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Biomedical subjects

C Guarnieri

Publications and source records attributed to C Guarnieri.

At least 91 records · Page 5Linked to original sources
PubMed1989

Involvement of thiol transferase- and thioredoxin-dependent systems in the protection of 'essential' thiol groups of ornithine decarboxylase.

Ornithine decarboxylase (ODC), an enzyme with 'essential' thiol group(s), may be inactivated in vitro by removal of thiol reducing agents and re-activated by soluble factors from rat liver in the presence of NADPH or GSH. The NADPH- and GSH-dependent reducing systems were separated and resolved into three components, called factors A, B1 and B2, by chromatographic techniques. Factor B1 (Mr 12,000) could reactivate ODC in the presence of GSH and co-purified with thiol transferase activity. Factor B2 (Mr 12,000) and factor A (Mr approx. 110,000) were both needed to re-activate ODC in the presence of NADPH, and co-purified with thioredoxin and thioredoxin reductase activity respectively. In an attempt to investigate the physiological role of the 'essential' thiol group(s) of ODC, erythroleukaemia cells were incubated with NN-bis-(2-chloroethyl)-N'-nitrosourea, t-butyl hydroperoxide and vinblastine, which are known to increase the cellular GSSG/GSH ratio, azelaic acid, an inhibitor of thioredoxin reductase, and sodium arsenite, a strong inhibitor of the ODC-re-activating factors. All these compounds were able to decrease significantly the ODC activity induced in these cells. These results suggest that the thiol transferase- and thioredoxin-dependent systems may be physiologically relevant in maintaining ODC in the active, reduced, state.

Animals↗
PubMed1989

Clinical pharmacology of the stereoisomers of leucovorin during repeated oral dosing.

As part of a clinical trial of cisplatin, 5-fluorouracil (5-FU), and leucovorin (LV) for treatment of patients with advanced head and neck cancer, patients received 100 mg of LV (d,l-5-formyltetrahydrofolate) orally every 4 hours for 5 days. On days 2 and 4 of treatment, plasma samples were obtained 2 hours after (peak) and 30 minutes before (trough) a dose of LV. Total LV and 5-methyltetrahydrofolate (THF) concentrations were measured with high-performance liquid chromatography (HPLC) analysis. LV stereoisomer concentrations were determined by chiral HPLC on a bovine serum albumin-bonded silica column. Thus far, plasma folate levels have been analyzed for ten cycles of treatment administered to 7 patients (40 samples). Administration of LV in divided oral doses approximates a plasma steady state with no significant differences noted between peak and trough concentrations. Mean (+/- SD) plasma concentrations for all samples were (mumol): LV, 3.2 +/- 1.3; l-LV, 0.28 +/- 0.21; d-LV, 2.9 +/- 1.2; and THF, 4.25 +/- 2.5. Plasma levels of d-LV and THF tended to be approximately 10% higher on day 4 than day 2, although mean differences were not significantly different due to substantial interpatient variability. Of note was that the sum of THF and l-LV exceeds that of d-LV which was consistent with selective absorption of the l-isomer of LV. Mean ratios of d-LV/l-LV and d-LV/l-LV and THF were 13.7 +/- 10 and 0.88 +/- 0.68, respectively. The authors conclude that oral administration of LV in divided dose (1) simulates a continuous intravenous infusion; (2) produces plasma levels of l-reduced folates in a range known to potentiate 5-FU cytotoxicity; and (3) results in low ratios of d/l-reduced folates that may be important in maximizing the effectiveness of 5-FU-LV chemotherapy.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1989

Effect of sodium arsenite on the induction and turnover of ornithine decarboxylase activity in erythroleukemia cells.

Sodium arsenite proved effective in preventing the induction of ornithine decarboxylase (ODC) activity elicited by dilution of Friend erythroleukemia cells in fresh medium. A 50 per cent inhibition was produced at approximately 1 microM arsenite and complete inhibition was obtained at concentrations above 10 microM. However, addition of arsenite 5 h after cell dilution, i.e. when ODC was already induced, appeared to stabilize the enzyme. The half-life of ODC activity, measured after cycloheximide treatment, increased almost six-fold after addition of sodium arsenite. Agents known to provoke oxidative alteration of the thiol-redox status in cells, also caused a similar effect on the induction and stability of ODC.

Animals↗
PubMed1989

Adaptive control of etoposide administration: impact of interpatient pharmacodynamic variability.

