纳洛酮对大鼠局灶性脑缺血再灌注损伤的保护作用

目的　观察纳洛酮对大鼠局灶性脑缺血再灌注损伤的保护作用。方法　用改良Longa法制作局灶性脑缺血再灌注损伤模型 ,观察不同剂量纳洛酮对模型大鼠神经功能障碍、脑梗死范围、脑组织病理改变以及血清中乳酸脱氢酶(LDH)和肌酸激酶 (CK)含量的影响。结果　纳洛酮能剂量依赖性地降低模型大鼠的血清LDH、CK ,减少梗死面积 ,降低神经功能行为评分 ,减轻脑组织病理改变。结论　纳洛酮对大鼠局灶性脑缺血再灌注损伤有一定的保护作用     

M型钾离子通道开放剂对脑梗死再灌注损伤的保护作用

目的探讨M型钾离子通道开放剂对缺血性脑卒中的脑保护作用及其可能机制。方法选取C57BL/6J小鼠60只,采用抽签法随机分为假手术组(10只,Sham组)、对照组(10只,MCAO组)、治疗组(40只,RTG组),按照给药时间的不同再将RTG组分为RTG 0h、RTG 1h、RTG 3h、RTG 6h四个亚组,每组10只。采用线栓法制作小鼠大脑中动脉缺血再灌注模型,于脑缺血2h后再灌注。RTG组给予瑞替加滨(10.5mg/kg),假手术组和缺血再灌注组给予等量生理盐水。采用TTC染色法检测脑梗死体积;Longa评分法进行神经功能评分;苏木素-伊红(HE)染色观察海马神经元的形态变化;免疫组化法和Western blotting法测定小鼠缺血半影区半胱氨酸蛋白酶-3(Caspase-3)和细胞膜蛋白CD40L的表达水平。结果假手术组未发现脑梗死病灶,海马神经元无明显改变,Caspase-3阳性细胞数少见,无CD40L表达。MCAO组和RTG组均可见大脑中动脉供血区梗死灶,但RTG四个治疗组梗死灶均较MCAO组明显缩小(P<0.05),RTG 6h组较RTG 0h、RTG 1h、RTG 3h给药组小鼠脑梗死体积增大,但差异无统计学意义(P>0.05)。RTG各组较MCAO组神经功能明显改善。MCAO组海马区脑组织肿胀与坏死明显,RTG各治疗组脑水肿和神经元坏死病理损害明显较轻。RTG组Caspase-3阳性细胞数较MCAO组明显减少(P<0.05),0、1、3h治疗组最为显著。RTG 0h、RTG 1h及RTG 3h组梗死周围区CD40L含量均较MCAO组明显下降(P<0.05),RTG6h下降不明显(P>0.05)。结论 M型钾离子通道开放剂瑞替加滨对缺血性脑卒中具有脑保护作用,其机制可能是降低神经细胞的兴奋性,减轻缺血半影区的炎症反应,从而抑制细胞凋亡。M型钾离子通道开放剂的脑保护作用具有时间依赖性,即超过一定的时间窗脑保护作用会减弱。     

