脑缺血预处理降低大鼠脑一氧化氮水平抗脑缺血损伤的实验研究

目的:探讨脑缺血预处理对大鼠缺血性脑损伤的保护作用及其与一氧化氮(NO)的关系。方法:缺血预处理组大鼠钳夹阻断两侧颈总动脉15min;3天后,线栓法制作大脑中动脉阻塞模型,线栓阻塞右大脑中动脉8h。非缺血预处理组除不钳夹阻断两侧颈总动脉外,其余步骤相同。右大脑中动脉阻塞8h后,对大鼠进行神经功能缺失评分及亚硝酸盐还原法测定大脑皮层及海马的NO含量。结果:(1)缺血预处理组大鼠神经功能缺失评分较非缺血预处理组有显著改善(P＜0．01);(2)缺血预处理组大鼠缺血侧和自身对照侧皮层、海马的NO含量较非缺血预处理组大鼠明显下降(P＜0．05)。结论:大鼠脑缺血预处理对随后的脑缺血损伤具有保护作用,其机制之一可能是降低了脑组织的NO水平。     

局灶预缺血诱导脑缺血耐受的动物模型

目的　建立一种简便可靠的 SD大鼠局灶性脑缺血预处理模型。方法　将大鼠随机分为 3组 ,分别给予 10 m in大脑中动脉缺血 ( MCAO)预处理 ( PC) ;10 min PC后 2 h MCAO( PC MCAO)及假手术 ( SS)后 2 hMCAO( SS MCAO) ,再灌注 2 2 h后处死 ,观察各组神经功能缺损、梗死体积及脑含水量变化。结果　 PC MCAO组神经功能评分、梗死体积及含水量均明显低于 SS MCAO组 ,PC组无神经功能缺损及梗死灶形成。结论　 2次线栓法建立的大鼠局灶脑缺血预处理模型 ,能有效减轻 MCAO所致的神经损伤 ,操作简便 ,稳定性好 ,是一种研究局灶脑缺血耐受的有用工具。     

Induction of 27-kDa heat shock protein following cerebral ischemia in a rat model of ischemic tolerance

Preconditioning the brain with sublethal cerebral ischemia induces tolerance to subsequent lethal periods of ischemia (ischemic tolerance). The purpose of this study is to investigate the role of low-molecular weight stress proteins, 27-kDa heat shock protein (HSP27) and αB crystallin, in ischemic tolerance. We measured the content of these proteins with enzyme immunoassay in the rat hippocampus and cerebral cortex following 6 min of ischemia with and without preconditioning with 3 min of ischemia and 3 days of reperfusion. We also visualized the localization of HSP27 immunohistochemically in comparison with that of HSP70. A 3-min period of ischemia caused a 2.4-fold increase in HSP27 content in the hippocampus after 3 days. Immunohistochemical localization of HSP27 was found in glial cells in all subregions of the hippocampus, whereas HSP70 immunostaining was seen only in CA1 pyramidal neurons. HSP27 content in the hippocampus decreased 2 h after 6 min of ischemia. HSP27 content progressively increased in the unpreconditioned hippocampus after 1 and 3 days, but returned to preischemic levels in the preconditioned hippocampus. HSP27 and HSP70 immunostaining was seen in CA1 pyramidal neurons after 1 day both with and without preconditioning. After 3 and 7 days, an intense HSP27 staining was observed in reactive glial cells in the CA1 without preconditioning, whereas the staining decreased in the preconditioned hippocampus. HSP70 staining was seen only in neurons at these time points. We observed no significant changes in HSP27 content in the cerebral cortex although neurons in the third and fifth layers were immunostained after 1 and 3 days. We observed no alterations in αB crystallin content after ischemia both in the hippocampus and the cortex. The present study demonstrated that cerebral ischemia induces HSP27 expression but not αB crystallin. Both HSP27 and HSP70 induction had a good temporal correlation with the induction of ischemic tolerance. However, different sites of action were suggested because the localization and cell types of HSP27 induction were quite different from those of HSP70 induction. The result suggests that it is unlikely that HSP27 is directly involved in the protection afforded by ischemic preconditioning.     

