绿茶多酚对脑缺血大鼠血脑屏障通透性的影响

目的探讨绿茶多酚对局灶性脑缺血大鼠血脑屏障的通透性及紧密连接蛋白occludin、claudin-5和蛋白激酶PKCα表达的影响。方法采用线栓法制作大鼠局灶性脑缺血模型,伊文思蓝法检测血脑屏障通透性的变化,免疫组化方法检测缺血脑组织occludin、claudin-5的表达,western blot方法检测脑缺血组织PKCα蛋白的表达。结果大鼠局灶性脑缺血60min、120min的脑组织血脑屏障通透性显著增加,同时点的脑组织BBB紧密连接开放;而绿茶多酚显著降低血脑屏障的通透性(P0.01),使BBB紧密连接处于关闭状态。大鼠局灶性脑缺血60min及120min脑组织紧密连接相关蛋白occludin、claudin-5的蛋白表达水平显著降低,绿茶多酚显著上调这些蛋白表达水平(P0.01)。大鼠局灶性脑缺血30min至120min脑组织中PKCα的蛋白表达水平显著升高,绿茶多酚显著降低PKCα蛋白的表达水平(P0.01)。结论绿茶多酚降低缺血大鼠脑组织血脑屏障通透性可能与上调紧密连接蛋白occludin、claudin-5的表达及下调蛋白激酶PKCα的表达有关。     

全反式维甲酸对大鼠脑缺血再灌注后血脑屏障通透性和claudin-5表达的影响

目的探讨全反式维甲酸(ATRA)对大鼠脑缺血再灌注后血脑屏障通透性和紧密连接蛋白claudin-5的影响。方法选取180只健康雄性大鼠,随机分为假手术组(Sham,60只),缺血再灌注组(I/R,60只)和全反式维甲酸干预组(ATRA,60只),每组又细分为1d、3d和7d三个亚组。采用Zea Longa法制作局灶性脑缺血再灌注模型,伊文思蓝渗透性实验检测血脑屏障通透性变化,免疫组织化学方法和Western blot方法检测各组大鼠脑组织claudin-5蛋白的表达,Real-time PCR方法检测各组大鼠脑组织claudin-5 m RNA的表达。结果缺血再灌注后,EB含量在1d时最高,3d时开始逐渐下降,而与相同时间点的I/R组相比,ATRA组EB含量显著降低(P0.01)。与Sham组相比,I/R组大鼠脑组织claudin-5蛋白及m RNA的表达水平均显著降低,而与相同时间点的I/R组相比,ATRA组则显著升高(P0.01)。结论全反式维甲酸降低大鼠脑缺血再灌注后血脑屏障通透性可能与上调脑缺血再灌注后脑组织中claudin-5的表达相关。     

全反式维甲酸对大鼠脑缺血再灌注后血脑屏障通透性和ZO-1表达的影响

目的探讨全反式维甲酸(ATRA)对大鼠脑缺血再灌注后血脑屏障通透性和紧密连接蛋白ZO-1的影响。方法选取180只健康雄性大鼠,随机分为假手术组(Sham,60只),缺血再灌注组(I/R,60只)和全反式维甲酸干预组(ATRA,60只),每组又细分为1d、3d和7d三个亚组。采用Zea Longa法制作局灶性脑缺血再灌注模型,伊文思蓝渗透性实验检测血脑屏障通透性变化,免疫组织化学方法法和Western blot方法检测各组大鼠脑组织ZO-1蛋白的表达,Real-time PCR方法检测各组大鼠脑组织ZO-1mRNA的表达。结果缺血再灌注后,EB含量在1d时含量最高,3d时开始逐渐下降,而与相同时间点的I/R组相比,ATRA组EB含量显著降低(P0.01)。与Sham组相比,I/R组大鼠脑组织ZO-1蛋白及mRNA的表达水平均显著降低(P0.01),而与相同时间点的I/R组相比,ATRA组大鼠脑组织ZO-1蛋白及mRNA的表达水平均显著升高。结论全反式维甲酸降低大鼠脑缺血再灌注血脑屏障通透性可能与上调脑缺血再灌注后脑组织中ZO-1的表达相关。     

