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Biomedical subjects

A Stern

Publications and source records attributed to A Stern.

At least 109 records · Page 6Linked to original sources
PubMed1993

Oxidative stress in fish cells: in vitro studies.

Bluegill sunfish BF-2 fibroblasts were used in the neutral red (NR) cytotoxicity assay to discern the toxicities of hydrogen peroxide (H2O2) and paraquat as indicated by their abilities to induce oxidative stress. The toxicity of H2O2 was markedly enhanced in BF-2 cells treated with the glutathione depleting agents, buthionine sulfoximine (BSO), maleic acid, and chlorodinitrobenzene; similar treatments did not sensitize the BF-2 cells to paraquat, a redox cycling xenobiotic. BSO treated BF-2 cells, however, were sensitized to nitrofurantoin, also a redox cycling chemical. Diethyldithiocarbamate, an inhibitor of superoxide dismutase, only weakly enhanced the sensitivity of the BF-2 cells to H2O2 and paraquat. 1,10-Phenanthroline, a chelator of Fe2+, reduced the cytotoxicity of H2O2 and paraquat, presumably by preventing hydroxyl radical formation in the Fenton reaction. Quin 2 AM, an intracellular chelator of Ca2+, markedly lessened the toxicity of H2O2, but not of paraquat; EGTA, an extracellular chelator of Ca2+, had no effect on the toxicity of H2O2 or paraquat. Apparently, perturbation of intracellular Ca2+ homeostasis is involved in H2O2 toxicity. For comparative purposes, some studies were performed with fathead minnow FHM epithelioid cells, BALB/c mouse 3T3 fibroblasts, and human HepG2 hepatoma cells. The BF-2 fibroblast/NR cytotoxicity red assay was shown to be a suitable model to study oxidative stress in fish.

3T3 Cells↗
PubMed1993

Ascorbic acid inhibits protein tyrosine phosphatases in NIH 3T3 cells expressing human epidermal growth factor receptors.

1. Physiological concentrations of ascorbic acid inhibited PTPase activity in HER 14 cells. 2. Higher concentrations of ascorbic acid produced a weaker inhibitory effect on PTPase activity in HER 14 cells. 3. EGF prevented the inhibitory effect of ascorbic acid on PTPase activity in HER 14 cells. 4. The inhibitory effect of physiological concentrations of ascorbic acid on PTPase activity depends on density of the cell culture, with less dense populations exhibiting greater inhibition of PTPase activity. 5. These observations suggest that ascorbic acid might have a modulatory role in cellular phosphorylation-dephosphorylation events.

3T3 Cells↗
PubMed1993

In vitro cytotoxicity of 1,4-naphthoquinone derivatives to replicating cells.

The acute cytotoxicities of a series of alkyl-1,4-naphthoquinones (NQ) and of 2-hydroxy-3-alkyl-1,4-NQs, as well as some amino derivatives, were evaluated with the neutral red cytotoxicity assay, using BALB/c mouse 3T3 fibroblasts. As compared to the unsubstituted 1,4-NQ: (i) Substitution at the 2 position reduced toxicity, with the extent of reduction following the sequence, hydroxyl >> dimethylamino >> C1-C5 alkyl group. (ii) Substitution with C2-C5 alkyl groups at position 3 enhanced toxicity. As noted with the n-alkyl-1,4-NQs, increasing the chain length of the 2-hydroxy-3-alkyl-1,4-NQs did not appreciably change potency of the test agent. (iii) Substitution with amino groups at position 3 had little effect on cytotoxicity. Some differences in cytotoxicity of specific test agent were noted between the 3T3 fibroblasts and isolated rat hepatocytes, as reported in the literature.

3T3 Cells↗
PubMed1993

To AZT or not to AZT.

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Emergency Medical Technicians↗
PubMed1993

Randomized comparative trial of imipenem/cilastatin versus aminoglycoside plus amoxycillin plus clindamycin in the treatment of severe intra- and post-operative infections.

