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Enhanced Eryptosis in Glucose-6-Phosphate Dehydrogenase Deficiency.
Bouguerra, G, Talbi, K, Trabelsi, N, Chaouachi, D, Boudriga, I, Abbès, S, Menif, S
Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology. 2021;(6):761-772
Abstract
BACKGROUND/AIMS: Defects in the Glucose-6-Phosphate Dehydrogenase (G6PD) enzyme enhance cellular oxidative damage, thus impairing erythrocytes and radically shortening their lifespan. We aimed to study programmed erythrocyte cell death in G6PD-deficient patients, describe the molecular genetics basis of G6PD and investigate phenotype-genotype correlations. METHODS We explored eryptosis using the annexin V-binding assay, taken as an indicator of PS exposure at the erythrocyte surface. We assessed reactive oxygen species (ROS) production, intracellular calcium concentrations and ceramide formation at the cell surface. Prior to and following treatments, cells were analyzed by flow cytometry. Finally, we explored G6PD gene mutations through PCR-Sanger sequencing. RESULTS Before stimulation, PS-exposing erythrocytes were significantly higher in G6PD-deficient patients than in healthy volunteers. This was paralleled by a significant increase in reactive oxygen species production, suggesting that oxidative stress is the main trigger of PS exposure in G6PD-deficient erythrocytes. Five previously described mutations were detected in our patients. Two genotypes correlated with a significantly higher percentage of PS-exposing cells. CONCLUSION Our study uncovers a novel effect detected in G6PD-deficient erythrocytes which is cell membrane scrambling with PS translocation to the erythrocyte surface. Our findings shed a light on the mechanisms of premature erythrocyte clearance in G6PD deficiency.
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Ethyl glucuronide, a marker of alcohol consumption, correlates with metabolic markers of oxidant stress but not with hemolysis in stored red blood cells from healthy blood donors.
D'Alessandro, A, Fu, X, Reisz, JA, Stone, M, Kleinman, S, Zimring, JC, Busch, M, ,
Transfusion. 2020;(6):1183-1196
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BACKGROUND Red blood cell (RBC) storage in the blood bank is associated with the progressive accumulation of oxidant stress. While the mature erythrocyte is well equipped to cope with such stress, recreative habits like alcohol consumption may further exacerbate the basal level of oxidant stress and contribute to the progress of the storage lesion. STUDY DESIGN AND METHODS RBC levels of ethyl glucuronide, a marker of alcohol consumption, were measured via ultra-high-pressure liquid chromatography coupled with high-resolution mass spectrometry. Analyses were performed on 599 samples from the recalled donor population at Storage Days 10, 23, and 42 (n = 250), as part of the REDS-III RBC-Omics (Recipient Epidemiology Donor Evaluation Study III Red Blood Cell-Omics) study. This cohort consisted of the 5th and 95th percentile of donors with extreme hemolytic propensity out of the original cohort of 13,403 subjects enrolled in the REDS-III RBC Omics study. Ehtyl glucuronide levels were thus correlated to global metabolomics and lipidomics analyses and RBC hemolytic propensity. RESULTS Ethyl glucuronide levels were positively associated with oxidant stress markers, including glutathione consumption and turnover, methionine oxidation, S-adenosylhomocysteine accumulation, purine oxidation, and transamination markers. Decreases in glycolysis and energy metabolism, the pentose phosphate pathway and ascorbate system were observed in those subjects with the highest levels of ethyl glucuronide, though hemolysis values were comparable between groups. CONCLUSION Though preliminary, this study is suggestive that markers of alcohol consumption are associated with increases in oxidant stress and decreases in energy metabolism with no significant impact on hemolytic parameters in stored RBCs from healthy donor volunteers.
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Stored RBC metabolism as a function of caffeine levels.
