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J R Mahoney

Publications and source records attributed to J R Mahoney.

11 recordsLinked to original sources
PubMed1991

A spectrophotometric assay for chlorine-containing compounds.

Determinations of hypochlorous acid and chloramine compounds are important in a number of areas. Several techniques are now available for such analyses, but most require unstable reagents and/or multiple steps in the analytical procedure. We have developed a simple, one-step spectrophotometric assay for reactive chlorine-containing compounds involving the oxidation of ascorbic acid by hypochlorous acid or chloramines. There is no interference from other nonhalide oxidants such as hydrogen peroxide or hypothiocyanous acid. Because small amounts of ascorbic acid will not damage biological materials, this method also allows continuous measurements of the generation of chlorine-containing compounds by activated neutrophils. This simple assay permits precise analysis of as little as 1 nmol of HOCl.

Ascorbic Acid↗
PubMed1991

Thiocyanate is the major substrate for eosinophil peroxidase in physiologic fluids. Implications for cytotoxicity.

The potent cytotoxic capacity of eosinophils for parasites and host tissue has in part been attributed to the catalytic action of eosinophil peroxidase (EPO), which preferentially oxidizes Br- to the powerful bleaching oxidant HOBr in buffers that mimic serum halide composition (100 mM Cl-, 20-100 microM Br-, less than 1 microM I-). However, serum also contains 20-120 microM SCN-, a pseudohalide whose peroxidative product, HOSCN, is a weak, primarily sulfhydryl-reactive oxidant. Because of its relative abundance and high oxidation potential, we hypothesized that SCN-, not Br- or I-, is the major substrate for EPO in physiologic fluids. We find that in Earle's buffer (100 mM Cl-) supplemented with 100 microM Br- and varying concentrations of SCN-, HOBr production by activated eosinophils and purified EPO, assayed by conversion of fluorescein to dibromofluorescein, was 50% inhibited (ID50) by only 1 microM SCN-. SCN- also blocked (ID50 10 microM) EPO oxidation of I- to HOI, assayed as iodofluorescein, despite the presence of 100 microM (i.e. grossly supraphysiologic) I-. Thionitrobenzoic acid oxidation kinetics indicate that SCN- is the initial species oxidized by EPO in equimolar mixtures of SCN- and Br- and in human serum. EPO also catalyzed the covalent incorporation of [14C]SCN- into proteins in buffers regardless of Br- concentration and in human serum. Comparing the cytotoxicity of HOSCN and HOBr for host cells, we find that even subphysiologic concentrations of SCN- (3.3-10 microM) nearly completely abrogate the potent Br(-)-dependent toxicity of EPO for 51Cr-labeled aortic endothelial cells and isolated working rat hearts, recently developed models of eosinophilic endocarditis. Thus, HOSCN, hitherto best known as a bacteriostatic agent in saliva and milk, is likely also the major oxidant produced by EPO in physiologic fluids, and the presence of SCN- averts damage to EPO-coated host tissues that might otherwise accrue as a result of HOBr generation. In view of these findings, the potential role of HOSCN in eosinophil killing of parasitic pathogens deserves close examination.

Animals↗
PubMed1991

Bromide-dependent toxicity of eosinophil peroxidase for endothelium and isolated working rat hearts: a model for eosinophilic endocarditis.

Eosinophilic endocarditis is a potentially lethal complication of chronic peripheral blood hypereosinophilia. We hypothesized that eosinophil peroxidase (EPO), an abundant eosinophil (EO) cationic granule protein, promotes eosinophilic endocarditis by binding to negatively charged endocardium, and there generating cytotoxic oxidants. Using an immunocytochemical technique, we demonstrated endocardial deposition of EPO in the heart of a patient with hypereosinophilic heart disease. Because EPO preferentially oxidizes Br- to hypobromous acid (HOBr) rather than Cl- to hypochlorous acid (HOCl) at physiologic halide concentrations, we characterized the Br(-)-dependent toxicity of both activated EOs and purified human EPO towards several types of endothelial cells and isolated working rat hearts. In RPMI supplemented with 100 microM Br-, phorbol myristate acetate-activated EOs, but not polymorphonuclear leukocytes, caused 1.8-3.6 times as much 51Cr release from four types of endothelial cell monolayers as in RPMI alone. H2O2 and purified human EPO, especially when bound to cell surfaces, mediated extraordinarily potent, completely Br(-)-dependent cytolysis of endothelial cells that was reversed by peroxidase inhibitors, HOBr scavengers, and competitive substrates. We further modeled eosinophilic endocarditis by instilling EPO into the left ventricles of isolated rat hearts, flushing unbound EPO, then perfusing them with a buffer containing 100 microM Br- and 1 microM H2O2. Acute congestive heart failure (evidenced by a precipitous decrement in rate pressure product, stroke volume work, aortic output, and MVO2 to 0-33% of control values) ensued over 20 min, which deletion of EPO, Br-, or H2O2 completely abrogated. These findings raise the possibility that EPO bound to endocardial cells might utilize H2O2 generated either by overlying phagocytes or endogenous cardiac metabolism along with the virtually inexhaustible supply of Br- from flowing blood to fuel HOBr-mediated cell damage. By this mechanism, EPO may play an important role in the pathogenesis of eosinophilic endocarditis.

