冷冻致兔脑水肿模型的建立与改进

目的:建立和改进一种冷冻诱导的兔局限性脑水肿模型。方法:采用液氮冷冻探头单点置于兔一侧大脑半球硬膜外,时间为60s,制作成局灶性脑冷冻伤后脑水肿模型。6h后断头取脑,计算脑含水量和脑组织伊文氏蓝含量,观察伊文氏蓝染色范围及其病理改变。结果:实验组冷冻侧脑含水量比对侧及对照组左、右半球明显增加(P<0.001)。伊文氏蓝含量(mg/g干脑重)也较对侧及对照组左半球和右半球显著增多(P<0.001)。冷冻灶周围有明显的蓝染带,局部脑组织稍膨隆。光镜检查见细胞肿胀明显,透亮度增加,部分细胞空泡样变性,血管周围间隙显著增宽,并可见灶性出血,部分区域液化。电镜检查见部分神经细胞内细胞器(线粒体、内质网)肿胀或消失。部分核碎裂或溶解,血管基底膜肿胀或断裂等表现。对照组无类似改变。结论:本实验模型重复性较强,稳定可靠,条件较易控制。冷冻伤脑水肿是一种典型的血管源性脑水肿,与创伤性脑水肿有极其相似之处,可广泛应用于创伤性脑水肿的各种实验研究。     

冷冻伤性脑水肿神经细胞内钙离子浓度变化及尼莫地平的治疗作用

报告大鼠冷冻伤性脑水肿突触体内游离钙离子浓度([Ca~(2 )]_i)和脑组织水分含量变化之间的规律,并采用L-型钙离子通道阻滞剂尼莫地平进行治疗,研究其对神经细胞[Ca~(2 )]_i和脑水肿的影响.结果表明:冷冻后4h神经细胞内即已发生钙离子超载.伴随脑组织水分含量的增加.脑含水量随[Ca~(2 )]_i的升高而增加,两者呈正相关关系.应用尼莫地平治疗后神经细胞突触体内[Ca~(2 )]_i.明显下降,脑水肿亦明显减轻.     

急性局灶性脑挫裂伤后大鼠血脑屏障改变的实验研究

目的:研究急性局灶性脑挫裂伤后大鼠血脑屏障的改变及对脑水肿的影响。方法:采用Feeney's自由落体撞击法建立急性局灶性脑挫裂伤模型。每组6只测量伤侧脑组织伊文思蓝(Evans Blue,EB)含量、脑组织含水量。每组3只电镜观察微血管内皮细胞超微结构改变。结果:急性局灶性脑挫裂伤后24h脑组织含水量为(79.79±0.83)%,与正常对照组(78.68±0.63)%相比有明显差异(P<0.01),脑损伤后6h脑组织中EB含量为(362.12±28.16)ug/g,与正常对照组(11.89±2.28)ug/g相比有明显差异(P<0.01);脑损伤后30min,微血管内皮细胞有轻度受损迹象,伤后3h毛细血管腔内有微绒毛形成,伤后6h微绒毛增多,伤后24~72h毛细血管腔明显狭窄。结论:脑含水量的变化与脑组织中EB含量变化不同步,BBB的开放先于脑水肿的形成。BBB的开放与微血管的机械性损伤、内皮细胞吞饮小泡增加、内皮细胞紧密连接中断有关,也与早期缺血、缺氧关系密切。     

冷冻诱导的脑外伤性脑水肿模型

本文报道用改良 Klatzo 法建立冷冻诱导的免脑外伤性脑水肿模型。通过测定脑组织H_2O、Na~ 、K~ 、Ca~(2 )含量,伊文思蓝染色、光镜、电镜检查,发现冷冻伤后早期出现脑细胞内水肿,继以血脑屏障破坏脑细胞外水肿,并随时相递剧。同时脑组织 Na~ 、Ca~(2 )含量渐升高、K~(?)减少,文中介绍建立稳定的兔脑冷冻伤脑水肿模型的经验,并建议供创伤性脑水肿脑细胞代谢障碍研究之用。     

