Kinetic and stoichiometric constraints determine the pathway of H 2 O 2 consumption by red blood cells

• Published in: Free radical biology & medicine 2018-06, Vol.121, p.231
• Main Authors: Orrico, Florencia, Möller, Matías N, Cassina, Adriana, Denicola, Ana, Thomson, Leonor
• Format: Article
• Language: English
• Subjects: Catalase - metabolism; Erythrocytes - metabolism; Humans; Hydrogen Peroxide - metabolism; Kinetics; Models, Theoretical; Oxidants - metabolism; Oxygen - metabolism; Peroxidases - metabolism
• ISSN: 1873-4596
• DOI: 10.1016/j.freeradbiomed.2018.05.006

Red blood cells (RBC) are considered as a circulating sink of H O , but a significant debate remains over the role of the different intraerythocyte peroxidases. Herein we examined the kinetic of decomposition of exogenous H O by human RBC at different cell densities, using fluorescent and oxymetric methods, contrasting the results against a mathematical model. Fluorescent measurements as well as oxygen production experiments showed that catalase was responsible for most of the decomposition of H O at cell densities suitable for both experimental settings (0.1-10 × 10 cell L ), since sodium azide but not N-ethylmaleimide (NEM) inhibited H O consumption. Oxygen production decreased at high cell densities until none was detected above 1.1 × 10 cell L , being recovered after inhibition of the thiol dependent systems by NEM. This result underlined that the consumption of H O by catalase prevail at RBC densities regularly used for research, while the thiol dependent systems predominate when the cell density increases, approaching the normal number in blood (5 × 10 cell L ). The mathematical model successfully reproduced experimental results and at low cell number it showed a time sequence involving Prx as the first line of defense, followed by catalase, with a minor role by Gpx. The turning points were given by the total consumption of reduced Prx in first place and reduced GSH after that. However, Prx alone was able to account for the added H O (50 µM) at physiological RBC density, calling attention to the importance of cell density in defining the pathway of H O consumption and offering an explanation to current apparently conflicting results in the literature.