小鼠局灶性脑梗死后半暗带的动态变化

脑血流完全阻断后，缺血中心区脑组织往往迅速产生不可逆性损害。缺血周边区，即半暗带，存活有一定的时限性，因此，研究半暗带持续时间就显得非常重要。随着基因治疗的深入研究，尤其转基因动物的使用，小鼠脑缺血模型的研究已成为热点。迄今，对其缺血半暗带的研究鲜见报道。我们选择昆明小鼠为观察对象，建立大脑中动脉栓线模型，应用MRI及TTC染色，观察缺血半暗带的动态变化。     

亚低温对大鼠局灶性脑缺血再灌注后半暗带区iNOS诱导细胞凋亡的影响

目的通过研究亚低温对大鼠局灶性脑缺血再灌注后诱导型一氧化氮合酶（iNOS）表达和细胞凋亡的影响，探讨亚低温脑保护的可能机制。方法雄性SD大鼠54只随机分为假手术组、常温缺血组和亚低温组。线栓法制备大脑中动脉闭塞再灌注模型。于缺血后48h取脑组织，邻片行HE染色，检测各组不同脑区iNOS蛋白表达和细胞凋亡情况，免疫双重染色研究iNOS蛋白表达与细胞凋亡间的关系，同时行NO含量测定。结果常温缺血组皮质缺血半暗带（IP）区iNOS免疫反应较强，TUNEL阳性细胞也主要位于皮质IP区，免疫双重染色发现TUNEL阳性细胞中存在着iNOS蛋白表达。亚低温组IP区iNOS蛋白表达明显下调，NO产生减少．IP区细胞凋亡的数目也减少。结论亚低温可能通过抑制IP区iNOS蛋白表达，减少NO产生．阻遏细胞凋亡，从而起到脑保护作用。     

尤瑞克林对大鼠局灶性脑缺血再灌注损伤后炎性反应的影响

目的研究尤瑞克林对大鼠局灶性脑缺血再灌注损伤后炎性反应的影响。方法将90只SD大鼠随机分为3组:假手术组,对照组,治疗组。采用线栓法建立大鼠大脑中动脉闭塞再灌注模型,缺血2 h后,拔出线栓,恢复灌注24 h,观察大鼠神经功能缺损症状、脑梗死体积、脑组织中白细胞浸润、MPO活性、IL-1和ICAM-1的表达。结果 (1)假手术组大鼠在神经功能缺损评分、脑梗死体积均低于对照组,有显著的统计学差异(P<0.01);脑组织中白细胞浸润程度、髓过氧化物酶(MPO)活性、ICAM-1和IL-1的表达均较对照组低,统计学差异明显(P<0.01);(2)治疗组与对照组相比,大鼠的神经功能缺损评分低、脑梗死体积小,有显著统计学差异(P<0.01);白细胞浸润程度、MPO活性、ICAM-1和IL-1的表达均较对照组减少(P<0.01)。结论尤瑞克林可通过抑制大鼠脑缺血再灌注损伤后的炎性反应来实现其神经保护作用。     

辛伐他汀对局灶性脑缺血大鼠半暗带区缓激肽受体mRNA表达的影响

目的：探讨辛伐他汀预处理对局灶性脑缺血大鼠半暗带缓激肽受体基因表达的影响。方法：72只雄性SD大鼠随机分为3组：辛伐他汀干预组（干预组,给予辛伐他汀10 mg.kg-1.d-1和生理盐水1 mL的悬浊液灌胃）,脑缺血再灌注模型组（模型组,给予等体积生理盐水灌胃）,假手术组（给予等体积生理盐水灌胃）,3组分别预处理14 d。参照Longa法将干预组和模型组建立大脑中动脉栓塞模型,假手术组仅暴露右侧颈总和颈内动脉主干,不栓塞。并将各组随机分为再灌注3、24和48 h 3个亚组（均n=8）。于对应时间点行神经功能评分,用苏木精-伊红染色检测脑组织形态学,荧光定量RT-PCR技术检测脑缺血半暗带缓激肽B1、B2受体（BK-1Rs、BK-2Rs）的基因表达水平。结果：与模型组相比,3、24和48 h干预组大鼠缺血再灌注后神经功能改善、功能评分值降低（分别P〈0.05,P〈0.05,P〈0.01）,脑组织病理形态学变化减轻。荧光定量RT-PCR检测发现,3 h模型组脑缺血半暗带BK-1Rs、BK-2Rs与假手术组比表达减低,差异有统计学意义（P〈0.05,P〈0.01）;3 h干预组BK-1Rs、BK-2Rs与模型组比表达增加,差异有统计学意义（P〈0.01,P〈0.05）,24和48 h模型组脑缺血半暗带BK-1Rs与假手术组比表达减低,差异有统计学意义（P〈0.01,P〈0.01）;24和48 h干预组BK-1Rs与模型组比表达增加,差异有统计学意义（P〈0.01,P〈0.01）。结论：辛伐他汀预处理可改善大鼠局灶性脑缺血再灌注神经功能缺损及组织病理形态学,其机制可能与增加脑缺血半暗带BK-1Rs、BK-2Rs的表达有关。     

