In situ detection of neuronal DNA strand breaks using the Klenow fragment of DNA polymerase I reveals different mechanisms of neuron death after global cerebral ischemia

Kunlin Jin, Jun Chen, Tetsuya Nagayama, Minzhi Chen, Jennifer Sinclair, Steven H. Graham, Roger P. Simon

Research output: Contribution to journalArticle

68 Scopus citations

Abstract

Ischemic cell injury in the brain may involve a cascade of programmed cell death. DNA damage may be either a catalyst or a consequence of this cascade. Therefore, the induction of DNA strand breaks in the rat brain following transient global ischemia was examined using (a) the Klenow labeling assay, identifying DNA single-strand breaks (SSBs) or double-strand breaks (DSBs) with protruding 5' termini, and (b) terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL), detecting DNA DSBs with protruding 3' termini or blunt ends. Klenow-positive staining occurred within 2 h of reperfusion and increased with increasing durations of reperfusion. DNA damage detected with the Klenow labeling assay preceded that of TUNEL expression in the caudate putamen, reticular thalamus, thalamus, and cortex. However, in CA1, DNA SSBs were not detected until 72 h of reperfusion and occurred simultaneously with DSBs. Thus, the time course and fragmentation characteristics of DNA damage differ between the hippocampal CA1 and other selectively vulnerable brain regions. This distinct pattern suggests that the delayed neuronal death in CA1 following transient global ischemia may occur via an apoptotic mechanism different from that of other brain regions.

Original languageEnglish
Pages (from-to)1204-1214
Number of pages11
JournalJournal of Neurochemistry
Volume72
Issue number3
DOIs
StatePublished - 24 Feb 1999

Keywords

  • Brain
  • DNA strand breaks
  • Ischemia
  • Klenow fragment-Terminal deoxynucleotidyl transferase
  • Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling

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