D'Alessandro, A, Fu, X, Reisz, JA, Kanias, T, Page, GP, Stone, M, Kleinman, S, Zimring, JC, Busch, M, ,
Transfusion. 2020;(6):1197-1211
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BACKGROUND Coffee consumption is extremely common in the United States. Coffee is rich with caffeine, a psychoactive, purinergic antagonist of adenosine receptors, which regulate red blood cell energy and redox metabolism. Since red blood cell (purine) metabolism is a critical component to the red cell storage lesion, here we set out to investigate whether caffeine levels correlated with alterations of energy and redox metabolism in stored red blood cells. STUDY DESIGN AND METHODS We measured the levels of caffeine and its main metabolites in 599 samples from the REDS-III RBC-Omics (Recipient Epidemiology Donor Evaluation Study III Red Blood Cell-Omics) study via ultra-high-pressure-liquid chromatography coupled to high-resolution mass spectrometry and correlated them to global metabolomic and lipidomic analyses of RBCs stored for 10, 23, and 42 days. RESULTS Caffeine levels positively correlated with increased levels of the main red cell antioxidant, glutathione, and its metabolic intermediates in glutathione-dependent detoxification pathways of oxidized lipids and sugar aldehydes. Caffeine levels were positively correlated with transamination products and substrates, tryptophan, and indole metabolites. Expectedly, since caffeine and its metabolites belong to the family of xanthine purines, all xanthine metabolites were significantly increased in the subjects with the highest levels of caffeine. However, high-energy phosphate compounds ATP and DPG were not affected by caffeine levels, despite decreases in glucose oxidation products-both via glycolysis and the pentose phosphate pathway. CONCLUSION Though preliminary, this study is suggestive of a beneficial correlation between the caffeine levels and improved antioxidant capacity of stored red cells.
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Absorption and Safety With Sustained Use of RELiZORB Evaluation (ASSURE) Study in Patients With Cystic Fibrosis Receiving Enteral Feeding.
Stevens, J, Wyatt, C, Brown, P, Patel, D, Grujic, D, Freedman, SD
Journal of pediatric gastroenterology and nutrition. 2018;(4):527-532
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OBJECTIVES Pancreatic insufficiency (PI) and malabsorption of fats lead to reduced caloric intake, inability to maintain weight, and increased gastrointestinal symptoms. Thus, enteral nutrition (EN) is used in patients with cystic fibrosis (CF) and poor nutritional status. The current study evaluated safety, tolerability, and improvement of fatty acid (FA) status in red blood cell (RBC) membranes, a marker of long-term FA absorption, with an in-line digestive cartridge (RELiZORB) that hydrolyzes fat in enteral formula. METHODS Patients with CF receiving EN participated in a multicenter, 90-day open-label study during which RELiZORB was used with overnight EN. The primary endpoint was change over time in RBC uptake of docosahexaenoic acid (DHA)+ eicosapentaenoic acid (EPA). Gastrointestinal symptoms were collected to evaluate safety and tolerability. Several clinical and anthropometric parameters were also assessed throughout the study. RESULTS A total of 36 subjects completed the study with a mean age of 13.8 years, body mass index of 17.7 and 6.2 years mean use of overnight EN. Fat absorption significantly improved as shown by increased RBC levels of DHA+EPA, improved ω-6/ω-3 ratio, and increased plasma levels of DHA+EPA. RELiZORB use was not associated with any unanticipated adverse events. CONCLUSIONS RELiZORB use was found to be safe, well tolerated, and resulted in increased levels of FAs in RBCs and plasma. This is the first prospective study to show EN can improve FA abnormalities in CF. Because improvement in omega-3 levels has been shown to help pulmonary and inflammatory status as well as anthropometric parameters in CF, RELiZORB may have important long-term therapeutic benefits in patients with CF.
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Effects of red blood cell (RBC) transfusion on sickle cell disease recipient plasma and RBC metabolism.