Neuronal damage in the rat hippocampus in a new model of repeated reversible transient cerebral ischemia

The influence of the interval of the repeated reversible transient cerebral ischemia on the neuronal damage in the hippocampal CA1 sector was investigated in the rats using a 4-vessel occlusion (4-VO) model. A single 3-min 4-VO did not produce any significant neuronal damage in the hippocampal CA1 sector, whereas the rats subjected to three 3-min 4-VO at 1-h intervals revealed a very severe neuronal damage which was much more severe than that in the rats subjected to a single 9-min 4-VO. In contrast, the rats subjected to three 3-min 4-VO at 6-h intervals revealed only a mild neuronal damage. The degree of the neuronal damage in the rats subjected to three 3-min 4-VO at 5-min intervals was similar to that in the rats subjected to a single 9-min 4-VO. The present study indicates that even such a brief, non-lethal ischemia as 3-min 4-VO can produce a severe neuronal damage if it occurs repeatedly at 1-h intervals.     

兔MCAO后脑组织NOS活性的变化

一氧化氮（ＮＯ）与脑缺血关系密切，对缺血性脑损害可能有直接的影响，一氧化氮的合成酶（ＮＯＳ）是ＮＯ生物合成的限速酶，本文在建立兔ＭＣＡＯ局灶脑缺血模型基础上，测定缺血后不同时间缺血区和正常脑组织的ＮＯＳ活性。结果证实缺血后早期（ＭＣＡＯ后１ｈ内）ＮＯＳ活性突然升高。     

不同脑缺血和再灌流过程中大鼠脑组织NO含量的动态变化

采用线栓法制成大鼠大脑中动脉梗塞 ( MCAO)模型 ,依 Hb O2 - NO法测定持续性脑缺血和缺血 /再灌流脑组织内 NO含量的变化 ,以探讨不同脑缺血和再灌流过程中 NO的变化规律及其意义。结果 :缺血 3小时受损脑组织 NO水平即增高 ,再灌流后 NO逐步升高 ,而持续性缺血状态下 NO则表现降低后再升高的变化。虽然两组 NO在 7天时均有明显降低 ,但仍高于缺血前水平。认为持续性脑缺血和缺血 /再灌流情况下 NO的变化规律有所不同 ,与缺血脑组织的缺氧及产生 NO所需底物供应缺乏有关 ,且可能与脑组织的损害密切相关     

脑缺血早期大脑皮质区神经元内一氧化氮合酶活性的变化

建立大鼠ＭＣＡＯ局灶脑缺血模型,利用ＮＡＤＰＨ－ｄ组织化学方法检测脑缺血早期大脑皮质缺血区神经元内一氧化氮合酶活性的变化。结果显示脑缺血早期３０ｍｉｎ神经元内一氧化氮合酶活性开始至高峰,随后下降,脑缺血后６０ｍｉｎ,降至正常。     

大鼠脑缺血再灌流后海马区一氧化氮合酶活性的变化

目的 :观察鼠全脑缺血再灌流后海马区NOS活性的变化。方法 :采用大鼠 4血管关闭方法制作全脑缺血再灌流模型。实验动物分为假手术组、缺血 10min组、再灌注 1、2、3d组 ,测定脑缺血再灌流后海马区NOS活性的变化。结果 :全脑缺血再灌注后海马组织NOS活性被激活上调。结论 :NO可能参与了海马CA1区迟发性神经元死亡 (DND)的发生。     

大鼠脑缺血时脑组织中NOS1阳性神经元变化的连续观察

目的探讨脑缺血时含神经型一氧化氮合成酶（ＮＯＳ）神经元变化及一氧化氮（ＮＯ）的相关作用。方法采用大脑中动脉梗塞模型，对３４只雄性ＳＤ大鼠脑缺血后不同时点ＮＯＳ１免疫组化、ＮＡＤＰＨ－ｄ染色及其病理学进行了对照研究。结果缺血后２～４８小时，梗塞侧的ＮＯＳ１神经元数量明显高于对侧，４小时达高峰。梗塞后２４～４８小时，神经元出现边界不清、膨胀，而含ＮＯＳ１神经元则保持形态学的完整性。１周时，大部分神经元死亡，但仍有完整的ＮＯＳ１阳性细胞散在。结论含ＮＯＳ１神经元对脑缺血有一定的抵抗能力，可能与ＮＯ的神经保护作用有关     

大鼠急性局灶性脑缺血再灌注脑组织NO含量和NOS活性的变化

目的探讨一氧化氮（ＮＯ）和神经元型ＮＯ合酶（ｎＮＯＳ）是否参与急性局灶性脑缺血再灌注的发病机理。方法采用栓红法建立大鼠大脑中动脉阻塞（ＭＣＡＯ）模型,观察脑组织ＮＯ含量和一氧化氮合酶（ＮＯＳ）活性的变化及ｎＮＯＳ抑制剂７－硝基吲唑（７－ＮＩ）对再灌注期两者的影响。结果缺血３０分种ＮＯ含量和ＮＯＳ活性显著升高,缺血３小进两者下降;再灌注３０分种ＮＯＴ和ＮＯＳ再次升高,而再灌注３小时两者又下降。７－Ｎ     