川芎嗪对脑缺血后不同脑区神经元型NO合酶表达的影响

目的 通过观察脑缺血后不同时间、不同脑区神经元型NO合酶(nNOS)的表达,探讨川芎嗪对脑缺血后nNOS表达的影响.方法 成年雄性SD大鼠,随机分为假手术组、缺血模型组及川芎嗪低(20mg/kg)、中(40mg/kg)、高(80mg/kg)剂量组5组,每组按缺血后时间又分为1、3、7、14、21d 5个亚组.线栓法制作左侧大脑中动脉阻塞模型,术后2h腹腔注射不同剂量的川芎嗪(1次/d),至处死前2h.免疫组化染色观察脑缺血后不同时间侧脑室室下区(SVZ)、胼胝体(CC)、梗塞区周围纹状体和大脑皮质、海马CA1区及齿状回(DG) nNOS表达.结果 假手术组各脑区不同时间nNOS的表达相近,差异均无统计学意义(P>0.05).在SVZ,模型组1～14d nNOS表达降低,21d 时增高;川芎嗪各剂量组均表现为3～14d nNOS表达明显降低,21d明显增高.在CC,模型组3～14d明显降低,21d有所回升;芎嗪各剂量组nNOS表达均表现为3～14d明显降低,以中、高剂量组降低最为明显,21d增高.在缺血周围皮质和纹状体,模型组nNOS表达均表现为3、7d明显降低,14、21d呈明显增加趋势;川芎嗪各剂量组nNOS的表达3～21d均较低,以中、高剂量组7～14d降低最为明显,21d时稍有回升.在DG、CA1区,模型组3、7d nNOS表达明显降低,14d时增高;川芎嗪各剂量组3～21d nNOS的表达均降低.各脑区不同时间点nNOS表达模型组与川芎嗪各组间均存在明显差异(P<0.05).结论 川芎嗪对脑缺血后3～14d各脑区nNOS的表达有明显抑制作用,提示川芎嗪可能通过抑制nNOS的表达、减少NO产生发挥其脑保护作用.     

血管活性肠肽对暂时性脑缺血大鼠梗死体积的影响

目的通过活体实验研究血管活性肠肽(VIP)对脑缺血损伤的神经保护作用。方法大鼠侧脑室内注射VIP后,采用大脑中动脉线栓(MCAO)法诱导建立暂时性局部脑缺血模型。通过神经学的评价对MCAO大鼠进行判定和筛选;TTC染色测定大鼠脑梗死体积;双抗夹心ELISA检测血清S100β的含量。结果MCAO后2 h大鼠呈现不同程度的神经功能损伤症状。评定为1-2级的仅有2例出现明显脑梗死区,3级的为全部。VIP注射后大鼠脑梗死体积明显缩小,较对照组减少约25%(P<0.05);血清S100β浓度也明显降低(P<0.05)。结论以神经学评分3级作为MCAO模型成功的判定标准是较为可靠的。VIP脑内注射可明显减小脑缺血后梗死体积,具有神经保护作用。VIP可以降低S100β的含量,这可能涉及到VIP的另一神经保护机制。     

川芎嗪对大鼠局部脑缺血后空间学习和记忆的影响

目的探讨川芎嗪对大鼠局部脑缺血后空间学习记忆能力障碍的作用。方法线栓法制作大鼠大脑中动脉阻塞(MCAO)模型,术后2周内腹腔注射川芎嗪,第15天开始采用Morris水迷宫装置评价大鼠的空间学习记忆能力,脑切片尼氏染色观察皮质和海马的神经元数量变化。结果脑缺血对照组大鼠在定向航行试验和空间探索试验中均表现出明显的空间认知功能的障碍,川芎嗪治疗组大鼠平均逃避潜伏期与缺血对照组比较明显缩短(P<0.01)。在空间探索试验中,川芎嗪治疗组大鼠原平台象限停留时间百分比以及穿过原平台位置次数均大于脑缺血对照组(P<0.01)。在形态学观察中,川芎嗪治疗组大鼠缺血侧顶叶皮质神经元数量明显多于脑缺血对照组(P<0.01),各组动物海马神经元数量上无统计学差异。结论川芎嗪可以明显改善大鼠永久性局部脑缺血后空间学习记忆能力,其机制可能与川芎嗪对神经元的保护作用有关。     

葛根素对大鼠局灶性缺血脑组织内海马神经元型一氧化氮合酶表达的影响

目的 观察葛根素对大鼠局灶性脑缺血海马神经元神经型一氧化氮合酶(nNOS)表达的影响.方法 局灶性脑缺血模型由线栓法阻塞大鼠大脑中动脉制成,葛根素腹腔注射干预治疗,采用氯化三苯基四氮唑染色方法测定脑梗死体积,免疫组织化学方法测定大鼠脑缺血后不同时间点前后海马区nNOS阳性神经元表达的变化.结果 2h和12h干预组脑梗死体积明显小于对照组(P<0.05),24h干预组与对照组比较无统计学差异(P>0.05).缺血2h组海马nNOS阳性细胞开始增加,12h组最高,24h组较前有所下降;干预组大鼠海马nNOS阳性细胞数与对照组比较降低明显,有统计学差异(P<0.01).结论 nNOS参与早期脑缺血损伤,葛根素通过抑制nNOS表达对大鼠脑神经元损伤起保护作用.     

