| A large number of free radicals are produced in a burst-like manner at the early stage of cerebral ischemia/reperfusion and mainly cause oxidative DNA damage. Neuronal DNA damage caused by oxidative stress consistes of the lesion of base, DNA single strand breaks (SSB) and DNA double strand breaks (DSB). Oxidative DNA damage after ischemia / reperfusion can be repaired by DNA repair system, but continuous or severe DNA damage against DNA repair could induce neuronal death, such as necrosis or apoptosis. It has been reported that early decrease of DNA repair enzyme probably has correlation with subsequent DNA-damaged cell death after cerebral ischemia / reperfusion. As neuronal death was the basis of brain injury, researches on DNA damage and repair would probably reveal the mechanism of ischemic neuronal death, and will bring a new strategy for treating ischemic cerebrovascular diseases. It has been proved that both ischemic preconditioning (IPC) and radix salviae miltiorrhizae (RSM) have significantneuroprotection against ischemic injury in the brain. A number of studies have been performed to clarify the protective effect and mechanism, however, whether IPC and RSM have any effects on the DNA damage and repair in the neuropathological process following cerebral ischemia/reperfusion still remains to be investigated.In this study, we made the model of rat middle cerebral artery occlusion/ reperfusion and used H&E stain method, immuno-histochemical technology and TUNEL detection, in order to explore the relationship between the expression of Ku70 protein-which is critical to the repair of DNA DSB and neuronal death after focal cerebral ischemia/reperfusion, and the possible molecular mechanisms by which IPC and RSM protect brain. Firstly, we observed the expression of Ku70 protein and neuronal apoptosis in the ischemic cortex and caudatum after MCAO 2h reperfusion 0.5h, 2h, 6h, 12h, 24h, 48h, and evaluated the temporal relationship between Ku70 protein and neuronal apoptosis. Then we investigated the influence of IPC and RSM on the expression of Ku70 protein after focal cerebral ischemia/reperfusion.Our results showed:(1) HE stain indicated the model of rat MCAO/R was credible, and the damaged regions included cortex and caudate putamen.(2) Immunohistochemical method showed the nuclear expression of Ku70 protein in the nomal control group, sham operation group and controlateral nonischemic hemisphere. The expression of Ku70 started to decrease at 0.5h of reperfusion in the caudate putamen and at 2h in the cortex (P<0.05), The number of Ku70 positive cells was decreased remarkably in the entire damaged area as early as 6hours after reperfusion and remained reduced until 48 hours. IPC (MCAO 20min) or RSM (15g/kg i.p.) significantly suppressed the early reduction of Ku70 expression in ischemic cortex and caudate putamen, compared with controls, the number of Ku70 positive cells was significantly higher at different reperfusion time (P<0.01).(3) There were no TUNEL positive cells in the nomal control group, sham operation group and controlateral nonischemic hemisphere. TUNEL positive cells begin to rise from 6h after reperfusion, reached peak at 24h and 48h (P<0.01). IPC (MCAO 20min) or RSM (15g/kg i.p.) reduced significantly TUNEL positive cells in ischemic cortex and caudate putamen at different reperfusion time after MCAO 2h (P<0.01).(4) The number of Ku 70 positive cells was decreased remarkably in the entire damaged area as early as 6 hours after ischemia/reperfusion and neuronal apoptosis occurred after the reduction of Ku expression.Our results demonstrate:(1) Expression of DNA repair protein Ku70 decreased remarkably in the damaged regions at early stage of cerebral ischemia/reperfusion, which results in DNA DSB unrepaired or incompletely repaired, It may involve the mechanism of subsequent neuronal apoptosis.(2) The neuroprotective mechanism of IPC is associated with upregulating expression or activity of DNA repair...
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