Increased cytochrome c-mediated DNA fragmentation and cell death in manganese-superoxide dismutase-deficient mice after exposure to subarachnoid hemolysate

Paul G. Matz, Miki Fujimura, Anders Lewen, Yuiko Morita-Fujimura, Pak H. Chan

Research output: Contribution to journalArticle

75 Citations (Scopus)

Abstract

Background and Purpose-We sought to investigate the mechanisms for oxidative injury caused by subarachnoid hemolysate, a pro-oxidant. Methods-Injection of 50 μL of subarachnoid hemolysate or saline was performed in CD1 mice (n=75), mutant mice deficient in Mn-superoxide dismutase (Sod2 +/-; n=23), and their wild-type littermates (n=23). Subcellular location of cytochrome c was studied by immunocytochemistry, immunofluorescence, and immunoblotting of cellular fractions. DNA fragmentation was assessed though DNA laddering and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL). Cell death was examined through basic histology. Results-Cytochrome c immunoreactivity was present in the cytosol of neurons at 2 hours after hemolysate injection and increased by 4 hours compared with saline-injected animals (P<0.02). Cytosolic cytochrome c was more abundant in Sod2+/- mutants. DNA fragmentation was evident at 24 hours, but not 4 hours, after hemolysate injection as determined by DNA laddering and TUNEL staining (P<0.02). DNA fragmentation colocalized to cells with cytosolic cytochrome c and iron. In Sod2+/- mutants, the extent of fragmentation was increased as determined by TUNEL staining (52% increase; P<0.02) and DNA laddering (optical density=0.819 versus 0.391; P<0.01). Cell death was evident on basic histology as early as 4 hours after hemolysate injection. No cell death was evident in controls. In Sod2+/- mutants, cell death was increased by 51% compared with wild-type littermates (P<0.05). Conclusions-These results demonstrate that subarachnoid blood products are associated with the presence of cytochrome c in the cytosol and subsequent cell death in neurons. It appears that Mn-superoxide dismutase plays a role in preventing cell death after exposure to subarachnoid blood products.

Original languageEnglish
Pages (from-to)506-515
Number of pages10
JournalStroke
Volume32
Issue number2
DOIs
Publication statusPublished - 2001 Jan 1

Keywords

  • Hemoglobin
  • Iron
  • Mice
  • Stroke, experimental
  • Subarachnoid hemorrhage
  • Superoxide dismutase

ASJC Scopus subject areas

  • Clinical Neurology
  • Cardiology and Cardiovascular Medicine
  • Advanced and Specialised Nursing

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