Isoflurane does not mimic ischaemic preconditioning in decreasing hydroxyl radical production in the rabbit.

BACKGROUND: Reactive oxygen species are an important mediator in isoflurane-induced myocardial preconditioning. However, hydroxyl radicals are also released during reperfusion after regional ischaemia. The purpose of the present study was to test whether ischaemic preconditioning and isoflurane would influence the production of hydroxyl radicals during reperfusion. METHODS: After i.v. administration of salicylate 100 mg kg(-1) and a 30 min stabilization period, New Zealand White rabbits were subjected to 40 min of regional myocardial ischaemia and 2 h of reperfusion. Ischaemic preconditioning was elicited by 5 min ischaemia followed by 10 min reperfusion (before the 40 min ischaemia). In another group, isoflurane (2.1%) was administered for 30 min, followed by 15 min washout, before the long ischaemia. Area at risk and infarct size were assessed by blue dye injection and tetrazolium chloride staining. We quantified the level of OH-mediated conversion of salicylate to its dihydrobenzoate derivatives (2,3- and 2,5-DHBAs). Normalized values of the DHBAs (ng DHBA per mg salicylate) were calculated. RESULTS: Mean (se) infarct size was 57 (6)% of the risk area in the untreated controls. This was significantly smaller in the ischaemic preconditioning and isoflurane groups: 22 (5) and 23 (6)% respectively. At 10 min of reperfusion, ischaemic preconditioning limited the mean increase in 2,3-DHBA to 24% from baseline, compared with 81% in control and 74% in the isoflurane group. Normalized 2,5-DHBA was maximally increased by 75% in the untreated group, 4 min after reperfusion. Ischaemic preconditioning significantly inhibited this increase (24% increase from baseline, P<0.01). However, the increase observed in the isoflurane group was not different from control (71%). CONCLUSIONS: As already known, ischaemic preconditioning and isoflurane markedly reduced infarct size. However, only ischaemic preconditioning decreased postischaemic production of hydroxyl radicals. These different effects suggest different protective mechanisms at the cellular level.

Ischaemic preconditioning but not isoflurane prevents post-ischaemic production of hydroxyl radicals in a canine model of ischaemia-reperfusion.

BACKGROUND AND OBJECTIVE: Isoflurane has been shown to mimic ischaemic preconditioning (IPC). The protective effect of IPC, or applying isoflurane or perfusion with the 'push-pull' complex zinc-desferrioxamine (Zn-DFO) in the canine heart, was investigated. METHODS: Thirty minutes after salicylate administration (100 mg kg(-1)) the heart was exposed. All dogs were subjected to a 10 min left anterior descending artery occlusion followed by 2 h of reperfusion. In Group I (n = 9) isoflurane (2.5%) was administered 10 min prior to and during ischaemia. In Group II (n = 8), IPC was elicited by 5 min coronary artery occlusion, followed by 5 min of reperfusion, prior to the 10 min ischaemia. In Group III (n = 9) Zn-DFO (2.5 mg kg(-1)) was given 10 min prior to ischaemia. The effects of these interventions were compared to control (n = 10). Coronary sinus blood concentrations of salicylate, 2,3-dihydroxybenzoic acid (DHBA), lactate, pH and oxygen content were monitored. RESULTS: In the control group, 2,3-DHBA increased by 32% above the pre-ischaemic value (P < 0.05). In contrast, in the IPC hearts, a significant decrease in the production of 2,3-DHBA was observed (40% lower than baseline, P < 0.01). In the isoflurane group only a 13% (and non-significant) decrease was noticed. In the Zn-DFO group a 33% decrease was found (P < 0.01). The increase in lactate concentrations in the IPC and Zn-DFO groups was significantly smaller than that of control and isoflurane groups. CONCLUSIONS: IPC protected the heart against the deleterious effects of reperfusion, possibly by amelioration of the level of oxygen-derived reactive species, and the complete inhibition of reactive hydroxyl radical production. Isoflurane did not prove to be as effective in reducing the free radical damage.

Synergistic cytotoxicity between pentachlorophenol and copper in a bacterial model.

