高压氧对大鼠脑缺血再灌注损伤后一氧化氮合酶阳性细胞分布和形态的影响

背景:一氧化氮在脑缺血损伤中起着很重要的作用,而高压氧能改善缺血再灌注引起的神经损伤,高压氧的这种作用与一氧化氮是否有关联?其机制有待探讨。目的:观察一氧化氮合酶阳性细胞在大鼠急性局灶性脑缺血再灌注损伤和高压氧治疗后表达的变化。设计:随机对照动物实验。单位:解放军第四军医大学唐都医院急诊科;西安高新医院检验中心;解放军空军总医院。材料:取健康SD雄性大鼠66只,随机分为5组:假手术组5只,假手术 高压氧组5只,模型组28只,模型 高压氧组28只,后2组又分缺血后5,12,24,72h4个时间点,每个时间点7只。方法:①造模:模型组和模型 高压氧组大鼠参照Koizum方法制备大脑中动脉缺血模型,并于插入栓子造成缺血1h后抽出栓子。其他2组手术,但不插入栓子。②高压氧治疗:假手术 高压氧组和模型 高压氧组大鼠分别在缺血后2,9,21,45,69h共5次将动物置于高压氧舱内,给予高压氧(0.25MPa绝对压)治疗1h。主要观察指标:各组于相应时间点处死取脑,黄递酶-NADPH组织化学方法观察一氧化氮合酶阳性细胞在视交叉平面梗死区皮质、视前区、纹状体外侧区和纹状体内侧区域分布及形态的变化。结果:经补充后66只大鼠进入结果分析。①缺血后一氧化氮合酶阳性细胞发生形态改变,主要变化为突起减少或消失,细胞由椭圆形、三角形变成圆形,细胞皱缩,胞体着色重,胞核和胞浆均染成深蓝色;形态改变的一氧化氮合酶阳性细胞在纹状体外侧区最多,其次是视前区和纹状体内侧区,而皮质区较少。假手术组和假手术 高压氧组未见有形态改变的一氧化氮合酶阳性细胞。②模型组脑内形态有改变的一氧化氮合酶阳性细胞表达随缺血再灌注时间延长而增多,模型 高压氧组各时间点在皮质、视前区和纹状体内侧区其表达均比模型组少,但都于缺血后72h至高峰[皮质:(15.46±3.02),(30.52±4.73)个/视野;视前区:(28.56±4.05),(68.81±7.84)个/视野;纹状体内侧区:(21.09±3.83),(45.71±5.24)个/视野;P均<0.01]。结论:高压氧可明显抑制大鼠急性局灶性脑缺血再灌注损伤区一氧化氮合酶阳性细胞的变性,部位主要在皮质、视前区和纹状体内侧区。

Hyperbaric oxygen for nitric oxide synthase-positive neurons of rats following cerebral ischemia/reperfusion injury

BACKGROUND: Nitric oxide (NO) plays an important role in the ischemic brain injury, and hyperbaric oxygen (HBO) can improve ischemia/reperfusion (I/R)-caused nerve injury. Whether the effect of HBO is associated with NO? Its mechanism needs to be further investigated.OBJECTIVE: To observe the changes of expression of nitric oxide synthase (NOS)-positive neurons of rats following acute focal cerebral I/R injury and HBO treatment.DESIGN: Randomized controlled animal experiment.SETTING: Department of Emergency, Tangdu Hospital, Fourth Military Medical University of Chinese PLA; Department of Laboratory Medicine, Xi'an Gaoxin Hospital; The General Hospital of the Air Force of Chinese PLA.MATERIALS : Sixty-six healthy male Sprague-Dawley rats were chosen and randomized into 5 groups: sham-operation group (n =5), sham-operation +HBO treatment group (n =5), model group (n =28), modeling +HBO treatment group (n =28). Ischemia 5,12, 24 and 72 hours four time points were set in the later 2 groups, 7 rats at each time point.METHODS: ①Rats in the model group and modeling+ HBO treatment group were created into models of middle cerebral artery ischemia according to the method from Koizum. Then, an embolus was inserted for ischemia; One hour later, the embolus was drawn out. Inserting embolus was omitted in the other two groups.②Rats in the sham operation + HBO treatment group and modeling + HBO treatment group were placed in HBO chamber at ischemia 2, 9, 21, 45 and 69 hours, separately, and given HBO treatment for 1 hour (0.25 MPa absolute pressure).MAIN OUTCOME MEASURES: The rats in each group were sacrificed at corresponding time points, and their brains were harvested. The distribution and morphology of NOS positive cells in cortical area, preoptic area, lateral and medial corpora striata of infarct region at the level of optic chiasma were observed with nicotinamide-adenine dinucleotide phosphate -diaphorase (NADPH-d) histochemical method.RESULTS: After supplement, 66 rats were involved in the final analysis. ①After ischemia, NOS-positive neurons changed in morphology, mainly presenting prominences were reduced or disappeared, neurons changed from ellipse or triangle into global shape, and shrank; Body of neuron darkly dyed; Both nucleus and cytoplasm were deeply dyed into dark blue; NOS-positive neurons with changed morphology were mostly in lateral corpora striatum, followed by preoptic area and medial corpora striatum, and those in the cortical area were few. NOS-positive neurons with changed morphology were not found in the sham-operation group and sham-operation + HBO treatment group. ②In the model group, NOS-positive neurons with changed morphology were increased with elongation of I/R time. At each time point, NOS-positive neurons in cortical area, preoptic area and medial corpora striatum in modeling + HBO treatment group were less than those in model group, but NOS-positive neurons in two groups both reached their peaks at ischemia 72 hours [Cortical area: (15.46±3.02) vs.(30.52±4.73)/visual field; Preoptic area:(28.56±4.05) vs. (68.81±7.84)/visual field; medial corpora striatum:(21.09±3.83) vs.(45.71±5.24)/visual field; all P＜0.01].CONCLUSION: HBO obviously inhibits the degeneration of NOS-positive neurons in acute focal cerebral I/R injury regions of rats, such as cortical area, preoptic area, medial corpora striatum, and so on