Two different pathways are involved in peroxynitrite-induced senescence and apoptosis of human erythrocytes.
CO(2) changes the biochemistry of peroxynitrite basically in two ways: (i) nitrating species is the CO(3)(-) / ()NO(2) radical pair, and (ii) peroxynitrite diffusion distance is significantly reduced. For peroxynitrite generated extracellularly this last effect is particularly dramatic at low cell density because CO(3)(-) and ()NO(2) are short-lived and decay mostly in the extracellular space or at the cell surface/membrane. This study was aimed to distinguish between peroxynitrite-induced extra- and intracellular modifications of red blood cells (RBC). Our results show that at low cell density and in the presence of CO(2) peroxynitrite induced the oxidation of surface thiols, the formation of 3-nitrotyrosine and DMPO-RBC adducts, and the down-regulation of glycophorins A and C (biomarkers of senescence). Reactivation of glycolysis reversed only the oxidation of surface thiols. Without CO(2) peroxynitrite also induced the oxidation of hemoglobin and glutathione, the accumulation of lactate, a decrease in ATP, the clustering of band 3, the externalization of phosphatidylserine, and the activation of caspases 8 and 3 (biomarkers of apoptosis). The latter biomarkers were all reversed by reactivation of glycolysis. We hypothesize that cell senescence could (generally) be derived by irreversible radical-mediated oxidation of membrane targets, while the appearance of apoptotic biomarkers could be bolstered by oxidation of intracellular targets. These results suggest that, depending on extracellular homolysis or diffusion to the intracellular space, peroxynitrite prompts RBCs toward either senescence or apoptosis through different oxidation mechanisms.
Pietraforte
Donatella
D
Departments of Cell Biology and Neurosciences, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161, Roma, Italy.
Matarrese
Paola
P
Straface
Elisabetta
E
Gambardella
Lucrezia
L
Metere
Alessio
A
Scorza
Giuseppe
G
Leto
Thomas L
TL
Malorni
Walter
W
Minetti
Maurizio
M
D017209:Apoptosis / Q000187:drug effects*; D002245:Carbon Dioxide / Q000378:metabolism; D016922:Cellular Senescence / Q000187:drug effects*; D004912:Erythrocytes / Q000187:drug effects*; D005434:Flow Cytometry; D006801:Humans; D010084:Oxidation-Reduction / Q000187:drug effects; D030421:Peroxynitrous Acid / Q000378:metabolism*
D002245:Carbon Dioxide; D030421:Peroxynitrous Acid