Ro25-6981对缺血再灌注大鼠脑室下区神经干细胞增殖的影响

目的:探讨N-甲基-D-天冬氨酸(NMDA)受体亚单位NR2B特异性拮抗剂Ro25-6981对缺血再灌注大鼠脑室下区神经干细胞增殖的影响及可能的机制。方法:线栓法制作大鼠大脑中动脉栓塞2 h再灌注模型,随机分成假手术组、缺血再灌注对照组和Ro25-6981干预组。免疫组织化学显色观察脑室下区巢蛋白和增殖细胞核抗原(PCNA)阳性细胞数,行免疫印迹对缺血侧脑组织脑源性神经营养因子(BDNF)蛋白表达水平进行定量分析。结果:缺血再灌注后脑室下区巢蛋白和PCNA阳性细胞较假手术组增加,而Ro25-6981干预组巢蛋白和PCNA阳性细胞较缺血再灌注对照组减少。缺血再灌注损伤促进了缺血侧脑组织BDNF蛋白的表达,Ro25-6981干预下调了缺血侧脑组织BDNF蛋白的表达。结论:NMDA受体亚单位NR2B能促进脑室下区神经干细胞的增殖,可能与调节BDNF表达有关。     

碱性成纤维细胞生长因子对全脑缺血再灌注大鼠脑皮质内源性神经干细胞的影响

探讨碱性成纤维细胞生长因子(bFGF)对全脑缺血再灌注大鼠脑皮质内源性神经干细胞增殖、迁移和分化的影响。本研究以"四血管法"制作全脑缺血再灌注模型,采用免疫组化方法检测BrdU和Nestin在不同组别各时间点的表达,用RT-PCR技术半定量检测大鼠脑皮质NSE mRNA的表达。结果显示假手术组大鼠脑皮质基本未见到BrdU和Nestin阳性细胞表达,NSE mRNA表达没有变化;模型组大鼠脑皮质的BrdU和Nestin阳性细胞在全脑缺血再灌注3d后开始增加,7d达到高峰,NSE mRNA表达无明显增加;bFGF治疗组BrdU、Nestin阳性细胞较模型组在缺血各时间点均明显增加(P0.05),11d达到高峰,NSE mRNA表达与假手术组和模型组相比第15d表达有明显增加(P0.05)。结果表明全脑缺血再灌注能够引起内源性神经干细胞的原位增殖,bFGF可促进全脑缺血再灌注大鼠内源性神经干细胞原位增殖,并延长增殖期,而且有明显促进内源性神经干细胞向神经元分化的作用。     

依达拉奉对永久性脑缺血大鼠脑内Nestin表达的影响

目的观察Nestin在永久性脑缺血大鼠脑内的变化及依达拉奉干预的影响。方法制作大鼠大脑中动脉闭塞(MCAO)SD大鼠模型,应用抗Nestin和抗Nestin/GFAP抗体进行免疫组化单标和免疫荧光双标染色。结果缺血后脑组织有大量Nestin阳性表达,阳性细胞主要分布于缺血侧侧脑室的室管膜下区(SVZ)、梗死灶周围大脑皮质和纹状体。细胞呈星状多突起,形似星形胶质细胞,且脑缺血后的大部分Nestin阳性细胞与星形胶质细胞标记物GFAP有共表达。缺血侧SVZ的Nestin阳性细胞数和免疫阳性表达强度于脑缺血后3 d达到高峰;梗死灶周围大脑皮质和纹状体区阳性细胞数和阳性表达强度分别于缺血后3 d和1周达到高峰;部分细胞聚集于梗死灶边缘,形成一个明显的梗死灶边缘带。经依达拉奉干预后,Nestin阳性细胞计数和表达强度与盐水组比较均升高(P<0.05),尤其在缺血治疗后3 d和1周(P<0.01)。结论脑缺血后,SVZ以及梗塞灶周围大脑皮质和纹状体区域大鼠Nestin阳性细胞数量增多、表达增强;经依达拉奉干预治疗后,Nestin阳性细胞的表达进一步增强,提示依达拉奉治疗可能促进脑缺血后神经干细胞的增生和分化,促进损伤组织的修复,发挥神经保护作用。     

