assay performed in 384-well microplate format using both SPA and imaging proximity as the detection methods. From this study, we conclude that a kinase assay can be performed in 384- and 1536-well format using imaging as the detection method, with significant time savings over standard scintillation counting. In addition, we show cost saving advantages of 1536- over 384-well format in terms of reagent usage, higher throughput, and waste disposal.
Amino Acid Sequence↗
Characterization of the promoter region of the rat CCAAT/enhancer-binding protein alpha gene and regulation by thyroid hormone in rat immortalized brown adipocytes.
CCAAT/enhancer binding proteins (C/EBP) are a family of transcription factors with a highly conserved basic/leucine zipper (bZIP) domain that has been implicated in the transcriptional control of genes involved in cell growth and differentiation. We have previously demonstrated that the expression of C/EBPalpha and C/EBPbeta genes is regulated by thyroid hormone in rat liver during development. The aim of the present study was to explore the molecular mechanisms underlying the control of C/EBPalpha gene expression by thyroid hormone. To achieve this goal, we isolated and characterized a genomic clone containing 1171 bp of the 5'-flanking region of the rat C/EBPalpha gene. This fragment was an active promoter in MB492 cells, an immortalized brown adipocyte cell line that expresses the endogenous C/EBPalpha gene in a T3-dependent manner. Sequence analysis suggested the presence of three thyroid hormone response elements, TRE-1 (-602/-589), TRE2 (-411/-396), and TRE3 (-376/-350). The results of deletion, mutagenesis, and gel mobility shift analysis disclosed that only TRE-1, an ER2-type response element, represented a functional T3 response element. Our results demonstrate that T3 is a factor that positively regulates C/EBPalpha gene expression in a direct fashion.
Adipose Tissue, Brown↗