P31: The effects of red blood cell age on nitric oxide and nitrite-dependent signaling: Implications for transfusion related toxicity

Background RBC are critical hubs for regulated NO-homeostasis, affecting both inhibition (NO and nitrite oxidation) and stimulatory (nitrite-reduction to NO) pathways. How RBC affect this balance, and whether this changes in disease remains unclear. Importantly, biophysical and biochemical properties of the RBC are key parameters, and which change as a function of RBC age in vivo and storage time ex vivo. Transfusion with older RBC is associated with increased morbidity and mortality in critically ill patients with dysfunction in NO-bioavailability underlying, in part, increased inflammation (e.g., in the lung leading to acute lung injury) and circulatory dysfunction. We hypothesized that changes in RBC morphology/ biochemical composition that occur during aging/ storage results in a shift in the balance presented above towards a greater inhibition of NO-signaling both by accelerated rates of NO-and nitrite oxidation. Method Comparisons of NO dioxygenation and nitrite oxidation kinetics were made between fresh and stored RBC as well as RBC of different ages. Functional assessment of changes in NO/nitrite reaction kinetics were evaluated by testing effects of RBC on NO-or nitrite-dependent vasodilation of aortic rings ex vivo and in vivo using a murine model of storage lesion in the context of trauma-hemorrhage and acute lung injury. Results NO-scavenging kinetics by intact oxygenated RBC were ∼40-fold higher with RBC stored for 42 days compared to freshly isolated RBC; an effect that translated to a greater inhibition of NO-dependent vasodilation. Similarly, RBC isolated from freshly drawn blood and separated as a function of endogenous age, showed that older RBC scavenged NO ∼2-fold faster compared to younger RBC, which also translated to a greater degree of inhibition of NO-dependent vasodilation. Both stored and older RBC exhibited increased rates of NO 2 - oxidation, which was not however associated with any deficit in NO 2 - -dependent vasodilation. Transfusion of fresh or stored RBC into mice after trauma-hemorrhage led to significant acute lung injury in the latter. Conclusion Collectively, these data suggest that endogenous NO-signaling will be modulated by the relative distribution of young vs. older RBC. Also the storage of RBC under current blood banking conditions results in cells with higher NO-scavenging activities that are predicted to complement increased hemolysis in contributing to pro-inflammatory stresses associated with transfusion. In addition, we posit that increased oxidation of nitrite, which would decrease an endogenous substrate for NO-formation during hypoxia, also contributes to storage lesion pathology and raises the concept of nitrite-based therapy to replete NO-signaling during trauma-hemorrhage. Disclosure Supported by NIH T32 HL007918.