INCREASES IN DNA LESIONS AND THE DNA-DAMAGE INDICATOR GADD45 FOLLOWING TRANSIENT CEREBRAL-ISCHEMIA

Transient global or focal ischemia leads to the production of several types of lesions in the DNA backbone including alkali-labile sites, an d both single-stranded (ss) and double-stranded (ds) breaks. The ds br eaks result in high molecular weight fragments of 10-50 kbp that conta in both 3'- and 5'-OH end groups, suggesting that more than one endonu clease is involved. This lesioning of DNA is followed by the appearanc e of the damage-response indicator Gadd45 in the ischemic hemisphere f ollowing middle cerebral artery occlusion. By 6 h, gadd45 mRNA was sho wn to increase by semi-quantitative reverse transcriptase-polymerase c hain reaction. In situ hybridization histochemistry indicated that the se increases in gadd45 mRNA occurred in pyramidal neurons located on t he edge of the infarcted cortex. Gadd45 immunostaining yielded similar findings with maximal protein staining detected at 18 h after occlusi on. In neurons, in the infarct core with frank DNA fragmentation shown by in situ TdT-mediated dUTP-biotin nick end labeling (TUNEL) at 24 h , the Gadd45 immunostaining was not visible. Taken together, these fin dings suggest that Gadd45 responds to DNA damage following ischemia as part of a repair response mounted by brain cells attempting to surviv e the insult.