大鼠局灶性脑缺血半暗带葡萄糖转运子3 mRNA的表达

目的 研究大鼠局灶性脑缺血不同缺血时间皮质半暗带和中心区葡萄糖转运子3（GLUT3）转录水平的表达规律。方法 用插线法建立大鼠局灶性脑缺血模型，剥取缺血半暗带及中心区皮质组织，采用逆转录-聚合酶链反应（RT-PCR），测定GLUT3mRNA水平的变化。结果 缺血半暗带GLUT3mRNA在缺血3小时升高，24小时达到高峰，96小时后基本恢复正常。缺血中心区GLUT3mRNA在缺血后3小时有一短暂的升高，随后迅速下降呈低水平表达。结论 GLUT3在缺血半暗带的表达上调，有可能是机体对缺血损伤的保护性反应。     

大鼠局灶性脑缺血损伤后半暗带区锌离子的变化

目的观察大鼠局灶性脑缺血再灌注后半暗带区锌离子的变化,探讨锌离子在脑缺血再灌注损伤中的可能作用。方法将28只SD大鼠随机分为假手术组(n=12)和大脑中动脉梗死(MCAO)组(n=16),以线栓法制作大鼠MCAO模型。分别于再灌注0h、3h、12h和24h时处死大鼠,取脑组织行TTC染色检测梗死体积,并制作脑组织冷冻切片,采用Newport Green(NG)染色法计数半暗带区NG阳性细胞数目并检测其平均荧光强度,分析NG阳性细胞数目与脑梗死体积的相关性。结果 (1)假手术组大鼠脑组织无梗死灶,也未见NG染色阳性细胞。MCAO组大鼠随再灌注时间延长脑梗死体积增大(均P<0.01),脑缺血半暗带区域NG阳性染色细胞数目随再灌注时间延长递增(均P<0.01)。各时间点间NG染色阳性细胞平均荧光强度无统计学差异(P>0.05)。(2)MCAO组大鼠脑切片NG阳性细胞数目与脑梗死体积比率呈正相关(r=0.88,P<0.01)。结论锌离子可能参与了脑缺血再灌注损伤的过程。     

人尿激肽原酶对大鼠局灶性脑缺血再灌注损伤的保护作用及对Caspase-3表达的影响

目的 观察人尿激肽原酶(HUK)对局灶性脑缺血再灌注损伤大鼠神经细胞凋亡及Caspase-3表达的影响. 方法 66只SD大鼠按随机数字表法分为假手术组(n=6)、缺血再灌注损伤组和HUK处理组,后两组又按不同观察时间点分为再灌注6h、12h、24 h、72 h、168 h共5个亚组(n=6).缺血再灌注损伤组和HUK处理组采用线栓法建立大鼠大脑中动脉局灶性脑缺血再灌注损伤模型,HUK处理组按浓度17.5×10-3PNAU/mL,1.0 mL/kg,于再灌注后3h尾静脉注射给药,1次/d.采用TUNEL法及免疫组化染色检测各组大鼠脑组织中凋亡细胞及Caspase-3阳性细胞的数量变化. 结果 脑缺血再灌注损伤后6h即有细胞凋亡,于24 h达到高峰,至168 h仍可见凋亡细胞.Caspase-3阳性细胞表达均于再灌注24 h达高峰,至168 h仍有较多表达.除168 h时间点外,其余各时间点HUK处理组大鼠神经细胞凋亡数量、Caspase-3阳性细胞数量均明显低于缺血再灌注损伤组,差异均有统计学意义(P＜0.05). 结论 HUK在大鼠局灶性脑缺血再灌注损伤早期(6～72h)时能抑制细胞凋亡,推测与其减少Caspase-3的表达有关.     