雌激素对大鼠脑缺血再灌注后血脑屏障作用研究

目的观察雌激素对大鼠脑缺血再灌注后血脑屏障(blood-brain barrier,BBB)的通透性、Occludin表达的影响,探讨雌激素在脑缺血中的作用。方法随机将去势雌性大鼠分为假手术组、模型组、雌激素预处理组,选取缺血再灌后4 h,24 h,3 d3个时间点作为观察点,对各个时间点脑水肿情况、Occludin蛋白表达、血脑屏障通透性改变进行考察分析,并选取24 h和3 d作BBB超微结构电镜观察,脑水肿改变情况采用脑含水量百分数测定;蛋白质表达采用western blot方法;血脑屏障通透性改变采用伊文思蓝(EB)比色法。结果与假手术组比较,局灶性脑缺血再灌4 h模型组大鼠脑组织含水量及EB含量均增加(P0.05),随缺血再灌时间延长,脑组织含水量、脑组织EB含量持续增加,至24 h,达到高峰(P0.01),与同时间点模型组比较,各治疗组大鼠脑组织含水量、EB含量均有不同程度降低(P0.05或P0.01),以24 h组降低最为显著(P0.01)。电镜观察雌激素预处理组较模型组同时间点比较BBB TJ开放减轻,星形胶质细胞足突及毛细血管管周水肿较轻,以3 d组显著。Western blot检测Occludin蛋白表达,发现模型组4 h时Occludin蛋白表达较假手术组减弱,但无显著性差异(P0.05),24 h组Occludin蛋白表达较假手术组减弱,有显著性差异(P0.05),3 d组Occludin蛋白表达进一步减弱,有明显差异(P0.01),雌激素组4 h时Occludin蛋白表达较同时间点模型组比较无显著性差异(P0.05),雌激素24 h及3 d组Occludin蛋白表达较同时间点模型组比较均升高,有显著性差异(P0.05)。结论局灶性脑缺血大鼠动物血脑屏障超微结构可见紧密连接发生断裂,内皮细胞内小泡数量增加及星形胶质细胞足突肿胀,可能是大鼠局脑缺血时血管源性脑水肿的重要因素。大鼠局灶性脑缺血,随缺血再灌时间延长,紧密连接相关蛋白occludin的表达明显下降,提示occludin在调节紧密连接通透性变化的过程中有重要作用。雌激素上调紧密连接Occludin蛋白的表达,有可能是其维护血脑屏障完整性减轻脑水肿的机制之一。     

地塞米松预处理对氧糖剥夺大鼠血脑屏障通透性的影响

目的探讨地塞米松(dexamethasone,DEX)对氧糖剥夺(oxygen glucose deprivation,OGD)条件下大鼠血脑屏障通透性的影响。方法提取纯化大鼠脑微血管内皮细胞,建立血脑屏障OGD模型。在OGD不同时间点(0、30、60 min),以及DEX预处理OGD 60min时,应用辣根过氧化物酶渗漏实验分析血脑屏障的通透性;应用RT-PCR和western blot法分析大鼠脑微血管内皮细胞的紧密连接相关蛋白claudin-5的mRNA和蛋白的表达变化;同时应用免疫荧光方法观察细胞膜上claudin-5的变化。结果与对照组相比,OGD 30、60min,辣根过氧化物酶流量均显著升高;与OGD 60min相比,DEX预处理显著降低了辣根过氧化物酶流量。与对照组相比,OGD30、60min,claudin-5的mRNA和蛋白表达水平均显著降低;与OGD 60min相比,DEX预处理显著增加了claudin-5的mRNA和蛋白表达。免疫荧光观察claudin-5在细胞膜上的表达,与对照组相比,OGD 60min时claudin-5的表达明显减少,DEX预处理后显著增加了claudin-5的表达。结论 DEX预处理降低了OGD条件下的血脑屏障通透性,其机制之一是DEX增加紧密连接相关蛋白claudin-5的表达。     