Both pentachlorophenol (PCP) and copper compounds have been widely used as wood preservatives, and are commonly found not only in the area near wood-preserving facilities, but also in body fluids and tissues of people who are not occupationally exposed to them. In this study, we found that exposing bacteria to a combination of PCP and copper at non- or sub-toxic concentrations resulted in enhanced cytotoxic effect in a synergistic mode as indicated by both the inhibition of growth and the lowering of the colony-forming ability. The toxicity of the combination PCP/Cu(II) was relieved by hydrophilic chelating agents, thiol compounds and adventitious proteins, but was markedly potentiated by low levels of the lipophilic metal chelating agents.

The push-and-pull mechanism to scavenge redox-active transition metals: a novel concept in myocardial protection.

OBJECTIVE: Traces of redox-active transition metals such as iron and copper play an important role in free radical formation during postischemic reperfusion of the heart. Two studies were conducted to assess the efficacy of the complexes of desferrioxamine with zinc or gallium to prevent this aspect of reperfusion injury. METHODS: In study I, isolated working rat hearts (n = 96) were subjected to 2 hours of hypothermic arrest at 10 degrees C induced by use of St Thomas' Hospital cardioplegic solution II supplemented with desferrioxamine, zinc-histidinate, zinc-desferrioxamine, gallium-nitrate, or gallium-desferrioxamine. In study II, isolated nonworking rat hearts (n = 23) were subjected to normothermic regional (10 minutes) or global (35 minutes) unprotected ischemia. In this study, the perfusate was supplemented with gallium-desferrioxamine during preischemic and postischemic periods. RESULTS: In study I, the addition of desferrioxamine, zinc-histidinate, or gallium-nitrate to St Thomas' Hospital solution II improved postischemic aortic flow recovery. When the binary complexes zinc-desferrioxamine or gallium-desferrioxamine were added, however, functional recovery was further enhanced significantly. In study II, high-performance liquid chromatography analyses of tissue from postischemic hearts exposed to unsupplemented perfusate revealed a marked increase of malondialdehydes. In hearts perfused with perfusate supplemented with gallium-desferrioxamine, however, tissue malondialdehyde concentrations were significantly smaller, indicating reduced free radical formation. CONCLUSIONS: The data suggest synergistic protection by the complexes of the iron chelator desferrioxamine with zinc or gallium. The single components neutralize transition metals by 2 different but complementary push-and-pull mechanisms, thereby leading to an inhibition of metal-mediated site-specific free radical formation and improvement of postischemic cardiac function.

Redox-active iron mediates amyloid-beta toxicity.

While amyloid-beta toxicity is mediated by oxidative stress and can be attenuated by antioxidants, the actual biochemical mechanism underlying neurotoxicity remains to be established. However, since aggregated amyloid-beta can interact with transition metals, such as iron, both in vitro and in vivo, we suspected that bound iron might be the mediator of toxicity such that holo- and apo-amyloid would have differential effects on cellular viability. Here we demonstrate that when amyloid-beta is pretreated with the iron chelator deferoxamine, neuronal toxicity is significantly attenuated while conversely, incubation of holo-amyloid-beta with excess free iron restores toxicity to original levels. These data, taken together with the known sequelae of amyloid-beta, suggest that the toxicity of amyloid-beta is mediated, at least in part, via redox-active iron that precipitates lipid peroxidation and cellular oxidative stress.

Antioxidant activity of Ferrozine-iron-amino acid complexes.

Amino acid-Fe(II)-chelator complexes exhibit strong antioxidant activity. Taking advantage of the unique spectral characteristics of the complexes formed when Ferrozine (Fz) is used as the chelator, we now show that the primary blue complex (epsilon(max) at 632 nm) decomposes by two independent pathways: (i) a nonoxidative pathway involving dissociation of the amino acid component and formation of a purple complex (epsilon(max) at 562 nm) and (ii) an oxidative pathway leading to Fe(III) and colorless products. Quantitative conversion of the blue to purple complex yields an isosbestic point (i.p.) at 601 nm, whereas no i.p. is formed during quantitative oxidation of the blue complex. However, under some experimental conditions, decomposition of the blue product occurs by both pathways, leading to occurrence of a clean i.p. at wavelengths varying from 601 to 574 nm. Results of simulation experiments, confirmed by direct analysis, demonstrate that shifts in the i.p. reflect differences in the fractions of blue compound that decompose by the oxidative and nonoxidative pathways. Indeed, the fraction of blue that is converted to the purple complex is readily deduced from the wavelength of the i.p. These results suggest that identification of a physiological chelator that can replace Ferrozine in amino acid-iron complexes might have important physiological and pharmacological applications.