We sought to use a previously derived pharmacodynamic model for 72-hour etoposide infusions to adaptively control administration of this agent and to demonstrate that more predictable toxicity could be obtained with this dosing scheme. A randomized crossover study design was used to compare "standard" dosing (125 mg/m2/day) to adaptive control, with dose adjustment at 28 hours based on the 24-hour plasma level. A total of 31 patients received 86 cycles of chemotherapy, 36 by standard dosing and 50 by adaptive control. However, there was no demonstrable advantage to the adaptive control scheme, because of apparent bias of the previous model. A new model was proposed that also included serum albumin, performance status, and prior RBC transfusions as measures of interpatient pharmacodynamic variability. We conclude that adaptive control dosing of etoposide is feasible but that the therapy must be individualized for both pharmacokinetic and pharmacodynamic variability.

Algorithms↗
PubMed1989

[Polymorphonuclear neutrophilic granulocytes and the defense and damage of periodontal tissues].

The relationship of neutrophil to the periodontal tissue can be equated to the image of a double-edged blade. The role of the polymorphonuclear cells in the containment of gingival bacteria and their products must be weighed against the localized tissue destruction that may be due to the extracellular release of lysosomal enzymes and oxygen derived radicals. It seems reasonable to suggest that the role of neutrophils in the gingival tissue is primarily a defensive one. Therefore, several factors such as the entity of intrinsic activation, the quantity and quality of stimuli, as well as many physiological and/or pathological conditions can determine whether the response of polymorphonuclear leukocytes is more protective or destructive of the parodontium.

Bacteria↗
PubMed1989

Anti oxy-radical properties of trimetazidine.

Trimetazidine at concentrations above 100 microM competed with cytochrome c in scavenging O2.- radicals formed by the reaction catalyzed by the xanthine oxidase enzyme upon xanthine. This scavenger effect was also observed when O2.- were generated by active human neutrophils in which the rate of O2.- formation was monitored by following the reduction of cytochrome c or the emission of luminol-dependent chemiluminescence. An additional scavenger effect of trimetazidine was measured in a OH. chemical generating system whereby the breakdown of deoxyribose by the thiobarbituric acid assay was detected. This study suggests that trimetazidine might function as an antioxy radical compound in conditions of increased oxy radical production.

Deoxyribose↗
PubMed1988

Beneficial effects of trimetazidine on mitochondrial function and superoxide production in the cardiac muscle of monocrotaline-treated rats.

The administration of a single dose of monocrotaline (105 mg/kg) after 21 days produced in rats a reduction of cardiac mitochondrial function at the level of complexes I, II and IV of the respiratory chain, associated with the formation of heart hypertrophy, prevalently of the right ventricle. Moreover, in these rats, the submitochondrial particles produced more O2- and in the cardiac tissue there was an elevation of malondialdehyde content. The repeated administration of trimetazidine (5 mg/kg/24 hr) improved the cardiac mitochondrial function, particularly in state 3 of respiration. In addition, the treatment with trimetazidine reduced, in the heart muscle, both the production of mitochondrial O2- and the content of tissue malondialdehyde. Trimetazidine added alone did not significantly change either the cardiac mitochondrial activity, or the mitochondrial O2- production in comparison to control rats. Also, the content of tissue malondialdehyde was not modified by the repeated administration of trimetazidine. In all the experimental conditions examined, the content of cardiac water-soluble fluorescence substrates did not significantly change in comparison to control rats.

Animals↗
PubMed1988

Rat liver cytosol contains NADPH- and GSH-dependent factors able to restore ornithine decarboxylase inactivated by removal of thiol reducing agents.

Removal of dithiothreitol (DTT) from partially purified ornithine decarboxylase (ODC) led to an almost complete inhibition of enzymic activity. The inactivation was reversed by addition of millimolar concentrations of DTT, whereas natural reductants such as NADPH or NADH were ineffective, and GSH had only a limited effect. Addition of rat liver cytosol to the incubation mixture resulted in a noticeable re-activation of ODC; however, dialysed cytosol had little effect unless NADPH or GSH was present. Fractionation of rat liver cytosol by gel filtration on Sephadex G-75 yielded two fractions involved in the NADPH- and GSH-dependent re-activation of ODC: one designated 'A', eluted near the void volume (Mr greater than or equal to 60,000), and the other designated 'B', eluted later (Mr approx. 12,000). The NADPH-dependent mechanism required both fractions A and B for maximal ODC re-activation; the most effective concentration of NADPH was 0.15 mM, although a significant effect was observed at a concentration more than 10-fold lower. The GSH-dependent mechanism involved the mediation of Fraction B only, and operated at millimolar concentrations of GSH. These results suggest the existence of reducing systems in the cytosol, which may play a role in maintaining, and potentially in regulating, ODC activity by modulation of its thiol status.

