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Biomedical subjects

J Ostrowski

Publications and source records attributed to J Ostrowski.

At least 91 records · Page 5Linked to original sources
PubMed1992

Exogenous nitric oxide stress on endothelial cells and macrophages.

Bovine endothelial cells (ECs, P1) and lipopolysaccharide/gamma-interferon-induced mouse macrophages (MMs) were incubated in the presence of SIN-1 and C 3754 (1 microM to 1 mM), sydnonimine metabolites of the antianginal predrugs molsidomine and pirsidomine, respectively up to 48 h. No change of the endogenous nitric oxide output from MMs and A23187- or adenosine triphosphate-stimulated ECs was found by means of the methemoglobin method. Data indicate that downregulation of the nitric oxide (NO) synthase is not obvious within the intact cells under exogenous NO stress supplied by high concentrations of the spontaneous NO donors. Cytosolic MM NO synthase extracts, however, revealed reduction in the enzymic [3H]arginine turnover to [3H]citrulline by SIN-1, but not by C 3786, the pharmacologically active metabolite of pirsidomine.

Amino Acid Oxidoreductases↗
PubMed1992

Induction of ornithine decarboxylase in normal and protein kinase C--depleted human colon carcinoma cells.

To examine the role of protein kinase C (PKC) in induction of human colon adenocarcinoma cell line, DETA/W, by polypeptide growth-promoting factors, ornithine decarboxylase activity (ODC) and DNA synthesis were determined in cells depleted of PKC. PKC depletion was achieved by prolonged cultivation (more than 30 passages) with 10(-6) M phorbol 12-myristate 13-acelate. Lack of PKC in studied cells was proved by measurements of PKC activity and immunoreactivity. Although ODC activities and DNA syntheses in PKC-depleted cells were decreased by about 40-50% compared to normal DETA/W cells, the percentage increase of these mitogen-responsive reactions was quantitatively similar in both cell sublines. These results raise the possibility that not all of the biological responses to growth factors are connected with the activation of calcium-dependent PKC.

Adenocarcinoma↗
PubMed1991

A serine/threonine kinase activity is closely associated with a 65-kDa phosphoprotein specifically recognized by the kappa B enhancer element.

The immunoglobulin kappa light chain enhancer, kappa B, is an important cis-acting transcriptional element. kappa B binds a number of proteins including the members of the ubiquitous NF-kappa B family of transcription factors. Agarose beads coupled to a double-stranded oligonucleotide containing the kappa B motif were used to isolate a 65-kDa predominantly nuclear phosphoprotein. Southwestern blot analysis demonstrated that this phosphoprotein can bind the kappa B element directly and specifically. This kappa B-associated protein was phosphorylated in vivo and in vitro by a nuclear serine/threonine kinase(s) which, in a number of different cell lines, appeared to be stimulated in response to interleukin-1 alpha and lipopolysaccharide treatment. In the B cell lines 70Z/3 and CH12 LX2B, and the T cell line EL-4 6.1 C10 the activity of the kappa B-associated kinase(s) correlated with the binding activity of nuclear NF-kappa B displayed in a gel shift assay. In vitro, the 65-kDa protein was phosphorylated in the absence of exogenously added kinase. The 65-kDa phosphoprotein and the kinase activity remained associated following sequential anion-exchange and hydrophobic interaction chromatography. These results suggest that the kappa B-associated phosphoprotein is either autophosphorylated or is phosphorylated by a closely associated kinase(s). Stimulation of a nuclear protein kinase which is closely associated with a sequence-specific DNA may reflect a novel mechanism by which growth factors regulate gene expression.

Base Sequence↗
PubMed1991

A small region of the beta-adrenergic receptor is selectively involved in its rapid regulation.

Plasma membrane receptors that couple to guanine nucleotide-binding regulatory proteins (G proteins) undergo a variety of rapid (minutes) and longer term (hours) regulatory processes induced by ligands. For the beta 2-adrenergic receptor (beta 2AR), the rapid processes include functional desensitization, mediated by phosphorylation of the receptor by the cAMP-dependent protein kinase and the beta-adrenergic receptor kinase, as well as a loss of hydrophilic ligand binding proposed to represent sequestration of receptors into a cellular compartment distinct from the plasma membrane. The slower processes include beta 2AR down-regulation (i.e., a decrease in the total cellular complement of receptors). It is not yet known whether beta 2AR phosphorylation and/or sequestration are prerequisites for down-regulation of the receptor. Like other G protein-coupled receptors, the beta 2AR molecule spans the plasma membrane seven times, and the cytoplasmic carboxyl-terminal domain has been proposed to contain molecular determinants for each of these regulatory processes. We replaced four serine and threonine residues located within a 10-amino acid segment of this domain of beta 2AR and thereby prevented agonist-promoted phosphorylation, sequestration, and rapid desensitization of the adenylyl cyclase response. In contrast, these mutations did not affect functional coupling to the stimulatory G protein Gs or long-term down-regulation. These findings thus define a small, hitherto unappreciated region of the receptor molecule that may selectively subserve its rapid regulation. In addition, with the demonstration that beta 2AR does not have to be phosphorylated or sequestered in order to enter the down-regulation pathway, the results suggest that the classical receptor endocytosis model may not be appropriate for beta 2AR regulation.