帕金森病模型大鼠十二指肠黏膜紧密连接蛋白表达下调

目的研究紧密连接蛋白在6-羟多巴(6-OHDA)制备的帕金森病(PD)大鼠模型十二指肠黏膜的表达变化。方法用6-OHDA损毁双侧中枢黑质多巴胺能神经元建立大鼠模型。用免疫荧光组织化学和蛋白免疫印迹检测紧密连接蛋白claudin-1、occludin和ZO-1肠黏膜的定位和表达。结果紧密连接蛋白claudin-1、occludin和ZO-1在PD大鼠模型十二指肠黏膜上均有表达,但仅ZO-1(P0.001)和occludin(P0.01)表达明显下调,而claudin-1无显著变化。结论 PD大鼠模型十二指肠黏膜紧密连接蛋白ZO-1、occludin的表达显著下调,可能与帕金森病十二指肠溃疡的发生发展相关。     

人工麝香对大鼠局灶性脑缺血再灌注后大脑MMP-9 mRNA及其蛋白表达的影响

目的观察人工麝香对大鼠脑缺血再灌注后脑组织基质金属蛋白酶-9（MMP-9）mRNA及其蛋白表达的影响。方法采用SD大鼠制作大脑中动脉局灶性脑缺血再灌注模型,用高、中、低浓度人工麝香治疗缺血2h再灌注24、48h大鼠。应用实时荧光定量PCR法检测缺血再灌注脑组织MMP-9 mRNA的表达水平,免疫组化方法检测脑组织MMP-9蛋白表达量。结果与假手术组相比,各组在缺血再灌注不同时间段脑组织MMP-9 mRNA及MMP-9蛋白表达水平均升高（P〈0.05）,其中,以模型组MMP-9 mRNA及MMP-9蛋白表达水平最高（P〈0.01）,高、中浓度人工麝香治疗组MMP-9 mRNA及MMP-9蛋白表达水平最低（P〈0.05）;与模型组相比,高、中浓度人工麝香治疗组脑组织MMP-9 mRNA及MMP-9蛋白表达水平降低最为显著（P〈0.01）。结论人工麝香能降低大鼠局灶性脑缺血再灌注后脑组织MMP-9 mRNA及MMP-9蛋白的表达水平。人工麝香可能通过抑制缺血再灌注后大鼠脑组织MMP-9的表达,降低脑组织血脑屏障基底膜细胞外基质的降解和血脑屏障（BBB）的通透性,减轻脑缺血后脑水肿。     

甘草甜素通过抑制炎症反应和氧化应激改善大鼠蛛网膜下腔出血后血脑屏障损伤

目的:探究甘草甜素对大鼠蛛网膜下腔出血(SAH)后血脑屏障损伤的改善作用以及与炎症反应、氧化应激反应的关系。方法:将大鼠分为假手术组、模型组、甘草甜素低剂量组、中剂量组、高剂量组(25 mg/kg、50 mg/kg、100 mg/kg)、阳性组(40 mg/kg尼莫地平),每组30只大鼠,对大鼠神经行为进行评分,给药结束后24、48、72 h测定大鼠脑组织含水量,采用伊文氏蓝渗出量(EB)评估法检测血脑屏障通透性,HE染色观察大鼠基底动脉血管形态学变化,透射电镜观察脑组织超微结构变化情况,酶联免疫法检测脑组织中IL-1β、IL-6、TNF-α、MDA、3-NT、8-OHDG表达;分别采用比色法、氮蓝四唑光还原法及硫代巴比妥酸法测定脑组织中氧化应激指标GSH-Px、SOD、MDA水平; RT-PCR检测脑组织中COX-2、iNOS、ICAM-1、VCAM-1 mRNA表达;蛋白免疫印迹法(WB)检测ZO-1,occludin、claudin-5、Bax、Bcl-2蛋白表达。结果:与假手术组相比,模型组大鼠神经行为学评分降低,给药后24、48、72 h脑含水量升高,EB含量升高;模型组大脑皮层细胞凋亡率升高;与模型组相比,甘草甜素组大鼠神经行为学评分升高,脑含水量、EB含量、细胞凋亡率降低,具有剂量依赖性(P 0. 05)。与假手术组相比,模型组IL-1β、IL-6、TNF-α、3-NT、8-OHDG、MDA水平升高,SOD、GSH-Px水平降低;与模型组相比,甘草甜素组和阳性组IL-1β、IL-6、TNF-α、3-NT、8-OHDG、MDA水平均降低,SOD、GSH-Px水平升高,具有剂量依赖性(P 0. 05)。与假手术组相比,模型组COX-2、iNOS、ICAM-1、VCAM-1 mRNA表达升高,ZO-1、occludin、claudin-5、Bcl-2蛋白表达降低,Bax蛋白表达升高;与模型组相比,甘草甜素组和阳性组COX-2、iNOS、ICAM-1、VCAM-1 mRNA表达降低,ZO-1、occludin、claudin-5、Bcl-2蛋白表达升高,Bax蛋白表达降低,具有剂量依赖性(P 0. 05)。结论:甘草甜素能够降低大鼠蛛网膜下腔出血后的血脑屏障通透性增加,缓解基底动脉血管痉挛以及减少内皮细胞紧密连接丢失,其机制可能与抑制炎症反应和氧化应激反应有关。     