Copper-mediated toxicity of 2,4,5-trichlorophenol: biphasic effect of the copper(I)-specific chelator neocuproine.

The lipophilic copper(I)-specific chelator neocuproine has been frequently used as an inhibitor of copper-mediated damage in biological systems. In this communication we report that the copper-mediated toxicity of 2,4,5-trichlorophenol is markedly potentiated by neocuproine at levels which are near-stoichiometric with respect to the copper concentration but is inhibited at higher concentrations. However, no potentiation was observed when neocuproine was substituted by bathocuproinedisulfonic acid, a negative charged ligand with essentially the same copper-binding characteristics as neocuproine. We found that the potentiation by neocuproine was due to the formation of a lipophilic copper complex, while the inhibition by bathocuproinedisulfonic acid was due to the formation of a hydrophilic one. Caution in the use of neocuproine to study copper-mediated toxicity is advised.

Antioxidant status and human health. Use of cyclic voltammetry for the evaluation of the antioxidant capacity of plasma and of edible plants.

The low molecular weight antioxidants (LMWA) play a major role in protecting biological systems against reactive oxygen-derived species (ROS), and reflect the antioxidant capacity of the system. The cyclic voltammetry (CV) has been conveniently used and validated for the quantitation of the antioxidant capacity of the LMWA of blood plasma, tissue homogenates, and plant extracts. The CV tracing provides the biological oxidation potential (E and E1/2 which relate to the nature of the molecule(s)), the intensity of the anodic current wave (Ia), and its area S (both relate to the concentration of the molecule(s)). The components of the first anodic wave of plasma were identified by comparison with HPLC-electrochemical detection. CV together with another plasma parameter R, which reflects the level of oxidized ascorbate, were used for the evaluation of the antioxidant status and the oxidative stress in healthy subjects and in chronic (diabetes mellitus) and acute patients (subjected to total body irradiation prior to bone marrow transplantation). These methodologies could be widely employed for rapid evaluation of subjects, in health and disease, for monitoring of their response to treatment and nutritional supplementation, and for screening of specific populations.

The use of cyclic voltammetry for the evaluation of antioxidant capacity.

Low-molecular weight antioxidants (LMWAs) play a major role in protecting biological systems against reactive oxygen-derived species and reflect the antioxidant capacity of the system. Cyclic voltammetry (CV), shown to be convenient methodology, has been validated for quantitation of the LMWA capacity of blood plasma, tissue homogenates, and plant extracts. Analysis of the CV tracing yields the values of (i) the biological oxidation potential, E and E(1/2), which relate to the nature of the specific molecule(s); (ii) the intensity (Ia) of the anodic current; and (iii) the area of the anodic wave (S). Both Ia and S relate to the concentration of the molecule(s). LMWA components of human plasma and animal tissues were identified and further validated by reconstruction of the CV tracing and by high-performance liquid chromatography-electrochemical detection. To reflect the oxidative stress status, the use of an additional parameter, R, has been proposed. R represents the level (%) of oxidized ascorbate (compared with total ascorbate) and is measured by high-performance liquid chromatography-electrochemical detection. All these parameters were monitored in healthy human subjects as well as in chronic (diabetes mellitus) and acute care patients (subjected to total body irradiation before bone marrow transplantation). The electroanalytical methodologies presented here could be widely employed for rapid evaluation of the status of subjects (in health and disease) for monitoring of their response to treatment and/or nutritional supplementation as well as for screening of specific populations.

Protection by desferrioxamine and other hydroxamic acids against tetrachlorohydroquinone-induced cyto- and genotoxicity in human fibroblasts.

