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Inhibition of glutathione reductase by isoproterenol oxidation products
F Remião1, H Carmo, F D Carvalho
1ICETA/CEQUP, Toxicology Department, Faculty of Pharmacy, Porto, Portugal. remiao@mail.ff.up.pt
Abstract:
Oxidative stress induced by catecholamines is a well recognized toxic event. This effect has been extensively observed in the heart, where high levels of catecholamines cause enzyme inhibition, lipid peroxidation, energy depletion and myocardial necrosis. Catecholamines can be converted into o-quinones and undergo cyclization into aminochromes. This process can occur enzymatically or through autoxidation and involves the formation of free radicals. Aminochromes are highly reactive molecules that can cause oxidation of protein sulfhydryl groups and deamination catalysis, among other deleterious effects; in addition, inhibition of some enzymes has been also reported. We have studied the effects of isoproterenol oxidation products (IOP) on glutathione reductase (GR) activity in vitro. Isoproterenol (ISO) autoxidation was conducted at 37 degrees C in the dark, for 4 h at pH 7.0 and this process was monitored by UV spectrophotometry at both 340 and 490nm. Addition of the autoxidized solution to GR in the presence of oxidized glutathione (GSSG) and NADPH showed that IOP inhibits GR in a competitive mode and that this effect increases during the 4 h incubation period. This inhibitory effect of IOP was partially prevented by the addition of reduced glutathione (GSH), L-cysteine and ascorbic acid to the reaction mixtures.
Insights
Oxidative stress from catecholamines can harm the heart. Isoproterenol oxidation products (IOP) were found to competitively inhibit glutathione reductase (GR) activity in vitro.
Area of Science:
- Biochemistry
- Cardiovascular Toxicology
Background:
- Catecholamines induce oxidative stress, leading to cellular damage, particularly in the heart.
- This oxidative stress involves the formation of reactive intermediates like o-quinones and aminochromes.
- Aminochromes can cause significant cellular damage through protein modification and enzyme inhibition.
Purpose of the Study:
- To investigate the in vitro effects of isoproterenol oxidation products (IOP) on glutathione reductase (GR) activity.
- To determine the mechanism and kinetics of IOP-induced GR inhibition.
Main Methods:
- Isoproterenol (ISO) was autoxidized at 37°C, pH 7.0 for 4 hours.
- UV spectrophotometry was used to monitor the autoxidation process.
- The inhibitory effect of IOP on GR activity was assessed in the presence of GSSG and NADPH, with varying concentrations of protective agents.
Main Results:
- Isoproterenol oxidation products (IOP) were found to inhibit glutathione reductase (GR) activity in a competitive manner.
- The inhibitory effect of IOP on GR increased with the duration of isoproterenol autoxidation.
- Reduced glutathione (GSH), L-cysteine, and ascorbic acid partially protected GR from IOP-induced inhibition.
Conclusions:
- Oxidative stress from catecholamines, exemplified by isoproterenol, can significantly impair crucial enzyme function like glutathione reductase.
- The findings highlight a potential mechanism for catecholamine-induced cellular dysfunction and provide insights into protective strategies.