远隔缺血预处理诱导大鼠缺血脑保护的基因表达谱分析

目的检测远隔缺血预处理对大鼠缺血半暗带脑组织基因表达的影响,探讨远隔缺血预处理诱导的内源性脑保护机制。方法雄性SD大鼠24只随机分为常规缺血组(n=12)和远隔预处理组(n=12)。常规缺血组应用远端大脑中动脉阻塞术(dMCAO)建立局灶性脑缺血模型;远隔预处理组给予左侧股动脉15 min夹闭/15 min再通3个循环缺血预处理后,再予以dMCAO。两组大鼠分别在缺血1、3、6和24 h后取脑,采用Agilent大鼠全基因组芯片检测两组脑缺血半暗带区基因表达的变化,获取差异表达基因,并对差异基因进行GO分析和Pathway功能注释(KEGG数据库)。结果远隔缺血预处理后:①缺血半暗带区脑组织基因出现差异表达(P0.05且差异倍数2),随着缺血时间延长差异基因个数逐渐增加,在缺血6 h达高峰;②GO分析结果提示差异基因涉及刺激反应、生长、生物调节、细胞杀伤等多个生物过程;③对差异基因进行KEGG通路分析,结果提示Jak-STAT信号通路、p53信号通路、丝裂原蛋白激酶(MAPK)信号通路、谷氨酸突触信号通路、血管内皮生长因子(VEGF)信号通路等多条分子通路出现差异调节,差异调节通路的数目也在缺血6 h达高峰。结论远隔缺血预处理诱导半暗带基因差异表达,差异基因通过多条分子通路影响多个生物过程。基因表达谱芯片为远隔缺血预处理脑保护机制提供了全面的数据和信息,对各个靶点、各条通路的进一步深入研究具有全面、广泛的指导意义。     

缺血预处理的脑保护机制

脑缺血预处理（ｃｅｒｅｂｒａｌ ｉｓｃｈｅｍｉｃ Ｐｒｅ－ｃｏｎｄｉｔｉｏｎｉｎｇ,ＣＩＰ）是指对脑预先进行短暂的亚致死性缺血预处理,以减轻随后发生的致死性脑缺血再灌注所造成的损伤,产生脑保护作用。１９９０年Ｋｉｔａｇａｗａ等在沙士鼠前脑缺血模型中首次观察到脑的ＣＩＰ现象［１］。近年来众多学者建立了各种动物模型,从多种角度对此现象进行了较为深入的研究,并取得一定进展,本文就缺血预处理的脑保护机制作一综述。１ ＣＩＰ现象的特点１．１ＣＩＰ的脑保护效应 ＣＩＰ的脑保护作用,主要体现在缩小局部脑缺血引起的…     

米诺环素对慢性脑缺血大鼠脑内Notch信号通路的影响

目的 观察慢性脑缺血后米诺环素是否可以通过调节Notch信号通路而发挥脑保护作用。方法 将健康雄性SD大鼠40只随机分为5组(n=8):假手术(Sham)组、缺血模型(Model)组、DAPT组、米诺环素(Min)组、DAPT+米诺环素(DAPT+Min)组; 双侧颈总动脉永久性结扎(2-VO)建立慢性脑缺血模型,给药1月后行为学检测大鼠的学习记忆能力,免疫组化和Western blot检测VEGF及Notch信号通路下游物质Hes1的表达水平。结果 与假手术组比较,缺血模型组大鼠的学习记忆能力降低(P<0.05); 米诺环素组与DAPT组比较,VEGF及Hes1的表达水平存在显著差异(P<0.01); 米诺环素组分别于与缺血模型组和DAPT+米诺环素组比较,DAPT+米诺环素组与DAPT组比较,学习记忆能力、VEGF及Hes1的表达水平存在显著差异(P<0.05)。结论 米诺环素可能通过对慢性脑缺血大鼠脑内Notch信号通路的调节来促进脑缺血后血管的新生,进而发挥脑保护作用。     

