局灶脑缺血脑水肿时强啡肽的含量变化

应用兔大脑中动脉阻断(MCAO)局灶脑缺血模型,将实验动物随机分成4组:对照组C、脑缺血后2h、4h、24h(I_2、I_4、I_(21))组。放射免疫法测定脑组织和血浆中强啡肽(Dynorphin,Dyn)含量的时相变化,同时测脑组织H_2O、Na~+含量。结果发现MCAO后脑组织和血浆Dyn含量显增,且随时相递增,并与脑组织H_2O、Na~+变化呈正相关,提示Dyn可能参与脑缺血、脑水肿的发生发展。     

脑缺血后β—内啡肽样免疫活性物质含量变化

利用兔大脑中动脉阻断(MCAO)模型,用放射免疫法检测脑皮质和血浆β-内啡肽样免疫活性物质的含量,并分别测定脑组织丙二醛(MDA)、Ca~(2 )及H_2O含量。实验动物随机分成3组:对照组(正常和假手术对照)、脑缺血后4h及24h。结果发现脑组织β-内啡肽、MDA、Ca~(2 )和H_2O含量均显著增加(P<0.05),且脑缺血后24h组高于4h组(P<0.05)。同时还发现β-内啡肽与脑组织H_2O、Ca~(2 )、MDA含量之间呈正相关,结果说明脑缺血后β-内啡肽含量的增加,参与了脑缺血脑损害,其机理可能与Ca~(2 )和自由基反应有关。     

NO对兔局灶脑缺血脑水肿和脑梗塞灶的影响

本文在兔MCAO局灶脑缺血模型基础上,利用外源性一氧化氮(NO)前体物质L-精氨酸和NOS抑制剂L-NNA研究NO对脑缺血后脑水肿和脑梗塞的影响。NZW兔42只随机分为7组,每组6只,计对照组、假手术组、MCAO组、MCAO+NS组、MCAO+L-Arg组、MCAO+D-Arg组及MCAO+L-Arg+L-NNA组。结果提示:与单纯MCAO组比较,L-Arg组的脑组织H_2O、Na~+、Ca~(2+)含量明显下降(P<0.01),脑梗塞灶亦明显缩小(P<0.01)。从而显示L-Arg系通过产生NO促使血管舒张而减轻脑水肿和缩小梗塞灶,为临床改进脑缺血的治疗提供依据。     

急性脑缺血后NO与ET代谢的相互关系

在兔大脑中动脉阻断(MCAo)型局灶脑缺血前后分别应用外源性-氧化氮合成酶(NOS)抑制剂L-NNA和一氧化氮(NO)合成底物L-Arginine,观察缺血4h后脑组织内皮素(ET)含量的变化。结果发现L-NNA组较缺血对照组脑组织ET含量明显增加(1262.9±387.6 vs789.3±188.4pg/mg·pro,P<0.01),脑水肿显著;而L-Arginine用药组脑组织ET含量较缺血对照组明显减少(P<0.05),且脑水肿减轻。本研究提示促进NO代谢能抑制缺血脑组织ET的产生。NO影响早期脑缺血可能与此途径有关。     

兔MCAO后脑组织和血浆内皮素的变化

建立兔ＭＣＡＯ脑缺血模型。兔４８只随机分成４组：对照组Ｃ、脑缺血后２ｈ、４ｈ、２４ｈ组。用放免法测兔脑组织和血浆的内皮素含量。发现脑缺血后，缺血侧脑组织和血浆内皮素含量均显著高于对照组（Ｐ＜０．００１），且随缺血后时间的延长而递增（Ｐ＜０．０１）。内皮素与脑水含量之间呈显著正相关（Ｐ＜０．０５）。提示内皮素可能参与缺血脑损害并加重缺血后脑水肿。     

脑缺血后脑细胞线粒体LPO、SOD的变化

我们利用兔MCAO模型,分别测定脑缺血后4h(I_4)、24h(I_(24))脑组织的H_2o、Na~ 、Ca~(2 )、LPO,SOD以及线粒体的LPO,SOD,经与对照组(包括假手术和正常对照)对比,结果发现脑缺血后脑组织H_2o、Na~ 、C_a~(2 )LPO及线粒体LPO均明显增加,而SOD活性变化则相反地降低。结果提示脑缺血后脑细胞线粒体的LPO含量增加,而SOD活性则下降,说明脑缺血后线粒体产生的自由基参与了脑缺血脑损伤机制。     

