蛋白脂蛋白在两型星形胶质细胞和少突胶质细胞中的表达

目的:探讨蛋白脂蛋白(PLP)在分化成熟的两型星形胶质细胞和少突胶质细胞中的表达。方法:用激光共聚焦双重免疫荧光标记技术检测PLP在分化成熟的两型星形胶质细胞和少突胶质细胞中的表达情况。结果:在O-2A谱系来源的成熟2型星形胶质细胞和少突胶质细胞中PLP抗体标记为阳性,而在T1A谱系来源的成熟1型星形胶质细胞中则未检测到PLP的表达,从而在蛋白质水平验证本室研究两型星形胶质细胞基因表达谱差异基因芯片结果中PLP mRNA在T2A中高表达,而在T1A中低表达的现象。结论:PLP等脂代谢相关基因可能在O-2A谱系发生和分化过程中起重要作用,O-2A谱系与神经髓鞘发生以及脑内脂质代谢内在机制密切相关。     

胰甘油三酯脂酶在大鼠两型星形胶质细胞中的表达

目的比较胰甘油三酯脂酶在1型星形胶质细胞(T1A)和2型星形胶质细胞(T2A)中的表达情况。方法取新生大鼠脑皮质,体外分离培养纯化的T1A和T2A,应用逆转录聚合酶链反应(RT-PCR))技术检测胰甘油三酯脂酶mRNA在两型星形胶质细胞中的表达差异,并且通过激光共焦免疫荧光双标技术从蛋白水平进一步证实。结果胰甘油三酯脂酶在T1A中不表达或低表达,主要定位于细胞核中;而在T2A中表达水平较高,分布广泛,在胞浆、胞核及细胞突起中均有表达。结论胰甘油三酯脂酶在两型星形胶质细胞中的表达存在差异,胰甘油三酯脂酶在T2A中高表达,提示T2A在中枢神经系统脂代谢方面起重要的作用。     

渗透压变化对缝隙连接蛋白在培养的大鼠下丘脑星形胶质细胞和神经元表达的影响

目的观察渗透压改变对培养的下丘脑星形胶质细胞的缝隙连接蛋白Connexin43（Cx43）和神经元的缝隙连接蛋白Cx32表达的影响。方法体外培养孕14d或新生1d SD大鼠下丘脑的神经元和星形胶质细胞并进行纯化和鉴定。实验各分两组,第1组：细胞由等渗培养液移至高渗培养液（含9％NaCl）分别放置1min、3min、5min、10min和15min后将液体吸出常规固定;第2组：细胞先置于高渗培养液中15min后换成等渗培养液,分别在1min、3min、5min和10min后将等渗培养液吸出常规固定。两者均进行抗Cx43或抗Cx32的免疫荧光染色,Confoeal显微镜观察。结果第1组,Cx43在刺激1min后在星形胶质细胞表达比对照组有所增加,3min达到高峰,以后逐渐降低,15min恢复正常;而Cx32在刺激后1min的神经元中表达比对照组增加,5min达到高峰,以后降低。15min恢复正常。第2组同样为Cx43在星形胶质细胞刺激1min后表达增加,3min达到高峰,以后降低,10min恢复正常;神经元的Cx32表达在刺激后5min达到高峰,以后降低,10min恢复正常。两组Cx43表达的高峰期均早于Cx32。结论Cx43和Cx32可能参与星形胶质细胞与神经元渗透压的调节。     

星形胶质细胞转分化为神经元在神经再生中的研究进展

<正>星形胶质细胞是广泛存在于中枢神经系统内的一类胶质细胞,在建立和维持正常脑功能以及脑损伤后修复中具有重要作用。近几年发现,特定区域脑损伤后星形胶质细胞发生活化,甚至转分化为神经元谱系,表现出类似于神经前体细胞的特性,这一重要的潜能使得星形胶质细胞成为神经再生领域研究的焦点。本文就星形胶质细胞的基本特性、其转分化为神经元的模型以及介导转分化发生的细胞与分子学     

星形胶质细胞在血脑屏障与神经元损伤中的作用

<正>星形胶质细胞(astrocyte,AST)是中枢神经系统重要的管家细胞,其在神经兴奋时清除多余的兴奋性递质、控制离子和水分子动态平衡、释放神经营养因子、转移代谢产物和废弃物、参与血脑屏障(blood-brain barrier,BBB)形成、调控     