阿托伐他汀对脑缺血再灌注大鼠血脑屏障的保护作用

目的:研究在脑缺血再灌注(ischemia-reperfusion,IR)损伤中阿托伐他汀对血脑屏障的保护作用及相关机制。方法:SD大鼠72只随机分为3组:假手术组、IR组和阿托伐他汀组。阿托伐他汀组大鼠术后予以阿托伐他汀(20 mg·kg-1·d-1)灌胃治疗,每天1次,连续3 d。利用线栓法制作脑IR模型,缺血2 h后再灌注72 h,对各组大鼠的神经功能进行评分,并检测梗死侧大脑半球脑组织含水量、伊文思蓝(Evans blue,EB)渗出量、紧密连接相关蛋白occludin和炎症因子磷脂酰肌醇3-激酶p110γ(PI3K-p110γ)的表达水平。结果:与假手术组相比,IR组大鼠脑组织含水量、EB的渗出量及神经功能评分均增加(P0.01),同时occludin表达下降(P0.01),PI3Kp110γ表达增加(P0.01);而阿托伐他汀治疗组大鼠脑水肿程度减轻(P0.01),EB渗出量减少(P0.01),occludin表达增加(P0.01),PI3K-p110γ表达减少(P0.01),神经功能评分下降,但差异无统计学意义(P0.05)。结论:阿托伐他汀可以减轻脑IR损伤,可能与抑制炎症反应、增加紧密连接蛋白表达从而维持血脑屏障稳定性有关。     

α硫辛酸对后肢缺血再灌注损伤大鼠血神经屏障的保护作用研究

目的:研究α硫辛酸(ALA)对大鼠后肢缺血再灌注(I/R)损伤后血神经屏障的保护作用。方法:54只成年雄性SD大鼠分为3组,分别为假手术组(sham)、缺血再灌注模型组(I/R)和ALA处理组(ALA),通过手术结扎大鼠左侧髂总动脉4 h制备后肢I/R注模型,ALA处理组大鼠于术前7 d连续给予ALA。伊文思蓝(EB)渗透实验检测血神经屏障的通透性,分别于再灌注后24 h检测各组大鼠坐骨神经组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)活性,以及丙二醛(MDA)的含量,应用Western Blot检测坐骨神经组织中紧密连接蛋白occludin和claudin-5表达变化。结果:与sham组相比,I/R组大鼠坐骨神经EB漏出增多(P 0. 05),SOD和GSH-Px活性降低(P 0. 05),MDA水平升高(P 0. 05),occludin和claudin-5蛋白表达下降(P 0. 05); ALA预处理的大鼠EB漏出减少(P 0. 05),GSH-Px和SOD活性增加(P 0. 05),MDA水平降低(P 0. 05),occludin和claudin-5蛋白表达增加(P 0. 05)。结论:ALA通过抑制机体的氧化应激水平对I/R损伤大鼠的血神经屏障起到保护作用。     

Time-course investigation of blood–brain barrier permeability and tight junction protein changes in a rat model of permanent focal ischemia

Permanent middle cerebral artery occlusion (pMCAO) is an animal model that is widely used to simulate human ischemic stroke. However, the timing of the changes in the expression of tight junction (TJ) proteins and synaptic proteins associated with pMCAO remain incompletely understood. Therefore, to further explore the characteristics and mechanisms of blood–brain barrier (BBB) damage during cerebral ischemic stroke, we used a pMCAO rat model to define dynamic changes in BBB permeability within 120 h after ischemia in order to examine the expression levels of the TJ proteins claudin-5 and occludin and the synaptic proteins synaptophysin (SYP) and postsynaptic density protein 95 (PSD95). In our study, Evans blue content began to increase at 4 h and was highest at 8 and 120 h after ischemia. TTC staining showed that cerebral infarction was observed at 4 h and that the percentage of infarct volume increased with time after ischemia. The expression levels of claudin-5 and occludin began to decline at 1 h and were lowest at 8 and 120 h after ischemia. The expression levels of SYP and PSD95 decreased from 12 to 120 h after ischemia. GFAP, an astrocyte marker, gradually increased in the cortex penumbra over time post-ischemia. Our study helps clarify the characteristics of pMCAO models and provides evidence supporting the translational potential of animal stroke models.     