Tetrachlorohydroquinone (TCHQ) has been identified as a major toxic metabolite of the widely used wood preservative pentachlorophenol and has also been implicated in its genotoxicity. We have recently demonstrated that protection by the trihydroxamate iron chelator desferrioxamine (DFO) on TCHQ-induced single-strand breaks in isolated DNA was not the result of its chelation of iron but rather of its efficient scavenging of the reactive tetrachlorosemiquinone (TCSQ) radical. In this study, we extended our research from isolated DNA to human fibroblasts. We found that DFO provided marked protection against both the cyto- and genotoxicity induced by TCHQ in human fibroblasts when it was incubated simultaneously with TCHQ. Pretreatment of the cells with DFO followed by washing also provided marked protection, although less efficiently compared with the simultaneous treatment. Similar patterns of protection were also observed for three other hydroxamic acids (HAs): aceto-, benzo-, and salicylhydroxamic acid. Dimethyl sulfoxide, an efficient hydroxyl radical scavenger, provided only partial protection even at high concentrations. In vitro studies showed that the HAs tested effectively scavenged the reactive TCSQ radical and enhanced the formation of the less reactive and less toxic 2,5-dichloro-3, 6-dihydroxy-1,4-benzoquinone (chloranilic acid). The results of this study demonstrated that the protection provided by DFO and other HAs against TCHQ-induced cyto- and genotoxicity in human fibroblasts is mainly through scavenging of the observed reactive TCSQ radical and not through prevention of the Fenton reaction by the binding of iron in a redox-inactive form.

Evidence for production of hydroxyl radicals by pentachlorophenol metabolites and hydrogen peroxide: A metal-independent organic Fenton reaction.

The production of hydroxyl radicals by tetrachlorohydroquinone (TCHQ, a major metabolite of the widely used biocide pentachlorophenol) in the presence of H(2)O(2) was studied by salicylate hydroxylation method. HPLC with electrochemical detection was used to measure the levels of 2,3- and 2,5-dihydroxybenzoic acid (DHBA) formed when the hydroxyl radicals react with salicylate. We found that TCHQ and H(2)O(2) could produce both 2,3- and 2,5-DHBA when incubated with salicylate. Their production was markedly inhibited by hydroxyl radical scavenging agents dimethyl sulfoxide and ethanol, as well as by tetrachlorosemiquinone radical scavengers desferrioxamine and other hydroxamic acids. In contrast, their production was not affected by the nonhydroxamate iron chelators diethylenetriaminepentaacetic acid (DTPA), bathophenanthroline disulfonic acid, and phytic acid, as well as the copper-specific chelator bathocuprione disulfonic acid. A comparison of product formation and distribution from the reaction of ferrous iron with hydrogen peroxide (the classic Fenton system) strongly suggests that the same hydroxyl radical adducts are formed as in the TCHQ/H(2)O(2) experiments. Taken together, we propose that hydroxyl radicals were produced by TCHQ in the presence of H(2)O(2), probably through a metal-independent organic Fenton reaction.

Gallium-desferrioxamine protects the cat retina against injury after ischemia and reperfusion.

This study sought to determine whether gallium-desferrioxamine (Ga/DFO) can curb free radical formation and mitigate biochemical and electrophysiological parameters of injury in the cat retina subjected to ischemia followed by reperfusion. For the biochemical studies, cat eyes were subjected to 90 min of retinal ischemia followed by 5 min of reperfusion, and enucleation of one eye of each cat was used to measure retinal reperfusion injury. Before enucleation of fellow eyes, 2.5 mg/kg Ga/DFO was injected intravenously 5 min before reperfusion. The flux of hydroxyl radicals, as measured directly by conversion of salicylate to 2,3- and 2,5-dihydroxybenzoic acid (2,3- and 2,5-DHBA), was significantly lower in Ga/DFO-treated eyes. The mean normalized level of 2,3-DHBA (considered a specific marker of hydroxyl radicals) was 3.5 times higher in untreated eyes. Ga/DFO caused a significant reduction, by 2.56-fold, in lipid peroxidation, as reflected by levels of malondialdehyde. Ascorbic acid, a natural antioxidant present in the retina, is severely depleted in untreated eyes. In contrast, in Ga/DFO-treated eyes, levels were 10 times higher than the control. Energy charge was 2.38 times higher in treated eyes. Levels of purine catabolites (hypoxanthine, xanthine, and uric acid) that reflect excessive metabolism of purine nucleotides were approximately twice higher in untreated retinas. Electroretionographic studies, performed on a different subset of animals, substantiated the biochemical results. In Ga/DFO-treated eyes the amplitude of the mixed cone-rod response b-wave (as compared with fellow nonischemic eyes) fully recovered within 24 h after ischemia (b-wave ratio 1.04 +/- 0.09, [mean +/- SEM]) whereas ischemic/reperfused and nontreated eyes recovered to only 0.33 +/- 0. 05. The results show that severe biochemical and functional retinal injury occurs in cat eyes subjected to ischemia and reperfusion. These severe changes were significantly reduced by a single administration of Ga/DFO just before reperfusion. We hypothesize that the protection afforded by Ga/DFO is due to a combined effect of "Push-Pull" mechanisms interfering with transition metal-dependent and free radical-mediated injurious processes.