人尿激肽原酶对急性局灶性脑缺血再灌注损伤大鼠细胞凋亡的影响

目的 研究人尿激肽原酶(HUK)对大鼠急性局灶性脑缺血再灌注(FCIR)损伤后细胞凋亡数量及B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2联合X蛋白(Bax)蛋白表达的影响.方法 84只雄性SD大鼠分为假手术组(12只)、脑缺血再灌注(IR)组(36只)、HUK处理组(36只),IR组和HUK处理组剩余大鼠又按照再灌注时间6 h、12 h、24 h、72 h、168 h分为5个亚组(均为6只).建立大鼠大脑中动脉FCIR模型.假手术组、IR组及HUK处理组中各取6只SD大鼠用于测定梗死体积,其余大鼠用于观察神经功能缺陷评分、TIJNEL法及免疫组化检测凋亡细胞数量及凋亡蛋白Bcl-2、Bax的表达.结果 HUK处理组神经功能缺陷评分、梗死灶体积、除168 h亚组外的各时 间点的凋亡细胞数及Bax蛋白表达均显著少于IR组(P＜0.05),除168 h亚组外的各时间点的Bcl-2蛋白表达均显著高于IR组(P＜0.05).结论 HUK对FCIR后的脑组织起保护作用,其机制可能为损伤后3 d内通过上调Bcl-2、下调Bax蛋白表达来抑制细胞凋亡.     

大鼠局灶性脑缺血模型缺血半暗带的定位研究

目的通过用组织病理学方法结合局部脑血流量(rCBF)测定,对大鼠局灶性脑缺血动物模型缺血半暗带的解剖定位进行初步探讨.方法用线栓法制成大鼠大脑中动脉(MCA)闭塞及再通模型、采用TTC染色法观察分组动物在MCA闭塞的不同时间再灌流48h后脑梗死灶的分布,并用氢清除法测定缺血区rCBF的变化.结果大鼠MCA闭塞1～2h,梗死灶主要位于缺血侧的外侧尾壳核和额顶叶皮质下部;MCA闭塞3h、梗死灶向周围扩大;MCA闭塞4h、梗死灶进一步扩展至大部分新皮质区,但与MCA闭塞6h时无明显区别.MCA闭塞后缺血侧的内侧尾壳核和额顶叶皮质上部rCBF下降至对照组的27%～45%.结论大鼠MCA闭塞后在2～3h的时间窗以内恢复血流,可使位于缺血边缘区的内侧尾壳核和额顶叶皮质上部脑组织被挽救,该区域可能相当于缺血半暗带的等值区.     

人尿激肽原酶对局灶性脑缺血再灌注大鼠VEGF表达影响的研究

目的 探讨人尿激肽原酶对局灶性脑缺血再灌注大鼠脑组织血管内皮生长因子(VEGF)表达的影响.方法 采用随机数字表法将56只雄性SD大鼠分为假手术组(8只)、生理盐水组(24只)、人尿激肽原酶组(24只),其中生理盐水组、人尿激肽原酶组依据再灌注后不同取材时间又分为6 h,12 h,24 h,72 h,7 d五个亚组.采用线拴法制备大鼠局灶性脑缺血再灌注模型,采用神经功能评分、TTC染色、脑梗死体积测定、光镜检测等方法对不同组大鼠予以评价.采用免疫组化技术观察缺血再灌注不同时间点大鼠脑组织梗死中心区及半影区VEGF表达变化情况.结果 人尿激肽原酶组大鼠神经功能评分低于生理盐水组大鼠(P<0.05);24 h脑梗死体积测定,人尿激肽原酶组平均值为(53 261.96±7 326.75)μm3,生理盐水组平均值为(92 715.84±13 755.44)μm3,差异有统计学意义(P<0.05);人尿激肽原酶组VEGF表达在不同时间点均明显强于生理盐水组(P<0.05).结论 人尿激肽原酶能减轻脑缺血再灌注模型大鼠的神经功能损伤程度,减少脑梗死体积,促进VEGF的表达,具有脑缺血后神经保护作用.     

