CO₂‐dependent metabolic modulation in red blood cells stored under anaerobic conditions

Abstract

Background Anaerobic RBC storage reduces oxidative damage, maintains ATP & 2,3-diphosphoglycerate (DPG) levels and has superior 24hr recovery at 6weeks compared to standard storage. This study will determine if removal of CO2 during O2 depletion by gas exchange may affect RBC during anaerobic storage. Methods This is a matched 3 arm study (n=14): control, O2&CO2 depleted with Ar (AN), O2 depleted with 95%Ar/5%CO2 (AN[CO2]). RBC in additives AS-3 or OFAS3 were evenly divided into 3 bags, and anaerobic conditions were established by gas exchange. Bags were stored 1-6°C in closed chambers under anaerobic conditions or ambient air, sampled weekly for up to 9weeks for a panel of in vitro tests. A full metabolomics screening was conducted for the first 4 weeks of storage. Results Purging with Ar (AN) results in alkalization of the RBC and increased glucose consumption. The addition of 5%CO2 to the purging gas prevented CO2 loss with an equivalent starting and final pH and lactate to control bags (p>0.5, days0-21). ATP levels are higher in AN[CO2] (p<0.0001). DPG was maintained beyond 2 weeks in the AN arm (p<0.0001). Surprisingly, DPG was lost at the same rate in both control and AN[CO2] arms (p=0.6). Conclusion Maintenance of ATP in the AN[CO2] arm demonstrates that ATP production is not solely a function of the pH effect on glycolysis. CO2 in anaerobic storage prevented the maintenance of DPG, and DPG production appears to be pH dependent. CO2 as well as O2 depletion provides metabolic advantage for stored RBC.

Metabolomics in transfusion medicine

“…Metabolomics studies have been performed to confirm that energy metabolism‐related variables are improved by anaerobic storage at the expense of the pentose phosphate pathway and general antioxidant potential, consistent with previous in vitro studies . More recently, we demonstrated a role for anaerobiosis versus anaerobiosis in presence of CO 2 in modulating metabolic responses upon deoxygenation (Fig. ).…”

Red blood cells in hemorrhagic shock: a critical role for glutaminolysis in fueling alanine transamination in rats

“…15,25,26,28,30,33,50 Surprisingly, decreases in ATP and DPG were observed in shock RBCs, suggesting that shock acidosis rather than high-energy phosphate compounds may be the main driver of oxygen off-loading through the Bohr effect in hemorrhaged rats. Increased intracellular acidification in shock RBCs is mirrored by increased NADH/NAD 1 ratios, which would in turn fuel antioxidant enzymes such as methemoglobin reductase to counter oxidative stress.…”

“…[25][26][27][28][29] Observations relevant to RBC physiological adaptations to high-altitude hypoxia have been recently exploited to propose anaerobic storage as an alternative strategy to mitigate the storage lesion of packed RBCs in the blood bank. [30][31][32][33] Factors influencing RBC hypoxic metabolic reprogramming have been identified in the hypoxia-induced stabilization of deoxyhemoglobin, which competes with glycolytic enzymes for the N-terminal cytosolic domain of band 3, thereby promoting enzyme release from the membrane to the cytosol with subsequent increases in the activity of glyceraldehyde 3-phosphate dehydrogenase and phosphofructokinase, increased glycolytic fluxes, and ATP/DPG generation to reinforce this feedback loop. [25][26][27][28][29] Of note, plasma adenosine levels, which increase significantly upon early hemorrhage even before severe HS ensues in rats, 12 are important upstream contributors to these mechanisms through signaling via the RBC adenosine receptor A2B.…”

The Proteome of the Red Blood Cell: An Auspicious Source of New Insights into Membrane-Centered Regulation of Homeostasis

“…It is striking that these enzymes seem to be organized in interacting multiprotein complexes, suggesting an intensive cross-talk between oxygen transport, metabolism, anti-oxidant activity, and protein breakdown [23,24]. Recent data have not only expanded the original models on the oxygen-dependent modulation of the red blood cell metabolome [25,26], but may also link the concentration of carbon dioxide, independent of its effect on intracellular pH, to the activity of these complexes [27]. The unexpected finding of a reduction in transketolase, an enzyme of the pentose phosphate pathway, in these same red blood cells [28] represents an illustration of the power of proteomic analysis in the unearthing of the cellular “interactome” [8,10,23].…”