沉默排斥性导向分子A对脑缺血再灌注损伤大鼠血脑屏障的保护作用

目的 观察沉默排斥性导向分子A（RGMa）对脑缺血再灌注损伤大鼠血脑屏障及紧密连接蛋白的影响。 方法 雄性成年 SD 大鼠立体定位侧脑室注射腺病毒后建立大脑中动脉闭塞(MCAO) /再灌注（I/R）模型。将成年雄性SD大鼠40只,分为空白对照组（sh-con）和RGMa干扰组（sh-RGMa）,分别在注射后1 d和3d观察腺病毒对RGMa的影响。将其余120只SD大鼠随机分为假手术组（sham）、缺血再灌注组（I/R）、空病毒组（I/R+sh-con）及病毒组（I/R+sh-RGMa）;I/R 72 h 后采用神经功能缺损评分评估各组大鼠神经功能恢复情况; 采用2,3,5-氯化三苯基四氮唑(TTC)染色测量脑梗死体积;采用静脉注射伊文思蓝观察血脑屏障通透性的改变;应用Western blotting和免疫组织化学染色检测RGMa的变化;应用Western blotting检测血脑屏障完整性相关紧密连接蛋白5（claudin-5）、基质金属蛋白酶9（MMP-9）和闭锁小带蛋白1（ZO-1）的表达。 结果 缺血再灌注后 72 h,I/R组较 sham 组神经功能评分降低,沉默RGMa能改善血脑屏障通透性,减少脑梗死体积;可下调MMP-9及上调claudin-5和ZO-1的表达。 结论 沉默RGMa对脑缺血再灌注损伤大鼠的血脑屏障有保护作用。     

Phosphorylation of claudin-5 and occludin by rho kinase in brain endothelial cells

Critical to the proper maintenance of blood-brain-barrier (BBB) integrity are the endothelial tight junctions (TJs). Posttranslational modifications of essential endothelial TJ proteins, occludin and claudin-5, contribute and possibly disrupt BBB integrity. Our previous work has shown that Rho kinase (RhoK) activation mediates occludin and claudin-5 phosphorylation resulting in diminished barrier tightness and enhanced monocyte migration across BBB in the setting of human immunodeficiency virus-1 encephalitis (HIVE). To determine whether RhoK can directly phosphorylate TJ proteins, we examined phosphorylation of cytoplasmic domains of recombinant claudin-5 and occludin by RhoK. We found that RhoK predominately phosphorylated two sites on occludin (T382 and S507) and one site on claudin-5 (T207). Specific anti-phosphopeptide antibodies were developed for these sites, allowing the detection of phosphorylated occludin at T382 and S507, and claudin-5 at T207 from full-length recombinant occludin and claudin-5 transiently expressed in COS-7 cells and mouse brain microvascular endothelial cells. Finally, these phosphospecific antibodies demonstrated enhanced staining of brain endothelial cells in the mouse model for HIVE and human HIVE brains featuring mononuclear cell infiltration across disrupted BBB. Our results demonstrated the direct phosphorylation of occludin and claudin-5 by RhoK at specific sites, which was increased in encephalitic brain tissue. These antibodies could be useful reagents for monitoring BBB dysfunction in vivo.     