Effects of dextromethorphan on regional cerebral blood flow in focal cerebral ischemia

Dextromethorphan (DM), a noncompetitive NMDA antagonist, has been demonstrated to reduce ischemic neuronal damage and edema, but DM's influence on cerebral blood flow has not been extensively studied. In this investigation, it is shown that DM has significant effects on regional cerebral blood flow (rCBF) patterns in a rabbit model of focal cerebral ischemia. rCBF was measured using radioactive microspheres following a 1 h permanent occlusion of the left internal carotid, anterior cerebral, and middle cerebral arteries in rabbits. Somatosensory evoked potentials (SEPs) were used to assess the degree of ischemia; only animals where SEPs were completely abolished were used for a frequency distribution analysis of rCBF. It was found that there were significantly more regions with lower flows in animals treated with normal saline (NS) (n = 7) compared to animals treated with DM (n = 7) (p less than 0.05, ipsilateral left side; p less than 0.001, contralateral right side). The frequency distribution medians were 27.5 ml 100 g-1 min-1 (left) and 70.0 ml 100 g-1 min-1 (right) in the NS group vs. 34.5 ml 100 g-1 min-1 (left) and 80.5 ml 100 g-1 min-1 (right) in the DM group. The left and right hemispheric regional means were 29.4 +/- 20 and 74.3 +/- 23 ml 100 g-1 min-1, respectively, in the NS group vs. 34.4 +/- 16 and 91.0 +/- 28 ml 100 g-1 min-1, respectively, in the DM group.(ABSTRACT TRUNCATED AT 250 WORDS)     

TMB—8对脑缺血大鼠脑血流量的作用

目的 研究8－（N,N－二乙胺）－n－辛基－3,4,5－三甲氧基苯甲酸酯（TMB－8）对局灶性脑缺血大鼠脑血流量（CBF）的作用。方法 用激光多谱勒血流仪测量大脑中动脉阻断（MCAO）大鼠脑血流量。分别于阻断前30分钟和阻断后20分钟给予TMB－8进行干预。结果 MCAO后,CBF迅速下降,维持恒定。阻断前30分钟给予TMB－8 0．5、1和2mg/kg,可剂量依赖性抑制CBF下降,阻断后20分钟给予TMB－8 1mg/kg,也能明显增加CBF。结论 TMB－8能预防和治疗MCAO局灶性脑缺血大鼠CBF减少,改善缺血区血供。     

Effects of kallikrein gene transfer on penumbral microvascular proliferation and on regional cerebral blood flow following cerebral ischemia/reperfusion injury

BACKGROUND： Recent findings have demonstrated that the kallikrein-kinin system （KKS） participates in the pathological process of cerebral ischemia/reperfusion injury. Kallikrein gene transfer exhibits neural protective effects following cerebral infarction. OBJECTIVE： To observe the effects of kallikrein gene transfer on vascular proliferation in the peripheral infarct focus and on regional cerebral blood flow （rCBF） following cerebral ischemia/reperfusion injury. DESIGN, TIME AND SETTING： The completely randomized, controlled experiment was performed at the Lin Baixin Laboratory Center, the Second Affiliated Hospital of Sun Yat-sun University between September 2007 and April 2008. MATERIALS： pUCI9-HTK plasmid was constructed and maintained in the Laboratory for Neurology, the Second Affiliated Hospital of Sun Yat-sen University, China. Mouse anti-human kallikrein 1 monoclonal antibody was purchased from R＆D Systems, USA. METHODS Ninety healthy, male, Sprague Dawley rats were used. Middle cerebral artery occlusion （MCAO） was established in all rats to induce cerebral ischemia/reperfusion injury. Following MCAO establishment, all rats were randomly divided into three groups （n = 30）： blank control, saline, and pAdCMV-HTK. The saline and pAdCMV-HTK groups were stereotactically micro-injected with 5μL of physiological saline or with pAdCMV-HTK [multiplicity of infection （MOI） = 20], respectively, into the ischemic penumbra. In the blank control group, only sham injection was performed. MAIN OUTCOME MEASURES： At 12, 24, and 72 hours after treatment, cerebral infarction volume was measured by 2, 3, 5-triphenyltetrazolium chloride （TTC） staining. Exogenous HTK expression, as well as regional vascular endothelial growth factor （VEGF） expression, was detected by immunohistochemistry. rCBF was examined by 14C-iodoantipyrine micro tracing. In addition, neurological severity score （NSS） was performed. Higher scores indicated more severe neurological deficits. RESULTS： NSS res     