镇静剂量异丙酚对大鼠脑缺血再灌注损伤保护作用的形态学研究

目的　研究异丙酚对局灶性脑缺血再灌注损伤的影响。方法　采用改良Longa法制成大鼠局灶性脑缺血再灌注模型 ,观察不同镇静剂量的异丙酚对脑缺血再灌注损伤的形态学变化的影响。结果　 1 5～ 30mg/kg的异丙酚能减少缺血再灌注脑组织的含水量 ,明显缩小脑梗塞范围 ,改善电镜下细胞超微结构的损害。结论　 1 5～ 30mg/kg的异丙酚可以通过改善缺血再灌注损害后脑组织的形态学变化起到脑保护作用。     

不同镇静剂量异丙酚对局灶性脑缺血损伤的保护机制

目的　观察不同镇静剂量异丙酚对大鼠脑缺血再灌注损伤的保护作用 ,研究其保护作用的机制。方法　用改良Longa法制作局灶性脑缺血再灌注损伤的模型 ,观察不同镇静剂量异丙酚对模型组大鼠神经症状 ,血清中乳酸脱氢酶 (LDH)和肌酸激酶 (CK)含量的影响 ,测定脑组织中超氧化物歧化酶 (SOD)活性 ,脂质过氧化产物丙二醛 (MDA)含量。结果　实验发现 ,镇静大剂量及中剂量异丙酚组大鼠血清中LDH ,CK及脑组织中MDA含量明显低于模型组 ,脑组织中SOD的活性明显高于模型组 ,且呈一定量效关系。结论　镇静剂量异丙酚对大鼠脑缺血再灌注损伤有保护作用 ,其机制与清除氧自由基有关。     

丁苯酞对大鼠脑缺血再灌注损伤中氧化应激炎性通路的影响

目的探讨丁苯酞对大鼠脑缺血再灌注损伤的保护作用及其相关机制。方法实验分为假手术组、模型组和低、中、高剂量丁苯酞组及全反式维甲酸(ATRA)组。神经功能损伤评分(NDS)评价神经功能,2,3,5-三苯基四唑氯化物(TTC)染色计算缺血脑组织体积,黄嘌呤氧化酶法检测超氧化物歧化酶(SOD),硫代巴比妥酸比色法检测丙二醛(MDA),ELISA检测白介素(IL)-6和肿瘤坏死因子(TNF)-α,Real-time PCR检测血红素加氧酶(HO-1)基因表达,Western blot检测NF-E2相关因子2(Nrf2)和HO-1蛋白表达。结果低、中、高剂量丁苯酞组大鼠NDS评分、脑梗死体积、MDA、IL-6和TNF-α表达水平、HO-1基因2~(-△△Ct)值、Nrf2和HO-1蛋白表达均明显低于模型组(P<0.05)。低、中、高剂量丁苯酞组大鼠SOD明显高于模型组(P<0.05)。低、中、高剂量丁苯酞组大鼠NDS评分、脑梗死体积、MDA、IL-6和TNF-α表达水平、HO-1基因2~(-△△Ct)值、Nrf2和HO-1蛋白表达呈计量依赖性减低,而SOD呈计量依赖性升高(P<0.05)。结论丁苯酞可同过上调Nrf2/HO-1通路发挥抗氧化作用,对脑缺血再灌注大鼠起到神经保护作用。     