高血压致缺血性脑卒中的脑保护机制研究

目的 探讨高血压致脑缺血过程中血浆GMP-140、tNF-α的含量与脑细胞损伤的关系以及脑蛋白水解物(元活苏)对缺血性脑卒中的保护作用。方法 将144例急性脑梗死患者随机分为两组,观察组(元活苏+常规药物)72例,对照组(常规药物)72例,观察两组患者治疗前后血浆GMP-140、TNF-α含量及缺损神经功能评分(ESS)的变化。结果 治疗后两组患者血浆GMP-140、TNF-α含量较治疗前均有明显下降(P<0.001);治疗后观察组血浆GMP-140、TNF—α含量较对照组显著下降,(P<0.05);治疗后两组患者缺损神经功能评分(ESS)较治疗前明显提高(P<0.01);而治疗后观察组ESS与对照组比较,有显著性差异(P<0.05)。结论 元活苏可降低血浆GMP-140、TNF-α含量,对高血压所致缺血性脑卒中具有脑保护作用。     

颈内静脉血浆ET、NO变化与脑缺血再灌注损伤相关性研究

目的观察兔全脑缺血再灌注(I/R)期间颈内静脉血浆ET、NO的动态变化及它们与脑I/R损伤的关系。方法取清洁级家兔24只,随机分A、B、C3组,每组8只,缺血15min后分别再灌注30min(A组),2h(B组)和4h(C组),C组在缺血前15min(I0)及再灌注30min(R1)、2h(R2)、4h(R3)4个时点、B组在前3个时点、A组在前2个时点采颈内静脉球部血样分别测定ET、NO及计算ET/NO比值;取左颞皮质作HE、尼氏体染色,光镜观察,对神经元进行图像分析,测量核截面积和尼氏体平均光密度。结果(1)与I0比较,NO在R1时显著增加(P<0.01),在R2、R3时缓慢下降;ET在R1~R3时持续上升,由317.58ng/L升至637.01ng/L;在R1时ET/NO比值与I0比较差异无显著性,在R2、R3时比值显著增大,由44.83×10-6升至224.30×10-6;(2)B、C组的病理损伤比A组严重,C组又比B组严重;B、C组的神经元核截面积比A组显著降低,C组的核截面积最小;尼氏体平均光密度A>B>C组;(3)ET分别与核截面积及尼氏体平均光密度呈显著负相关;ET/NO比值与核截面积及尼氏体平均光密度呈非常显著负相关;结论ET增加及ET与NO之间平衡的破坏可能是脑I/R损伤的主要原因之一;检测颈内静脉血ET和NO的动态变化可以反映脑I/R损伤程度。     

β阻滞剂对急性期实验性脑出血大鼠的神经保护作用

目的测定脑出血Wistar大鼠血肿周围神经细胞内Ca2+浓度、血浆降钙素基因相关肽(CGRP)、内皮素(ET)含量变化及美托洛尔(β-阻滞剂)对其影响,探讨β-阻滞剂对出血后神经细胞的保护机制。方法 Wistar大鼠84只,随机分为正常组、模型组(24h、48h、72h组)、药物治疗组(24h、48h、72h组)。采用大鼠断尾取血注入尾状核建立脑出血模型,模型组造模后即刻给予生理盐水1.5ml灌胃,药物治疗组造模后即刻给予美托洛尔100g/kg灌胃。采用Fura-2/AM荧光探针法测定血肿周围神经细胞内Ca2+浓度、放射免疫方法(RIA)测定各组血浆CGRP及ET含量。结果与正常组比较,模型组血肿周围神经细胞内Ca2+浓度、血浆ET与CGRP含量增高(P0.05);与模型组比较。药物治疗组各时间点血肿周围神经细胞内Ca2+浓度降低(P0.05),药物治疗组24h ET、CGRP含量增高;48、42h ET及CGRP含量低于模型组(P0.05);3d内ET/CGRP比值药物治疗组比模型组偏低。结论 Ca2+、CGRP、ET参与了脑出血的病理生理过程,美托洛尔(β-阻滞剂)可能是对血肿周围神经细胞内Ca2+浓度及血浆CGRP、ET含量调节,使血肿周围神经细胞内Ca2+浓度降低、血浆ET/CGRP比值接近正常,达到脑保护作用。     