轴突成束和延伸蛋白ζ-1基因在1型和2型星形胶质细胞中的表达

目的 探讨轴突成束和延伸蛋白ζ-1(FEZ1)基因在1型和2型星形胶质细胞(T1A、T2A)中的表达差异,及其在新生SD大鼠中枢神经系统中的表达. 方法培养、分离、纯化T1A和T2A,提取总RNA,分别以Cy3-dCTP和cy5-dCTP标记T1A和T2A的mRNA,进行基因表达谱芯片检测分析.选取其中与轴突延伸相关的FEZ1基因,应用半定量RT-PCR法研究FEZ1在新生大鼠中枢神经系统多个部位的表达. 结果 T1A和T2A中差异表达基因138条,其中60条在T1A中高表达,78条在T2A中高表达.FEZ1基因在T2A中高表达.RT-PCR结果显示,FEZ1在新生大鼠中枢神经系统大脑皮层、嗅球和脊髓组织中均有表达. 结论 T1A和T2A中存在多个差异表达基因,其中与轴突延伸相关的FEZ1基因表达明显差异,提示T1A和T2A可能在诱导轴突生长方面有不同功能;FEZ1基因与中枢神经系统的发育和再生相关.     

星形胶质细胞在大鼠下丘脑的分布

目的:观察星形胶质细胞在正常大鼠下丘脑的分布。方法:利用免疫组织化学技术,观察胶原纤维酸性蛋白(Glial fibrillary acidic protein,GFAP)在下丘脑的表达。结果:GFAP阳性细胞在第三脑室室周区、视上核、下丘脑室旁核外侧大细胞部、正中隆起大量分布;下丘脑其他区域或核团,如前区、外侧区、前外侧核、视交叉上核、室旁核小细胞部、弓状核、腹内侧核等内分布稀疏。结论:星形胶质细胞在正常大鼠下丘脑的分布有明显区域性,可能与其相应区域的生理活动与调节功能有关。     

星形胶质细胞在体内直接重编程为神经元的研究进展

人类神经元几乎不具有再生能力,这也意味着神经元一旦受损或退化,神经系统的自我修复将无计可施。研究发现,星形胶质细胞作为一种广泛分布于中枢神经系统(CNS)中的神经胶质细胞,具备诱导转化为神经元的潜力。事实上,各国研究学者已经成功利用多种技术手段将来源不同的星形胶质细胞重编程为神经元。本文综述了脊髓损伤(SCI)、脑卒中(CS)、帕金森病(PD)、阿尔茨海默病(AD)和亨廷顿病(HD)、肌萎缩性脊髓侧索硬化症(ALS)等神经系统相关疾病中星形胶质细胞直接重编程为功能性神经元的再生策略。     

大鼠腰骶髓和延髓星形胶质细胞及神经元对慢性结肠炎的反应

目的探讨大鼠腰骶髓和延髓星形胶质细胞及神经元对慢性结肠炎的反应,及反应性星形胶质细胞和反应性神经元之间的关系.方法成年雄性SD大鼠,实验组（n=17）给予三硝基苯磺酸（TNBS）灌肠诱导结肠炎;对照组（n=16）给予生理盐水灌肠.免疫组织化学法显示大鼠腰骶髓和延髓内胶质原纤维酸性蛋白（GFAP）阳性星形胶质细胞和Fos阳性神经元.结果TNBS灌肠后,GFAP阳性星形胶质细胞主要分布在脊髓背角浅层（Ⅰ～Ⅱ层）、中间外侧核（Ⅴ层）、后连合核（Ⅹ层）和腹角外侧核（Ⅸ层）.Fos阳性神经元集中分布在背角深层（Ⅲ～Ⅳ,Ⅴ～Ⅵ层）.在延髓,两者均主要分布在由孤束核、中间网状带和腹外侧区组成的延髓内脏带（MVZ）.TNBS灌肠后3、7、14 d,脊髓中GFAP阳性细胞密度明显高于对照组（P＜0.05）.TNBS灌肠后3 d,延髓中GFAP阳性细胞密度明显高于对照组（P＜0.05）.TNBS灌肠后28 d,脊髓和延髓中GFAP阳性细胞密度下降,与对照组无显著性差异（P＞0.05）.结论结肠炎性刺激引起脊髓和延髓中星形胶质细胞激活.随着结肠炎的恢复,星形胶质细胞的反应性下降.在延髓内脏带,反应性星形胶质细胞与反应性神经元关系密切.     

