BACKGROUND: Numerous studies have shown that apoptosis, caused by cerebral ischemia/reperfusion injury, is a dynamic evolving process that occurs in a time-dependent fashion. Non-apoptotic cells can result in apoptosis, with a prolonged reperfusion period and an accumulation of DNA damage. Recent studies have indicated that theanine has a protective effect on the brain. Nonetheless, there is no research relating to whether theanine is neuroprotective following brain injury. OBJECTIVE: To investigate the effects of theanine on apoptosis in the cerebral cortex and mRNA expression of the DNA repair protein, XRCC1, and DNA repair enzyme, Ku70, at various time points following global cerebral ischemia/reperfusion in rats. DESIGN, TIME AND SETTING: A randomized, controlled animal study was performed at the Pharmacological Laboratory of Animal Experimental Center of Xi’an Jiaotong University, China, from April to October 2007. MATERIALS: Theanine (Wuhan Remote Technology Co., Ltd., China), XRCC1, Ku70 and GAPDH primers (Shanghai Sangon Biological Engineering Technology & Services Co., Ltd., China) were used in this study. METHODS: A total of 108 healthy, male Sprague Dawley rats were randomly divided into sham operation, ischemia/reperfusion and theanine groups. Each group was further divided into 6 sub- groups, with six rats in each, according to reperfusion time (2, 6, 12, 24, 48 and 72 hours). Rat models of global cerebral ischemia/reperfusion were established by the four-vessel occlusion method. Theanine (1 000 mg/kg) was injected into the caudal vein of rats in the theanine group immediately after surgery. Saline (4 mL/kg) was used in the sham operation and ischemia/reperfusion groups. The same dose was administered every 24 hours. MAIN OUTCOME MEASURES: Flow cytometry was employed to determine apoptosis of cerebral cortical neurons. Real-time reverse transcription-polymerase chain reaction was applied to quantify the expression of XRCC1 and Ku70 mRNA in the cerebral cortex. RESULTS: The apoptotic rate in the ischemia/reperfusion group was significantly greater compared with the sham operation group at various time points (P < 0.05). The apoptotic rate in the theanine group was significantly diminished compared with the ischemia/reperfusion group, but significantly increased compared with the sham operation group at 6, 12, 24, 48 and 72 hours (P < 0.05). Ex- pression of XRCC1 and Ku70 mRNA was significantly decreased in the ischemia/reperfusion group compared with the sham operation group at 2, 6, 12, 24, 48 and 72 hours (P < 0.05). The relative expression of XRCC1 mRNA in the theanine group was significantly increased compared with the ischemia/reperfusion group, but significantly reduced compared with the sham operation group at 6, 12, 24, 48 and 72 hours (P < 0.05). The relative expression of Ku70 mRNA was significantly lower in the theanine group compared with the sham operation group at 2, 6 and 12 hours (P < 0.05) and significantly greater than the ischemia/reperfusion group at 6, 12, 24, 48 and 72 hours (P < 0.05). CONCLUSION: The apoptotic rate in rats following global cerebral ischemia/reperfusion injury reaches a peak at 24 hours after reperfusion; however, the levels of DNA repair protein began to decrease after only 2 hours of reperfusion. Theanine antagonizes the decrease in XRCC1 and Ku70 mRNA expression in the rat cerebral cortex during global cerebral ischemia/reperfusion injury.
