Impaired antioxidant capacity causes a disruption of metabolic homeostasis in sickle erythrocytes.

This study examined particularly relevant redox pathways such as glycolysis, pentose phosphate pathway (PPP), metHb reductase and nucleotide metabolism, in order to better address how sickle cells deal with redox metabolism disruption. We also investigated the generation of specific oxidative lesions, and the levels of an unexplored antioxidant that could act as a candidate biomarker for oxidative status in sickle cell anemia (SCA). We adopted rigorous exclusion criteria to obtain the studied groups, which were composed by 10 subjects without hemoglobinopathies and 10 SCA patients. We confirmed that sickle cells overwhelm the antioxidant defense system, leading to an impaired antioxidant capacity that significantly contributed to the increase in cholesterol oxidation (ChAld) and hemolysis. Among the antioxidants evaluated, ergothioneine levels decreased in SCA (two-fold). We found strong correlations of ergothioneine levels with other erythrocyte metabolism markers, suggesting its use as an antioxidant therapy alternative for SCA treatment. Moreover, we found higher activities of MetHb reductase, AChE, G6PDH, HXK, and LDH, as well as levels of NADPH, ATP and hypoxanthine in sickle cells. On this basis, we conclude that impaired antioxidant capacity leaves to a loss of glycolysis and PPP shifting mechanism control and further homeostasis rupture, contributing to a decreased lifespan of sickle cells.

Prolonged erythrocyte auto-incubation as an alternative model for oxidant generation system.

This study investigated the effects of incubation period and melatonin treatment on red blood cell (RBC) metabolism in an auto-incubation model of H2O2-induced oxidative stress. The study was carried out on three healthy adult donors by incubating RBCs in their own plasma at 37 °C, or under the influence of 1 mM H2O2 with and without 100 µM melatonin at different times (0, 1, 3 and 6 h). We assessed incubation period, treatment, as well as any interaction effects between these predictors on erythrocyte osmoregulation, hemolytic rate, oxidative stress markers, and adenylate nucleotide levels. We did not find any relevant effects of both incubation period and treatments on osmotic, antioxidant and adenylate parameters. On the other hand, hemolysis degree and biomolecule oxidation levels in the plasma increased over time, 3-fold and about 25%, respectively, regardless any treatment influence. H2O2 treatment more than doubled protein carbonyl groups, regardless time in plasma, and in a time-depending way in erythrocyte membrane extract, effects that were neutralized by melatonin treatment. Through multivariate analyses, we could expand the understanding of energy and redox metabolisms in the maintenance of cellular integrity and metabolic homeostasis. Another interesting observation was the 65-75% contribution of the oxidative lesion markers on hemolysis. Hence, these findings suggested a new and more intuitive RBC suspension model and reinforced the beneficial use of melatonin in human disorders.