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C M Arroyo

Publications and source records attributed to C M Arroyo.

33 records · Page 2Linked to original sources
PubMed1991

Difficulties encountered in the detection of nitric oxide (NO) by spin trapping techniques. A cautionary note.

The spin trapping technique was used in an attempt to detect the free radical nitric oxide (NO) in solution. Five different spin traps were examined, alpha-phenyl-N-tert butyl nitrone (PBN), alpha-(4-pyridyl-N-oxide) N-tert-butylnitrone (POBN), 5,5-dimethyl-pyrroline-N-oxide (DMPO), 2-methyl-2-nitrosopropane (MNP) and 3,5-dibromo-4-nitrosobenzene sulfonate (DBNBS). Our results suggest that the nitroso spin traps (MNP, DBNBS) are better suited for the identification of NO-related signals, than the nitrones, DMPO, PBN and POBN. In addition, it is shown that spin trapping of NO-related signals with nitroso and nitrone spin traps is subject to many artifacts.

Cyclic N-Oxides↗
PubMed1990

Receptor-mediated generation of an EDRF-like intermediate in a neuronal cell line detected by spin trapping techniques.

We have studied receptor-mediated generation of an activator of soluble guanylate cyclase in cultured mouse neuroblastoma cells (clone N1E-115) by ESR/spin trapping spectroscopy. A spin adduct was detected during the activation of muscarinic receptors by carbamylcholine in the presence of the spin trap 3,5-dibromo 4-nitrosobenzene sulphonate (DBNBS). The spin adduct does not correspond to that originating from the free radical nitric oxide or hydroxylamine. The same adduct was generated in cytosol preparations from N1E-115 cells incubated with L-arginine, NADPH, in the presence of calcium. The use of isotopically labelled guanidino-N15-L-arginine supported the generation of a DBNBS spin trapped adduct originating from the guanidino moiety of L-arginine. Superoxide dismutase (SOD) stabilized the precursor of the spin adduct as well as the activator of soluble guanylate cyclase derived from L-arginine. Our results provide direct evidence for the receptor-mediated formation of a diffusible precursor of NO. derived from L-arginine.

Animals↗
PubMed1990

ESR study of electron transfer reactions between gamma-irradiated pyrimidines, adriamycin and oxygen.

Solid pyrimidine nucleic acid bases (cytosine, thymine, and uracil) were gamma-irradiated (50 KGy) and dissolved in deaerated solutions of adriamycin in water and dimethylsulfoxide (DMSO). Analogous experiments using unirradiated pyrimidines as controls were also performed. In water only gamma-irradiated cytosine showed a reaction with the adriamycin yielding a single ESR peak (g = 2.0033) consistent with the adriamycin semiquinone radical. Since the unirradiated cytosine gave no reaction, the result suggests an electron transfer from cytosine radicals (generated by gamma-radiolysis) to adriamycin. In DMSO the three gamma-irradiated and unirradiated pyrimidines reacted with adriamycin yielding the adriamycin semiquinone radical observed by ESR. These results suggest that in DMSO an electron is transferred to adriamycin from the pyrimidine radicals and from the parent pyrimidine molecules. However, the process is on the order of 10(5) times more efficient for the pyrimidine radicals. Superoxide radicals (O2-.) were formed following addition of oxygen to the deaerated DMSO solutions containing adriamycin semiquinone radicals. O2-. was spin trapped using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). The results show a possible reaction sequence in which an electron transferred to adriamycin, by pyrimidine radicals and parent pyrimidine molecules, is subsequently transferred to dissolved oxygen.

Dimethyl Sulfoxide↗
PubMed1990

Endothelial cells as a source of oxygen-free radicals. An ESR study.

Endothelial cells were subjected to anoxia/reoxygenation in order to simulate some of the free radical mechanisms occurring in ischaemia/reperfusion. With ESR and spin trapping using the spin traps 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) and 3,3,5,5-dimethyl-1-pyrroline-1-oxide (M4PO), the results show that upon reoxygenation of endothelial cells, following a period of anoxia, these cells generate superoxide (O2-.). Cytotoxicity of the spin traps was measured by standard trypan blue exclusion methods. Cell injury or death was measured at various times during reoxygenation by lactate dehydrogenase (LDH) release. Experiments using oxyupurinol, SOD, CAT and a combination of SOD and CAT show that while oxypurinol partially prevents spin adduct formation, the combination of SOD and CAT is more effective in doing so. These results suggest that the majority of the oxygen radicals produced by endothelial cells are done so exogenously. The results also suggest that endothelial cells are not only a source of oxygen radicals but also a target.

Animals↗
PubMed1989

Inhibition of sarcolemmal carbon-centered free radical formation by propranolol.