活血效灵丹对家兔脑出血后脑水肿和脑血肿的影响

目的　研究脑出血后应用活血效灵丹对脑水肿和脑血肿的作用。方法　以胶原酶Ⅶ诱导家兔脑出血模型 ,利用干湿重法检测脑组织含水量 ,研究不同时间给药 (6h、2 4h)对脑组织含水量的影响及脑血肿的治疗效果。结果造模后血肿侧脑组织含水量在第 4天、7天均较对照组增高 (P <0 .0 5～ 0 .0 0 1) ,且血肿侧高于非血肿侧 (P <0 .0 1～0 .0 0 1) ;经过活血效灵丹治疗后 ,6h和 2 4h治疗组血肿侧脑组织含水量均较模型组明显减少 ,其中 6h治疗组含水量减少略优于 2 4h治疗组 (P <0 0 5 )。治疗组在第 4天、7天 ,脑血肿的体积明显小于模型组 (P <0 .0 1～ 0 .0 5 ) ,但 6h和 2 4h治疗组血肿体积无明显差异。结论　脑出血后早期应用活血效灵丹对脑水肿和脑血肿具有治疗作用     

THE EFFECT OF LIGUSTRAZINE ON NEUROGENESIS IN CORTEX AFTER FOCAL CEREBRAL ISCHEMIA IN RATS

Objective To explore the effect of Ligustrazine on neurogenesis in cortex after focal cerebral ischemia in rats. Methods Focal cerebral ischemia was induced by left middle cerebral artery occlusion with a suture. Two hours later, injection of Ligustrazine （80 mg/kg, 1 time/d） was performed peritoneally. Four hours after the ischemia, 5-bromodeoxyuridine （BrdU） （50 mg/kg, 1 time/d） was injected peritoneally. At 7 d, 14 d and 21 d after ischemia, BrdU positive cells in the cortex were observed by cal staining. Results In ischemic model group, at 7 day, sparsely-distributed BrdU positive cells were observed in the Ⅱ-- Ⅵ layers of the ipsilateral cortex, with a bandlike distribution in ischemic penumbra. With the prolongation of ischemia, the number of BrdU positive cells increased. In Ligustrazine group, BrdU positive cells were also observed in theⅡ-- Ⅵ layers of the cortex, with an intense distribution in ischemic penumbra. The numbers of BrdU positive cells at 7 d, 14 d and 21 d were more than those in ischemic model group respectively. Conclusion Ligustrazine increases the proliferated cells in cortex after focal cerebral ischemia in rats. The results suggest that it may be useful for promoting self-repair after ischemia.     

PROLIFERATION AND MIGRATION OF EPENDYMAL CELLS IN THE BRAIN OF ADULT RATS AFTER PERMANENT FOCAL CEREBRAL ISCHEMIA

Objective Ependymal cells are thought to be the primary source of neural stem cells in the adult central nervous system. The purpose of this study is to examine spatial and temporal profiles of ependymal cell proliferation and migration after focal cerebral ischemia. Methods Eighty male Sprague Dawley rats underwent permanent middle cerebral artery occlusion after injection of 10/μL of 0.2% Dil into the lateral ventricle. Rats were sacrificed and brain sections were acquired for pathological evaluation and laser confocal imaging at day 1,3,7,11,14,21 and 28 after ischemia. Results The density of Dil-labeled cells in the ischemic ipsilateral subventricular zone was significantly higher than that in the control group and these labeled cells dispersed in the ischemic ipsilateral subventricular zone and/or were located in ependyma from day 1 to 11. In the ischemic ipsilateral cortex, some Diilabeled cells occurred in peri-infarction and infarction of parietal region at day14 and peaked at day 21 when some Dil-labeled cell nodules were found in this region. During postischemic day 14--28, a significant decrease in labeled celldensity in the ischemic ipsilateral subventricular zone was coincident with a significant increase in labeled cells density in the cortex (peri-infarction and infarction). Conclusion The results indicate that ependymal cells proliferate and migrate after focal cerebral ischemia in the adult rat brain.     