白发性缺血预处理对大脑中动脉区脑梗死体积及预后的影响

目的评价自发性缺血预处理(IP)对大脑中动脉(MCA)脑梗死的体积和预后的影响。方法对160例MCA脑梗死病例按在一周内有无TIA史分为A组(无TIA史)及B组(有TIA史),A组按全前循环脑梗死(TACI)、部分前循环脑梗死(PACI)、腔隙性前循环脑梗死(LACI)的脑梗死亚型分为A1组,A2组,A3组；B组按TACI、PACI、LACI脑梗死亚型分为B1组,B2组、B3组,对患者入院第1天及发病后第90天后的神经功能进行NIHSS评分,对发病后第90天脑梗死CT体积按Streiner公式进行计算。分别比较A1组与B1组,A2组与B2组及A3组与B3组在发病第1天NIHSS评分,发病后第90天NIHSS评分及发病后第90天脑梗死体积三方面的差异性。结果A1与B1组的脑梗死体积及NIHSS评分无显著差异(P＞0.05),A2、A3组的脑梗死体积及NIHSS评分明显高于B2、B3组(P＜0.05)。结论自发性IP能缩小MCA区PACI型及LACI型脑梗死范围,减少致残率,减轻临床症状,改善预后,但对TACI型脑梗死的体积和预后的影响在本研究中无显著差异。     

脑缺血预处理的脑保护作用

脑缺血预处理(NIPC)可启动机体自身保护机制,提高脑组织对缺血、缺氧的耐受性,减少脑的缺血性损害,其保护作用是明确的。本文分析了脑缺血预处理现象,根据其形成、特点、时间过程和对重复缺血性损害的反应,结合功能恢复情况以及临床上短暂性脑缺血发作的影响,对于IPC的机制和其他的预处理方法进行了比较,试图找到一种安全有效、能够直接用于临床的预处理手段,通过“防病”的方式预防缺血性脑卒中的发生,减轻缺血性脑损伤。     

丙酮酸乙酯通过Notch1和NF-κB信号修复大脑 中动脉闭塞诱导的神经损伤

目的 探讨丙酮酸乙酯（EP）对大脑中动脉闭塞（MCAO）诱导的神经功能损伤的作用及 其潜在的分子机制。方法 将40 只SD 雄性大鼠随机均分为4 组：假手术组（Sham）,大脑中动脉闭塞组 （MCAO）,大脑中动脉闭塞+5 mg/kg 丙酮酸乙酯组（MCAO+5EP）,大脑中动脉闭塞+10 mg/kg 丙酮酸乙酯 组（MCAO+10EP）。神经损伤严重程度评分（mNSS）及旋转试验检测大鼠神经损伤及恢复,实时定量PCR 检测脑源性神经营养因子（BDNF）和神经生长因子（NGF）、Notch1和核因子κB（ NF-κB）信号的表达。 结果 与MCAO 组相比,2 个剂量的EP均可降低mNSS 评分（均P＜ 0.05）,提高大鼠在15 r/min 旋转轴上 的持续时间（均P ＜ 0.05）,上调BDNF 和NGF mRNA 的表达（均P ＜ 0.05）,下调Notch1 和NF-κB mRNA 的表达（均P＜ 0.05）,且10 mg/kg 的EP 作用效果更好。结论 10 mg/kg 的EP具有较好的神经保护作用, 其机制可能与抑制Notch1 和NF-κB 信号有关。     

脑缺血预处理对脑功能保护的实验研究

目的:本实验探讨脑缺血预处理对脑功能的保护作用。方法:将39只大鼠随机分为5组:A组(空白对照组)、B组(实验对照组)、C组(致死性缺血组)、C1组及D2组(均为缺血预处理组)。对各组动物的双侧颈总动脉分别用微型动脉瘤夹予以夹闭或松开,以达到实验所需的脑缺血或再灌注时间;各组于不同缺血阶段后再灌注3天或7天取材观察指标。结果D1组、D2组海马CA1区神经元大部分保持完好,神经元密度高于未行预处理的C组(P<0.01)。结论:脑缺血预处理对脑功能具有明显的保护作用。     