Mechanism of the synergistic cytotoxicity between pentachlorophenol and copper-1,10-phenanthroline complex: the formation of a lipophilic ternary complex.

When non- or sub-toxic levels of pentachlorophenol (PCP) and bis-(1, 10-phenanthroline)cupric complex, Cu(II)(OP)(2), were combined, a remarkable synergistic toxicity was observed as indicated by growth inhibition and bacterial inactivation. Similar synergistic cytotoxic effects were observed with other polychlorinated phenols and other positively charged cupric complexes. The synergism observed for these chemicals and similar reactive pairs of chemicals was found to be due to the formation of lipophilic ternary complexes which facilitated copper transport into the bacterial cells. The formation of ternary complexes of similar lipophilic character could be of relevance as a general mechanism of toxicity.

Human studies related to protein oxidation: protein carbonyl content as a marker of damage.

Proteins constitute the major 'working force' for all forms of biological work. Their exact conformation and pattern of folding are tightly connected to their activity and function. Reactive oxygen and nitrogen species (ROS and RNS) are formed during normal metabolism and in higher fluxes under pathological conditions. They cause cellular damage, an important part of which is the oxidation of amino acid residues on proteins, forming protein carbonyls. Other direct modifications of protein side chains, such as o-tyrosine, chloro-, nitrotyrosine, and dityrosine, have been identified. In addition, carbohydrate and lipid derivatives can react with proteins to form adducts that can be analyzed. Protein carbonyl content (PCC) is the most widely used marker of oxidative modification of proteins. There are several methodologies for the quantitation of PCC; in all of them 2,4-dinitrophenyl hydrazine is allowed to react with the protein carbonyls to form the corresponding hydrazone, which can be analyzed optically by radioactive counting or immunohistochemically. Using PCC as a marker, it could be demonstrated that oxidative damage to proteins correlates well with aging and the severity of some diseases. A critical evaluation of PCC and other markers of protein oxidation is presented, together with examples of protein oxidation in diabetes, neurodegenerative diseases, and aging.

The use of Zn-desferrioxamine for radioprotection in mice, tissue culture, and isolated DNA.

Redox-active metals mediate oxidative injury and might also potentiate radiation damage. The iron chelator desferrioxamine (DFO), which diminishes oxidative damage in many chemical and biological systems as well as in human subjects, has a controversial role in radiobiology and reportedly acts both as a radiosensitizer and a radioprotector. The present research focused on the radioprotective activity of its zinc complex. Zn-DFO was studied using three test systems differing by their complexities: isolated DNA from pUC 19 plasmid, cultured V79 Chinese hamster cells, and C3H mice. Zn-DFO (0.5-2 mM) protected isolated DNA against gamma-radiation better than each of its components alone; however, neither Zn-DFO nor DFO (50-100 microM) alone affected the radiation sensitivity of cultured cells. With total body irradiation, Zn-DFO, but not DFO alone at 100 micromol/kg body weight, administered to mice 30 min before irradiation provided significant radioprotection (P < 0.01). Zn-DFO had an LD(50/30) of 10.3 Gy, whereas DFO and vehicle alone had LD(50/30) of 8.03 Gy and 7.91 Gy, respectively. The effect of Zn-DFO on the hemodynamic parameters in mice did not differ from that of the vehicle (saline) alone. This excludes the explanation that the radioprotective activity of Zn-DFO results from its effect on oxygen levels. In addition to the possible direct effect of Zn, other potential modes of action underlying the radioprotective activity of Zn-DFO might involve a displacement of iron and its substitution by zinc, a greater proximity of the drug to DNA, and less likely an improved penetration of the drug into cells because of its structure. The failure of Zn-DFO to protect cells in tissue cultures indicates that it has some systemic role in the whole animal, possibly due to a prolonged half-life in the animal's circulation.