Adenylyl Cyclases↗
PubMed1991

Negative autoregulation of cysB in Salmonella typhimurium: in vitro interactions of CysB protein with the cysB promoter.

CysB protein positively regulates genes of the Salmonella typhimurium cysteine regulon and negatively autoregulates cysB. The cysB promoter was characterized by primer extension of cellular RNA, which gave products identifying a major in vivo transcription start site located 95 bp upstream of the cysB start codon and two minor sites located 9 and 10 bp downstream of the major site. Gel shift binding studies and DNase I footprinting experiments showed that CysB protein binds to the cysB promoter from position -10 to +36 relative to the major transcription start site. We have designated this binding site CBS-B. CysB protein inhibited transcription initiation at the cysB promoter in an in vitro runoff assay, indicating that cysB is negatively autoregulated by the binding of CysB protein to the cysB promoter, where it acts as a repressor. N-Acetyl-L-serine, an inducer of the cysteine regulon, inhibited the binding of CysB protein to the cysB promoter and partially reversed the ability of CysB protein to inhibit transcription initiation. These effects are in contrast to those observed in studies of positively regulated cys promoters, in which N-acetyl-L-serine stimulates binding and causes CysB protein to activate transcription initiation.

Amino Acid Sequence↗
PubMed1991

Metabolism of vitamin A in inflammatory bowel disease.

The aim of this study was to determine serum retinol levels in patients with inflammatory bowel disease and to attempt to elucidate the mechanism of changes in vitamin A metabolism in these disorders. It was found that in 15 patients with active ulcerative colitis, 14 patients with active Crohn's disease and in 3 operated patients with recurrent Crohn's disease serum retinol levels and retinol-binding protein were significantly lower than in controls. Concentrations of vitamin A did not depend on the localization of inflammatory bowel disease, previous ileal resections, duration of the disease or age and sex of the patients. During successful treatment of active ulcerative colitis normalization of serum retinol levels without substitution of vitamin A was observed. Repeated determinations in patients with Crohn's disease who had low serum retinol levels in an active phase of disease revealed normal vitamin A levels in an inactive phase. The absorption of vitamins A and E in patients with inflammatory bowel disease was normal. The normal serum retinol concentrations in patients with diarrhea due to irritable bowel syndrome, and in those with anorexia nervosa exclude the influence of diarrhea and body weight itself on vitamin A levels. The results of this study indicate that serum retinol levels in patients with active inflammatory bowel disease are secondary to the decreased serum retinol-binding protein concentrations, and probably depend on the increased protein catabolism in these disorders.

Adult↗
PubMed1991

The relationship between extracellular K+ and Ca2+ on aminopyrine accumulation in rat parietal cells.

The effects of extracellular K+ in relation to extracellular Ca2+ on acid production were studied. Studies were performed in vitro using isolated cells from rat stomachs, and acid production was indirectly determined by 14C-aminopyrine (AP) accumulation. In the absence of K+ in the incubation medium histamine-stimulated AP accumulation ratios were significantly decreased independently in the presence or absence of extracellular Ca2+. Under basal conditions, in the absence of extracellular Ca2+, increasing concentrations of extracellular K+ enhanced AP accumulation ratios to significantly higher than those found in the presence of Ca2+. In histamine-, cAMP-, and carbachol-stimulated parietal cells, high K+ concentrations increased AP accumulation significantly less in Ca(2+)-free than in Ca(2+)-containing media. High K+ also induced significantly both an increase in cytosolic free Ca2+ concentration and 45Ca2+ uptake. The present results confirmed the importance of K+ in gastric acid production and suggested a role for Ca2+ as a modulator of mechanisms of parietal cell stimulation.

Aminopyrine↗
PubMed1991

Does the membrane potential control incorporation of tubulovesicles into the secreting apical membrane of the rat parietal cell?