龟板对局灶性脑缺血再灌注后Nestin表达的影响

目的 :探讨补肾中药龟板对局灶性脑缺血再灌注后神经干细胞Nestin表达的作用。方法 :采用大脑中动脉线栓法造成局灶性脑缺血再灌注模型 ,应用免疫组织化学技术检测神经干细胞巢蛋白 (Nestin)的表达 ,观察龟板对脑缺血后神经干细胞巢蛋白表达的影响。结果 :脑缺血再灌注后 7d ,龟板组神经病学评分明显低于缺血对照组 :龟板组缺血侧室管膜、室管膜下区、皮层和纹状体Nestin阳性细胞数显著多于缺血对照组 (P <0 .0 5 )。结论 :补肾中药龟板能上调脑缺血再灌注后Nestin的表达 ,这可能是其治疗缺血性脑血管病的机理之一。     

Ro25-6981对脑缺血再灌注模型大鼠海马齿状回颗粒下层nestin和BrdU表达的影响

目的:研究Ro25-6981对成年大鼠短暂性脑缺血再灌注海马齿状回颗粒下层(subgranular zone,SGZ)神经干细胞增殖的影响。方法:健康成年雄性SD大鼠在进行侧脑室置管7 d后,随机分为正常组、假手术组和脑缺血再灌注组,每组又分别再分为Ro25-6981组和生理盐水组。缺血再灌注组大鼠采用四动脉阻断前脑缺血再灌注模型,即缺血15 min再灌注1、3、7、14、21、28 d和35 d,缺血前15 min经侧脑室套管注射Ro25-6981或生理盐水。各组大鼠均采用免疫组织化学显示SGZ区Brd U、nestin阳性细胞的表达情况,采用免疫荧光双重标记法检测再灌7 d时SGZ区Brd U/nestin双标阳性细胞数量的改变。结果:脑缺血再灌注生理盐水组SGZ区nestin阳性细胞表达再灌1 d时开始增多,7 d时达到峰值,14 d时急剧下降,至35 d时降至正常水平;缺血前给予Ro25-6981,各时间点的nestin阳性细胞表达均有所下降,其中在3、7 d和14 d时减少明显,差异有统计学意义(P0.05)。脑缺血再灌注生理盐水组SGZ区Brd U阳性细胞再灌1 d开始增殖,7 d达到峰值,35 d降至正常水平;缺血前给予Ro25-6981,各时间点的Brd U阳性细胞表达均有所下降,其中在3、7、14 d和21 d时减少明显,差异有统计学意义(P0.05)。与脑缺血再灌注生理盐水组相比,Ro25-6981可以降低脑缺血再灌7 d海马SGZ区的Brd U/nestin双标阳性细胞密度,差异有统计学意义(P0.05)。结论:Ro25-6981可以抑制前脑缺血再灌注后海马SGZ区神经干细胞的增殖,提示NR2B参与并促进了前脑缺血再灌注引起的神经干细胞的增殖。     

