茶氨酸对脑缺血损伤大鼠自由基代谢的影响及机制研究:缺血再灌注时自由基产生的主要来源

　　【中图分类号】R285 【文献标识码】A 【文章编号】1672－3783(2011)07－0241－01 　　【摘要】目的：探讨茶氨酸对脑缺血损伤的保护作用及机制。方法： 45只SD雄性大鼠，线栓阻塞大鼠大脑中动脉制备局灶性脑缺血模型，观察脑缺血损伤大鼠神经症状变化，测定脑组织含水量、脑组织病理及钙离子含量：观察其血清和脑组织超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量变化以及茶氨酸对上述变化的影响。结果：脑缺血组大鼠的神经症状评分、脑组织含水量及钙离子含量均显著增高，病理损伤明显，正常神经元细胞数目减少；茶氨酸可显著降低神经症状评分、脑组织含水量及钙离子含量，增加正常神经元细胞数目。脑缺血组大鼠的血清、脑组织SOD活性显著降低，荼氨酸组大鼠血清、脑组织SOD活性较脑缺血组显著增高：脑缺血组大鼠的血清、脑组织MDA含量显著增高，茶氨酸组脑组织MDA含量较脑缺血组下降明显。结论：茶氨酸对大鼠脑缺血损伤具有显著的保护作用，可能与调节脑缺血损伤引起的自由基代谢失调有关。
　　
　　Effects of theanine on free radical metabolism in rats with ischemic brain injury
　　SHI Xing-bing
　　【Abstract】Object To explore the mechanism of protective action of theanine in cerebral ischemic injury via detecting changes in free radical metabolism. Methods 45 male SD rats were randomized to sham-operated group ischemia-reperfusion group or theanine group. the rat model of focal cerebral ischemia were constructed by nylon thread occlusion of the middle cerebral artery. The changes of neural symptoms in the rats were observed; the contents of water and Ca2+ and the patholog?cal alterations in the brain tissues were determined;the changes of serum superoxide dismutase (SOD)activityand malondialdehyde (MDA) content and the effects oftheaninewere observed. Results In the ischemia- reperfusion group neurolog?cal deficit scores and the contents of water and Ca2 + were significantly increased, pathologic injure was obvious and the number of normal neurons was reduced . However theanine significantly decreased the scores and the contents of water and Ca2+ and increased the number of normal neurons, The SODactivities in both serum and brain tissues were markedly declined in the ischemia-reperfusiongroup but significantly increased in thetheanine group as compared to the ischemia-reperfusion group. The MDA contents in both serum and brain tssues were notably elevated in the ischemia-reperfusion group while signjficantly decreased in the theanine group as compared to the ischemia-reperfusion group.Conclions Theanine has significant protective effect on cerebral ischemic injury in rats which may be associated withregulatory effect on the imbalance of free radical metabolism due to ischemic injury.
　　【Key words】 Brain ischemiaReperfusionTheanine Free radicals
　　
　　近年来有文献［1］报道茶氨酸对脑缺血再灌注损伤有保护作用，本实验采用大鼠局灶性缺血再灌注模型，观察茶氨酸对脑缺血损伤的作用，并从对自由基代谢影响方面探讨其机制。
　　1 材料与方法
　　1．1材料选用SD雄性大鼠。SPF级，3月龄，30只，体重(300 4-20)g，由湖北省实验动物中心提供。主要试剂：茶氨酸购自杭州浙江大学泛科化工有限公司，粉剂由广州军区武汉总医院制剂室配制为25％的水溶液，置4℃冰箱备用，美国产Rabbit NSE试剂盒由上海天呈科技公司提供。
　　1．2 动物分组按随机数字表法将大鼠分为假手术组、脑缺血组和茶氨酸组，每组15只大鼠。假手术组手术进行到分离血管为止，不结扎血管；茶氨酸组于再灌注开始时将25％的茶氨酸溶液2 mL/kg耳缘静脉注射：脑缺血组不作特殊药物处理。实验过程中保持大鼠肛温(37．0±0．5)℃、室温(26±1)℃。采用改进的线栓法制备大脑中动脉闭塞局灶性脑缺血模型［2］。
　　1．3 观察指标及测定方法(1)神经症状评分：按徐淑云等［3］药理实验方法进行神经症状评分，记录分值。(2)超氧化物歧化酶(SOD)活性：采用黄嘌呤氧化酶法，单位为NU/mL(血清)或NU/mg蛋白(脑组织)。(3)丙二醛(MDA)含量：采用比色法，单位NU/mE(血清)或NU/mg蛋白(脑组织)。(4)脑组织含水量：切取的脑组织用电子天平称重后放入(100±2)℃干燥箱中干燥24 h。再称重。脑组织含水量=(脑湿重一脑干重)/脑湿重×100％。(5)光镜观察：切取脑组织用100 g/L甲醛周定，常规脱水．包埋，切片，经苏木精一伊红染色．光镜下观察神经细胞的损伤情况以及每视野下正常神经元细胞数目，每组每只大鼠分别观察10个视野。计算每视野正常神经元细胞个数。取其均值。(6)脑组织ca2+含量：用电感耦合等离子体发射光谱仪测定Ca2+含量，单位以Pg/g表示。主要结局观察指标：(1)神经症状评分；(2)脑组织含水量；(3)脑组织病理光镜观察神经细胞的损伤情况以及正常神经元细胞数目。
　　1．4 统计学分析数据用SPSS 17．0统计分析软件由笔者处理。计量资料均数±标准差表示。用单因素方差分析检测组间有无统计学差异。P＜0.05表示组间差异有统计学意义。
　　2 结果
　　2．1实验动物数量分析共纳人大鼠45只．因造模不成功或死亡的共有6只，进入结果分析的共有39只。
　　2．2 神经症状评分和脑组织含水量脑缺血组大鼠的神经症状评分较假手术组显著增高(P