普伐他汀对慢性脑缺血大鼠脑海马区早老素-1表达的影响

目的 观察降血脂药普伐他汀对慢性脑缺血(CVI)大鼠脑海马区早老素-1（PS-1）蛋白水平的影响。方法 36只雄性Wistar大鼠随机分为正常对照组(7只),高脂饮食组随机分为空白对照组(6只)、普伐他汀组(7只)、单纯脑缺血组(7只)及脑缺血+普伐他汀组 (9只)。采用永久性结扎双侧颈总动脉建立CVI模型。于术后2个月测血脂,行酶联免疫吸附试验(ELISA)检测血浆及脑左侧海马区匀浆中β淀粉样蛋白(Aβ)水平,蛋白免疫印迹检测海马区PS-1改变。结果 ⑴高脂饮食导致大鼠血浆胆固醇（TC）、三酰甘油(TG)、低密度脂蛋白(LDL-C)显著升高。普伐他汀可使TC降至接近正常水平,LDL-C有不同程度下降,TG无明显改善。⑵各实验组血浆Aβ水平无统计学差异。脑海马区匀浆中,空白对照组、普伐他汀组Aβ轻度升高,而脑缺血组显著升高(P <0.01),普伐他汀可以有效降低脑缺血组Aβ水平(P <0.01)。蛋白免疫印迹结果显示高脂饮食组、空白对照组及脑缺血组PS-1全长蛋白明显上调(P <0.05),而普伐他汀组、脑缺血+普伐他汀组PS-1蛋白可降至接近正常水平。结论 降血脂药物普伐他汀在降低脑缺血后Aβ产生的同时抑制了PS-1的表达上调,可能对阿尔茨海默病有一定的防治作用     

NADPH-oxidase activity is elevated in penumbral and non-ischemic cerebral arteries following stroke

Reactive oxygen species play a role in neuronal damage following cerebral ischemia-reperfusion. We tested whether activity of the superoxide-generating enzyme, NADPH-oxidase, is enhanced in cerebral arteries within, adjacent and distant from the ischemic core. The right middle cerebral artery (MCA) of conscious rats was temporarily occluded by perivascular injection of endothelin-1 to induce stroke (ET-1; n=19). Control rats were injected with saline (n=9). At 24 h or 72 h post-administration of ET-1, the MCA and its branches within the ipsilateral penumbra and infarcted core, corresponding arteries in the contralateral hemisphere, and basilar artery were excised. Anatomically similar arteries were excised from saline-injected rats. At 24 h after stroke, NADPH-stimulated superoxide production by arteries from the infarcted core did not differ from levels generated by arteries from control rats, whereas levels were significantly lower 72 h after stroke. However, at both time points after stroke, superoxide production by arteries from the ischemic penumbra was 8-fold greater than levels generated by arteries from control rats. Surprisingly, even in the non-ischemic arteries from the contralateral hemisphere and in the basilar artery, superoxide production was increased approximately 4- to 6-fold at 24 h, but had returned to normal 72 h after stroke. The NADPH-oxidase inhibitor, diphenyleneiodonium, virtually abolished superoxide production by all arteries. Thus, the activity of NADPH-oxidase is enhanced in cerebral arteries from the ischemic penumbra at 24 h and 72 h following cerebral ischemia. Additionally, NADPH-oxidase activity is temporarily enhanced after cerebral ischemia within arteries from non-ischemic parts of the brain.     

局灶脑缺血后早,晚期一氧化氮合成酶的活性变化

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,测定缺血后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(1h内)NOS活性突然升高,随之下降;脑缺血后24h NOS活性又升高,至48h、96h明显升高。     

亚低温对大鼠脑缺血再灌注后炎性反应的影响

目的：观察亚低温的不同时程对大鼠脑缺血再灌注后炎性反应的影响。方法：24只SD大鼠随机分为4组：常温组，亚低温1／2h组，亚低温1h组和亚低温3h组。每组各6只大鼠。参照Zea Longa的方法建立脑缺血再灌注动物模型，于再灌注24h断头取脑，行HE染色，观察脑组织中自细胞浸润及神经细胞的变化情况。结果：常温组在缺血梗死灶周围区可见白细胞浸润明显及深染受损的神经元；随亚低温时间的延长，白细胞浸润逐渐减少，神经元亦表现为淡染的轻度损伤。结论：亚低温可抑制脑缺血再灌注后的白细胞浸润，具有神经保护作用。     

Changes in Gene Expression in the Rat Hippocampus after Focal Cerebral Ischemia

Objective

The rat middle cerebral artery thread-occlusion model has been widely used to investigate the pathophysiological mechanisms of stroke and to develop therapeutic treatment. This study was conducted to analyze energy metabolism, apoptotic signal pathways, and genetic changes in the hippocampus of the ischemic rat brain.