局灶性脑缺血后侧脑室室下区神经发生及其与血管内皮生长因子关系的研究

为了研究成年大鼠局灶性脑缺血后侧脑室室下区(SVZ)神经发生的情况及其与血管内皮生长因子(VEGF)的关系,探讨脑缺血后神经发生及其调控机制,本研究通过大脑中动脉阻断法(MCAO)建立大鼠局灶性脑缺血模型,5-溴-2-脱氧尿核苷(BrdU)标记增殖的神经前体细胞,用免疫荧光双标记法动态检测BrdU、TuJ1、MAP-2、GFAP的表达,同时观察增殖细胞表达VEGF及其受体情况。结果显示:与对照组相比,大鼠SVZ的BrdU阳性细胞数在脑缺血后4 d组明显增加,14 d组达到高峰;Br-dU/TuJ1、BrdU/MAP-2阳性双标细胞数在脑缺血后14 d组开始增加,28 d组达到高峰;但BrdU/GFAP阳性双标细胞数则无明显变化;增殖的BrdU阳性细胞同时表达VEGF及其受体FLK-1。以上结果提示:大鼠局灶性脑缺血可激活SVZ自体神经前体细胞原位增殖、分化,且增殖的细胞同时表达VEGF及其受体可能是脑缺血后神经发生增强的调节机制之一。     

NDRG2对成年大鼠室管膜下区神经干细胞增殖的影响

目的:探讨NDRG2(N-myc downstream regulated gene 2)对成年大鼠侧脑室室管膜下区(subventricular zone,SVZ)神经干细胞增殖的影响。方法:成年雄性Sprague-Dawley大鼠20只,随机分为对照组(n=10)、实验组(n=10)两组。实验组给予侧脑室注射过表达NDRG2的腺病毒,对照组给予侧脑室注射腺病毒空载体。采用Brd U免疫荧光染色法检测细胞增殖情况,Tunel染色检测细胞凋亡情况。结果:Brd U染色结果显示:实验组大鼠侧脑室室管膜下区Brd U阳性细胞显著少于对照组(P0.05);Tunel染色提示:两组大鼠侧脑室室管膜下区凋亡的细胞数未见明显差异(P0.05)。结论:上调NDRG2在成年大鼠脑内的表达,会抑制其侧脑室室管膜下区神经干细胞的增殖,但不影响其细胞的存活。     

去松果体后成年大鼠学习记忆及侧脑室室管膜下区神经干细胞增殖的变化

目的观察松果体切除对学习记忆及侧脑室室管膜下区(SVZ)神经干细胞增殖的影响。方法将30只成年健康雄性Sprague-Dawley大鼠随机分为非手术、假手术及去松果体组,每组大鼠10只。在建立动物模型16d后,连续5d测定大鼠在Morris水迷宫的学习记忆能力,继之用免疫组织化学方法观察SVZ的增殖细胞核抗原(PCNA)阳性细胞的变化。结果去松果体组大鼠在Morris水迷宫泳游的逃避潜伏期及在原平台象限游泳距离的百分比均较非手术组或假手术组大鼠的明显延长或减少(P<0.01)。去松果体大鼠SVZ的PCNA阳性细胞数也明显低于非手术组或假手术组大鼠(P<0.01)。结论本研究首次观察到,去松果体使体内褪黑素减少,可导致学习记忆功能及SVZ神经干细胞增殖能力出现相似的明显下降趋势,说明褪黑素是确保学习记忆及神经发生得以正常进行的重要激素之一;提示褪黑素可能直接通过作用于神经干细胞上的相应受体以及间接通过提高基底前脑胆碱能系统功能来促进神经干细胞增殖,进而提高嗅觉记忆功能。     

NMDA受体在正常大鼠颈动脉体的表达

应用免疫组织化学方法观察了离子型谷氨酸受体——NMDA受体的NR1及NR2A亚单位在正常成年雄性SD和Wistar大鼠颈动脉体的表达,并对阳性产物的表达强度进行了灰度分析及统计学处理。结果表明:正常成年SD大鼠及Wistar大鼠的颈动脉体内均存在NMDA NR1免疫反应阳性细胞,从阳性细胞的形态和分布特点判断这些阳性细胞为主细胞,两种大鼠之间无明显差异(P>0.05);而两种大鼠的颈动脉体内几乎不存在NMDA NR2A免疫反应阳性细胞。本实验的结果提示:在正常成年大鼠颈动脉体的主细胞上有NMDA受体的分布,并且该受体的二聚体构成不同于经典受体,这可能与谷氨酸在颈动脉体发挥的特殊功能有关。     