Animals↗
PubMed1988

Lipid peroxidation in normoxic and ischaemic-reperfused hearts of fish oil and lard fat fed pigs.

The in situ and in vitro rate of lipid peroxidation of hearts were determined in two groups of pigs which had been fed diets which differed only in fatty acid composition for 8 weeks. During the dietary period venous plasma levels of malondialdehyde and lipofuscin were not higher in pigs receiving the highly unsaturated fatty acid-containing mackerel oil than those receiving lard fat. Malondialdehyde was produced in the coronary system of the mackerel oil fed animals. After the heart was subjected to a sequence of short periods of ischaemia (5 min) and reperfusion (10 min), myocardial malondialdehyde production in the mackerel oil fed pigs did not increase. Contribution of prostaglandin synthesis products to myocardial malondialdehyde formation is probably of minor importance. Recovery of regional heart function after the ischaemic periods was similar for both dietary groups. In the phospholipids of sarcolemmal preparations isolated from the left ventricle of mackerel oil fed animals 18:2 n-6 and 20:4 n-6 were partially replaced by 20:5 n-3 and 22:6 n-3. Ischaemia-reperfusion did not alter sarcolemmal fatty acid composition and Ca2+ pumping ATPase activity. Sarcolemmal membrane from mackerel oil fed pigs exposed in vitro to a free radical generating system showed a higher malondialdehyde production than that from lard fat fed pigs. Thus, in spite of the increased susceptibility of heart membranes to free radical generated peroxidation in mackerel oil fed animals, recovery of left ventricular function was similar following multiple short-term periods of ischaemia.

Animals↗
PubMed1987

External GSSG enhances intracellular glutathione level in isolated cardiac myocytes.

The addition of external GSSG at concentrations in the range 50-500 microM produces in isolated adult rat heart myocytes an increase of GSH level and only a slight increase of GSSG level. On the contrary, external GSH at the above same indicated concentrations did not change the cell glutathione pool. The pretreatment of the cells with diethylamaleate depleted the myocytes of glutathione and enhanced the GSSG-induced replenishment effect on GSH level. On the contrary, the addition of GSH did not increase the concentration of cell glutathione. The level of cell GSH in diethylmaleate-treated myocytes was not increased after 30 min of incubation with cysteine, or acetylcysteine. The GSSG induced-stimulation on GSH level was not inhibited by buthionine sulfoximine, an inhibitor of glutathione synthesis. On the contrary, this stimulatory effect was inhibited by N, N-bis(2-chloroethyl)-N-nitrosourea, an inhibitor of glutathione reductase, or partially, by the remotion of glucose from the incubation medium. These results support the idea that the isolated adult rat heart myocytes are able to utilize external GSSG in order to increase the intracellular glutathione pool, probably through the reduction of the imported GSSG to GSH.

Acetylcysteine↗
PubMed1987

Polyamines stimulate superoxide production in human neutrophils activated by N-fMet-Leu-Phe but not by phorbol myristate acetate.

The polyamines putrescine, spermidine and spermine, at concentrations of 10 microM, stimulated superoxide generation by human polymorphonuclear leukocytes induced by fMet-Leu-Phe in the presence of Ca2+. This positive effect was not evident in the absence of Ca2+ or when the polymorphonuclear leukocytes were stimulated by phorbol myristate acetate. Spermidine in the range of 10-100 microM showed a dose-dependent stimulatory effect on the superoxide generation induced by fMet-Leu-Phe, whilst at doses above 25 mM it produced an inhibitory effect. At this concentration, spermidine did not reduce the phorbol myristate acetate-neutrophil-induced O2-. generation, while an inhibitory effect by the polyamine was evident at concentrations above 50 mM. In addition, 100 microM spermidine increased the amount of superoxide generated and enhanced the ability of the chemotactic peptide to stimulate superoxide generation. The polyamines in the range of 10 microM-25 mM did not modify the activity of purified NADPH oxidase, nor the rate of reduction of cytochrome c as supported by the xanthine/xanthine oxidase reaction. These results indicate that physiological concentrations of polyamines can stimulate superoxide formation by polymorphonuclear leukocyte cells produced by the chemotactic peptide fMet-Leu-Phe, probably by increasing the availability of external calcium.