In a prospective, randomized, controlled study, clinical and bacteriological efficacy of imipenem/cilastatin (I/C) was compared with a standard combination of aminoglycoside + amoxycillin + clindamycin (C) in patients (pts) with severe intra- and postoperative infections. A total of 84 pts were randomly separated into two groups of 42 pts. Diagnoses were pneumonia n = 21 (14 in I/C group and 7 in C), peritonitis n = 45 (16 in I/C group and 29 in C), septicaemia n = 12 (9 in I/C group and 3 in C), and 7 other infections (3 in I/C group and 4 in C). Doses used were imipenem/cilastatin 1 g q 8 h and amoxycillin 2 g q 8 h plus clindamycin 0.6 g q 6 h, plus netilmicin according to serum concentrations. Success rates were 85.4% (n = 35: 34 cured and one improved) in the I/C group and 83.3% (n = 35: 30 cured and five improved) in the C group. Six pts in group I/C and 7 in group C failed to respond to treatment. One patient in the I/C group was not assessable. 62% of the bacterial isolates were eradicated in the I/C group and 55% in group C. 7% were suppressed in I/C and 5% in C. It is concluded that imipenem/cilastatin is an effective and well-tolerated alternative to antibiotic combinations in severe intra- and post-operative infections. It offers the advantages of fewer drug doses and less renunciation of serum drug concentration monitoring.

Aminoglycosides↗
PubMed1992

Effects of ligands on reduction of oxygen by vanadium(IV) and vanadium(III).

V(IV) and V(III) reduce molecular oxygen with increasing rates as the pH is raised from 6.0 to 7.4. Under all conditions tested, V(IV) is the more efficient reductant. EDTA and ATP generally inhibit the reduction of oxygen by V(III) and V(IV). In contrast, desferrioxamine accelerates the reduction of oxygen by V(IV) but with decreasing effectiveness at pH 7.4 compared to pH 6.0, while desferrioxamine accelerates the reduction of oxygen by V(III) only at pH 6.0. Histidine enhances the reduction of oxygen by V(IV) at pH 7.0 and 7.4. The observed rates of oxygen reduction by V(III) and V(IV) imply that the intracellular distribution of vanadium among its redox states reflects not an equilibrium but a steady state.

Adenosine Triphosphate↗
PubMed1992

Binding of recombinant variants of human tissue-type plasminogen activator (t-PA) to human umbilical vein endothelial cells.

Endothelial cells synthesize and secrete hemostatic components like tissue-type plasminogen activator (t-PA) which is thought to be the major determinant of fibrinolytic activity in the blood. Most recently, a receptor protein for t-PA on human umbilical vein endothelial cells (HUVEC) in culture has been described (1); there are, however, in addition low affinity binding sites for t-PA on HUVEC. The sites of binding are of particular interest, because they are potential regulators of t-PA activity and clearance. We analysed the low affinity binding of recombinant t-PA (rt-PA) to normal diploid HUVEC and to the permanent human cell lines Jurkat, Daudi, HL 60 and K562 by flow cytometry applying t-PA specific monoclonal antibodies. Using this test system binding of both recombinant glycosylated human t-PA produced in Chinese hamster ovary cells (CHO-t-PA) and of nonglycosylated t-PA, produced in E. coli (BM 06.021) was investigated. Analysis of the binding pattern to HUVEC and other cell lines revealed that deglycosylation of full length rt-PA increases non-specific binding. Additionally, we investigated the binding properties of an unglycosylated t-PA deletion variant which comprises the kringle 2 and the protease domains (BM 06.022). Data obtained show that deletion of these domains most drastically reduces non-specific binding to HUVEC and other human cell lines.

Antibodies, Monoclonal↗
PubMed1992

Inhibition of hemin-induced hemolysis by desferrioxamine: binding of hemin to red cell membranes and the effects of alteration of membrane sulfhydryl groups.