幼年大鼠脑室下区内NMDA受体亚单位NR1的表达变化

目的:观察N-甲基-D-天冬氨酸受体亚单位1(N-Methyl-D-aspartate receptor subunit1,NR1)在幼年不同时期大鼠脑室下区(subventricular zone,SVZ)相同部位以及生后14d(表达高峰期)大鼠侧脑室不同部位旁的SVZ表达状况,为研究NR1是否在神经发生中起作用提供组织学依据。方法:生后7,14,21,28d的SD大鼠脑6μm厚石蜡切片,免疫组化ABC法显色,图像分析及统计处理。结果:NR1在生后7,14,21,28d大鼠SVZ均有表达,14d时表达至高峰;NR1在14d大鼠侧脑室不同部位旁SVZ内也均有表达,且侧脑室外侧角旁及下角旁SVZ表达最高。NR1阳性细胞有多种形态,均以胞膜着色为主,胞浆浅淡,偶可见核仁。结论:NR1在幼年不同时期大鼠SVZ及14d大鼠侧脑室不同区域旁SVZ均有表达,但表达高低因时间及部位而异,提示NR1在幼年大鼠SVZ的表达具有时空特异性,并可能在神经发生过程中发挥作用。     

2010年广州某医疗中心儿童夏季细菌性腹泻的病原学调查

目的研究广州地区小儿夏季细菌性腹泻的病原菌分布。方法采集2010年5～7月广州市妇女儿童医疗中心珠江新城院区腹泻患儿的大便标本进行常规病原菌的分离培养,通过生化反应和血清凝集试验进行鉴定和分型,并使用金标法对空肠弯曲菌抗原进行检测。结果从110份标本中检出44株病原菌,检出率为40.0%。其中致病性大肠埃希菌17株,2岁以下患儿检出15株;空肠弯曲菌12株,2岁以下患儿检出10株;沙门菌6株;念珠菌纯生长6株;产气荚膜杆菌3株。结论广州地区夏季儿童细菌性腹泻的主要病原菌是致病性大肠埃希菌及空肠弯曲菌,两者的易感人群以2岁以下婴幼儿为主。     

Changes in activity of neutral proteases in tissues of ground squirrels in the dynamics of hibernation

Total activities of neutral proteases in the cerebral, hepatic, and myocardial tissues of ground squirrel vary during hibernation: in autumn (before hibernation) activities of the enzymes in the brain and myocardium start increasing, while in the liver they do not change. A common feature for all tissues is minimum activity of active neutral proteases in the middle of hibernation month 1 bout, while the maximum activity is recorded before awakening. Translated from Byulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 146, No. 9, pp. 278–280, September, 2008     

MiRNA调控肿瘤细胞中P-gP表达的研究进展

微小RNA（miRNA）是一类高度保守的非编码单链RNA分子,与靶基因mRNA3’端非编码区结合,抑制mRNA的翻译,促进靶基因降解。而这种调控机制与很多肿瘤细胞中P型糖蛋白（P,glycoprotein,p-gp）的表达有着密切的联系。p-gp是与耐药相关的跨膜蛋白,过表达可导致肿瘤细胞产生多药耐药（multidrugresistance,MDR）。就miRNA的生物学特性、p-gP的功能进行综述,关系。肿瘤MDR是临床化学治疗失败的主要原因,本文并重点介绍其调控肿瘤细胞中P-gP表达和MDR的     

293细胞检测致癌物的应用研究

目的 探讨应用293细胞对致癌物进行细胞转化及致瘤性等研究,为检测可疑致癌物提供实验依据.方法 应用微囊藻毒素LR(Microcystin LR,MC-La)对293细胞进行转化培养,取转化后的细胞进行软琼脂克隆实验、血清依赖性实验以及免疫缺陷小鼠体内致瘤实验来验证MC-LR转化的293细胞的致癌性.结果 293细胞在MC-LR的作用下,逐渐表现出转化细胞的特性:血清依赖程度显著降低,在软琼脂培养基中锚着独立生长并形成细胞集落,在SCID小鼠体内形成浸润性肿瘤组织.结论 293细胞易于培养并对环境致癌物敏感,可用于可疑致癌物的检测.     