An experimental study of pioglitazone in treating vascular dementia

Objective To compare the therapeutic effects of different doses of pioglitazone, a kind of peroxisome proliferator-activated receptor γ (PPARγ) agonist, on vascular dementia and explore how pioglitazone affects cerebral ischemia. Methods Modified Pulsinelli's vessel ligation was used to establish a vascular dementia model in rats. Recognition, learning and memory were evaluated by Morris's water maze test. Immunoenzyme staining was used to determine the number of nerve cells. Immunofluorescence double-staining was used to examine the expression of PPARγ/nerve cells and PPARγ/astrocytes in different groups. Results Both in pioglitazone groups and sham-operation group, the latency was reduced significantly compared to that in control group (P<0.01). Sham-operation group had the largest number of neurons in the cortex, followed by low-dose pioglitazone group and high-dose pioglitazone group, and control group came last. Compared with control group, pioglitazone groups had more PPARγ expression in nerve cells, and the fluorescence intensity of PPARγ was stronger. Conclusion Pioglitazone can induce the expression of PPARγ in neuron endochylema and astrocyte endochylema to protect nerve cells, and then to improve spatial learning and memory function in VD rats.     

大鼠脑缺血淋巴细胞浸润的免疫组化研究

目的 探讨脑缺血后淋巴细胞浸润情况。方法 用免疫组化方法检测了30只大鼠大脑中动脉局部脑缺血12h、24h、3d、7d和10d后损伤区CD3、CD4和CD8阳性淋巴细胞。结果 缺血12～24h即可见CD3阳性和CD8阳性细胞浸润，3d和7d组阳性细胞最多。这些浸润的淋巴细胞位于反应带以及坏死中心和反应带的交界处。位于坏死区周围的CD3阳性和CD8阳性细胞主要为圆形，有大细胞和小细胞两种形态；位于反应带的淋巴细胞为小圆细胞，呈弥散分布，也可在微血管周围形成血管套。CD4阳性淋巴细胞数量较少，形态多样，大小不一。结论 淋巴细胞及亚群可能参与缺血性脑损伤的病理过程。     

脑缺血组织nestin蛋白重新表达及其与碱性成纤维细胞生长因子表达的关系

目的　研究缺血脑组织nestin蛋白重新表达和碱性成纤维细胞生长因子 (bFGF)表达的关系。方法　建立永久性脑缺血大鼠模型 ,采用免疫组化染色方法 ,观察脑缺血后 1、3、7、14、2 8d脑组织nestin蛋白和bFGF的表达。结果　缺血后 1～ 3d缺血灶附近nestin免疫阳性细胞大量出现 ,缺血 7d开始减少 ;而bFGF免疫阳性细胞在缺血 14d大量出现 ,缺血 2 8d开始减少。结论　缺血脑组织重新表达nestin蛋白不需要bFGF的营养和支持 ,缺血脑组织周围的nestin免疫阳性细胞来源于重演胚胎发育过程的星形胶质细胞     

参芎化瘀胶囊通过PI3-K/Akt信号通路改善大鼠缺血性脑损伤

目的探讨参芎化瘀胶囊对缺血性脑损伤的治疗作用及其机制。方法 SD大鼠分为假手术组、模型组及参芎化瘀胶囊高、低剂量组。改良的Pulsineli 4血管阻断(4-VO)法制作全脑缺血模型;HE染色观察海马区神经细胞的形态变化;免疫组织化学法检测海马区磷脂酰肌醇3-激酶(PI3-K)、蛋白激酶B(PKB/Akt)的表达;原位缺口末端标记法(TUNEL)检测凋亡细胞;八臂迷宫法测试动物学习记忆功能。结果与假手术组比较,模型组大鼠海马区神经元结构损伤明显,PI3-K、Akt表达增高,凋亡神经细胞数量增加,大鼠的学习记忆功能下降;与模型组比较,参芎化瘀胶囊组海马神经元形态结构损伤减轻,PI3-K、Akt表达进一步增多,凋亡神经细胞数量减少,动物学习记忆功能改善,上述变化在高剂量参芎化瘀胶囊更为明显。结论参芎化瘀胶囊对脑缺血损伤有很好的治疗作用,其机制可能与调控PI3-K/Akt信号途径有关。     

