Quantitative time-course metabolomics in human red blood cells reveal the temperature dependence of human metabolic networks

Abstract

The temperature dependence of biological processes has been studied at the levels of individual biochemical reactions and organism physiology ( basal metabolic rates) but has not been examined at the metabolic network level. Here, we used a systems biology approach to characterize the temperature dependence of the human red blood cell (RBC) metabolic network between 4 and 37 °C through absolutely quantified exo- and endometabolomics data. We used an Arrhenius-type model () to describe how the rate of a biochemical process changes with every 10 °C change in temperature. Multivariate statistical analysis of the metabolomics data revealed that the same metabolic network-level trends previously reported for RBCs at 4 °C were conserved but accelerated with increasing temperature. We calculated a median coefficient of 2.89 ± 1.03, within the expected range of 2-3 for biological processes, for 48 individual metabolite concentrations. We then integrated these metabolomics measurements into a cell-scale metabolic model to study pathway usage, calculating a median coefficient of 2.73 ± 0.75 for 35 reaction fluxes. The relative fluxes through glycolysis and nucleotide metabolism pathways were consistent across the studied temperature range despite the non-uniform distributions of coefficients of individual metabolites and reaction fluxes. Together, these results indicate that the rate of change of network-level responses to temperature differences in RBC metabolism is consistent between 4 and 37 °C. More broadly, we provide a baseline characterization of a biochemical network given no transcriptional or translational regulation that can be used to explore the temperature dependence of metabolism.

Metabolomics evaluation of early‐storage red blood cell rejuvenation at 4°C and 37°C

“…Since NAD + is required by GAPDH to sustain glycolytic activity, regeneration of NADH back to NAD + may thus favor the reactivation of late glycolysis. In this view, the presence of pyruvate in rejuvenation solutions further impacts ATP and diphosphoglycerate generation capacity, more so in RBCs rejuvenated at 37°C, consistent with the temperature dependency of enzymatic kinetics in glycolysis . However, the levels of ATP, diphosphoglycerate, and glycolytic intermediates as well as pyruvate/lactate ratios suggest that cold rejuvenation may be sufficient to prime reactivation of RBC metabolism while maintaining a logistic advantage over standard rejuvenation.…”

“…RBC storage in the blood bank is metabolically characterized by the progressive impairment of energy and redox metabolism . Briefly, it can be summarized that stored RBCs have a decreased capacity to generate ATP as a result of (1) temperature‐dependent effects on enzyme activities; (2) intracellular pH acidification via lactate accumulation; (3) negative feedback on phosphofructokinase activity; (4) reversible and irreversible oxidation of GAPDH; (5) DPG breakdown to sustain ATP generation for the first 2 weeks of storage; and (6) decreased glucose uptake and switch to other metabolic substrates, if available (e.g., citrate, other sugars) . These metabolic changes are functionally relevant in that, by depleting high‐energy phosphate compounds, RBCs lose their capacity to cope with osmotic/mechanical stressors and to regulate oxygen transport/off‐loading.…”

“…2 Briefly, it can be summarized that stored RBCs have a decreased capacity to generate ATP as a result of (1) temperature-dependent effects on enzyme activities; (2) intracellular pH acidification via lactate accumulation; (3) negative feedback on phosphofructokinase activity; (4) reversible and irreversible oxidation of GAPDH; (5) DPG breakdown to sustain ATP generation for the first 2 weeks of storage; and (6) decreased glucose uptake and switch to other metabolic substrates, if available (e.g., citrate, other sugars). 1,11,37,40,41 These metabolic changes are functionally relevant in that, by depleting high-energy phosphate compounds, RBCs lose their capacity to cope with osmotic/mechanical stressors and to regulate oxygen transport/off-loading. Depletion of ATP further impairs a glutathione synthesis that is already slowed under refrigerated storage conditions, ultimately resulting in the reduction of the total pool of glutathione (both GSH and GSSG) and accumulation of the metabolic dead-end product 5-oxoproline (because mature RBCs lack oxoprolinase).…”

“…In this view, the presence of pyruvate in rejuvenation solutions further impacts ATP and diphosphoglycerate generation capacity, more so in RBCs rejuvenated at 378C, consistent with the temperature dependency of enzymatic kinetics in glycolysis. 41 However, the levels of ATP, diphosphoglycerate, and glycolytic intermediates as well as pyruvate/lactate ratios suggest that cold rejuvenation may be sufficient to prime reactivation of RBC metabolism while maintaining a logistic advantage over standard rejuvenation. Of note, reactivation of energy metabolism corresponded to a restored total glutathione pool upon (cold or standard) rejuvenation, but not washing 15-day-old RBCs.…”

Heterogeneity of blood processing and storage additives in different centers impacts stored red blood cell metabolism as much as storage time: lessons from REDS‐III—Omics

“…4 Since then, omics technologies have helped to elucidate the complexity of the storage lesion, especially through the use of metabolomics, which provides a comprehensive overview of RBC metabolism. 5 While limited in the number of biological replicates and conditions tested in a single study, metabolomics investigations have flooded the literature over the past few years in the attempt to document the effect of different storage additives (SAGM, AS-1, AS-3, AS-5, AS-7, PAGGSM, PAG3M, ESOL-5), 6–13 rejuvenation procedures, 14 anaerobic storage conditions, 15–18 temperature effects (>4°C 19 or cryopreservation 20 ) or novel storage additives (e.g. supplementation with adenine, alternative sugars or antioxidants).…”