贵州黔东南地区育龄人群地中海贫血基因类型分析

目的了解贵州黔东南州育龄人群地中海贫血基因类型及其分布。方法 2017年1月~2019年3月对930例疑似地中海贫血的育龄人群采用PCR结合导流杂交进行α、β地中海贫血基因检测。结果 930例病例中检出地中海贫血422例,检出率45.38%,α地中海贫血193例(20.75%),β地中海贫血212例(22.80%),α复合β地中海贫血23例(1.83%)。12种α地中海贫血基因类型主要为--SEA/αα(67.87%)、-α3.7/αα(9.84%)和-α4.2/αα(7.25%),8种β地中海贫血基因类型以CD41-42/N(61.79%)、CD17/N(26.89%)和IVS-Ⅱ-654/N(6.13%)常见,8种α合并β地中海贫血基因类型主要为-α3.7/CD41-42(41.19%)和--SEA/CD41-42(23.53%)。结论贵州黔东南州β地中海贫血检出率高于α地中海贫血,主要基因类型为CD4142/N和--SEA/αα。     

Factor of selectivity in organization of complex behavior in primates

Selectivity in the activity of the mammalian brain is examined on the basis of an analysis of a substantial experimental material obtained in our laboratory during the investigation of the conditioned reflex and delayed behavior in lower and higher primates.Translated from Fiziologicheskii Zhurnal SSSR imeni I. M. Sechenova, Vol. 73, No. 2, pp. 269– 276, February, 1987.     

Induction of high frequency activity in the somatosensory thalamus of rats in vivo results in long-term potentiation of responses in SI cortex

Summary Extracellular single-unit techniques were employed to record unitary activity simultaneously from the thalamic ventral posterior medial (VPM) nucleus and the ipsilateral primary somatosensory cortex of adult rats. Cross-correlation analysis triggered by the spontaneous firing of thalamocortical relay neurons in VPM and the discharge of layer IV neurons in the corresponding ipsilateral cortical barrel indicated that the paired-units included in this study were strongly correlated in their activity. The baseline responses of highly correlated cortical/thalamic pairs to a 10 ms deflection of a vibrissa on the contralateral side were measured using poststimulus time histograms. After establishing the baseline response, high frequency activity in VPM was induced in one of two ways: i) direct electrical stimulation of thalamic neurons or ii) whisker stimulation in the presence of bicuculline methiodide (BIC) released near the thalamic neurons. Both methods resulted in a conditioning stimulus (CS) paradigm consisting of bursts of high-frequency activity (50–100 Hz) with an inter-burst interval of 150 ms (7 Hz). Almost immediately following the presentation of the CS, the response of layer IV cortical neurons to vibrissa stimulation increased by 37–62% over baseline values, which was maintained after the effects of BIC had worn off in VPM. This enhancement in the response of the cortical neurons was not accompanied by a concomitant increase in the thalamic responses. Thus, these results strongly suggest that the potentiation first occurred at the thalamocortical synapse.     

Role of Thrombin in Activation of Neurons in Rat Hippocampus

The effect of thrombin, an agonist of proteinase-activated receptor (PAR) family, was studied on cultured rat hippocampal neurons. Thrombin in a concentration range of 1 pM - 10 nM induced a transitory dose-dependent increase in intracellular free calcium concentration. Involvement of PAR1 in neural response to thrombin was corroborated in experiments with TFLLRN, a selective synthetic peptide agonist of these receptors. In a calcium-free medium and after treatment with cyclopiazonic acid (inhibitor of Ca(2+)-ATPase in the endoplasmic reticulum) activation of PAR not only mobilized Ca(2+) from intracellular stores, but also induced Ca(2+) entry into the cells. Thrombin decreased Ca(2+) signal triggered by activation of NMDA-subtype glutamate receptors.     