Hemin binds to red cell membranes during hemin-induced hemolysis but the precise mechanism of hemolysis has not been characterized. Desferrioxamine (DFO), an iron chelator, inhibited hemin-induced hemolysis. DFO partially prevented hemin binding to red cell membranes and partially removed previously bound hemin. Glutathione, an intracellular sulfhydryl compound, also inhibited hemin-induced hemolysis but was only about one tenth as potent as DFO. Decrease of membrane sulfhydryl groups by treatment of cells with either N-ethylmaleimide (NEM) or diamide (azodicarboxylic acid bis [dimethylamide]) enhanced hemin-induced hemolysis. Enhancement of hemin-induced hemolysis by NEM and diamide and inhibition of hemolysis by DFO were independent with no evidence of synergism or interference between the two processes. Red cell membranes were saturated with hemin at approximately 75 nmol per mg protein. DFO decreased the hemin saturation level to 25 nmol per mg protein. In the presence of DFO, hemin was bound as the DFO-hemin complex since membranes preferentially removed DFO-hemin complexes from mixtures of complexed and free hemin while free DFO was not bound by the membranes. Access to the inner surface of the membrane was required for binding of the DFO-hemin complex since DFO completely prevented hemin binding in intact cells but not in cells undergoing hemolysis or red cell ghosts. Approximately 50 x 10(6) molecules of hemin were bound to the membrane of one red cell following hemin-induced hemolysis.

Deferoxamine↗
PubMed1992

Pharmacokinetic and thrombolytic properties of unglycosylated recombinant tissue-type plasminogen activator (BM 06.021) produced in Escherichia coli.

Recombinant tissue-type plasminogen activator (rt-PA) was produced in Escherichia coli cells in order to obtain an unglycosylated rt-PA (BM 06.021) with increased thrombolytic potency due to altered pharmacokinetic properties. The pharmacokinetics were studied in rabbits upon intravenous infusion of 200 kU/kg over 30 min. The thrombolytic dose-response effects were evaluated in a rabbit model with 125I-labeled venous thrombi upon intravenous infusion over 4 h. The thrombolytic effects after intravenous bolus injection of 200 kU/kg BM 06.021 were investigated in a canine model of coronary artery thrombosis. All studies were performed comparing BM 06.021 with glycosylated rt-PA (alteplase). BM 06.021 demonstrated a longer (p less than 0.05) half-life (5.6 +/- 2.6 vs. 2.1 +/- 0.3 min) and a lower (p less than 0.05) clearance rate (7.5 +/- 0.8 vs. 22.2 +/- 3.1 ml.min-1.kg-1) than alteplase in rabbits upon intravenous infusion. The dose-response curve of BM 06.021 for thrombolysis in a rabbit model of jugular vein thrombosis was located to the left of that for alteplase with a 2.1-fold lower effective dose of 50% thrombolysis (ED50) of BM 06.021 (207 vs. 436 kU/kg). Intravenous bolus injection of 200 kU/kg BM 06.021 induced the same reperfusion rate (4/6) as intravenous infusion of 800 kU/kg alteplase over 90 min in a canine model of coronary artery thrombosis. The residual thrombus wet weight did not significantly differ between BM 06.021 and alteplase (5.7 +/- 1.8 vs. 6.3 +/- 1.1 mg). The results indicate that unglycosylated rt-PA (BM 06.021) has a higher in vivo thrombolytic potency than glycosylated rt-PA (alteplase).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

In vitro cytotoxicity of methylated phenylenediamines.

The acute cytotoxicities of methylated phenylenediamines (PDs) were evaluated with the neutral red assay, using BALB/c 3T3 mouse fibroblasts as the bioindicators. When the test agents were grouped according to their degree of methylation, good correlations were noted between their in vitro cytotoxicity and their in vivo myotoxicity to experimental animals, as well as to their in vitro autoxidation rates. For test agents of comparable methylation, the sequence of potency was ring-methylated p-PD > N-methylated p-PD >> N-methylated o-PD > N-methylated m-PD.

