Team: Laboratory on Repair and Transcription in hematopoiestic Stem Cells
Metabolomic analysis of normal and pathological human red blood cells
The laboratory of research on Replication and Transcription in Stem cells is part of the institute of Cellular and Molecular Radiobiology located at the Fontenay-aux-Roses CEA center. In close collaboration with the Drug Metabolism Research Laboratory, located at the Saclay CEA center, we have developed new tools to analyze the metabolic profiling of human red blood cells (RBC).
Emerging metabolomic tools can now be used to establish metabolic signatures of specialized circulating hematopoietic cells in physiological or pathological conditions and in human hematological diseases. As human RBC have no nucleus and no organelle such as mitochondria, RBC might represent a simple model to study metabolomic deficiencies associated with hereditary disease. To determine metabolomes of normal and pathological erythrocytes, we use a sample treatment method suitable for the extraction erythrocytes metabolites, and a liquid chromatography-mass spectrometry based metabolite-profiling method to detect, identify and quantify metabolites.
We started the metabolomic studies on human RBC by comparing the metabolomes of normal and sickle cells RBC. This study highlighted major concentration changes in metabolites produced by endogenous glycolysis (accumulation of many glycolytic intermediates), metabolites involved in endogenous glutathione and ascorbate metabolisms (accumulation of ascorbate metabolism intermediates such as diketogulonic acid and decreased levels of both glutathione and glutathione disulfide), in markers for membrane lipids turnover such as carnitine, in some amino acids (suggesting alterations of membrane transport systems in RBC from sickle cell disease patients), and in exogenous arginine and NO metabolisms (i.e., metabolites such as spermine, spermidine or citrulline).
We conducted a similar study of Overhydrated Hereditary Stomatocytosis (OHSt) RBC. OHSt, clinically characterized by a hemolytic anemia, is a rare disorder of the erythrocyte membrane permeability to monovalent cations, associated with mutations in the Rh-associated glycoprotein gene. In OHSt RBC, glycolysis is exhausted with accumulation of ADP, pyruvate, lactate, and malate. Ascorbate metabolic pathway is altered probably due to a limited entry of dehydroascorbate. Although no major oxydative stress has been reported in patients with overhydrated hereditary stomatocytosis, we found decreased amounts of oxidized glutathione, creatine and ergothioneine, suggesting transporter abnormalities and/or uncharacterized oxidative stress. In these two studies, metabolomic analysis of young and old normal red blood cells indicates metabolites whose levels are directly related to sickle cell disease or to Overhydrated Hereditary Stomatocytosis. These results show the relevance of metabolic profiling for the follow-up of sickle cell patients or other red blood cell diseases and pinpoint the importance of metabolomics to further depict the physiopathology of human hematological diseases. We will continue the research on the metabolic profiling of normal and sickle cell RBC and enlarge this profiling by lipidomic studies. In addition to cognitive data on the metabolome of a human cell, this might lead to the definition of metabolic signatures that will be relevant for clinical applications. We will also study the metabolic profiling of RBC from patients with chronic diseases such as diabetes to determine if a metabolic signature can be drawn and be useful to predict any complications of such disease.
- Metabolomic analysis of human red blood cells.
- Darghouth D, Koehl B, Madalinski G, Heilier JF, Bovee P, Xu Y, Olivier MF, Bartolucci P, Benkerrou M, Pissard S, Colin Y, Galacteros F, Bosman G, Junot C and Roméo PH. Physiopathology of sickle cell disease is mirrored by red blood cells metabolome. Blood, 2011, 117 : e57-e66.
- Darghouth D, Koehl B, Heilier JF, Madalinski G, Bovee P, Bosman G, Delaunay J, Junot C and Roméo PH. Alterations of red blood cells metabolome in overhydrated hereditary stomatocytosis. Hematologica, 2011, 96 : 1861-1865.