Animals↗
PubMed1990

Recovery of postischemic myocardial ATP levels and hexosemonophosphate shunt activity.

Recovery of myocardial ATP levels after a period of ischemia is slow, requiring several days to return to normal. The biochemical limitation for ATP recovery appears to be the availability of the adenine nucleotide (AN) precursor, phosphoribosylpyrophosphate (PRPP), which is produced by the phosphorylation of ribose in the hexosemonophosphate shunt (HMP). In fact, ATP precursors, in particular ribose, have been shown to enhance the rate of postischemic ATP recovery. Infusion of exogenous ribose bypasses the rate-limiting steps in the HMP and speeds up adenine nucleotide (AN) biosynthesis. I propose another method for augmenting the rate of postischemic ATP recovery; increase the flux of substrate through the HMP. This would have the effect of making more PRPP available for AN biosynthesis. Effective physiologically and biologically tolerable means for enhancing HMP activity are presently available. These may be of significant utility in facilitating postischemic myocardial energy recovery.

Adenine Nucleotides↗
PubMed1989

A comparison of different carbohydrates as substrates for the isolated working heart.

Ribose has been shown to greatly enhance ATP recovery in situations such as postischemia when total adenine nucleotides have been depleted by catabolism. In addition, metabolic studies have reported that both five carbon sugars and alcohols (ribose and xylitol) can support energy metabolism presumably after conversion to substrates for glycolysis. Because of the importance of these two aspects of energy metabolism to myocardial function, we compared the ability of ribose and xylitol with glucose and pyruvate as exclusive substrates for the isolated working rat heart. Our studies revealed, however, that the utilization of ribose or xylitol as substrates by the myocardium is not sufficiently rapid to rely on these as exclusive oxidizable substrates. In fact, ribose or xylitol are no more effective than substrate-free medium in this regard. Myocardial glycogen was depleted in these groups and after a lag period consumption of oxygen also decreased. In contrast to the postischemic situation the total adenine nucleotide levels were preserved during ribose, xylitol or substrate-free perfusion. Consequently, the energy charge in these hearts fell significantly. In hearts perfused with ribose, xylitol or no substrate, the rate pressure product and the stroke volume rapidly declined after an initial brief stable period corresponding to glycogen depletion. Glycogen levels were 6% of the average control value in ribose- and xylitol-perfused hearts and were undetectable in substrate-free perfused hearts. In contrast, either glucose or pyruvate supported steady levels of ATP and myocardial oxygen consumption; maintained the energy charge; and supported the stroke volume, rate pressure product, and cardiac work. In glucose-perfused hearts the glycogen was reduced to 21% of control values, while in pyruvate-perfused hearts the average glycogen levels were 76% of control. Thus, although the heart is able to metabolize ribose and xylitol through the hexose monophosphate pathway, the rate of utilization through glycolysis and presumably the TCA cycle is not sufficient for these compounds to serve as exclusive substrates for the isolated working heart.

Adenine Nucleotides↗
PubMed1989

Enhanced high energy phosphate recovery with ribose infusion after global myocardial ischemia in a canine model.

High energy phosphate levels are depressed following global ischemia and require several days to completely recover. Short-term methods to enhance ATP recovery have included infusion of ATP precursors, inhibition of enzymes that catabolize AMP, and membrane transport stabilization. Several precursors have been used to augment adenine nucleotide synthesis including adenosine, inosine, adenine, and ribose. Because of the short-term nature of previous experiments, recovery had been incomplete and the effects in the intact animal unknown. The purpose of this study was to determine the effects of ribose infusion in a long-term model of global ischemia and attempt to identify the precursor which limits myocardial ATP regeneration in the intact animal. Global myocardial ischemia (20 min, 37 degrees C) was produced in dogs on cardiopulmonary bypass. With reperfusion either ribose (80 mM) in normal saline or normal saline alone was infused at 1 ml/min into the right atrium and the animals were followed for 24 hr. Ventricular biopsies were obtained through an indwelling ventricular cannula prior to ischemia, at the end of ischemia, and 4 and 24 hr postischemia and analyzed for adenine nucleotides and creatine phosphate levels. Radiolabeled microspheres were used to measure myocardial and renal blood flows and no significant difference was found between ribose-treated control groups. In both groups, myocardial ATP levels fell by at least 50% at the end of ischemia. No significant ATP recovery occurred after 24 hr in the control dogs, but in the ribose-treated animals, ATP levels rebounded to 85% of control by 24 hr.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1989

Acute iron poisoning. Rescue with macromolecular chelators.