大脑中动脉狭窄脑深部小梗死发病机制的研究

目的 研究大脑中动脉粥样硬化性狭窄患者脑深部小梗死的发生频率和发病机制。方法86例发病时间〈1周的急件脑梗死患者，行经颅多普勒超声、彩色超声、磁共振血管造影及功能磁共振成像等检查方法被明确诊断为症状性大脑中动脉粥样硬化性狭窄，并排除颈内动脉病变、心源性栓塞以及非动脉粥样硬化性狭窄。利用磁共振扩散加权成像观察梗死灶的形态学表现及特点，分析其发病机制。结果37例（43．02％）症状性大脑中动脉粥样硬化性狭窄患者存在深部小梗死，多呈孤立性单发病灶。其中内囊纹状体梗死及巨大腔隙者18例（20．93％），梗死灶体积多超过两个层面，少数病灶呈多灶分布的特点；直径较小的腔隙性梗死19例（22．09％），梗死灶体积〈15mm，多位于一个层面内。伴有大脑中动脉粥样硬化性狭窄的深部小梗死的患者，病情多不稳定，预后较差。所有脑深部小梗死患者均伴有大脑中动脉主干支狭窄，且梗死灶体积与大脑中动脉粥样硬化性狭窄程度有关，内囊纹状体梗死多见于大脑中动脉重度狭窄者。结论约50％大脑中动脉粥样硬化性狭窄患者存在脑深部小梗死，其发病机制与大脑中动脉粥样硬化斑块或斑块残端血栓蔓延堵塞深穿支动脉入口有关。对此类患者的治疗不同于经典的腔隙性脑梗死。     

Notch信号途径对胶质瘤干细胞的调控机制研究

神经胶质瘤是中枢神经系统最常见的恶性肿瘤,大多恶性程度高、侵袭性强。手术治疗目前仍是首选甚至是唯一的治疗手段,但远期效果往往不佳,因而复发率和病死率较高。目前已经发现,在人脑神经胶质瘤中存在具有神经干细胞特征的肿瘤细胞,即胶质瘤干细胞。这些肿瘤于细胞具有增殖快、可分化、易成瘤、耐药耐辐射等特点,因此被视为是胶质瘤恶性表型的根源和潜在的治疗靶点。     

大鼠脑缺血耐受与脑自体神经干细胞的增殖

背景：脑缺血耐受与脑自体神经干细胞均具有脑保护作用,但前者能否促使脑自体神经干细胞增殖,学者们报道不一致。 目的：明确缺血预处理与大鼠脑梗死后7 d海马区自体神经干细胞增殖的关系,以及其对大鼠脑梗死后神经行为学评分的影响。 方法：采用二次线栓法建立局灶-局灶性SD大鼠脑缺血耐受模型,40只SD雄性大鼠随机分为：假手术组,缺血组,假手术+缺血组,预缺血+缺血组,每组10只。脑梗死后3,7 d采用Zea-Longa评分方法进行神经行为学评分,运用荧光免疫组织化学技术检测大鼠脑缺血侧海马区BrdU标记阳性细胞数量。 结果与结论：脑梗死后3,7 d的Zea-Longa神经行为学评分,预缺血+缺血组低于缺血组、假手术+缺血组(P < 0.01),而缺血组和假手术+缺血组间差异无显著性意义(P > 0.05)。脑梗死后7 d缺血侧海马区BrdU标记阳性细胞数,缺血组、假手术+缺血组、预缺血+缺血组高于假手术组(P < 0.01);预缺血+缺血组高于缺血组、假手术+缺血组(P < 0.01);而缺血组和假手术+缺血组间差异无显著性意义(P > 0.05)。结果表明缺血预处理可促进大鼠脑梗死后海马区齿状回颗粒下层成体神经干细胞的增殖,并能改善其神经功能缺损症状。     

