OP-17 - Kinetic and stoichiometric constraints determine the pathway of H2O2 consumption by red blood cells

Abstract

Red blood cells (RBC) are considered as a circulating sink of H2O2, but a significant debate remains over the role of the different erythrocyte peroxidases. Herein we examined the kinetic of decomposition of exogenous H2O2 by human RBC at different cell densities, using fluorescent and oxymetric methods, contrasting the results against a mathematical model. Fluorescent measurements of H2O2 as well as oxygen production experiments showed that catalase was responsible for most of the decomposition of H2O2 at low cell number (0.1-1 x 1010 cell L-1), since sodium azide but not N-ethylmaleimide (NEM) inhibited H2O2 consumption. Oxygen production decreased at high cell densities until none was detected at ≥ 1.1 x 1012 cell L-1, being recovered after inhibition of the thiol dependent systems. When comparing total H2O2 consumed by the different pathways, most H2O2 is metabolized by catalase at low cell densities, while Prx predominates at high cell densities. At intermediate cell densities there is a transition region where Gpx accounts for up to 50% of H2O2 being consumed. Our findings underscore the importance of considering the cell number carefully because it will determine the effective concentration of enzymes during analysis performed on complex cell systems.