大鼠脑缺血再灌流时神经元损伤与星形胶质细胞的反应

为探讨星形胶质细胞在缺血性神经元损伤中的作用及其与神经元损伤的关系 ,本实验阻塞大鼠大脑中动脉 2 h,再灌流0 .5～ 48h建立短暂局灶性脑缺血模型 ,进行 H-E染色 ;通过胶质原纤维酸性蛋白和细胞核增殖抗原免疫组化单重或双重反应 ,TU NEL和胶质原纤维酸性蛋白免疫组化双重反应观察了神经元和星形胶质细胞的反应。结果表明 :再灌流 2 4h缺血区面积最大 ,再灌流 6h开始出现神经元不可逆变性 ,2 4h梗塞成熟 ;星形胶质细胞表现为反应性、营养不良性和退形变三种不同的形态特点。再灌流 48h时星形胶质细胞数量开始增多。 48h之内星形胶质细胞无增生 ,且有少量星形胶质细胞凋亡。这些结果提示脑缺血时星形胶质细胞反应与神经元损伤密切相关 ,反应性星形胶质细胞是其积极应答神经元损伤的结果 ,在维持神经元存活中起作用。     

Expression of paxillin in laryngeal squamous cell carcinoma and its prognostic value

Paxillin (PXN) gene has been reported to act as an oncogene in many malignancies and play important roles in the development of human carcinomas. However, the relationship between the expression of PXN and clinicopathological characteristics in human laryngeal carcinoma remains unclear. This study aimed to examine the expression of PXN, and to evaluate the clinical significance of its expression in human laryngeal squamous cell carcinoma (LSCC). Real-time quantitative PCR (qRT-PCR), Western blotting and immunohistochemistry were performed to analyze the expression of PXN in LSCC tissues and corresponding paracancerous normal tissues. Kaplan-Meier survival and Cox regression analyses were performed to evaluate the prognosis of patients with LSCC. The expression of PXN was significantly higher in LSCC than in matched paracancerous normal tissues. Immunohistochemical analysis was performed in human LSCC samples and the data were correlated with clinicopathologic features. Levels of PXN in LSCC were related to histopathological grade (P = 0.016), Lymph node metastasis (P = 0.029) and TNM stage (P < 0.001). Kaplan-Meier analysis revealed that survival curves of the overall survival of patients with high PXN expression was significantly worse than that of low PXN expression (P = 0.035). Cox regression analysis revealed that PXN expression level was an independent prognostic factor of the overall survival rate of patients with LSCC (P = 0.002). These findings suggest that PXN expression has potential use as a novel biomarker of LSCC patients and may serve as an independent predictive factor for prognosis of LSCC patients.     

LPS对大鼠脊髓背角星形胶质细胞CXCL1和CCL2表达的影响

目的:观察脂多糖(LPS)诱发的大鼠脊髓背角星形胶质细胞趋化因子CXCL1和CCL2的表达及释放,分析Toll样受体2(TLR2)以及Toll样受体4 (TLR4)的作用。方法:培养新生SD大鼠(<3d)脊髓背角星形胶质细胞,免疫荧光鉴定纯度达95%之后分为空白对照组、LPS处理组、TAK-242+LPS处理组、LPS-RS+LPS处理组。在LPS(1μg/ml)作用下,采用real time RT-PCR法检测SD大鼠(<3d)脊髓背角星形胶质细胞CXCL1和CCL2 mRNA表达; Western Blot用于检测胶质纤维酸性蛋白(GFAP)、TLR2、TLR4、CXCL1以及CCL2表达;ELISA检测CXCL1和CCL2释放。结果:与正常对照组相比,LPS作用6 h,背角星形胶质细胞CXCL1和CCL2在mRNA和蛋白水平的表达均上调(P <0. 05),CXCL1和CCL2释放增加(P <0. 05)。LPS刺激CXCL1和CCL2表达和释放增加的效应可以被TLR4受体拮抗剂TAK-242(50 nmol)以及TLR2/TLR4受体拮抗剂LPS-RS(...     

Protein phosphatase-2A modulates the serine and tyrosine phosphorylation of paxillin in Lewis lung carcinoma tumor variants

Cellular adhesion and motility, processes regulated by focal adhesion assembly and disassembly, can influence a tumor cell's ability to metastasize. Focal adhesion dynamics are, in turn, influenced by the serine and tyrosine phosphorylation state of paxillin. Using Lewis lung carcinoma (LLC) tumor variants, this study shows the importance of the serine/threonine protein phosphatase-2A (PP-2A) in maintaining adherence and restricting tumor cell motility, and modulating the serine and tyrosine phosphorylation of paxillin. Treating non-metastatic LLC-C8 tumor variants with okadaic acid to inhibit PP-2A activity resulted in cell rounding and increased motility. These effects on motility and adherence were accompanied by increased serine and decreased tyrosine phosphorylation of paxillin. These results suggest PP-2A regulation of paxillin phosphorylation may have a role in controlling tumor cell adherence and motility. This revised version was published online in July 2006 with corrections to the Cover Date.     