Calcium↗
PubMed1987

Location of the phosphorylation site for casein kinase-2 within the amino acid sequence of ornithine decarboxylase.

The sizes of the radiolabeled fragments obtained by CNBr and DMSO/HBr digestion of 32P-labeled ornithine decarboxylase phosphorylated by rat liver casein kinase TS (type-2) are consistent with the location of the phosphorylation site within the sequence(303-309) Ser-Asp-Asp-Glu-Asp-Glu-Ser. Parallel experiments with synthetic peptides rule out the suitability of Ser-309, as well as of other serines of ornithine decarboxylase having just two or three acidic residues close to their C terminal side. Ser-303 appears, therefore, to be the main if not the only target for casein kinase-2.

Amino Acid Sequence↗
PubMed1987

Altered thiol group status in the heart ornithine decarboxylase inactivated following perfusion with t-butylhydroperoxide.

1. The perfusion for 15 min of isolated rat hearts with 100 microM t-butylhydroperoxide leads to a 75% diminuition of the tissue GSH/GSSG ratio. 2. After t-butylhydroperoxide infusion, the isoproterenol-stimulated heart ODC was strongly inhibited. The addition of 2 mM DTT in the assaying buffer removed the ODC inactivation. 3. The inhibited ODC had an eluition profile similar to active ODC when chromatographed on a Sephacryl S-200 column; moreover, the ODC activity recovered after a thiol affinity chromatography as unbound fraction, was two times increased in the t-butylhydroperoxide perfused hearts in comparison to control. 4. The hearts perfused with 1 mM acetylcysteine after 15 min of perfusion with t-butylhydroperoxide recovered almost completely the initial ODC activity.

Acetylcysteine↗
PubMed1987

The effect of treatment with coenzyme Q10 on the mitochondrial function and superoxide radical formation in cardiac muscle hypertrophied by mild aortic stenosis.

A 40% reduction of the diameter of the ascending aorta maintained for 60 days induced the formation of a compensate cardiac hypertrophy in rabbits without changing the value of the azide insensitive Ca2+-ATPase activity in comparison to control hearts. The cardiac mitochondria isolated from constricted animals assayed in presence of glutamate and succinate did not show a change in the R.C.I. and ADP/O values in comparison to the controls, whilst the QO2 value enhanced or decreased respectively when determined with glutamate or succinate. The intramuscular injections of CoQ10 (12 mg/kg body weight/48 h) enhanced the mitochondrial CoQ10 concentrations both in the control and in the constricted animals and further increased the QO2 value determined in both groups of animals when glutamate was used as the substrate. The production of O2.- radicals by the level of the complexes I and III of the respiratory chain, did not change in the constricted animals, nor in the animals administered with CoQ10 in comparison to the control. CoQ10 augmented the rate of oxygen consumption by the submitochondrial particles only in the constricted animals. Moreover, the treatment with the coenzyme or the constriction of the aorta, did not modify the cardiac superoxide dismutase activity, but increased the glutathione peroxidase activity only in the banded animals. In addition, in the CoQ10 treated animals there was a reduction of NADH-diaphorase activity both in the control and constricted animals, while the malondialdehyde, generated during the thiobarbituric acid test, and the cardiac content of lipofuscin were decreased.

Animals↗
PubMed1986

Modulation of ornithine decarboxylase activity and ornithine decarboxylase-antizyme complex in rat heart by hormone and putrescine treatment.

Ornithine decarboxylase was present in a cryptic, complexed form in an amount approximately equivalent to that of free ornithine decarboxylase activity in adult rat heart. Addition of isoproterenol (10 mg/kg) caused a notable rise in ornithine decarboxylase activity and a simultaneous decrease in the amount of the complexed enzyme. During the period of ornithine decarboxylase decay, when cardiac putrescine content had reached high values, the level of the complex increased above that of the control. Administration of putrescine (1.5 mmol/kg, twice) or dexamethasone (4 mg/kg) produced a decrease of heart ornithine decarboxylase activity, while it did not remarkably affect the level of complexed ornithine decarboxylase, therefore raising significantly the ratio of bound to total ornithine decarboxylase. Putrescine also elicited the appearance of free antizyme, concomitantly with the disappearance of free ornithine decarboxylase activity after 3-4 h of treatment. These results indicate that a significant amount of ornithine decarboxylase occurs in an inactive form in the heart under physiological conditions and that its absolute and relative levels may vary following stimuli which affect heart ornithine decarboxylase activity.

Animals↗