title: DMT1-mutant erythrocytes have shortened life span, accelerated glycolysis and increased oxidative stress.
abstract: OBJECTIVE
  Deficiency of the divalent metal transporter 1 (DMT1) leads to hypochromic microcytic anemia. We have previously shown that DMT1 deficiency impairs erythroid differentiation and induces apoptosis of erythroid cells. Here we analyzed metabolic processes and survival of mature erythrocytes in order to address potential involvement of erythrocyte defect in the pathophysiology of the disease.
  
  
  METHODS
  FACS analysis was used to determine the half-life of erythrocytes (CFSE fluorescence), phosphatidylserine exposure (Annexin V binding), cytosolic Ca(2+) (Fluo3/AM fluorescence) and reactive oxygen species (ROS; DCF fluorescence). Enzyme activities were determined by standard biochemical methods. The concentration of ATP and ADP was measured on HPLC-MS/MS.
  
  
  RESULTS
  We observed an accelerated clearance of CFSE-labeled DMT1-mutant erythrocytes from circulating blood when compared to wild-type erythrocytes. In vitro, DMT1-mutant erythrocytes showed significantly increased Annexin V binding after exposure to hyperosmotic shock and glucose depletion. Despite exaggerated anti-oxidative defense, higher ROS levels were present in DMT1-mutant erythrocytes. Accelerated anaerobic glycolysis and reduced ATP/ADP ratio detected in DMT1-mutant erythrocytes indicate enhanced demand for ATP.
  
  
  CONCLUSIONS
  We propose that DMT1 deficiency negatively affects metabolism and life span of mature erythrocytes; two other aspects of defective erythropoiesis which contribute to the pathophysiology of the disease.
authors.affiliation: Department of Biology, Faculty of Medicine and Dentistry Palacky University, Olomouc, Czech Republic.
authors.forename: Zuzana
authors.initials: Z
authors.lastname: Zidova
authors.forename: Katarina
authors.initials: K
authors.lastname: Kapralova
authors.forename: Pavla
authors.initials: P
authors.lastname: Koralkova
authors.forename: Renata
authors.initials: R
authors.lastname: Mojzikova
authors.forename: Dalibor
authors.initials: D
authors.lastname: Dolezal
authors.forename: Vladimir
authors.initials: V
authors.lastname: Divoky
authors.forename: Monika
authors.initials: M
authors.lastname: Horvathova
mesh_terms: D000244:Adenosine Diphosphate / Q000097:blood; D000255:Adenosine Triphosphate / Q000097:blood; D000747:Anemia, Hypochromic / Q000097:blood / Q000235:genetics* / Q000473:pathology; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D027682:Cation Transport Proteins / Q000097:blood / Q000172:deficiency / Q000235:genetics*; D004195:Disease Models, Animal; D004912:Erythrocytes / Q000378:metabolism* / Q000473:pathology; D004920:Erythropoiesis; D006019:Glycolysis; D006801:Humans; D051379:Mice; D009154:Mutation; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000097:blood; D000099236:Solute Carrier Family 11, Member 2
chemical_list: D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D027682:Cation Transport Proteins; D017382:Reactive Oxygen Species; D000099236:Solute Carrier Family 11, Member 2