Culp-Hill, R, Srinivasan, AJ, Gehrke, S, Kamyszek, R, Ansari, A, Shah, N, Welsby, I, D'Alessandro, A
Transfusion. 2018;(12):2797-2806
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BACKGROUND Exchange transfusion is a mainstay in the treatment of sickle cell anemia. Transfusion recipients with sickle cell disease (SCD) can be transfused over 10 units per therapy, an intervention that replaces circulating sickle red blood cells (RBCs) with donor RBCs. Storage of RBCs makes the intervention logistically feasible. The average storage duration for units transfused at the Duke University Medical Center is approximately 2 weeks, a time window that should anticipate the accumulation of irreversible storage lesion to the RBCs. However, no metabolomics study has been performed to date to investigate the impact of exchange transfusion on recipients' plasma and RBC phenotypes. STUDY DESIGN AND METHODS Plasma and RBCs were collected from patients with sickle cell anemia before transfusion and within 5 hours after exchange transfusion with up to 11 units, prior to metabolomics analyses. RESULTS Exchange transfusion significantly decreased plasma levels of markers of systemic hypoxemia like lactate, succinate, sphingosine 1-phosphate, and 2-hydroxyglutarate. These metabolites accumulated in transfused RBCs, suggesting that RBCs may act as scavenger/reservoirs. Transfused RBCs displayed higher glycolysis, total adenylate pools, and 2,3-diphosphoglycerate, consistent with increased capacity to deliver oxygen. Plasma levels of acyl-carnitines and amino acids decreased, while fatty acids and potentially harmful phthalates increased upon exchange transfusion. CONCLUSION Metabolic phenotypes confirm the benefits of the transfusion therapy in transfusion recipients with SCD and the reversibility of some of the metabolic storage lesion upon transfusion in vivo in 2-week-old RBCs. However, results also suggest that potentially harmful plasticizers are transfused.
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Red Blood Cell Folate Insufficiency among nonpregnant Women of Childbearing age in Guatemala 2009 to 2010: Prevalence and predicted Neural Tube Defects risk.
Rosenthal, J, Reeve, ME, Ramirez, N, Crider, KS, Sniezek, J, Vellozzi, C, Devine, O, Lopez-Pazos, E
Birth defects research. Part A, Clinical and molecular teratology. 2016;(7):587-95
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BACKGROUND The World Health Organization recently released recommendations stating that red blood cell (RBC) folate concentrations should be above 400 ng/L (906 nmol/L) for optimal prevention of folate-sensitive neural tube defects (NTDs). The objective of this study was to determine the distribution of folate insufficiency (FI) (<906 nmol/L) and potential risk of NTDs based on RBC folate concentrations among nonpregnant women of child-bearing age in Guatemala. METHODS A national and regional multistage cluster probability survey was completed during 2009 to 2010 among Guatemalan women of child-bearing age 15 to 49 years of age. Demographic and health information and blood samples for RBC folate analyses were collected from 1473 women. Prevalence rate ratios of FI and predicted NTD prevalence were estimated based on RBC folate concentrations comparing subpopulations of interest. RESULTS National FI prevalence was 47.2% [95% confidence interval, 43.3-51.1] and showed wide variation by region (18-81%). In all regions, FI prevalence was higher among indigenous (27-89%) than among nonindigenous populations (16-44%). National NTD risk based on RBC folate concentrations was estimated to be 14 per 10,000 live births (95% uncertainty interval, 11.1-18.6) and showed wide regional variation (from 11 NTDS in the Metropolitan region to 26 NTDs per 10,000 live births in the Norte region). CONCLUSION FI remains a common problem in populations with limited access to fortified products, specifically rural, low income, and indigenous populations. However, among subpopulations that are most likely to have fortified food, the prevalence of FI is similar to countries with well-established fortification programs. Birth Defects Research (Part A) 106:587-595, 2016. © 2016 Wiley Periodicals, Inc.
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Effects of glucomannan-enriched, aronia juice-based supplement on cellular antioxidant enzymes and membrane lipid status in subjects with abdominal obesity.