大鼠脑挫裂伤半暗带水肿与血-脑屏障破坏的关系

目的研究大鼠局灶性脑挫裂伤半暗带的水肿变化与血-脑屏障(BBB)破坏的关系。方法将126只雄性SD大鼠随机分为3组:假手术组、挫裂伤组和给药组,采用Feeney法制作脑挫裂伤模型,给药组采用曲克芦丁脑蛋白水解物合剂腹腔给药,分别用伊文思蓝(EB)染色法和干湿法观察BBB的变化和脑组织的水肿情况,同时电镜观察超微结构。结果EB染色显示:挫裂伤组和给药组在伤后1 h EB开始漏出,6 h最严重。水含量测定结果显示:挫裂伤组和给药组伤后6 h水含量开始增加,72 h增至高峰。与挫裂伤组比较,给药组脑组织EB溢出量、水含量降低(P<0.05)。结论大鼠脑挫裂伤半暗带BBB通透性改变早于脑水肿的发生,提示BBB破坏可能是早期创伤性脑水肿的结构基础。曲克芦丁脑蛋白水解物合剂可通过改善BBB等多种途径治疗脑挫裂伤。     

高渗盐复合液治疗大鼠冷冻性脑水肿的实验研究

目的探讨高渗盐复合液(HSH)对大鼠冷冻性脑水肿的治疗作用。方法运用大鼠冷冻性脑水肿动物模型,分别以三种剂量(2ml/kg、4ml/kg、8ml/kg)的HSH、20%甘露醇(MNT,4ml/kg)和0.9%NaCl(4ml/kg)进行治疗,其中0.9%NaCl组为假治疗组,其余为治疗组;监测颅内压,测定脑含水量。结果治疗组给药2h内均有同样的降颅压作用,其中2ml/kgHSH组脱水作用时间明显短于其它三组、降颅压效果弱于其它三组;HSH4ml/kg和8ml/kg组具有相似的降颅压作用,无显著差异。治疗组健侧大脑半球含水量与假治疗组比较明显降低;治疗组伤侧大脑半球含水量与假治疗组对应侧相比稍减少,差异不显著。结论HSH可降低颅内压、减少脑含水量,其降颅压作用与MNT类似,作用呈量效关系,临床推荐的最佳剂量为4ml/kg。     

黄体酮对局灶性脑缺血再灌注大鼠水通道蛋白-4表达及血脑屏障通透性的影响

目的研究黄体酮对大鼠局灶性脑缺血再灌注后脑组织内水通道蛋白-4(AQP4)的表达及血脑屏障通透性的影响。方法健康雄性SD大鼠96只,随机分为4大组:假手术组、手术组、溶剂治疗组和黄体酮治疗组,建立大脑中动脉栓塞再灌注(MCAO/R)模型,分别在缺血2h再灌注6h、1d、3d、5d 4个时间点将大鼠麻醉后断头取脑,采用Western blot法、伊文氏蓝(EB)渗出量及干湿重法分别测定脑组织AQP4的表达、血脑屏障的通透性及脑含水量。结果手术组AQP4的表达、EB的含量及脑组织含水量明显高于假手术组;黄体酮组AQP4的表达、EB的含量及脑组织含水量较手术组及溶剂组明显降低,差异有统计学意义。结论黄体酮可以降低缺血再灌注大鼠脑组织AQP4的表达,从而降低血脑屏障的通透性,减轻脑水肿。     

