| Background and purpose: Large amount of free radicals are produced in a burst-like manner after cerebral ischemia reperfusion and mainly cause neuronal DNA damage. Neuronal DNA damage caused by oxidative stress especially the DNA single strand breaks and DNA double strand breaks if unrepaired will lead to neuronal death, oxygen free radicals produced after cerebral ischemia and reperfusion will attack neuronal DNA base and produce 8-ohdg which is regarded as the best marker of DNA oxidative damage. 8-ohdg can also reflect the degree of oxidative stress. Although the oxidative DNA damage is regarded as a important mechanism in cerebral ischemia-reperfusion, there are some problems that are still not completely known:How about the change of neuronal DNA single and double stand breaks and the result of DNA-damaged neurons after cerebral ischemia -reperfusion?DNA-PK is critical to the repair of DNA dsb,what is the change of Ku as the regulative subunit of DNA-PK?In order to further study these problems, We have observed the change of neuronal DNA single and double strand breaks, Ku70 and 8-ohdg in ischemic area after cerebral ischemia and reperfusion. The DNA ladder is also used to judge the outcome of DNA-damaged neuron. The relationships between oxidative stress and neuronal DNA strand breaks, Ku70 expression and DNA double strand breaks have been explored so as to provide a theoretical basis for clinical therapy of cerebral ischemia.Methods: Male Wistar rats (weighing 240-330g) were randomlydivided into 2 groups: false operation group (Group FO) and ischemia-reperfusion group (Group IR) in which the rats were subjected to 2h of middle cerebral artery occlusion (MCAO) following 1, 6, 12, 24, 48 and 72h of reperfusion. DNA single strand breaks, DNA double strand breaks, Ku70 expression and 8-ohdg, cell apoptosis were examined by Single-cell gel eiectrophoresis (SCGE), TUNEL method, immnunohistochemistry, gel eiectrophoresis method respectively. The relationships between oxidative stress and neuronal DNA strand breaks, Ku70 expression and DNA double strand breaks were analyzed by linear analysis method.Results: 1.8-ohdg positive neurons in Group IR marketly increased at1h-12h after reperfusion, peaked at 12h after reperfusion and began to decreaseat 24h-72h after reperfusion. There are some 8-ohdg positive neurons in theouter area of the ischemic zone at 24h-72h after reperfusion.2. DNA single strand breaks in ischemic brain issue began to increase at1h after reperfusion, peaked at 12h after reperfusion and began to decrease at24h-72h after reperfusion.3.TUNEL positive cells began to rise from 6h after reperfusion, reachedpeak at 24h and became rehabilitated gradually after 72h in cortex and basalganglia of ischemic brain tissue.4.Ku70 positive cells in cortex and basal ganglia decreased at 1h afterreperfusion, reduced to the lowest level at 24h and keeped in this level aboutlyat 24h-72h after reperfusion.5. There were no DNA ladder which represent cell apotosis at 1h-12h after reperfusion. DNA ladder began to appear at 24h-72h after reperfusion .6. According to the statistics, a significantly positive correlation existed between neuronal DNA single strand breaks in basal ganglia and 8-ohdg, a significantly negative correlation existed between neuronal DNA double strandbreaks and Ku70 expression(P<0.05).Conclusions: 1 .Neuronal DNA damage occurs in the early stage after reperfusion due to the oxidative stress, neuronal DNA single strand breaks occurs earlier than neuronal DNA double strand breaks. The neuronal DNA in basal ganglia is more easy damaged than that in cortex.2. Overwhelming and continously DNA damage exceeds the ability of neuron to repair or the repairing systems are destroyed by the reperfusion injury lead to the occurring of unrecovered DNA damage,which result in neuron death.3.The reduction of Ku after reperfusion may be the main reason that the DNA double strand breaks cannot be repaired in the ischemic region.
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