Therapeutic strategy using DNA repair enzyme after ischemic stroke in mice

Abstract

[한글]

[영문]Previous investigations demonstrated a nice correlation between the level of the DNA base excision repair protein, apurinic/apyrimidinic endonuclease/redox factor-1 (APE/Ref-1), and the degree of oxidative DNA damage. However, there has been no attempt undertaken yet to directly address the neuroprotective effects of modulating DNA repair functions in an in vivo model of cerebral ischemia.This study was performed to test the hypothesis that the augmentation of APE/Ref-1 is an effective neuroprotective measure in the ischemic neuronal death and the role of APE/Ref-1 on neuroprotection is mainly related to its DNA-repairing function. I also investigated the temporal relationship of the therapeutic strategy using the DNA repairing mechanisms to the postischemic state in a model of cerebral Ischemia/Reperfusion (I/R)Mice were subjected to intraluminal suture occlusion of the middle cerebral artery for 1 hour followed by reperfusion. Pre-ischemic treatment of the adenoviral vector harboring an entire APE/Ref-1 gene sequence and post-ischemic treatment of the small peptide containing only the APE functional domain were introduced intracerebroventricularly. Western blot analysis and immunohistochemistry were performed. Assays in AP sites, single-strand DNA breaks, caspase-3 activity, and apoptotic cell death were performed and quantified.The reduction of APE/Ref-1 occurred before DNA fragmentation after I/R, which was shown by the temporal and spatial analysis of their correlations. Overexpression of APE/Ref-1 significantly decreased oxidative DNA damages and the volume of cerebral infarction after I/R. I found that post-ischemic administration of the APE peptide up to 4 hours after I/R significantly inhibits the induction of apoptotic DNA fragmentation and subsequent infarct volume at 24 hours after I/R.In conclusion, augmentation or restoration of decreased APE/Ref-1 exerted significant protective effects on ischemic stroke in both pre- and post-ischemic treatment, which was encouraging for the clinical development of APE peptide as a novel therapeutic strategy.