Acute iron intoxication is a frequent, sometimes life-threatening, form of poisoning. Present therapy, in severe cases, includes oral and intravenous administration of the potent iron chelator, deferoxamine. Unfortunately, high dose intravenous deferoxamine causes acute hypotension additive with that engendered by the iron poisoning itself. To obviate this problem, we have covalently attached deferoxamine to high molecular weight carbohydrates such as dextran and hydroxyethyl starch. These macromolecular forms of deferoxamine do not cause detectable decreases in blood pressure of experimental animals, even when administered intravenously in very large doses, and persist in circulation much longer than the free drug. These novel iron-chelating substances, but not deferoxamine itself, will prevent mortality from otherwise lethal doses of iron administered to mice either orally or intraperitoneally. Further reflecting this enhanced therapeutic efficacy, the high molecular weight iron chelators also abrogate iron-mediated hepatotoxicity, suppressing the release of alanine aminotransferase. We conclude that high molecular weight derivatives of deferoxamine hold promise for the effective therapy of acute iron intoxication and may also be useful in other clinical circumstances in which control of free, reactive iron is therapeutically desirable.

Acute Disease↗
PubMed1988

The role of oxygen in toxic shock syndrome.

The etiology of toxic shock syndrome (TSS) has been extensively investigated in recent years. It is generally accepted that the causative organism is Staphylococcus aureus. Certain strains of this bacterium produce one or more toxins which are thought to be responsible for the symptoms associated with TSS. There is no general agreement, however, as to why menstruating women who use tampons are of particular risk for developing this disease. More recently, TSS has also been associated with the use of absorbent contraceptive sponges. In this paper I will present evidence that the introduction of oxygen into the vaginal cavity during tampon insertion is responsible in part for the development of TSS. Furthermore, I will discuss several methods for the safe removal of this exogenous oxygen.

Female↗
PubMed1985

Lipopolysaccharide-treated RAW 264.7 cells produce a mediator that inhibits lipoprotein lipase in 3T3-L1 cells.

RAW 264.7 cells upon stimulation with lipopolysaccharide secrete a protein mediator(s) that suppresses lipoprotein lipase activity in differentiated 3T3-L1 cells. The mediator(s), which is absent from unstimulated culture supernatants, is nondialyzable and thermolabile. Preliminary characterization suggests that this mediator(s) may be the same as that previously found in medium from lipopolysaccharide-treated thioglycollate-elicited mouse peritoneal macrophage cultures.

Adipose Tissue↗
PubMedDate not supplied

Hemolytic and microbicidal actions of diethyldithiocarbamic acid.

Micromolar concentrations of diethyldithiocarbamic acid (DDC) kill fungi, bacteria and malaria. DDC forms chelates with copper and the microbicidal effectiveness of this drug is enhanced greatly by small amounts of copper. DDC, in the presence of at least 1 molar equivalent of copper, also causes lysis of human erythrocytes. To explore the cytocidal actions of DDC and copper, we have used human erythrocytes and Escherichia coli as models. We found that: (1) the combination of DDC and copper also lysed E. coli spheroplasts, suggesting a possible common mechanism of hemolytic and microbicidal action; (2) higher ratios of drug: metal (greater than 4:1) diminished hemolytic and, as observed earlier, microbicidal effects; (3) cobalt, known to suppress the microbicidal effects of DDC:Cu, also prevented red cell lysis; (4) despite the necessary involvement of copper in DDC-mediated hemolysis, there was no evidence of oxidative damage to erythrocytes, and both lysis of erythrocytes and killing of E. coli were undiminished in the absence of oxygen; (5) the DDC:Cu chelate preferentially located in organic solvents and in membranes of erythrocytes. The chelate was quite soluble in chloroform but much less so in a C-16 hydrocarbon (hexadecane) which resembled erythrocyte membrane lipid. In hexadecane and at greater than 10(-4) M DDC and 5 x 10(-5) copper, an amphipathic drug:metal complex accumulated at the organic:aqueous interface; and (6) this amphipathic complex may permeabilize the lipid bilayer, causing leakage of ions and cell water and eventuating in colloid osmotic lysis. Red cells and E. coli exposed to the chelate showed early loss of intracellular rubidium (86Rb+). Furthermore, lysis of erythrocytes and E. coli spheroplasts was suppressed by the inclusion of either dextran or sucrose. Thus, it appears that DDC:Cu chelates are cytocidal by virtue of concentrating in the lipid bilayer and, perhaps, forming amphipathic complexes which disrupt membrane integrity. Drugs with similar behavior hold promise for therapy of malaria because metals capable of forming such complexes may accumulate within parasitized red cells.

Cell Membrane Permeability↗