title: Stimulation of suicidal erythrocyte death by ipratropium bromide.
abstract: OBJECTIVE
  Ipratropium bromide, an anticholinergic agent widely used in obstructive lung disease, has previously been shown to trigger suicidal death of nucleated cells or apoptosis. Despite their lack of mitochondria and nuclei, key organelles in the execution of apoptosis, erythrocytes may similarly undergo suicidal cell death, which is characterized by cell shrinkage and by cell membrane scrambling with phosphatidylserine-exposure at the cell surface. Triggers of eryptosis include increase of cytosolic Ca(2+)-activity ([Ca(2+)](i)). The present study explored whether ipratropium bromide triggers eryptosis.
  
  
  METHODS
  [Ca Ca(2+)](i) was estimated utilizing Fluo3 fluorescence, cell volume from forward scatter, phosphatidylserine-exposure from annexin-V-binding, and hemolysis from hemoglobin release.
  
  
  RESULTS
  A 48 h exposure to ipratropium bromide (1 nM) significantly increased [Ca(2+)](i), decreased forward scatter and increased annexin-V-binding. Ipratropium bromide treatment was followed by slight but significant increase of hemolysis. Removal of extracellular Ca(2+) or inhibition of Ca(2+) permeable cation channels with amiloride (1 mM) virtually abolished cell membrane scrambling. Ca(2+) ionophore ionomycin (1 µM, 30 min) increased the percentage of phosphatidylserine exposing erythrocytes to similarly high levels in the absence and presence of ipratropium bromide (1 nM).
  
  
  CONCLUSIONS
  Ipratropium bromide triggers suicidal erythrocyte death or eryptosis, an effect mainly due to stimulation of Ca(2+)-entry.
authors.affiliation: Department of Physiology, University of Tuebingen, Tuebingen, Germany.
authors.forename: Nazneen
authors.initials: N
authors.lastname: Shaik
authors.forename: Eyad
authors.initials: E
authors.lastname: Alhourani
authors.forename: Anastasia
authors.initials: A
authors.lastname: Bosc
authors.forename: Guilai
authors.initials: G
authors.lastname: Liu
authors.forename: Syeda
authors.initials: S
authors.lastname: Towhid
authors.forename: Adrian
authors.initials: A
authors.lastname: Lupescu
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D018938:Caco-2 Cells; D020013:Calcium Signaling; D048429:Cell Size / Q000187:drug effects; D018680:Cholinergic Antagonists / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects / Q000502:physiology*; D005434:Flow Cytometry; D006801:Humans; D009241:Ipratropium / Q000494:pharmacology*; D010718:Phosphatidylserines / Q000378:metabolism
chemical_list: D018680:Cholinergic Antagonists; D010718:Phosphatidylserines; D009241:Ipratropium