缺血预处理诱导脑缺血耐受时间依赖性的实验研究

目的探讨缺血预处理诱导缺血耐受的时间依赖性及可能的机制。方法健康SD大鼠110只,采用随机数字表法分为假手术(SS)+大脑中动脉阻塞(MCAO)组(简称SS组)和缺血预处理(IP)+MCAO组(简称IP组),每组50只,另10只备用。将两组大鼠再随机分为5个亚组(n=10),IP组给予预缺血10 min后分别于6 h、24 h、3 d、7d、14 d后给予大脑中动脉完全阻塞22 h,再灌注2 h;SS组未进行缺血预处理,而单纯暴露颈总动脉处的解剖结构10 min,余步骤同IP组。采用Swanson间接法和Nick&Spot图像采集分析系统计算各组大鼠梗死体积,光镜下观察各组脑组织病理变化,采用免疫组化方法检测大鼠脑组织胶质纤维酸性蛋白(GFAP)、脑源性神经营养因子(BDNF)表达。结果给予10 min的大脑中动脉短暂的缺血预处理能够明显减少24 h、3 d、7 d1、4 d后再次缺血后的梗死体积(P<0.05),而6 h时间点脑梗死体积与SS组比较无统计学差异(P>0.05)。与SS组比较,IP组各时间点GFAP和BNDF阳性表达增多(P<0.05),且间隔3 d、7 d组增多明显(P<0.05),14 d后开始逐渐降低。结论 IP不但能引起神经系统损害,而且能够诱导缺血耐受的产生;IP诱导的缺血耐受对IP后3~7 d发生的再缺血损害保护作用最强。BDNF表达基本与缺血耐受的保护作用时间规律一致,提示BDNF可能是脑缺血耐受的重要机制之一。     

TLR9 bone marrow chimeric mice define a role for cerebral TNF in neuroprotection induced by CpG preconditioning

Systemic preconditioning with the TLR9 ligand CpG induces neuroprotection against brain ischemic injury through a tumor necrosis factor (TNF)-dependent mechanism. It is unclear how systemic administration of CpG engages the brain to induce the protective phenotype. To address this, we created TLR9-deficient reciprocal bone marrow chimeric mice lacking TLR9 on either hematopoietic cells or radiation-resistant cells of nonhematopoietic origin. We report that wild-type mice reconstituted with TLR9-deficient hematopoietic cells failed to show neuroprotection after systemic CpG preconditioning. Further, while hematopoietic expression of TLR9 is required for CpG-induced neuroprotection it is not sufficient to restore protection to TLR9-deficient mice that are reconstituted with hematopoietic cells bearing TLR9. To determine whether the absence of protection was associated with TNF, we examined TNF levels in the systemic circulation and the brain. We found that although TNF is required for CpG preconditioning, systemic TNF levels did not correlate with the protective phenotype. However, induction of cerebral TNF mRNA required expression of TLR9 on both hematopoietic and nonhematopoietic cells and correlated with neuroprotection. In accordance with these results, we show the therapeutic potential of intranasal CpG preconditioning, which induces brain TNF mRNA and robust neuroprotection with no concomitant increase in systemic levels of TNF.     

Protective effects of remote ischemic preconditioning in rat hindlimb on ischemia-reperfusion injury

Three cycles of remote ischemic pre-conditioning induced by temporarily occluding the bilateral femoral arteries (10 minutes) prior to 10 minutes of reperfusion were given once a day for 3 days before the animal received middle artery occlusion and reperfusion surgery. The results showed that brain infarct volume was significantly reduced after remote ischemic pre-conditioning. Scores in the forelimb placing test and the postural reflex test were significantly lower in rats having undergone remote ischemic pre-conditioning compared with those who did not receive remote ischemic pre-conditioning. Thus, neurological function was better in rats having undergone remote ischemic pre-conditioning compared with those who did not receive remote ischemic pre-conditioning. These results indicate that remote ischemic pre-conditioning in rat hindlimb exerts protective effects in ischemia-reperfusion injury.     