大鼠创伤性脑内出血中红细胞对脑水肿的影响

目的研究红细胞对创伤性脑内出血(TICH)后脑水肿的影响。方法自由落体打击造成大鼠脑外伤,借助鼠脑立体定向仪向伤区脑皮质内注射全血(WB)、溶解红细胞(LRBC)或压积红细胞(PRBC),制作TICH模型。于伤后1、3、5d处死大鼠,取伤区脑组织测含水量及Na 、K 、Ca2 含量,并作病理检查。结果(1)TBI、WB和PRBC3组第3d的脑含水量最高,LRBC组第1d的含水量最高;4组间比较,第1d时LRBC含水量最高,第3d时PRBC和WB组含水量最高。(2)Na 的变化与含水量的变化相一致。结论TICH所形成的脑水肿较单纯脑外伤重。在TICH中,红细胞对早期脑水肿的形成无明显作用,而对迟发性脑水肿的形成有重要作用。     

bFGF对脑缺血再灌注大鼠ICAM-1表达及脑组织含水量的影响

目的探讨bFGF对局灶性缺血再灌注大鼠脑组织含水量及脑组织ICAM-1水平的影响。方法SD大鼠48只,随机分为假手术组（n=16）、缺血再灌注组（n=16）和bFGF组（n=16）。应用线栓法制作大鼠局灶性脑缺血再灌注模型,大脑中动脉阻塞1h再灌注损伤24h,bFGF组伤后即刻一次性经腹腔注射bFGF（10g／kg）,假手术组和损伤组以相同方法给予0．9％的生理盐水。采用干湿法检测各组大鼠脑组织含水量,采用伊文思蓝（evansblue,EB）法检测脑毛细血管通透性,采用免疫组化法检测大鼠脑组织ICAM-1水平。结果与假手术组比较,缺血再灌注组脑含水量、脑皮质EB含量及ICAM-1表达显著增加（P〈0．05）,与缺血再灌注组比较,bFGF组脑含水量、脑皮质EB含量及ICAM-1表达较模型组显著性降低（P〈0．05）。结论ICMA-1表达增加是脑缺血再灌注后脑水肿形成和缺血性损伤的重要原因之一,减少ICAM-1表达和脑组织含水量推测是bFGF脑保护作用机制之一。     

雪莲黄酮对大鼠创伤性脑水肿的治疗作用及其机制的初步研究

目的研究雪莲黄酮对大鼠创伤性脑水肿的治疗作用及其机制。方法采用液压冲击法制作大鼠脑创伤模型,用雪莲黄酮注射液进行治疗。治疗后不同时间取伤灶及其周围脑组织进行含水量和与脂质过氧化有关的一些指标进行检测。结果脑创伤后6h、24h及3d伤灶及其周围脑组织中含水量及丙二醛含量明显增加(P<0.01),而超氧化物岐化酶含量明显减少(P<0.01),但雪莲黄酮能逆转上述变化。结论雪莲黄酮对大鼠创伤性脑水肿有一定的治疗作用,作用机制可能与其抗脂质过氧化作用有关。     

大鼠重型颅脑损伤急性期水通道蛋白4的表达

目的探讨水通道蛋白（AQP4）在大鼠重型脑外伤急性期的表达变化及其与脑水肿间的关系。方法49只成年雄性SD大鼠,随机分为对照组及实验组（伤后4h、8h、12h、24h、5d共5组）。制作重度冲击加速性损伤模型,分别于伤后4h、8h、12h、24h、72h、5d采用干湿比重法测脑组织含水量,原子吸收分光光度法测定Na^＋、K^＋含量,Evans Blue（EB）测定法观察大鼠血-脑屏障（BBB）通透性变化,半定量逆转录聚合酶链反应（RT-PCR）检测脑组织AQP4 mRNA表达及其变化。结果脑组织AQP4 mRNA在伤后4h开始表达上调,8h、12h依次增高,24h达到峰值（P〈0.05）,3d时仍维持较高水平,伤后5d有所降低。脑含水量、Na^＋含量的变化与AQP4 mRNA表达变化一致。经相关性分析,AQP4 mRNA的表达与脑含水量及脑EB含量均呈正相关（P〈0.05）。结论重型脑损伤急性期,AQP4 mRNA表达的变化与颅脑损伤后BBB的破坏及脑水肿的形成和发展密切相关。AQP4可能参与重型脑损伤后脑水肿的形成并起重要作用。     