Kardum, N, Petrović-Oggiano, G, Takic, M, Glibetić, N, Zec, M, Debeljak-Martacic, J, Konić-Ristić, A
TheScientificWorldJournal. 2014;:869250
Abstract
The aim of this study was to analyze the effects of a 4-week-long consumption of glucomannan-enriched, aronia juice-based supplement on anthropometric parameters, membrane fatty acid profile, and status of antioxidant enzymes in erythrocytes obtained from postmenopausal women with abdominal obesity. Twenty women aged 45-65 with a mean body mass index (BMI) of 36.1 ± 4.4 kg/m(2) and waist circumference of 104.8 ± 10.1 cm were enrolled. Participants were instructed to consume 100 mL of supplement per day as part of their regular diet. A significant increase in the content of n-3 (P < 0.05) polyunsaturated fatty acids in membrane phospholipids was observed, with a marked increase in the level of docosahexaenoic fatty acid (P < 0.05). Accordingly, a decrease in the n-6 and n-3 fatty acids ratio was observed (P < 0.05). The observed effects were accompanied with an increase in glutathione peroxidase activity (P < 0.05). Values for BMI (P < 0.001), waist circumference (P < 0.001), and systolic blood pressure (P < 0.05) were significantly lower after the intervention. The obtained results indicate a positive impact of tested supplement on cellular oxidative damage, blood pressure, and anthropometric indices of obesity.
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Neither folic acid supplementation nor pregnancy affects the distribution of folate forms in the red blood cells of women.
Hartman, BA, Fazili, Z, Pfeiffer, CM, O'Connor, DL
The Journal of nutrition. 2014;(9):1364-9
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It is not known whether folate metabolism is altered during pregnancy to support increased DNA and RNA biosynthesis. By using a state-of-the-art LC tandem mass spectrometry technique, the aim of this study was to investigate differences in RBC folate forms between pregnant and nonpregnant women and between nonpregnant women consuming different concentrations of supplemental folic acid. Forms of folate in RBCs were used to explore potential shifts in folate metabolism during early erythropoiesis. Total RBC folate and folate forms [tetrahydrofolate; 5-methyltetrahydrofolate (5-methyl-THF); 4α-hydroxy-5-methyl-tetrahydrofolate (an oxidation product of 5-methyl-THF); 5-formyl-tetrahydrofolate; and 5,10-methenyl-tetrahydrofolate] were measured in 4 groups of women (n = 26): pregnant women (PW) (30-36 wk of gestation) consuming 1 mg/d of folic acid, and nonpregnant women consuming 0 mg/d (NPW-0), 1 mg/d (NPW-1), and 5 mg/d (NPW-5) folic acid. The mean ± SD RBC folate concentration of the NPW-0 group (890 ± 530 nmol/L) was lower than the NPW-1 (1660 ± 350 nmol/L) and NPW-5 (1980 ± 570 nmol/L) groups as assessed by microbiologic assay (n = 26, P < 0.0022). No difference was found between the NPW-1 and NPW-5 groups. We detected 5-methyl-THF [limit of detection (LOD) = 0.06 nmol/L] in all groups and tetrahydrofolate (LOD = 0.2 nmol/L) in most women regardless of methylenetetrahydrofolate reductase genotype. Most women consuming folic acid supplements had detectable concentrations of 5,10-methenyl-tetrahydrofolate (LOD = 0.31 nmol/L). However, there was no difference in the relative distribution of 5-methyl-THF (83-84%), sum of non-methyl folates (0.6-3%), or individual non-methyl folate forms in RBCs across groups. We conclude that although folic acid supplementation in nonpregnant women increases RBC total folate and the concentration of individual folate forms, it does not alter the relative distribution of folate forms. Similarly, distribution of RBC folate forms did not differ between pregnant and nonpregnant women. This trial was registered at clinicaltrials.gov as NCT01741077.
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Intra-erythrocyte infusion of dexamethasone reduces neurological symptoms in ataxia teleangiectasia patients: results of a phase 2 trial.