Role of candesartan against cerebral ischemia and oxidative damage in normotensive rats

Objective Angiotensin Ⅱ (Ang Ⅱ ) contributes to modulating blood pressure by stimulation of Ang Ⅱ AT1 receptors. We devised a rat transient middle cerebral artery occlusion (MCAO) model to assess whether oxidative damage is decreased after pretreatment with Angiotensin Ⅱ AT1 receptor blocker (ARB). Methods After 2 weeks pretreatment with ARB 0. 5 and 1 mg/kg, the male Wister rats were subjected to 2 h middle cerebral artery occlusion (MCAO). At 24 h, the lumen diameter of middle cerebral artery, the plasma level of 8-hydroxy-2'-deoxyguanosine (8-OHdG), and HIF-1 a levels were recorded and compared. Results After pretrcatment with ARB 0.5 and 1 mg/kg, blood pressure did not significantly change compared with that of controls. In the group of candesartan at 1 mg/(kg· day), the lumen diameter was significantly increased compared to that in control group [(86.0±5.0) μm vs. (69.0± 2.1) μm; P<0. 01, n = 6- 8]. The plasma 8-OHdG levels of ARB pretreatment groups were decreased. In immunohistochemical findings, 8-OHdG- and HIF-1α-containing cells in ARB pretreatment groups were decreased. Conclusion Brain ischemia and oxidative damage can be reversed by AT1 receptor blockade in normotensive rats after transient cerebral artery occlusion.     

Pretreatment with candesartan protects brain against ischemia in normotensive rats

Objective Angiotensin Ⅱ （Ang-Ⅱ ） increases NADPH oxidase activity and stimulates the production of reactive oxygen species （ROS） including superoxide anion through Ang Ⅱ AT1-receptor （AT1-R） activation. ROS is involved in various pathological processes in brain ischemia. We investigated whether the AT1-R blocker （ARB） candesartan can protect normotensive rats against brain ischemia. Methods After 2-week pretreatment with candesartan, rats were subjected to 2 hours middle cerebral artery occlasion-reperfusion （MCAO-R） and 24 hours later, the infarct volume, iNOS, and eNOS mRNA in the internal carotid artery was recorded and compared. Results Candesartan pretreatment reduced cerebral ischemia and oxidative brain damage after MCAO-R in normotensive rats, resulting in a decreased cortical infarct volume [0.5 mg/kg candesartan, （46. 8±13.2）mm^3 ; 1.0 mg/kg candesartan, （ 19.3± 15.3） mm^3 vs. control, （ 111.7 ±14.3 ） mm^3 ; P〈 0.05, P〈 0.01, respectively]. Candesartan pretreatment increased the eNOS mRNA level in the internal carotid artery. Conclusion In normotensive rats exposed to MCAO-R, candesartan protectes against brain ischemia. This effect may represent a significant therapeutic advantage and may induce end-organ protection even at normal blood pressure.     

自发性高血压大鼠不同脑区神经元的变化

目的观察自发性高血压大鼠(SHR)脑皮质和海马CA1,CA3区及齿状回(DG)神经元的变化。方法采用尾动脉测压法测定SHR和Wistar大鼠(对照组)收缩压、平均动脉压、舒张压和心率;甲苯胺兰染色观察额叶、顶叶、颞叶及海马CA1、CA3区及DG神经元数量改变。结果SHR组收缩压、舒张压和平均动脉压明显高于对照组(P<0.05),而心率两组无明显差异(P>0.05)。SHR组与对照组额叶、顶叶、颞叶神经元数量无明显差异(P>0.05),但海马CA1、CA3区及DG神经元较对照组明显减少(P<0.05,P<0.01)。结论SHR海马CA1、CA3区及DG神经元损害明显,存在神经元丢失,而脑皮质神经元丢失不明显。