创伤性脑水肿的发生机制及Iloprost的治疗作用

目的 探讨创伤性脑水肿的发生机制及前列环素的衍生物Ｉｌｏｐｒｏｓｔ对创伤性脑水肿的治疗作用。方法 干湿重法测定脑组织含水量,甲酰胺法测定伊思蓝（ＥＢ）含量,Ｆｕｒａ－２／ＡＭ荧光标记法测定突触全内「Ｃａ＾２＋」ｉ,并采用前列腺环素稳定的衍生物Ｉｌｏｐｒｏｓｔ进行治疗,研究其对ＥＢ含量、「Ｃａ＾２＋」ｉ及脑水肿的影响。结果 创伤后２ｈ即已发生钙超载,脑组织水含量及ＥＢ含量的增加,Ｉｌｏｐｒｏｓｔ治     

Peripheral thermal injury causes early blood-brain barrier dysfunction and matrix metalloproteinase expression in rat

High mortality incidence after serious systemic thermal injury is believed to be linked to significant increases in cerebral permeability, ultimately leading to irreversible blood-brain barrier (BBB) breakdown. The aim of this study was to investigate whether disruption of microvascular integrity in a rat thermal injury model is associated with early matrix metalloproteinase (MMP) expression. A total of 35 Sprague-Dawley rats were studied in thermal injury and control groups, each group containing two subgroups, one for brain edema and Evans blue analysis and another for MMP mRNA analysis. Thermally injured animals were anesthetized and submerged vertically in 85 degrees C water to the neck for 6 seconds producing a third degree burn affecting 70% of the total body surface area. BBB integrity was determined by measuring amount of Evans blue after 7 hours of injury with a spectrophotometer. Brain edema was detected by calculating water content. Brain mRNA levels were determined with real-time PCR 3 and 7 hours post-injury. Brain water content was significantly increased after peripheral injury at hour 7. Evans blue leakage was also significantly increased at the same time, suggesting an impaired BBB function after injury. Expressions of MMP-2 and MMP-9 mRNA in brain were increased as early as 3 hours after injury and remained at hour 7. Our study demonstrated a significant increase in cerebral permeability that occurs after serious systemic thermal injury. The underlying mechanisms could be related to early expression of MMPs.     

Brain edema and tumor necrosis factor-like weak inducer of apoptosis in rats with cerebral ischemia

BACKGROUND： Recent studies have demonstrated that tumor necrosis factor-like weak inducer of apoptosis （TWEAK） participates in brain edema. However, it is unclear whether blood-brain barrier （BBB） disruption is associated with TWEAK during the process of brain edema OBJECTIVE： To investigate the effects of TWEAK on BBB permeability in brain edema. DESIGN, TIME AND SETTING： An immunohistochemical observation, randomized, controlled animal experiment was performed at the Laboratory of Neurosurgical Anatomy, Xiangya Medical College, Central South University ＆ Central Laboratory, Third Xiangya Hospital, Central South University between January 2006 and December 2007. MATERIALS： A total of 48 adult Wistar rats were randomly divided into three groups： normal control （n = 8）, sham-operated （n = 8）, and ischemia/reperfusion （n = 32）. Rats from the ischemia/reperfusion group were randomly assigned to four subgroups according to different time points, i.e., 2 hours of ischemia followed by 6 hours （n = 8）, 12 hours （n = 8）, 1 day （n = 8）, or 12 days （n = 8） of reperfusion. METHODS： Focal cerebral ischemia/reperfusion injury was induced by middle cerebral artery occlusion （MCAO） using the suture method in rats from the ischemia/reperfusion group. Thread was introduced at a depth of 17-19 mm. Rats in the sham-operated group were subjected to experimental procedures similar to the ischemia/reperfusion group; however, the introducing depth of thread was 10 mm. The normal control group was not given any intervention. MAIN OUTCOME MEASURES： TWEAK expression was examined by immunohistochemistry; brain water content on the ischemic side was calculated as the ratio of dry to wet tissue weight; BBB permeability was measured by Evans blue extravasation. RESULTS： A total of eight rats died prior to and after surgery and an additional eight rats were randomly entered into the study. Thus 48 rats were included in the final analysis. In the ischemia/reperfusion group, TWEAK-positiv     