NOS活性在局灶脑缺血后的时相变化

一氧化氮(NO)在脑缺血中起重要作用,一氧化氮合成酶(NOS)作为NO合成的关键酶,其活性变化直接调节NO的生成量及生物学效应,本文在建立兔MCAO局灶脑缺血模型基础上,测定缺血不同时间缺血区和正常脑组织的NOS活性。结果显示脑缺血早期(1小时内)NOS活性突然升高,随之下降;脑缺血后24小时NOS活性又回升,48小时、96小时明显升高。     

诱导脑缺血耐受影响脑组织抗氧化活力

目的建立诱导脑缺血耐受的缺血预处理模型并研究其对脑组织超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-PX)活性的影响.方法动物分别给予假预处理(SHAM)、一侧颈内动脉注射生理盐水(SI)、双侧颈总动脉夹闭(BCAO)和双侧颈总动脉夹闭同时一侧颈内动脉注射生理盐水(BS),24h后腔内线栓法制作大脑中动脉阻断(MCAO)模型.观察MCAO后72h4组脑梗死体积,观察各组预处理24h和MCAO 24h、48h缺血侧脑组织SOD和GSH-PX活力.结果BS 组脑梗死体积明显低于其它3组(P<0.05),预处理后24h BS组缺血侧脑组织SOD和GSH-PX活力明显高于其它组(P<0.01);MCAO24h、48h 4组缺血侧脑组织SOD和GSH-PX活力均明显降低,BS组SOD和GSH-PX活力明显高于其它3组(P<0.01).结论双侧颈总动脉夹闭同时一侧颈内动脉注射生理盐水能诱导缺血耐受,该缺血预处理增高脑组织抗氧化活力,可能是缺血耐受产生的机制之一.     

Protection of rat hippocampus against ischemic neuronal damage by pretreatment with sublethal ischemia

We examined whether preconditioning with sublethal ischemia protects against neuronal damage following subsequent lethal ischemic insults. Forebrain ischemia for 3 min in Wistar rats increased heat shock protein-70 immunoreactivity in the hippocampal CA1 subfield but produced no neuronal damage. Preconditioning with 3 min of ischemia followed by 3 days of reperfusion protected against hippocampal CA1 neuronal damage following 6 and 8 min of ischemia but not damage after 10 min of ischemia. The result strongly suggests that stress response induced by sublethal ischemia protects against ischemic brain damage.     

依达拉奉对脑缺血再灌注大鼠海马一氧化氮的影响

目的研究依达拉奉对脑缺血再灌注大鼠海马一氧化氮(NO)产生的影响。方法大鼠脑缺血采用四血管阻断法,选择性测定电极检测的浓度。实验分为生理盐水组、依达拉奉组(Edaravone)和7-Nitroindazole(7-NI)组。结果依达拉奉和7-NI皆未影响大鼠的血压和海马血流量,均显著减少了缺血再灌注时海马内NO的产生(均P<0.001)。结论依达拉奉可能通过抑制神经型一氧化氮合酶(nNOS)或减少NO而起到神经保护作用。     

大鼠局灶缺血预处理诱导的脑缺血耐受中热休克蛋白70的表达及其意义

目的　研究局灶脑缺血预处理对热休克蛋白 70 (HSP70 )表达和脑缺血耐受的影响。方法　SD大鼠随机分为 3组 :预缺血组、假手术组及对照组 ,前两组分别在 2小时大脑中动脉缺血 (MCAO)前 3天给予10分钟的预缺血或假手术 ,MCAO后 2 4小时处死 ,对照组给予两次相隔 3天的假手术 ,比较各组梗死体积及HSP70的表达。结果　预缺血组梗死体积较假手术组减少 5 2 5 4 % (P <0 0 1) ,HSP70表达高于假手术组及对照组 (P <0 0 1)。结论　 10分钟大脑中动脉预缺血可有效诱导缺血耐受 ,增加HSP70表达。HSP70表达上调可能是局灶性脑缺血耐受产生的分子机制之一     