血液光量子疗法对局灶性脑缺血大鼠大脑皮层局部血流量和脑水肿的影响

本文动态观测了光量子疗法治疗血栓栓塞性脑缺血大鼠,大脑皮层局部血流量(rCBF)及脑水肿的变化。结果:缺血6小时经光量子血液治疗2小时后,rCBF于注后30min明显增加,达19.78％(P<0.01),以后各点维持在一定水平,2小时增加到19.30(P<0.05)。脑组织比重:缺血中心区未见明显改善(P>0.1)而半暗区组织比重非常明显增加(P<0.001),可见水肿明显减轻。说明光量子疗法增加rCBF,减轻半暗区水肿。     

Sequential assessment of regional cerebral blood flow, regional cerebral blood volume, and blood-brain barrier in focal cerebral ischemia: a case report

Regional CBF (rCBF) and regional cerebral blood volume (rCBV) were evaluated by N,N,N'-trimethyl-N'-(2)-hydroxy-3-methyl-5-[123I]iodobenzyl-1, 3-propanediamine-2 HCl- and 99mTC-labeled red blood cells, respectively, and single-photon emission computerized tomography (SPECT) in a patient with focal cerebral ischemia. Sequential transmission computerized tomography (TCT) and SPECT functional data were compared with clinical findings to monitor the pathophysiological events occurring in stroke. A lack of correlation between rCBF-rCBV distributions and blood-brain barrier (BBB) breakdown was found in the acute phase. In the face of more prolonged alteration of BBB, as seen by TCT enhancement, a rapid evolution of transient phenomena such as luxury perfusion was shown by SPECT studies. Follow-up of the patient demonstrated a correlation between the neurological recovery and a parallel relative improvement of the cerebral perfusion.     

Interstitial pO2 in ischemic penumbra and core are differentially affected following transient focal cerebral ischemia in rats.

Stroke causes heterogeneous changes in tissue oxygenation, with a region of decreased blood flow, the penumbra, surrounding a severely damaged ischemic core. Treatment of acute ischemic stroke aims to save this penumbra before its irreversible damage by continued ischemia. However, effective treatment remains elusive due to incomplete understanding of processes leading to penumbral death. While oxygenation is central in ischemic neuronal death, it is unclear exactly what actual changes occur in interstitial oxygen tension (pO2) in ischemic regions during stroke, particularly the penumbra. Using the unique capability of in vivo electron paramagnetic resonance (EPR) oximetry to measure localized interstitial pO2, we measured both absolute values, and temporal changes of pO2 in ischemic penumbra and core during ischemia and reperfusion in a rat model. Ischemia rapidly decreased interstitial pO2 to 32% +/- 7.6% and 4% +/- 0.6% of pre-ischemic values in penumbra and core, respectively 1 hour after ischemia. Importantly, whilst reperfusion restored core pO2 close to its pre-ischemic value, penumbral pO2 only partially recovered. Hyperoxic treatment significantly increased penumbral pO2 during ischemia, but not in the core, and also increased penumbral pO2 during reperfusion. These divergent, important changes in pO2 in penumbra and core were explained by combined differences in cellular oxygen consumption rates and microcirculation conditions. We therefore demonstrate that interstitial pO2 in penumbra and core is differentially affected during ischemia and reperfusion, providing new insights to the pathophysiology of stroke. The results support normobaric hyperoxia as a potential early intervention to save penumbral tissue in acute ischemic stroke.