Enhanced susceptibility to suicidal death of erythrocytes from transgenic mice overexpressing erythropoietin

Abstract

Eryptosis, a suicidal death of mature erythrocytes, is characterized by decrease of cell volume, cell membrane blebbing, and breakdown of cell membrane asymmetry with phosphatidylserine exposure at the cell surface. Triggers of eryptosis include increased cytosolic Ca2+ activity, which could result from activation of Ca2+-permeable cation channels. Ca2+ triggers phosphatidylserine exposure and activates Ca2+-sensitive K+ channels, leading to cellular K+ loss and cell shrinkage. The cation channels and thus eryptosis are stimulated by Cl− removal and inhibited by erythropoietin. The present experiments explored eryptosis in transgenic mice overexpressing erythropoietin (tg6). Erythrocytes were drawn from tg6 mice and their wild-type littermates (WT). Phosphatidylserine exposure was estimated from annexin binding and cell volume from forward scatter in fluorescence-activated cell sorting (FACS) analysis. The percentage of annexin binding was significantly larger and forward scatter significantly smaller in tg6 than in WT erythrocytes. Transgenic erythrocytes were significantly more resistant to osmotic lysis than WT erythrocytes. Cl− removal and exposure to the Ca2+ ionophore ionomycin (1 μM) increased annexin binding and decreased forward scatter, effects larger in tg6 than in WT erythrocytes. The K+ ionophore valinomycin (10 nM) triggered eryptosis in both tg6 and WT erythrocytes and abrogated differences between genotypes. An increase of extracellular K+ concentration to 125 mM blunted the difference between tg6 and WT erythrocytes. Fluo-3 fluorescence reflecting cytosolic Ca2+ activity was larger in tg6 than in WT erythrocytes. In conclusion, circulating erythrocytes from tg6 mice are sensitized to triggers of eryptosis but more resistant to osmotic lysis, properties at least partially due to enhanced Ca2+ entry and increased K+ channel activity.

Erythrocyte programmed cell death

“…Moreover, the mechanisms underlying the enhanced sensitivity of newly formed erythrocytes to suicidal cell death remain elusive. However, it is tempting to associate the phenomenon of neocytolysis with the observation of enhanced eryptosis of erythrocytes drawn from erythropoietinoverexpressing transgenic mice (134). Those erythrocytes were significantly more resistant to osmotic lysis than WT erythrocytes but more sensitive to the eryptotic effects of Cl 2 removal and exposure to the Ca 21 ionophore ionomycin (134).…”

“…However, it is tempting to associate the phenomenon of neocytolysis with the observation of enhanced eryptosis of erythrocytes drawn from erythropoietinoverexpressing transgenic mice (134). Those erythrocytes were significantly more resistant to osmotic lysis than WT erythrocytes but more sensitive to the eryptotic effects of Cl 2 removal and exposure to the Ca 21 ionophore ionomycin (134). In view of those observations the possibility was considered that erythropoietin stimulates the expression of genes in progenitor cells which render the erythrocytes more sensitive to eryptosis and leads to enhanced erythrocyte death as soon as the erythropoietin concentrations decline.…”

“…Along those lines neocytolysis (133), the accelerated death of young erythrocytes following a limited exposure to high altitude or space flight, may reflect the death of those erythrocytes, which have been generated under high erythropoietin concentrations and are thus more vulnerable to eryptosis. Clearly, additional experiments are needed to prove or disprove this speculation (134).…”

Erythropoietin and Oxidative Stress

“…In addition, by assuring EC integrity, EPO prevents ischemic cardiac demise by reducing myocardial injury and cardiomyocyte apoptosis (Burger, et al, 2006), lessening myocardial ischemia (Bullard, et al, 2005), modulating cardiac remodeling (Miki, et al, 2006, Toma, et al, 2007, reducing ventricular dysfunction (Parsa, et al, 2004, Parsa, et al, 2003, and improving cardiac function (Gao, et al, 2007, Westenbrink, et al, 2007. Therefore, EPO plays a critical role in the vascular and renal systems with the maintenance of erythrocyte (Foller, et al, 2007) and podocyte (Eto, et al, 2007) integrity, regulates the survival of ECs , Chong, et al, 2002b, and may act as a powerful endogenous protectant during cardiac injury (Asaumi, et al, 2007). EPO can protect against myocardial cell apoptosis and decrease infarct size, resulting in improved left ventricular contractility.…”

Triple play: Promoting neurovascular longevity with nicotinamide, WNT, and erythropoietin in diabetes mellitus

“…Serum levels of EPO also may function as a biomarker for cardiovascular injury [250]. Work from experimental studies illustrates that EPO plays a critical role in the vascular and renal systems with the maintenance of erythrocyte [251] and podocyte [252] integrity, regulates the survival of ECs [61,110], and may act as a powerful endogenous protectant during cardiac injury [253].…”