Methods

Focal transient cerebral ischemia was induced by obstructing the middle cerebral artery for two hours. After 24 hours, the induction of ischemia was confirmed by the measurement of infarct size using 2,3,5-triphenyltetrazolium chloride staining. A cDNA microarray assay was performed after isolating the hippocampus, and was used to examine changes in genetic expression patterns.

Results

According to the cDNA microarray analysis, a total of 1,882 and 2,237 genes showed more than a 2-fold increase and more than a 2-fold decrease, respectively. When the genes were classified according to signal pathways, genes related with oxidative phosphorylation were found most frequently. There are several apoptotic genes that are known to be expressed during ischemic brain damage, including Akt2 and Tnfrsf1a. In this study, the expression of these genes was observed to increase by more than 2-fold. As energy metabolism related genes grew, ischemic brain damage was affected, and the expression of important genes related to apoptosis was increased/decreased.

Conclusion

Our analysis revealed a significant change in the expression of energy metabolism related genes (Atp6v0d1, Atp5g2, etc.) in the hippocampus of the ischemic rat brain. Based on this data, we feel these genes have the potential to be target genes used for the development of therapeutic agents for ischemic stroke.     

脑血管痉挛大鼠脑微区血流量和血浆内皮素1观察

应用非开颅大鼠蛛网膜下腔出血(SAH)模型,观察24h内脑微区血流量和颅内血清内皮素1(ET1)动态变化,并测量基底动脉(BA)管径。发现SAH后脑微区血流量迅速降低,1h达最低值,24h内无明显恢复趋势。SAH后1h开始至24h血浆ET1浓度显著增高。SAH后0.5h BA管径明显缩小。结果提示SAH后ET1的增多与脑血管痉挛及其缺血性脑损害的产生有关。     

脑缺血和再灌注后脑组织内皮素－1基因表达及丹参的影响

为研究脑缺血和再灌注后脑组织内皮素－１（ＥＴ－１）基因表达的变化以及丹参对它的影响，采用线栓法制成大鼠大脑中动脉缺血和再灌注模型，并用地高辛精标记ＥＴ－１基因进行原位杂交。结果显示，缺血组（缺血２４ｈ）和再灌注组（缺血１．５ｈ再灌注２４ｈ）缺血侧皮层及尾壳核ＥＴ－１基因表达均显著高于健侧相应的脑区（Ｐ＜０．０１，Ｐ＜０．０５），经丹参治疗后缺血或再灌注鼠缺血侧皮层及尾壳核ＥＴ－１基因表达均显著低于生理盐水（ＮＳ）对照组（Ｐ＜０．０５，Ｐ＜０．０１），但仍比健侧脑区显著增高。本实验表明，缺血和再灌注均可诱导脑组织ＥＴ－１基因的异常表达，进一步加重缺血和再灌注引起的脑损伤，丹参对缺血诱导的ＥＴ－１基因表达有部分抑制作用，这可能是丹参防治缺血性脑血管病的分子机理之一。     

缺血预处理（IPC）对大鼠局灶性脑梗死后HSP70和FOS表达的影响

目的 :研究缺血预处理 (IPC)对局灶性脑梗死后HSP70和FOS表达的影响 ,探讨IPC的脑保护作用机制。方法 :利用线栓法建立局灶性脑缺血 大脑中动脉闭塞模型 (MCAO)。MCAO 10min作为IPC ,IPC后 48h制作永久性大脑中动脉梗死 (PMCAO)模型。了解IPC对PMCAO后大脑神经功能和脑组织学损害的影响 ,免疫组化法研究PMCAO后 3hFOS表达变化以及 2 4h后HSP 70表达变化。结果 :IPC显著减轻PMCAO后大鼠神经功能损害和组织学损害 ,减少PMCAO后FOS、HSP 70的表达。结论 :IPC对其后PMCAO有明显的保护作用 ,能诱导脑缺血耐受 (IT)的产生 ,脑IT的神经保护作用与脑梗死后HSP 70和FOS表达改变密切相关。