Animals↗
PubMed1992

Desferrioxamine enhances the reactivity of vanadium (IV) and vanadium (V) toward ferri- and ferrocytochrome c.

Ligands, especially desferrioxamine, affect the rate at which vanadium reduces or oxidizes cytochrome c. Whether reduction or oxidation occurs, and how fast, depends on the nature of the ligand, the state of reduction of the vanadium, the pH (6.0, 7.0, or 7.4), and the availability of oxygen. In general, oxidation of ferrocytochrome c was favored by (1) low pH, (2) an oxidized state of the vanadium, (3) the presence of oxygen, and (4) more strongly binding ligands (desferrioxamine much greater than histidine = ATP greater than EDTA greater than albumin greater than aquo). Thus, at pH 6.0, desferrioxamine accelerated the V(V)-catalyzed ferrocytochrome c oxidation 160-fold aerobically, and 3500-fold anaerobically. In general, strongly binding ligands slowed oxidations, especially at higher pH. Desferrioxamine was unique among the five ligands in that it not only accelerated oxidation of ferrocytochrome c at pH 6.0, but at pH 7.4 the redox balance shifted to the point where it paradoxically reduced ferricytochrome c. V(V) is an improbable electron donor, but desferrioxamine will reduce cytochrome c, and V(V) accelerates this process. Oxidation of cytochrome c by V(V):desferrioxamine was faster anaerobically, and reduction by V(IV):desferrioxamine was faster aerobically. Although V(V) did not oxidize ferrocytochrome c at pH 7.4, V(IV) did, provided oxygen and desferrioxamine were both present. V(IV):desferrioxamine almost completely reduced ferricytochrome c, and this reduction was followed by a slow, progressive oxidation. This latter oxidation of cytochrome c is mediated by active species generated in the reaction between V(IV):desferrioxamine and oxygen, because none of these reagents alone can induce oxidation at a comparable rate. The mediating species were transient, and generated in reactions with oxygen.(ABSTRACT TRUNCATED AT 250 WORDS)

Catalase↗
PubMed1992

Biochemical properties of the kringle 2 and protease domains are maintained in the refolded t-PA deletion variant BM 06.022.

BM 06.022 is a t-PA deletion variant which comprises the kringle 2 and the protease domain. Production of BM 06.022 in Escherichia coli leads to the formation of inactive inclusion bodies, which have to be refolded by an in vitro refolding process to achieve activity and proper structure of the domains. We analysed the biochemical properties of BM 06.022 to obtain some information about the structure of kringle 2 and the protease as compared with the structure of these domains in the intact t-PA molecule. The kinetic analysis of the amidolytic activity of BM 06.022 and CHO-t-PA yielded similar values for kcat (13.9 s-1 and 11.4 s-1 for the single chain forms and 33.9 s-1 and 27.1 s-1 for the two chain forms of BM 06.022 and CHO-t-PA, respectively) and for Km (2.5 mM and 2.1 mM for the single chains forms and 0.5 mM and 0.3 mM for the two chain forms of BM 06.022 and CHO-t-PA, respectively). BM 06.022 and CHO-t-PA have the same plasminogenolytic activity in the absence of CNBr fragments of fibrinogen. However, BM 06.022 has a lower plasminogenolytic activity in the presence of CNBr fragments of fibrinogen and a lower affinity to fibrin as compared with CHO-t-PA. The affinity of BM 06.022 for fibrin is completely suppressed by 0.3 mM epsilon-aminocaproic acid, while the intact t-PA has a residual affinity of approximately 30%. The dissociation constants for the interaction with the lysine analogue epsilon-aminocaproic acid are 0.10 mM and 0.09 mM for BM 06.022 and the intact t-PA, respectively. Furthermore, BM 06.022 and CHO-t-PA are inhibited by PAI-1 in a similar manner.

Amidohydrolases↗