The mechanism of propranolol-inhibited sarcolemmal membrane lipid peroxidation was investigated by electron spin resonance spin-trapping technique using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) and 2-methyl-2-nitrosopropane (MNP). Highly purified canine myocytic sarcolemma were peroxidized by a superoxide-driven (from dihydroxyfumarate) and Fe3+-catalyzed free radical-generating system. Hydroxyl radicals (.OH), identified by electron spin resonance signals as DMPO-OH adducts, were generated in the aqueous phase. Propranolol up to 500 microM did not effectively reduce the intensity of the DMPO-OH adducts. When the sarcolemma were incubated with MNP before the addition of free radicals, MNP adducts characteristic of carbon-centered radicals were produced. Pretreatment of the membranes with propranolol (3-100 microM) decreased the intensity of the MNP adducts in a log concentration-dependent manner; the EC50 is about 7 microM. D- and L-propranolol were found equally effective. When protein-depleted sarcolemmal lipids were similarly incubated with MNP and the free radical system, identical MNP adducts were observed; this finding suggests that the adducts were lipid-derived products arising from lipid peroxidation. Furthermore, their formation was also inhibited by propranolol pretreatment. Since propranolol is not an effective scavenger of oxygen radicals in the aqueous phase, the data suggest that the antiperoxidative effect of propranolol is due to its lipophilic interaction with the membrane and thus subsequent interruption of the free radical chain reactions.

Animals↗
PubMed1989

Spin trapping of free radicals formed during peroxidation of sarcolemmal membranes (SLM).

The generation of free radicals in a superoxide (O2-.) driven Fe+3 catalysed reactions with isolated myocytic sarcolemma using electron spin resonance was investigated. Incubation of highly purified canine myocytic sarcolemma in the presence of the spin trap, 2-methyl-2-nitrosopropane (MNP), followed by the addition of dihydroxyfurmarate (DHF) and Fe+3-ADP resulted in the generation and detection of radical adducts of this spin trap. Spin trapping of the alkyl radicals with 2-methyl-2-nitrosopropane led to the identification of methyl radical adduct following exposure to DHF/Fe+3-ADP. With sarcolemma and the alkyl nitroso compound, the only radical product trapped was the methyl radical formed by beta-scission of alkoxyl radical. The participation of hydroperoxide-derived radicals in this system verified that the decomposition of unsaturated hydroperoxy fatty acid does proceed via a free radical mechanism.

Animals↗
PubMed1988

Reaction of disodium cromoglycate with hydrated electrons.

A possible mechanism by which disodium cromoglycate (DSCG) prevents a decrease in regional cerebral blood flow but not hypotension in primates following whole body gamma-irradiation was studied. Several studies have implicated superoxide radicals (O2-.) in intestinal and cerebral vascular disorders following ischemia and ionizing radiation, respectively. O2-. is formed during radiolysis in the reaction between hydrated electrons (e-aq) and dissolved oxygen. For this reason, the efficiency of DSCG to scavenge e-q and possibly prevent the formation of O2-. was studied. Hydrated electrons were produced by photolysis of potassium ferrocyanide solutions. The rate constant, k = 2.92 x 10(10) M-1s-1 for the reaction between e-aq and DSCG was determined in competition experiments using the spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). This spin trap reacts rapidly with e-aq followed by protonation to yield the ESR observable DMPO-H spin adduct. The results show that DSCG is an efficient e-aq scavenger and may effectively compete with oxygen for e-aq preventing the radiolytic formation of O2-..

Cromolyn Sodium↗
PubMed1988

Effect of 4-hydroxypyrazolo (3,4-d) pyrimidine (allopurinol) on postirradiation cerebral blood flow: implications of free radical involvement.

In an attempt to elucidate mechanisms underlying the irradiation-induced decrease in regional cerebral blood flow (rCBF) in primates, hippocampal and hypothalamic blood flows of rhesus monkeys were measured by hydrogen clearance, before and after exposure to 100 Gy, whole body, gamma irradiation. Systemic blood pressures were monitored simultaneously. Compared to control animals, the irradiated monkeys exhibited an abrupt decline in systemic blood pressure to 35% of the preirradiation level within 10 min postirradiation, falling to 12% by 60 min. A decrease in hippocampal blood flow to 32% of the preirradiation level was noted at 10 min postirradiation, followed by a slight recovery to 43% at 30 min and a decline to 23% by 60 min. The hypothalamic blood flow of the same animals showed a steady decrease to 43% of the preirradiation levels by 60 min postirradiation. The postradiation systemic blood pressure of the allopurinol treated monkeys was not statistically different from the untreated, irradiated monkeys and was statistically different from the control monkeys. However, the treated, irradiated monkeys displayed rCBF values that were not significantly different from the nonirradiated controls. These findings suggest the involvement of free radicals in the postirradiation decrease in regional cerebral blood flow but not necessarily in the postirradiation hypotension seen in the primate.

Allopurinol↗
PubMed1987

Identification of free radicals in myocardial ischemia/reperfusion by spin trapping with nitrone DMPO.