Poly-IC preconditioning protects against cerebral and renal ischemia-reperfusion injury

Preconditioning induces ischemic tolerance, which confers robust protection against ischemic damage. We show marked protection with polyinosinic polycytidylic acid (poly-IC) preconditioning in three models of murine ischemia-reperfusion injury. Poly-IC preconditioning induced protection against ischemia modeled in vitro in brain cortical cells and in vivo in models of brain ischemia and renal ischemia. Further, unlike other Toll-like receptor (TLR) ligands, which generally induce significant inflammatory responses, poly-IC elicits only modest systemic inflammation. Results show that poly-IC is a new powerful prophylactic treatment that offers promise as a clinical therapeutic strategy to minimize damage in patient populations at risk of ischemic injury.     

远隔缺血预适应对脑小血管病患者认知功能障碍及血管内皮生长因子表达水平的影响

目的 探讨远隔缺血预适应(RIPC)对脑小血管病患者认知功能障碍及血管生长因子(VEGF)表达水平的影响.方法 收集145例脑小血管病患者为研究对象,随机分为RIPC组及对照组,RIPC组给予双上肢远隔缺血预适应干预及常规内科药物治疗,对照组仅给予常规内科药物治疗,观察2组患者开始干预24 h内、第1个月、第6个月的蒙...     

远端肢体缺血预处理抑制缺血性脑水肿的水通道蛋白磁共振分子成像(AQP MRI)评价

目的探讨经肢体远端缺血预处理后脑缺血大鼠模型在AQP MRI上的改变。方法 51只成年雄性SD大鼠随机分为3组:预处理组、脑缺血组和假手术组。预处理组和脑缺血组分别经LRP预处理或阴性预处理后经历1 h的大脑中动脉栓塞建立脑缺血模型。在恢复再灌后48 h行MRI检查采集T_2FLAIR、常规DWI及AQP MRI图像,并行TTC染色。结果预处理组大鼠的梗死面积较脑缺血组显著减少(P0.05)。预处理组病灶的T_2信号较假手术组显著升高(P0.001),但较脑缺血组显著降低(P0.001);预处理组的AQP ADC值较假手术组显著降低(P0.001),但相比脑缺血组有所增高(P0.05)。预处理组AQP ADC值的改变呈多样化,而脑缺血组的AQP ADC值表现为均匀地降低。预处理组大鼠AQP ADC的差值百分比变化与T_2值及传统ADC值的差值百分比变化之间均无明显相关(P0.05),而脑缺血组大鼠AQP ADC的差值百分比变化与T_2值的差值百分比变化呈负相关(r=-0.832,P0.01),与传统ADC的差值百分比变化呈正相关(r=0.822,P0.01)。结论 AQP MRI能够在体显示AQPs的分布情况,是帮助深入探索LRP的脑缺血保护作用分子机制的重要手段。     

缺血预处理对大鼠脑缺血再灌注后ICAM-1表达的影响

目的 通过研究脑缺血预处理对大鼠脑缺血再灌注后细胞间粘附分子-1(Intowellular adhesion molecule-1，ICAM-1)表达及多形核白细胞(Ploymorphonuclear leukocytes，PMNLs)浸润变化的影响．探讨预处理的延迟保护作用机制。方法采用四血管阻断法建立大鼠全脑-全脑缺血预处理模型。应用HE染色和免疫组化染色技术，观察脑缺血预处理后ICAM-1表达及多形核白细胞浸润的变化。结果(1)脑缺血再灌注后ICAM-1表达的阳性血管数量、PMNLs浸润数量均明显增加；(2)脑缺血预处理可明显减少缺血再灌注后ICAM-1的表达阳性血管数及PMNLs的浸润数量。结论一次性缺血预处理3min后48h可以少再次长时间缺血(30min)／再灌注(24h)的ICAM-1的表达和多形核白细胞浸润。ICAM-表达的下调和PMNLs浸润减少参与缺血预处理的保护作用。     