Halothane modifies oxygen free radical activity on the voltage-sensitive calcium channels in canine myocardial membranes.

BACKGROUND: The functional derangements in the myocyte cell membrane, the sarcolemma, during short myocardial ischaemia and reperfusion are attributed to excessive influx of Ca2+ ions via the voltage-sensitive calcium channels (VSCC) and to the free radical-related injury. However, it is unclear whether the primary changes in the VSCC should be attributed to the ischaemic effect or to free radical action on channel constituents. Under these circumstances of ischaemia and reperfusion, volatile anaesthetics have exhibited protective properties on the myocardium. The present study is aimed at characterizing the effect of artificially-generated oxygen free radicals on the VSCC in canine sarcolemma, independently of the effect of ischaemia, and the effect of halothane on the membranes during the surge of the free radicals. METHODS: Selective production of free radicals (O2-, CO2-) was made by gamma irradiation of isolated sarcolemma membranes with 137 Cesium (Cs), in the presence of 20 mM sodium formate. Control studies were performed without formate in the aqueous solution. In an additional group, liquid halothane (3 microl. 1.9 vol%) was added to the sarcolemma / formate preparation immediately prior to irradiation. The effects of free radicals on the VSCC was evaluated by redioligand binding studies of the calcium channel blocker [3H] isradipine to the sarcolemma. RESULTS: In six control studies, the rediolytic aqueous species produced by 137 Cs irradiation resulted in unchanged [3 H] isradipine binding. In the presence of formate [n=9], the free radicals have caused a 23% to 25% decrease, both, in density and dissociation constant (P=0.05) of [3 H]isradipine to the VSCC binding sites. When superoxide radicals were generated in the presence of 1.9% halothane and formate (n=6), a significant increase in maximal binding capacity (by 55% +/- 2; P<0.01) and in the dissociation constant (by 209% +/- 35, P<0.01) occurred. CONCLUSION: Oxidative free radicals which are generated by gamma irradiation exerted minimal changes on the normal function of the VSCC as reflected by the non-significant changes in [3 H] isradipine specific binding. Introduction of halothane into free radical generating system causes acute perturbations to the VSCC kinetics, and does not provide protection to the cardiac membranes.

Antioxidant capacity of edible plants: extraction protocol and direct evaluation by cyclic voltammetry.

Reactive oxygen-derived species are produced in cells under physiological conditions and in response to stress. Among the various antioxidant systems responsible for protection against these species, the low-molecular-weight antioxidants (LMWA), such as ascorbate, play an important role. Cyclic voltammetry (CV) has been proposed as a tool for quantitation of the total antioxidant capacity of plasma. It has also been shown that biological oxidation potentials, as determined from the anodic current waves of the CV tracings, are specific characteristics of the various LMWA components, and that the amplitude of each wave can be used for quantitation of the specific component. The adaptation of CV for evaluation of the total antioxidant capacity of edible plants is demonstrated here. The area under the anodic current wave is proposed as a better indicator for the content of LMWA, compared with the amplitude. This distinction could prove valuable when more than a single molecule contributes toward a specific anodic wave and when the identities of the components of a wave are not known. Vegetables and fruits that are commonly consumed in the U.S. diet were used. They were extracted with either water, aqueous acetic acid (30%), or a mixture of water, acetic acid, and acetonitrile (40:30:30). The LMWA contents were evaluated by CV. In three to five steps the LMWAs were completely extracted from the edible foods, and their amounts were translated into equivalents of ascorbate.

Novel protection strategy against X-ray-induced damage to salivary glands.