Neuroprotective potential of ceftriaxone in in vitro models of stroke

Astrocytic glutamate transporters are considered an important target for neuroprotective therapies as the function of these transporters is abnormal in stroke and other neurological disorders associated with excitotoxicity. Recently, Rothstein et al., [Rothstein JD, Patel S, Regan MR, Haenggeli C, Huang YH, Bergles DE, Jin L, Dykes Hoberg M, Vidensky S, Chung DS, Toan SV, Bruijn LI, Su ZZ, Gupta P, Fisher PB (2005) Beta-lactam antibiotics offer neuroprotection by increasing glutamate transporter expression. Nature 433:73-77] reported that beta-lactam antibiotics (including ceftriaxone, which easily crosses the blood-brain barrier) increase glutamate transporter 1 (GLT-1) expression and reduce cell death resulting from oxygen-glucose deprivation (OGD) in dissociated embryonic cortical cultures. To determine whether a similar neuroprotective mechanism operates in more mature neurons, which show a different pattern of response to ischemia than primary cultures, we exposed acute hippocampal slices obtained from rats treated with ceftriaxone for 5 days (200 mg/kg; i.p.) to OGD. Whole-cell patch clamp recording of glutamate-induced N-methyl-d-aspartate (NMDA) currents from CA1 pyramidal neurons showed a larger potentiation of these currents after application of 15 microM dl-threo-beta-benzyloxyaspartic acid (TBOA; a potent blocker of glutamate transporters) in ceftriaxone-injected animals than in untreated animals, indicating increased glutamate transporter activity. Western blot analysis did not reveal GLT-1 upregulation in the hippocampus. Delay to OGD-induced hypoxic spreading depression (HSD) recorded in slices obtained from ceftriaxone-treated rats was longer (6.3+/-0.2 vs. 5.2+/-0.2 min; P<0.001) than that in the control group, demonstrating a neuroprotective action of the antibiotic in this model. The effect of ceftriaxone was also tested in organotypic hippocampal slices obtained from P7-9 rats (>14 days in vitro). OGD or glutamate (3.5-5.0 mM) damaged CA1 pyramidal neurons as assessed by propidium iodide (PI) fluorescence. Similar damage was observed after pre-treatment with ceftriaxone (10-200 microM; 5 days) and ceftriaxone exposure did not result in GLT-1 upregulation as assayed by Western blot. Treatment of slice cultures with dibutyryl cAMP (100-250 microM; 5 days) increased GLT-1 expression but did not reduce cell damage induced by OGD or glutamate. Thus we confirm the neuroprotective effect of antibiotic exposure on OGD-induced injury, but suggest that this action is related to independent modulation of transporter activity rather than to the level of GLT-1 protein expression. In addition, our results indicate that the protective effects of beta-lactam antibiotics are highly dependent on the experimental model.     

内毒素血症在肝癌发生发展中的作用

目的:探讨内毒素血症在肝癌发生发展中作用。方法:利用饮水中加入0.03%TAA,4个月形成肝硬化,6个月形成肝癌动物的模型。TAA+LPS组从第5个月开始皮下注射内毒素,至实验结束。在肝癌发生发展过程中进行血浆内毒素水平、γ-GT活性、DNA指数、增殖指数,并对N-ras、p53基因点突变进行分析。结果:内毒素可增加bcl-2与p53蛋白过度表达与增加N-ras、p53基因的点突变,增加自由基生成与降低抗氧化酶活性,加重DNA损伤。结论:内毒素能够促进TAA诱发肝癌变的过程。