Peripheral thermal injury causes early blood–brain barrier dysfunction and matrix metalloproteinase expression in rat

Abstract

High mortality incidence after serious systemic thermal injury is believed to be linked to significant increases in cerebral permeability, ultimately leading to irreversible blood–brain barrier (BBB) breakdown. The aim of this study was to investigate whether disruption of microvascular integrity in a rat thermal injury model is associated with early matrix metalloproteinase (MMP) expression.

A total of 35 Sprague–Dawley rats were studied in thermal injury and control groups, each group containing two subgroups, one for brain edema and Evans blue analysis and another for MMP mRNA analysis. Thermally injured animals were anesthetized and submerged vertically in 85°C water to the neck for 6 seconds producing a third degree burn affecting 70% of the total body surface area. BBB integrity was determined by measuring amount of Evans blue after 7 hours of injury with a spectrophotometer. Brain edema was detected by calculating water content. Brain mRNA levels were determined with real-time PCR 3 and 7 hours post-injury.

Brain water content was significantly increased after peripheral injury at hour 7. Evans blue leakage was also significantly increased at the same time, suggesting an impaired BBB function after injury. Expressions of MMP-2 and MMP-9 mRNA in brain were increased as early as 3 hours after injury and remained at hour 7.

Our study demonstrated a significant increase in cerebral permeability that occurs after serious systemic thermal injury. The underlying mechanisms could be related to early expression of MMPs.     

脑血疏口服液对大鼠脑缺血再灌注损伤后血脑屏障的影响

目的 探讨脑血疏口服液对大鼠脑缺血再灌注损伤后血脑屏障的影响。方法 将120只SD大鼠随机分为3组:假手术组、对照组,脑血疏组; 采用线栓法建立大鼠左侧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; 采用Longa FZ 5级评分法进行大鼠神经功能缺损评分; TTC染色计算脑梗死体积百分比; 运用干-湿重法测脑含水率; 通过伊文思蓝( EB)含量反映血脑屏障的损伤程度; 免疫组化检测基质金属蛋白酶-9(MMP-9)的表达水平。结果(1)假手术组大鼠在神经功能缺损评分、脑梗死体积、脑含水率均低于对照组(P<0.01); 脑组织中EB含量和MMP-9表达水平较对照组低(P<0.01);(2)脑血疏组大鼠的神经功能缺损评分较低、脑梗死体积较小,脑水肿程度较轻; EB含量和MMP-9表达水平均较对照组明显减少(P<0.01)。结论 脑血疏口服液对大鼠脑缺血再灌注损伤后血脑屏障具有保护作用,其机制可能是通过抑制MMP-9的表达。     

H-7在脑出血后脑损伤中作用机制的研究

目的探讨蛋白激酶C(PKC)抑制剂异喹啉磺酰类(H-7)在脑出血后脑损伤中的作用机制。方法SD雄性大鼠随机分为对照组、假手术组、脑出血组、H-7治疗组,采用自体血注入法建立大鼠脑出血模型,治疗组应用H-7每12 h腹腔注射一次。分别用伊文思蓝(EB)测血脑屏障(BBB)通透性,干-湿重法测脑组织含水量,同时进行组织病理学观察。结果H-7治疗组可以明显改善大鼠脑出血后BBB通透性(P<0.05),血肿周围脑水肿明显减轻(P<0.05),并且减少脑出血后血肿周围炎症细胞浸润。结论H-7通过抑制PKC,减轻脑出血后脑水肿?血脑屏障的开放和炎症细胞浸润,揭示PKC参与了脑出血后脑损伤。     