局灶性脑缺血时脑组织中nNOS对神经元的作用研究

探讨在局灶性脑缺血时脑组织中一氧化氮合成酶（ｎＮＯＳ）对神经元的作用。方法建立大鼠大脑中动脉栓塞的动物模型,在不同的时间点进行相邻切片的ｎＮＯＳ免疫组化及ＨＥ染色,并将它们进行比较以估计神经元的不同受损情况。结果缺血开始后１～４８ｈ,栓塞侧皮质和纹状体中ｎＮＯＳ阳性神经元数量明显高于对侧,第８ｈ达到最高值。栓塞后２４ｈ,神经元出现边界不清、膨胀,但这些细胞在ｎＮＯＳ免疫组化染色的切片上保持形态的完整。７２ｈ后,在ＨＥ染色的切片上所有的神经元都已死亡,但在ｎＮＯＳ免疫组化染色的切片上仍存在一些形态完整的ｎＮＯＳ阳性细胞。结论在大脑缺血时,ｎＮＯＳ阳性神经元具有抵抗缺血缺氧的功能。     

Ischemic tolerance to memory impairment associated with hippocampal neuronal damage after transient cerebral ischemia in rats

When rats were trained preoperatively with a three-panel runway task and were then exposed to 10-min ischemia by the method of 4-vessel occlusion, they showed no increase in the number of errors (attempts to pass through two incorrect panels of the three panel-gates at four choice points), having normal retention of memory performance learned before the ischemic insult. Next, we investigated the abilities of ischemic rats to acquire the three-panel runway task and to learn a subsequent reversal task, where the correct panel-gate locations were changed. Rats with 5-min ischemia exhibited performance as good as that of control rats, but rats exposed to 10- and 20-min ischemia showed more errors than control rats during 10 acquisition sessions and 5 subsequent reversal sessions, each of which (consisting of 6 trials) was given once a day. Marked neuronal degeneration was observed in the hippocampal CA1 sector from the rats with 10- and 20-min ischemia. Exposure to sublethal 5-min ischemia followed by 10-min ischemia at a 2-h interval had no effect on either the memory impairment during acquisition and reversal tests or the hippocampal CA1 damage. When rats were exposed to 5-min ischemia 2 days before lethal 10-min ischemia, they showed acquisition and subsequent reversal learning as good as that of control rats. Preconditioning with sublethal 5-min ischemia followed by 2 days of reperfusion also prevented the neuronal destruction of the hippocampal CA1 sector induced by 10-min ischemia. These findings suggest that postischemic hippocampal CA1 neuronal damage does not affect retention of spatial memory acquired before ischemia, but produces a significant impairment of acquisition and subsequent reversal learning. The present results also demonstrate that preconditioning with sublethal ischemia can develop tolerance to subsequent lethal ischemia to prevent the learning impairment related to the hippocampal CA1 neuronal damage.     

亚低温对大鼠局灶性脑缺血再灌注后不同脑区iNOS表达的影响

目的 探讨亚低温对大鼠局灶性脑缺血再灌注后不同脑区诱导型一氧化氮合酶(iNOS)表达的影响.方法 雄性SD大鼠,随机分为假手术组、常温缺血组和亚低温组.采用线栓法制作大脑中动脉闭塞再灌注模型,于缺血后48h,观察不同组间组织形态学变化,检测不同脑区iNOS蛋白表达、iNOS活性和产物NO含量.结果 常温缺血后48h,纹状体和皮质均检测到iNOS活性升高和免疫阳性反应,且皮质缺血半暗带区iNOS免疫反应明显强于纹状体和皮质缺血核心区.亚低温明显缩小梗死面积,抑制皮质和纹状体iNOS活性,明显下调半暗带区iNOS蛋白表达,减少NO产生.结论亚低温可能通过减少半暗带区iNOS蛋白表达,抑制iNOS活性,减少NO产生而起到脑保护作用.     

低氧诱导因子-1α表达上调在大鼠局灶性脑缺血耐受中的意义

目的:观察局灶脑缺血预处理对低氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)表达的影响,探讨其在脑缺血耐受中的意义。方法:SD大鼠随机分为预缺血组、假手术组及对照组,前两组分别在2h大脑中动脉缺血(MCAO)前3d给予10min的预缺血或假手术,MCAO后24h处死,对照组给予两次相隔3d的假手术,比较各组梗死体积及HIF-1α的表达。结果:对照组未见HIF-1α表达,预缺血组HIF-1α表达高于假手术组,梗死体积较假手术组减少53.15％(P<0.01)(P<0.01)。结论:病灶性脑缺血可诱导HIF-1α表达。HIF-1α表达上调可能是脑缺备耐受产生的分子机制之一。