Peripheral vascular disease as remote ischemic preconditioning,for acute stroke

Objectives

Remote ischemic preconditioning (RIPC) is a powerful endogenous mechanism whereby a brief period of ischemia is capable of protecting remote tissues from subsequent ischemic insult. While this phenomenon has been extensively studied in the heart and brain in animal models, little work has been done to explore the effects of RIPC in human patients with acute cerebral ischemia. This study investigates whether chronic peripheral hypoperfusion, in the form of pre-existing arterial peripheral vascular disease (PVD) that has not been surgically treated, is capable of inducing neuroprotective effects for acute ischemic stroke.

Methods

Individuals with PVD who had not undergone prior surgical treatment were identified from a registry of stroke patients. A control group within the same database was identified by matching patient's demographics and risk factors. The two groups were compared in terms of outcome by NIH Stroke Scale (NIHSS), modified Rankin scale (mRS), mortality, and volume of infarcted tissue at presentation and at discharge.

Results

The matching algorithm identified 26 pairs of PVD-control patients (9 pairs were female and 17 pairs were male). Age range was 20–93 years (mean 73). The PVD group was found to have significantly lower NIHSS scores at admission (NIHSS ≤ 4: PVD 47.1%, control 4.35%, p < 0.003), significantly more favorable outcomes at discharge (mRS ≤ 2: PVD 30.8%, control 3.84%, p < 0.012), and a significantly lower mortality rate (PVD 26.9%, control 57.7%, p = 0.024). Mean acute stroke volume at admission and at discharge were significantly lower for the PVD group (admission: PVD 39.6 mL, control 148.3 mL, p < 0.005 and discharge: PVD 111.7 mL, control 275 mL, p < 0.001).

Conclusion

Chronic limb hypoperfusion induced by PVD can potentially produce a neuroprotective effect in acute ischemic stroke. This effect resembles the neuroprotection induced by RIPC in preclinical models.     

Notch信号通路在神经干细胞发生中的作用

Notch基因首先由Metz等发现,后因证实该基因功能缺失的果蝇其翅膀边缘会造成一些缺刻表型而得名.研究表明,Notch是一个高度保守的信号通路,广泛表达于无脊椎动物和哺乳动物等多个物种,在多种器官及细胞的发育过程中作为主要的仲裁信号通路决定细胞的命运,并且通过细胞间相互作用的方式精确地调节着细胞的生长、分化及凋亡[1].     

NMDA preconditioning protects against seizures and hippocampal neurotoxicity induced by quinolinic acid in mice

PURPOSE: N-methyl D-aspartate (NMDA) preconditioning has been used to prevent cellular death induced by glutamate or NMDA in cultured neurons. Quinolinic acid (QA)-induced seizures are used to average NMDA receptors-evoked neurotoxicity in animal models. The purpose of this study was to investigate the potential neuroprotective effects of NMDA preconditioning against QA-induced seizures and hippocampal damage in vivo. METHODS: Mice were pretreated with nonconvulsant doses of NMDA for different times before i.c.v. QA infusion and observed for the occurrence of seizures. Hippocampal slices from mice were assayed to measure cellular viability. RESULTS: NMDA preconditioning presented 53% protection against QA-induced seizures, as well as QA-induced cellular death in the hippocampus. The NMDA receptor antagonist, MK-801, prevented the protection evoked by NMDA preconditioning. The adenosine A1 receptor antagonist, CPT, prevented the protection evoked by NMDA preconditioning against QA-induced seizures, but not against QA-induced hippocampal cellular damage. The adenosine A1 receptor agonist, CPA, did not mimic the NMDA preconditioning-evoked protective effects. CONCLUSIONS: These results suggest that in vivo preconditioning with subtoxic doses of NMDA protected mice against seizures and cellular hippocampal death elicited by QA, probably through mechanisms involving NMDA receptors operating with adenosine A1 receptors.