Correlation of aquaporin-4 expression to blood-brain barrier permeability in rats with focal cerebral ischemia

BACKGROUND: Ischemic cerebrovascular disease causes injury to the blood-brain barrier. The occurrence of brain edema is associated with aquaporin expression following cerebral ischemia/reperfusion. OBJECTIVE: To analyze the correlation of aquaporin-4 expression to brain edema and blood-brain barrier permeability in brain tissues of rat models of ischemia/reperfusion. DESIGN, TIME AND SETTING: The randomized control experiment was performed at the Jiangsu Province Key Laboratory of Anesthesiology, Xuzhou Medical College, China from December 2006 to October 2007. MATERIALS: A total of 112 adult, male, Sprague-Dawley rats, weighing 220-250 g, were used to establish rat models of middle cerebral artery occlusion and reperfusion by the suture method. Rabbit anti-aquaporin-4 (Santa Cruz, USA) and Evans blue (Sigma, USA) were used to analyze the tissue. METHODS: The rats were randomized into sham-operated (n = 16) and ischemia/reperfusion (n = 96) groups. There were 6 time points in the ischemia/reperfusion group, comprising 4, 6, 12, 24, 48, and 72 hours after reperfusion, with 16 rats for each time point. Rat models in the sham-operated group at 4 hours after surgery and rat models in the ischemia/reperfusion group at different time points were equally and randomly assigned into 4 different subgroups. MAIN OUTCOME MEASURES: Brain water content on the ischemic side and the control side was measured using the dry-wet weight method. Blood-brain barrier function was determined by Evans Blue. Aquaporin-4 expression surrounding the ischemic focus, as well as the correlation of aquaporin-4 expression with brain water content and Evans blue staining, were measured using immunohistochemistry and Western blot analysis. RESULTS: Brain water content on the ischemic side significantly increased at 12 hours after reperfusion, reached a peak at 48 hours, and was still high at 72 hours. Brain water content was greater on the ischemic hemispheres, compared with the control hemispheres at 6, 12, 24, 48, and 72 hours after reperfusion, as well as both hemispheres in the sham-operated group (P<0.05). Evans blue content significantly increased on the ischemic side at 4 hours after ischemi',dreperfusion, and reached a peak at 48 hours. Evans blue content was greater on the ischemic hemispheres, compared with the control hemispheres at various time points, as well as both hemispheres in the sham-operated group (P<0.05). Aquaporin-4-positive cells were detected in the cortex and hippocampus, surrounding the ischemic penumbra focus, at 4-6 hours after ischemia/reperfusion. The number of positive cells significantly increased at 12 hours and reached a peak at 48-72 hours. Aquaporin-4 was, however, weakly expressed in the control hemispheres and the sham-operated group. The absorbance ratio of aquaporin-4 to β-actin was greater at 12, 24, 48, and 72 hours following cerebral ischemia/reperfusion, compared with the sham-operated group (P<0.05). Aquaporin-4 expression positively correlated to brain water content and Evans blue staining following cerebral ischemia/reperfusion (r1 = 0.68, r2= 0.81, P<0.05). CONCLUSION: Aquaporin-4 is highly expressed in brain tissues, participates in the occurrence of ischemic brain edema, and is positively correlated to blood-brain barrier permeability following cerebral ischemia/reperfusion.     

Effect of hypothermia on blood-brain barrier permeability following traumatic brain injury in chronically ethanol-treated rats

The objective of this study was to investigate the effect of hypothermia on the blood-brain barrier (BBB) disruption caused by traumatic brain injury (TBI) in chronically ethanol-treated rats. BBB permeability was measured using Evans blue (EB) dye. Arterial blood pressure levels of animals in hypothermic groups decreased significantly. The EB dye extravasation into the brain significantly increased in hypothermia and at 6 and 24 h after TBI. In ethanol-treated rats that were subjected to TBI, hypothermia led to a significant decrease in EB dye content in the brain at 24 h but not at 6 h after TBI when compared with TBI alone.