Exposure of the major salivary glands to ionizing radiation often results in severe alterations in structure and function. The mechanism of these effects is still unknown, and no adequate prevention or treatment is yet available. The purpose of this study was to examine a mechanism based on the assumption that redox-active metal ions, which propagate the production of highly reactive free radicals, are responsible for the unique radiosensitivity of salivary glands. Zinc-desferrioxamine (Zn-DFO) was recently reported to be a very potent protector against the injuries induced by such metal ions in the vicinity of sensitive cellular targets. We chose to examine its protective potential against the damage to salivary glands induced by X rays. Head and neck irradiation (15 Gy) was delivered to rats 90 min after the intraperitoneal administration of 20 mg/kg Zn-DFO. This group was compared to two control groups, irradiated and nonirradiated. At 2 months after irradiation, both systemic and salivary parameters were analyzed. The results demonstrated that X irradiation induced a profound attenuation of body weight (30%) and a reduction of parotid gland saliva flow rate (74%), parotid gland weight (36%), submandibular gland/sublingual gland saliva flow rate (46%), and submandibular/sublingual gland weight (24%) (P < 0.01 for all parameters). The content of potassium in parotid gland saliva was increased by 46% (P < 0.01), while the protein content was unaltered. The increase in the potassium concentration of the saliva is considered to be another indication of salivary gland hypofunction. Administration of Zn-DFO prior to irradiation resulted in partial protection against radiation-induced injury to the parotid gland but not the submandibular gland. In the Zn-DFO-treated and irradiated group, the parotid gland saliva flow rate was reduced by 42%, the weight of the parotid gland was reduced by 13%, and the potassium concentration in the parotid gland saliva was increased by 21% (P < 0.05 for all parameters). These results give credence to the validity of the hypothesis which correlates radiation-induced damage of the salivary glands with the injurious role of intracellular redox-active metal ions. Furthermore, the results offer prospects in the clinical setting, as Zn-DFO is a modification of DFO, which is a clinically approved and widely used medication. Further examination of the clinical use of Zn-DFO is currently under way, focusing on its beneficial protective effect on healthy non-neoplastic tissue.

Topical use of zinc desferrioxamine for corneal alkali injury in a rabbit model.

PURPOSE: To evaluate the efficacy of topical zinc desferrioxamine in acute corneal alkali injury in rabbits. METHODS: Twenty rabbits were anesthetized and a standardized alkali burn (1N NaOH) was performed in the center of the cornea (7.5-mm diameter). The animals were randomly divided into two groups and treated (double-masked) with topical zinc desferrioxamine, 220 microM, (group 1) or its vehicle (group 2). Drops were applied 7 times/day for 28 days. Topical gentamicin, 0.3%, was instilled twice a day. Animals were evaluated twice a week. At each examination (using the slit-lamp), the depth of corneal ulcer was graded as follows: 0, no ulcer; 1, tissue loss less than one third of corneal thickness; 2, one third to two thirds tissue loss; 3, more than two thirds tissue loss; 4, descemetocele; or 5, perforation. Ulceration area, vascularization, and epithelial defects also were measured. RESULTS: During the study period, the grading of mean corneal ulcerations in group 1 ranged from 0.2 to 1.00, whereas in group 2, it ranged from 1.4 to 2.7. The mean grade and area of ulceration in group 2 were greater than those in group 1 (p < 0.05). CONCLUSION: Topical zinc desferrioxamine may be an adjunctive treatment in protecting the cornea against induced alkali injury.

New modes of action of desferrioxamine: scavenging of semiquinone radical and stimulation of hydrolysis of tetrachlorohydroquinone.

Desferrioxamine (DFO) is a common drug used in the treatment of iron overload. In addition to its iron-chelation, other properties have been identified. Alas, DFO has demonstrable effects which cannot be explained by its classically established properties; i.e., DFO protects against DNA single strand breaks induced by tetrachlorohydroquinone (TCHQ), while other iron chelators such as DTPA (diethylenetriaminepentaacetic acid) do not. The autooxidation process of TCHQ yielding the tetrachlorosemiquinone radical (TCSQ.) intermediate, was studied here in the presence of chelators. DFO led to a marked reduction in both concentration and life span of TCSQ. via formation of DFO-nitroxide radical (DFO.). In contrast, DTPA had no detectable effect on TCHQ autooxidation. Present studies indicate that the protective effects of DFO on TCHQ-induced DNA damage were not due to the binding of iron, but rather to scavenging of the reactive TCSQ. and the formation of the less reactive DFO.. An additional mode of action of DFO was identified, via stimulation of the hydrolysis (dechlorination) of tetrachloro-1,4-benzoquinone (chloranil), which is the oxidation product of TCHQ, to form 2,5-dichloro-3,6-dihydroxy-1,4-benzoquinone (chloranilic acid). The results of this study demonstrate two new modes of action for DFO: the scavenging of deleterious semiquinone radical, and the stimulation of the hydrolysis of halogenated substituents on the quinone structure. Both modes might prove highly relevant to the biological activities of DFO.