The spin trapping ESR technique was applied to investigate oxygen-derived radicals in ischemic and post-ischemic rat hearts. Using 5,5'-dimethyl-l-pyrroline-N-oxide, carbon-centered radicals were identified during ischemia and oxy-radical adducts (superoxide anion radical, O.-2 and hydroxyl radicals, .OH) in post-ischemic rat heart. The formation of these spin adducts was inhibited by superoxide dismutase, suggesting that superoxide plays a role in the adducts' formation. The results demonstrate that oxygen derived free radicals are important byproducts of abnormal oxidative metabolism during myocardial ischemic and reperfusion injuries.

Animals↗
PubMed1987

Spin-trapping evidence that graded myocardial ischemia alters post-ischemic superoxide production.

The spin trapping agent 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) was used to investigate oxy-radical production in post-ischemic rat hearts previously exposed to 20, 30, or 40 minutes of global ischemia. A hydroxyl spin adduct (DMPO-OH) was identified in coronary effluent during the initial seconds of reperfusion by Electron Spin Resonance (ESR) Spectroscopy. The intensity of the ESR signal in post-ischemic effluent increased as ischemic duration was prolonged; however, regardless of the duration of ischemia, maximal spin adduct detection occurred 3 minutes after initiation of reperfusion. Superoxide dismutase inhibited the formation of DMPO-OH, suggesting that superoxide anion was initially generated and is the principle source for the production on the hydroxyl adduct. Our investigations indicate that superoxide anion is produced during the early moments of reperfusion and that its production in the post-ischemic heart is related to the severity of ischemia.

Animals↗
PubMed1987

Spin trapping of oxygen and carbon-centered free radicals in ischemic canine myocardium.

Oxygen free radical injury has been postulated to occur during myocardial ischemia. We have used Electron Spin Resonance and Spin Trapping techniques to directly demonstrate the production of carbon-centered (R.) and oxygen-centered lipid radical (RO.) in ischemic canine heart. In addition, venous effluent from the ischemic region showed that conjugated dienes (lipid peroxidation products) increased with ischemic duration. Our results suggest that the formation of the oxygen-centered and carbon-centered lipid radical species during ischemia are a consequence of oxy-radical peroxidation of myocardial membrane lipids.

Animals↗
PubMed1986

Neutron-induced free radicals in oriented DNA.

Samples of oriented DNA containing 30 per cent water were irradiated with neutrons at 77 K. The electron spin resonance (e.s.r.) spectra obtained from these irradiated DNA samples show that the formation of radicals is different when the incident neutrons are parallel or perpendicular to the DNA helix. When the incident neutrons are perpendicular to the DNA helix the e.s.r. spectra of thymine and guanine ionic radicals (T-., G+.) are observed. An additional e.s.r. spectrum corresponding to the hydrogen addition radical on thymine (TH.) is observed when the incident neutrons are parallel to DNA helix. The TH. radical appears to be formed by protonation of T-. .

DNA↗
PubMedDate not supplied

The role of interleukin-6 (IL-6) in human sulfur mustard (HD) toxicology.

The authors applied in vitro models of controlled damage to human epidermal keratinocytes (HEKs), human skin fibroblasts (HSFs), and human breast skin tissue (HBST) to examine the mechanism responsible for sulfur mustard (HD)-induced interleukin-6 (IL-6) alterations. Treatment with 100 microM HD for 24 hours resulted in a significant increased amount of IL-6 being secreted by HEKs (HD-exposed to control ratio [E/C] = 4.15 +/- 0.07) and by HSFs (E/C = 7.66 +/- 0.04). Furthermore, the HD-induced secretion of IL-6 in HEKs was neutralized with monoclonal human IL-6 antibodies. The secretion of IL-6 in HBST supernatant exposed to HD produced conflicting results. Although an increase of IL-6 was observed in control superfusion media from HBST, IL-6 levels were observed to decrease as the concentration of HD increased. Time course of IL-6 mRNA levels were performed using a competitive polymerase chain reaction (PCR) and human IL-6 mRNA assay detection kit in control and HD (100 microM)-treated HEKs cells. IL-6 mRNA transcripts in HD-exposed HEKs were first observed within 2 hours, dropped at 5 to 6 hours, and increased by approximately 2.2-fold and 8.5-fold at 24 to 48 hours after HD exposure, respectively, as detected by the Xplore mRNA Quantification System. Surface-enhanced laser desorption ionization (SELDI) mass spectrometry was also applied to study the secretion pattern of IL-6 on lysate preparations of HBST. A peak in the area of 23,194 to 23,226 Da was detected using antibody coupled to the chip. This peak was assigned to correspond to the mass of the IL-6 glycoprotein. Recombinant human IL-6 (rhIL-6) exposed to HD lacked the second disulfide bridge and was partially unfolded, as determined by nuclear magnetic resonance-nuclear Overhauser enhancement and exchange spectroscopy (NMR-NOESY). The disappearance of the resonance peak at 3.54 ppm and the appearance of a new chemical shift at 1.85 ppm suggested that a change in structure had occurred in the presence of HD. From the data, the possibility cannot be excluded that IL-6 might be involved in the early event of structural changes of the signal transducer glycoprotein that indirectly initiates the cascade of events such as skin irritation and blister formation observed in the pathophysiology of HD injury.

Adult↗