Defining the effects of storage on platelet bioenergetics: The role of increased proton leak

Abstract

The quality of platelets decreases over storage time, shortening their shelf life and potentially worsening transfusion outcomes. The changes in mitochondrial function associated with platelet storage are poorly defined and to address this we measured platelet bioenergetics in freshly isolated and stored platelets. We demonstrate the hypotonic stress test stimulates both glycolysis and oxidative phosphorylation and the stored platelets showed a decreased recovery to this stress. We found no change in aggregability between the freshly isolated and stored platelets. Bioenergetic parameters were changed including increased proton leak and decreased basal respiration and this was reflected in a lower bioenergetic health index (BHI). Mitochondrial electron transport, measured in permeabilized platelets, showed only minor changes which are unlikely to have a significant impact on platelet function. There were no changes in basal glycolysis between the fresh and stored platelets, however, glycolytic rate was increased in stored platelets when mitochondrial ATP production was inhibited. The increase in proton leak was attenuated by the addition of albumin, suggesting that free fatty acids could play a role in increasing proton leak and decreasing mitochondrial function. In summary, platelet storage causes a modest decrease in oxidative phosphorylation driven by an increase in mitochondrial proton leak, which contributes to the decreased recovery to hypotonic stress.

Systems analysis of metabolism in platelet concentrates during storage in platelet additive solution

“…Disruption of oxidative metabolism increases the rate of glycolysis to compensate for a decrease in ATP turnover (57). The decrease in oxidative metabolism in long term stored platelets has previously been demonstrated (58). We have described the shifts in metabolic pathway utilization that take place in buffy coat and apheresis platelets during storage through cell scale modelling.…”

Mitochondrial damage and senescence phenotype of cells derived from a novel frataxin G127V point mutation mouse model of Friedreich's ataxia

“…Mitochondrial stress tests were performed by sequentially injecting oligomycin (Oligo, 1 µg/ml), carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP, 1.5 µM) and antimycin A (Anti A, 10 µM) ( Chacko et al, 2013 ). To record fatty acid-mediated OCR measurements, BSA-palmitate substrate (25 µg/ml) was incubated with MEFs for 30 min before the assay in XF-DMEM medium ( Ravi et al, 2015 ). Etomoxir (10 µM) was added to determine fatty acid-sensitive OCR ( Ravi et al, 2015 ).…”

“…To record fatty acid-mediated OCR measurements, BSA-palmitate substrate (25 µg/ml) was incubated with MEFs for 30 min before the assay in XF-DMEM medium ( Ravi et al, 2015 ). Etomoxir (10 µM) was added to determine fatty acid-sensitive OCR ( Ravi et al, 2015 ). The data were normalized to the total protein content in each well, measured using the Lowry HS protein assay (Bio-Rad) and expressed as the mean.…”

Integrative metabolomics and transcriptomics signatures of clinical tolerance to Plasmodium vivax reveal activation of innate cell immunity and T cell signaling

“…Collection of human platelets was approved by the University of Alabama at Birmingham Institutional Review Board, and performed as described [36] . Briefly, platelet-rich plasma, obtained from individual donors from the blood bank at the University of Alabama at Birmingham, was centrifuged at 1500 g for 10 min, and washed with PBS containing prostaglandin I2 (1 µg/ml).…”