FTY720 Reduces Endothelial Cell Apoptosis and Remodels Neurovascular Unit after Experimental Traumatic Brain Injury

Traumatic brain injury (TBI) is a major cause of death and disability worldwide. A sequence of pathological processes occurred when there is TBI. Previous studies showed that sphingosine-1-phosphate receptor 1 (S1PR1) played a critical role in inflammatory response in the brain after TBI. Thus, the present study was designed to evaluate the effects of the S1PR1 modulator FTY720 on neurovascular unit (NVU) after experimental TBI in mice. The weight-drop TBI method was used to induce TBI. Western blot (WB) was performed to determine the levels of SIPR1, claudin-5 and occludin at different time points. FTY720 was intraperitoneally administered to mice after TBI was induced. The terminal deoxynucleotidyl transferase-dUTP nick end labeling (TUNEL) assay was used to assess endothelial cell apoptosis. Immunofluorescence and WB were performed to measure the expression of tight junction proteins: claudin-5 and occludin. Evans blue (EB) permeability assay and brain water content were applied to evaluate the blood-brain barrier (BBB) permeability and brain edema. Immunohistochemistry was performed to assess the activation of astrocytes and microglia. The results showed that FTY720 administration reduced endothelial cell apoptosis and improved BBB permeability. FTY720 also attenuated astrocytes and microglia activation. Furthermore, treatment with FTY720 not only improved neurological function, but also increased the survival rate of mice significantly. These findings suggest that FTY720 administration restored the structure of the NVU after experimental TBI by decreasing endothelial cell apoptosis and attenuating the activation of astrocytes. Moreover, FTY720 might reduce inflammation in the brain by reducing the activation of microglia in TBI mice.     

Different changes in the expression of multiple kinds of tight-junction proteins during ischemia-reperfusion injury of the rat ileum

Dysfunction of tight junctions (TJs), located at the most apical part of the intestinal epithelium, is believed to result in various complications in intestinal disease. However, the behaviors of multiple kinds of TJ proteins during ischemia-reperfusion injury are not understood in detail. To determine changes in expression and localization of TJ proteins during intestinal-barrier recovery, we induced intestinal ischemia-reperfusion injury in rats, measured mucosa-to-blood permeability of fluorescein isothiocyanate-dextran-4 kDa, and compared it with spatiotemporal changes of ZO-1, occludin, and claudin-1, -2, -3, -4, and -5 by immunoconfocal microscopy. At 1 hour post-reperfusion, villi were denuded and intestinal-barrier function was lost. From 6 to 24 hours post-reperfusion, villous epithelium was restored by cell migration, and barrier function together with reticular pattern expression of ZO-1, occludin, and claudin-1, -3, and -5, recovered time-dependently. To the contrary, after ischemia-reperfusion injury, the localized expression of claudin-2 and claudin-4 observed in the non-treated control was lost and replaced with broader expression from crypts to villi with increased basolateral claudin-4 expression in epithelial cells. These data demonstrated that recovery of intestinal barrier function is associated with expression of ZO-1, occludin, and claudin-1, -3, and -5, whereas claudin-2 and claudin-4 show unique changes in expression and localization.     

Alterations of the blood-brain barrier in cerebral white matter lesions in the ageing brain

White matter lesions (WML) are associated with dementia and are common in brain ageing. In order to determine whether alteration of the blood-brain barrier (BBB) may contribute to the pathogenesis of WML we assessed albumin leakage and expression of the tight junction (TJ) proteins claudin-5 (Cln-5), zona occludin-1 (ZO-1) and occludin in cases derived from the Medical Research Council Cognitive Function and Ageing Study. Albumin extravasation was widespread in the ageing brain and enhanced in WML, suggesting dysfunction of the BBB may contribute to the pathogenesis of WML. This was not accompanied by significant changes in the endothelial expression of TJ proteins. However, ZO-1 and occludin were expressed by glial cells throughout the parenchyma of both control white matter and WML, suggesting these TJ proteins may have other functions in the brain.     