The present studies were designed to examine the effect of changes in membrane potential by means of protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) and variations in the pH of the medium on the secretory response of parietal cells. Studies were performed in vitro using isolated cells from rat stomachs and acid production was indirectly determined by 14C-aminopyrine (AP) accumulation. CCCP affected both basal and histamine-stimulated AP accumulation in a concentration-dependent manner. The AP accumulation ratios depended on pH of the incubation medium; the ratio was lowest at pH 6.6, and increased progressively as the pH of the medium increased to 7.8. Moreover, the decreases in AP accumulation ratios caused by simultaneous addition of CCCP and AP to cell suspensions compared to those in which CCCP was added to incubated cells after achieving the steady-state of AP accumulation were quantitatively similar. These findings suggest that the decrease in AP accumulation due to CCCP treatment is a consequence of an activation of acid secretion rather than an inhibitor of acid production. From the present and previously published data, we propose a working hypothesis: membrane recycling is dependent on changes in apical membrane potential.

Aminopyrine↗
PubMed1991

Dose-related antiarrhythmic effects of nicainoprol in patients with chronic ventricular arrhythmias--a double-blind, placebo-controlled, cross-over multicentre trial. Nicainoprol Study Group.

One hundred and three patients with a history of ventricular arrhythmia were screened by baseline 24-h Holter monitoring for a minimal mean number of 30 premature ventricular beats (PVB) per hour. One hundred and one patients were subsequently enrolled in a double-blind, cross-over trial, including four 4-day periods with administration of placebo (PL) or nicainoprol (N.) in doses of 200, 400, 600 mg three times daily in randomized order. Antiarrhythmic efficacy could be evaluated in 61 patients who met all requirements of the protocol, including a complete sequence of valid Holter recordings (greater than 18 h each). Total PVBs were significantly reduced by the 400-mg dose (65%) and the 600-mg dose (71%) as compared to baseline. Two hundred mg N. and PL led to a non-significant reduction of PVBs by 41% and 24%, respectively. An individual response, assumed where there was simultaneous reduction of total PVBs greater than 75%, of ventricular pairs greater than 90% and elimination of ventricular runs, was found in 6 (10%), 10 (16%), 21 (34%), and 22 (36%) patients with PL, 200 mg N., 400 mg N. and 600 mg N., respectively. Four hundred and 600 mg N, significantly prolonged the PR interval (0.17 s and 0.18 s; median values) as compared with baseline (0.16 s), while the QRS duration was significantly increased only by the 600-mg dose (from 0.10 to 0.11 s). Heart rate, mean blood pressure, QT and JTc intervals were not significantly influenced by any dose of N.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1990

Weak bases increase surface IgM expression in 70Z/3 B lymphoid cell line without increasing kappa gene expression.

A murine B lymphoid cell line, 70Z/3, has proven to be a useful model to study the developmental transition from membrane immunoglobulin (mIgM)-negative to mIgM-positive B cells. 70Z/3 cells have normal intracellular levels of mu heavy chain but kappa mRNA is transcribed at a very low rate. Because both kappa and mu chains are required for IgM to localize on the external membrane the cells remain essentially mIgM negative. A number of biologic agents can stimulate kappa mRNA transcription leading to increased surface IgM expression. Pharmacologic agents which cause cytoplasmic alkalinization have also been shown to increase the level of surface IgM expression when assessed by FITC-labeled anti-IgM antibodies. This increase has been used to argue that cytoplasmic alkalinization is sufficient to stimulate 70Z/3 differentiation but previous studies did not measure kappa mRNA levels or rate of synthesis. In this study we set out to determine whether the increased FITC staining that follows cytoplasmic alkalinization results from increased surface IgM expression and whether that increase results from activation of kappa mRNA transcription. When cells were treated with methylamine, NH4Cl, or monensin there was transient cytoplasmic alkalinization. The levels of surface IgM expression were measured by flow cytometry of cells stained with FITC-labeled anti-mouse kappa and mu. Using this criterion, a 24-hr treatment with either weak bases or monensin increased the level of surface IgM. The increase in fluorescence was not the result of nonspecific binding or uptake by the cell because there was no increase in fluorescence when methylamine-treated cells were stained with FITC-labeled antibody directed against an antigen not found on these cells. Iodination of surface proteins confirmed that the increase in fluorescence was the result of increased levels of IgM protein on the cell surface. However, exposure of the cells to weak bases or monensin caused no increase in either the steady-state level of kappa light chain mRNA, or in the level of kappa protein. We conclude that the transient alkalinization is not sufficient to induce differentiation of the 70Z/3 cells.

Ammonium Chloride↗
PubMed1990

Determination of the active metabolite of molsidomine in human plasma by reversed-phase high-performance liquid chromatography.