Chessa, L, Leuzzi, V, Plebani, A, Soresina, A, Micheli, R, D'Agnano, D, Venturi, T, Molinaro, A, Fazzi, E, Marini, M, et al
Orphanet journal of rare diseases. 2014;:5
Abstract
BACKGROUND Ataxia Teleangiectasia [AT] is a rare neurodegenerative disease characterized by early onset ataxia, oculocutaneous teleangiectasias, immunodeficiency, recurrent infections, radiosensitivity and proneness to cancer. No therapies are available for this devastating disease. Recent observational studies in few patients showed beneficial effects of short term treatment with betamethasone. To avoid the characteristic side effects of long-term administration of steroids we developed a method for encapsulation of dexamethasone sodium phosphate (DSP) into autologous erythrocytes (EryDex) allowing slow release of dexamethasone for up to one month after dosing. Aims of the study were: the assessment of the effect of EryDex in improving neurological symptoms and adaptive behaviour of AT patients; the safety and tolerability of the therapy. METHODS Twenty two patients (F:M=1; mean age 11.2 ± 3.5) with a confirmed diagnosis of AT and a preserved or partially supported gait were enrolled for the study. The subjects underwent for six months a monthly infusion of EryDex. Ataxia was assessed by the International Cooperative Ataxia Rating Scale (ICARS) and the adaptive behavior by Vineland Adaptive Behavior Scales (VABS). Clinical evaluations were performed at baseline and 1, 3, and 6 months. RESULTS An improvement in ICARS (reduction of the score) was detected in the intention-to-treat (ITT) population (n=22; p=0.02) as well as in patients completing the study (per protocol PP) (n=18; p=0.01), with a mean reduction of 4 points (ITT) or 5.2 points (PP). When compared to baseline, a significant improvement were also found in VABS (increase of the score) (p<0.0001, ITT, RMANOVA), with statistically significant increases at 3 and 6 months (p<0.0001). A large inter-patient variability in the incorporation of DSP into erythrocytes was observed, with an evident positive effect of higher infusion dose on ICARS score decline. Moreover a more marked improvement was found in less neurologically impaired patients. Finally, a 19 month-extension study involving a subgroup of patients suggested that Erydex treatment can possibly delay the natural progression of the disease.EryDex was well tolerated; the most frequent side effects were common AT pathologies. CONCLUSIONS EryDex treatment led to a significant improvement in neurological symptoms, without association with the typical steroid side effects. TRIAL REGISTRATION Current Controlled Trial 2010-022315-19SpA.
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Storing red blood cells with vitamin C and N-acetylcysteine prevents oxidative stress-related lesions: a metabolomics overview.
Pallotta, V, Gevi, F, D'alessandro, A, Zolla, L
Blood transfusion = Trasfusione del sangue. 2014;(3):376-87
Abstract
BACKGROUND Recent advances in red blood cell metabolomics have paved the way for further improvements of storage solutions. MATERIALS AND METHODS In the present study, we exploited a validated high performance liquid chromatography-mass spectrometry analytical workflow to determine the effects of vitamin C and N-acetylcysteine supplementation (anti-oxidants) on the metabolome of erythrocytes stored in citrate-phosphate-dextrose saline-adenine-glucose-mannitol medium under blood bank conditions. RESULTS We observed decreased energy metabolism fluxes (glycolysis and pentose phosphate pathway). A tentative explanation of this phenomenon could be related to the observed depression of the uptake of glucose, since glucose and ascorbate are known to compete for the same transporter. Anti-oxidant supplementation was effective in modulating the redox poise, through the promotion of glutathione homeostasis, which resulted in decreased haemolysis and less accumulation of malondialdehyde and oxidation by-products (including oxidized glutathione and prostaglandins). DISCUSSION Anti-oxidants improved storage quality by coping with oxidative stress at the expense of glycolytic metabolism, although reservoirs of high energy phosphate compounds were preserved by reduced cyclic AMP-mediated release of ATP.