title: Plasmalemma permeability and necrotic cell death phenotypes after intracerebral hemorrhage in mice.
abstract: OBJECTIVE
  Traumatic and ischemic brain injury induce plasmalemma permeability and necrosis; however, no studies have examined these aspects of cellular injury in intracerebral hemorrhage models.
  
  
  METHODS
  In vivo propidium iodide (PI) and YOYO-1 were used to assess plasmalemma damage after collagenase-induced intracerebral hemorrhage in mice. Ex vivo aspartylglutamylvalylaspartic acid, terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling, and electron microscopy were used to assess the relationship between plasmalemma permeability and mode of cell death. Cell types vulnerable to plasmalemma damage were determined by immunohistochemistry.
  
  
  RESULTS
  Plasmalemma permeability was first detected in the lesion at 1 to 3 hours and peaked at 48 to 72 hours. Neurons and IBA-1-positive cells with morphological features of monocytes were sensitive, whereas resident microglia and astrocytes were resistant to plasmalemma permeability. PI+ cells colocalized with fluorescent-labeled caspase substrates and terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling beginning at 3 to 6 hours. At 48 hours, greater than half of injured cells were PI+/aspartylglutamylvalylaspartic acid- or PI+/terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling- suggesting necrosis, and <5% were PI-/terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling+ or PI-/aspartylglutamylvalylaspartic acid+. Electron microscopy confirmed ultrastructural features of necrosis at 24 hours after intracerebral hemorrhage, high mobility group box protein-1 was released from permeable cells, and mice deficient in receptor interacting protein kinase (RIPK) 3, a known necrosis trigger, had 50% less PI+ cells at 24 hours. Permeable cells remained in the brain for at least 24 hours with <10% spontaneous resealing.
  
  
  CONCLUSIONS
  Necrosis contributes to cell demise after intracerebral hemorrhage. Programmed necrosis and plasmalemma damage may represent novel therapeutic targets to prevent cell death or rescue injured cells after intracerebral hemorrhage.
authors.affiliation: Neuroscience Center, Department of Pediatrics, Massachusetts General Hospital, Charlestown, MA 02129, USA.
authors.forename: Xiaoxia
authors.initials: X
authors.lastname: Zhu
authors.forename: Luyang
authors.initials: L
authors.lastname: Tao
authors.forename: Emiri
authors.initials: E
authors.lastname: Tejima-Mandeville
authors.forename: Jianhua
authors.initials: J
authors.lastname: Qiu
authors.forename: Juyeon
authors.initials: J
authors.lastname: Park
authors.forename: Kent
authors.initials: K
authors.lastname: Garber
authors.forename: Maria
authors.initials: M
authors.lastname: Ericsson
authors.forename: Eng H
authors.initials: EH
authors.lastname: Lo
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Whalen
mesh_terms: D000818:Animals; D020169:Caspases / Q000378:metabolism; D002452:Cell Count; D016923:Cell Death / Q000502:physiology*; D002460:Cell Line; D002462:Cell Membrane / Q000473:pathology* / Q000648:ultrastructure; D002470:Cell Survival; D002543:Cerebral Hemorrhage / Q000139:chemically induced / Q000473:pathology*; D017364:Collagenases; D004396:Coloring Agents; D049452:Green Fluorescent Proteins; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D008297:Male; D051379:Mice; D008854:Microscopy, Electron; D009336:Necrosis; D010539:Permeability; D010641:Phenotype; D011419:Propidium
chemical_list: D004396:Coloring Agents; D049452:Green Fluorescent Proteins; D011419:Propidium; D020169:Caspases; D017364:Collagenases
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