血脑屏障氧糖剥夺体外模型中缺氧诱导因子-1α的表达及血脑屏障通透性的变化

目的研究血脑屏障氧糖剥夺体外模型中缺氧诱导因子-1α和紧密连接相关蛋白claudin-5的表达及血脑屏障通透性变化。方法提取纯化大鼠脑微血管内皮细胞,建立血脑屏障氧糖剥夺模型,模拟在体缺血,在氧糖剥夺不同时间点(0、30、60、120、240min),应用EVOM测定仪测定培养的脑微血管跨内皮细胞内皮阻抗值,分析血脑屏障的通透性;应用辣根过氧化物酶渗漏实验分析血脑屏障的通透性;应用Westernblot法分析大鼠脑微血管内皮细胞的缺氧诱导因子-1α和紧密连接相关蛋白claudin-5的表达。结果与正常对照组相比,氧糖剥夺30min,缺氧诱导因子-1α表达增加,辣根过氧化物酶流量升高;氧糖剥夺60min达峰值;氧糖剥夺120min、240min,逐渐降低,但仍高于正常对照组。氧糖剥夺30min,跨内皮阻抗值降低,claudin-5表达减少,在60min最低;120min、240min表达逐渐升高,但仍低于正常对照组。结论氧糖剥夺30min至240min,缺氧诱导因子-1α和claudin-5表达及血脑屏障通透性改变明显,氧糖剥夺60min达峰值。     

树鼩脑缺血时海马微环境与血脑屏障通透性改变的可能机制

目的观察树鼩血栓性脑缺血时海马血脑屏障（blood-brain barrier，BBB）通透性的变化，并探讨其与微环境相互作用的可能机制。方法通过光化学反应诱导树鼩血栓性脑缺血模型，采用单泵等速微灌流系统及荧光分光法检测脑缺血后4、24及72h海马细胞外液异硫氰基荧光素标记的葡聚糖（fluorescein thiocarbamoyl dextrans，FITC-D）含量及血浆FITC-D的清除率（Clearancerate of FITC-D，Cf），Cf的高低反映海马BBB的通透性。用高效液相色谱技术检测海马灌流液的氨基酸递质水平，并观察缺血海马微血管超微结构。结果树嗣血栓性脑缺血后4h海马BBB通透性开始升高，缺血24h最为明显，Cf=（0．526±0．130）／μl（P〈0．01），72h通透性有所恢复。脑缺血后24h海马细胞外Glu及GABA水平明显升高，分别为（5．71±0．39）μmol·L^-1和（3．81±0．14）μmol·L^-1，均具有统计学意义（P〈0．01）。海马微血管超微结构观察显示星形胶质细胞肿胀，毛细血管周围水肿形成。结论脑缺血可导致海马BBB通透性增高，而海马微环境紊乱对BBB通透性具有促进作用。     

局灶性脑缺血后125Ⅰ-神经生长因子对血脑屏障的通透性

目的 通过大鼠局灶性脑缺血实验,探讨神经生长因子(NGF)在大脑局部脑缺血情况下通过血脑屏障（BBB）的最佳时间及机制,为NGF在临床上的应用提供新的用药途径。方法 对65只健康雄性SD大鼠采用改良的线栓法制备局灶性脑缺血模型,按缺血后不同时间点,给予每只大鼠注入125 Ⅰ-NGF,利用γ射线计数仪检测γ计数量,观察缺血不同时间BBB通透性的改变。脑缺血后BBB通透性最大的时间点（缺血3 d）大鼠为实验组（15只）,不制造脑缺血动物为对照组（15只）。对大鼠行脑冠状切片,通过放射自显影观察成影部位、面积及亮度。利用透射电子显微镜观察脑缺血不同时间BBB的变化。结果 随脑缺血时间延长,BBB通透性逐渐增加,其中脑缺血后第3天BBB通透性最大,而后又逐渐减弱。在BBB相同通透性情况下,γ计数显示,伴随用药时间的延长对照组和实验组γ计数均有所增加,实验组各时间点γ计数高于对照组,两组比较差异均有显著性（P＜0.05）,以用药 4 h γ计数最高。放射自显影结果显示,对照组给药1 h整个大脑不显影,4 h及7h脑室及室周脑组织显影,面积约占脑冠状切面的4.1%,亮度明显;实验组给药1 h右侧大脑表面显影,面积约占脑冠状切面的6%,亮度较强,4 h及7h除了脑室及室周脑组织显影外,在右侧脑额叶（FL）、顶叶（AL）和颞叶皮质和髓质中有大面积显影,面积占脑冠状切面的36.2%～47.3%,亮度很强。结论 随着大鼠脑缺血时间延长BBB通透性增加,其中缺血3 d通透性最强;大鼠局灶脑缺血3 d用药4 h 125Ⅰ-NGF进入BBB量最大,在额叶、顶叶及颞叶显影最明显。NGF能通过局灶性脑缺血的血脑屏障,可能为临床给药途径提出新思路。