低温对沙土鼠脑缺血再灌注期间海马CA1区微管运动蛋白活性的影响

Effects of hypothermia on the activity of microtubule based motor proteins in hippocampus CA1 sector in gerbils during cerebral ischemia/reperfusion

目的 观察脑缺血再灌注期间海马CA1区锥体细胞微管运动蛋白Kinesin活性的变化及低温对其的影响 ,以探讨其活性的改变与延迟性神经元死亡的关系和低温脑保护作用的机制。方法 沙土鼠前脑缺血再灌注模型 ,脑缺血时间为 10min。应用免疫组织化学染色方法结合计算机图象分析技术测定海马微管运动蛋白Kinesin的活性 ,应用组织学检查进行神经元计数。结果 在常温缺血再灌注 6h、48h和 96h ,海马CA1区微管运动蛋白Kinesin的活性分别降至假手术组的 49%、32 %和 12 % (P <0 0 1) ,在缺血 96h出现大量的神经元死亡 ,正常神经元数目仅为假手术组的 5 % ,而低温组在再灌注 6h、48h和 96h后 ,微管运动蛋白Kinesin活性及神经元计数均明显高于常温组。

Objective To investigate the effect of hypothermia on the activity of kinesin ( a microtubule based motor proteins) in the hippocampus CA1 sector in gerbils during cerebral ischemia/reperfusion,and to explore the relationship between the change in kinesin and delayed neuronal death(DND).Methods Gerbils weighing 50-60g were anesthetized with intraabdominal pentobarbital sodium 40 mg/kg.Bilateral carotid arteries were dissected and isolated and temporarily clamped for 10 min and then released for reperfusion.The study was divided into 3 groups.In group Ⅰ (sham operation) bilateral carotid arteries were dissected and isolated but not occluded.Group Ⅱ (normothermia) and group Ⅲ (hypothermia) were further divided into 3 subgroups (n=10) according to the duration of reperfusion: 6h,48h and 96h reperfusion.In normothermia group brain temperature was maintained at 37℃±0.2℃.In hypothermia group brain temperature was maintained at 37℃±0.2℃ during cerebral ischemia ,15 min after reperfusion being started brain temperature was rapidly reduced to 32℃±0.2℃ and maintained for 6h and then returned 37℃±0.2℃.The activity of kinesin in the hippocampus CA1 sector was determined after 6h, 48h and 96h reperfusion by immunohistochemical staining with computer image analysis system.DND was assayed by histological examination.Results The activity of kinesin in the hippocampus CA1 sector in normothermia groups decreased significantly after 6h, 48h and 96h reperfusion and was about 49%, 32% and 12% of that in sham-operation group.However hypothermia obviously mitigated the decreament of the activity of kinesin in the hippocampus CA1 sector.The number of neurons in hippocampus CA1 sector after 96h reperfusion in normathermia group was about 5% of that in sham-operation group, whereas in hypothermia group after 96h reperfusion the number of neurons was higher than that in normothermia group.Conclusions Hypothermia significantly reduces delayed neuronal death after celebral ischemia.It may result from the improved activity of kinesin after cerebral ischemia-reperfusion by hypothermia.