A reversed-phase high-performance liquid chromatographic method, with ultraviolet detection, is proposed for the plasma determination of SIN-1, the active metabolite of molsidomine, which involves propoxycarbonyl derivatization. The internal standard is the ethoxycarbonyl derivative of SIN-1 (i.e. molsidomine). Derivatization and extraction are each performed in one step (2 min) with 70% yield. The nature of a by-product is discussed. The method provides rapid elution (less than 15 min), linearity over the range 0.4-200 ng/ml, day-to-day precision between 2.5 and 11.3% and a limit of determination of 0.5 ng/ml. This method is also suitable for the simultaneous determination of molsidomine and SIN-1. In this case the internal standard is an ethoxycarbonyl derivative of a piperazino-3-sydnonimine, a SIN-1 analogue.

Antihypertensive Agents↗
PubMed1990

[Comparison of the quantitative determination of blood in stools with the Hemoccult test in various digestive tract diseases].

The reported investigations of haemoglobin (Hb) concentration in stools of healthy subjects and patients with digestive tract diseases were carried out for determining the physiological value of blood loss into the digestive tract, and for comparing the sensitivity of quantitative and qualitative tests for demonstration of occult blood in stools in cases of colonic neoplasms. The qualitative Haemoccult test was compared to the quantitative test based on heme conversion to porphyrins by iron decoupling, which makes possible determination of total Hb content in stools and measurement of the so called intestinal fraction which develops as a result of Hb conversion to porphyrins in the upper digestive tract.

Adult↗
PubMed1990

In vitro interactions of CysB protein with the cysK and cysJIH promoter regions of Salmonella typhimurium.

The cysteine regulons of Salmonella typhimurium and Escherichia coli are positively regulated by CysB protein and either O-acetyl-L-serine or N-acetyl-L-serine, both of which act as inducers. Gel mobility shift assays and DNase I footprinting experiments showed that CysB protein binds to the S. typhimurium cysK promoter at two sites, one, designated CBS-K1, at positions -78 to -39 relative to the major transcription start site, and the other, designated CBS-K2, at positions -115 to -79. The S. typhimurium cysJIH promoter was found to contain a single binding site, designated CBS-JH, at positions -76 to -35. Acetyl-L-serine stimulated binding to CBS-K1 and CBS-J and inhibited binding to CBS-K2. In the absence of acetyl-L-serine, CysB protein bound to both CBS-K1 and CBS-K2 and gave a complex that migrated more slowly during gel electrophoresis than did that formed in the presence of acetyl-L-serine, in which case CysB protein bound only to CBS-K1. Complexes formed with DNA containing the two binding sites either at the middle or at one end of the fragment migrated differently, suggesting that DNA was bent in the slow complex formed in the absence of acetyl-L-serine and that DNA in the fast complex was less bent or not bent at all. An analysis of upstream deletions of the cysK promoter showed that only CBS-K1 is required for in vivo promoter activity. CBS-J is analogous in position to CBS-K1 and is probably also required for activity of the cysJIH promoter. CBS-K2 has no known function but may help sequester CysB protein at the cysK promoter.

Base Sequence↗
PubMed1990

In vitro interactions of CysB protein with the cysJIH promoter of Salmonella typhimurium: inhibitory effects of sulfide.

The cysteine regulon of Salmonella typhimurium is positively regulated by the CysB protein and an inducer, which can be either O-acetyl-L-serine or N-acetyl-L-serine. In vivo experiments confirmed that sulfide and L-cysteine (supplied as L-cystine) interfere with induction by exogenously supplied O-acetyl-L-serine and also showed the same effects when N-acetyl-L-serine was used as an inducer. In a gel shift assay, purified CysB protein bound specifically to a 278-base-pair DNA fragment containing the S. typhimurium cysJIH promoter region. Binding occurred in the absence of inducer but did not stimulate in vitro transcription initiation, indicating that binding alone is insufficient to cause formation of a transcription initiation complex. Addition of N-acetyl-L-serine or O-acetyl-L-serine was required for transcription initiation and also stimulated binding three- to eightfold. Sulfide inhibited both transcription initiation and binding by interfering with the stimulatory effects of inducer in a competitive manner. These findings indicate that sulfide is an anti-inducer and may explain why full expression of the cysteine regulon requires sulfur limitation. L-Cysteine did not affect in vitro transcription initiation or binding of CysB protein to the cysJIH promoter region. The in vivo effects of L-cysteine may be secondary to its degradation to sulfide by the inducible enzyme cysteine desulfhydrase.

Bacterial Proteins↗