Oligodendrocyte progenitor cells derived from mouse embryonic stem cells give rise to type-1 and type-2 astrocytes in vitro

Oligodendrocyte progenitor cells (OPCs) in primary culture can give rise to mature oligodendrocytes and type-2, but not type-1 astrocytes depending on the culture conditions. The OPCs thus are called oligodendrocyte-type-2 astrocyte (O2-A) progenitor cells. Mouse embryonic stem cells (mESCs) have been efficiently differentiated into OPCs; however, the fate plasticity of mESC-derived OPCs is not well characterized. In the present study, using GFP-Olig2 mESC line, we showed that the Olig2(+)/GFP(+)/A2B5(+)/NG2(+) OPCs derived from GFP-Olig2 mESCs can mature into oligodendrocytes when co-cultured with mESC-derived neurons. Interestingly, when induced to astrocytic differentiation by bone morphogenetic protein-4, these mESC-derived OPCs can not only generate type-2 astrocytes, but also type-1 astrocytes. These results challenge the dogma that OPCs in culture can only generate type-2, but not type-1 astrocytes, and support the in vivo finding that during perinatal development, OPCs can give rise to a subset of type-1 astrocytes.     

腺苷A1受体拮抗剂DPCPX对低氧复氧脑星形胶质细胞影响的实验研究

目的:观察腺苷A1受体拮抗剂1,3-二丙基-环戊黄嘌呤(8-cyclopentyl-1,3-dipropylxanthine, DPCPX)对体外培养低氧复氧(hypoxia and reoxygenation, H/R)大鼠脑星形胶质细胞促红细胞生成素(erythropoietin, EPO)释放量、谷氨酰胺合成酶(glutamine synthase, GS)、超氧化物歧化酶(superoxide dimutase, SOD)和谷胱甘肽过氧化物酶(glutathione peroxidase, GSH-PX)活性的影响,探讨腺苷A1受体在H/R星形胶质细胞神经保护中的作用.方法:对大鼠大脑皮层星形胶质细胞进行体外培养并建立H/R模型,在低氧同时加入终浓度为100 nmol/L DPCPX 12 h后复氧1,6,12,24 h,检测EPO释放量,GS,SOD和GSH-PX活性的变化.结果:与对照组比较,DPCPX组H/R星形胶质细胞的EPO释放量显著减少;GS,SOD和GSH-PX的活性降低.结论:腺苷A1受体参与H/R星形胶质细胞的神经保护作用.     

Morphological and physiological properties of neostriatal neurons: An intracellular horseradish peroxidase study in the rat

The physiological and morphological (light and electron microscopic) properties of four categories of neostriatal neurons (two types of medium spiny cells and two types of aspiny cells) were analyzed using the technique of intracellular recording and intracellular labeling with horseradish peroxidase. All of the neurons in this study had excitatory responses following stimulation of the cortex and substantia nigra except for the large aspiny neuron for which only substantia nigra inputs were tested. Morphologically, these neurons differed with respect to the size and shape of their somata, density and distribution of dendritic spines and distribution of their axons and axon collaterals. Ultrastructurally, observed somatic differences included the quantity and distribution of organelles and conformation of the nuclear envelope.The axons of one type of medium spiny neuron and the large aspiny neuron were myelinated. Unmyelinated axon collaterals arose from the axons of both types of medium spiny neurons and formed synapses on the dendritic shafts and possibly with the necks of spines of other neostriatal neurons. The parent axons of the most common type of medium spiny neurons were followed to the globus pallidus and, in some cases, to the internal capsule.     

Expression and function of the ST2 gene in a murine model of allergic airway inflammation

BACKGROUND: We have recently reported that soluble ST2 protein levels are elevated in the sera of patients with asthma, and correlate well with the severity of asthma exacerbation. However, the role, function, and kinetics of soluble ST2 expression in asthma remain unclear. OBJECTIVE: The objective of the present study was to clarify the function and kinetics of soluble murine (m) ST2 expression in a murine asthma model. METHODS: We analyzed the kinetics of gene and protein expression of mST2 in sera or lung tissue after allergen (ovalbumin; OVA) challenge in a murine model of allergic airway inflammation, the effects of mST2 protein on OVA-induced Th2 cytokine production in vitro from splenocytes of sensitized mice, and the effects of soluble mST2 on Th2-dependent allergic airway inflammation by in vivo gene transfer of mST2. RESULTS: Serum mST2 protein levels increased to the maximal level 3 h after the allergen challenge, before serum IL-5 levels peaked. The mRNA expression of mST2 in lung tissue was induced after the allergen challenge, while that in the spleen was constitutively detected. Furthermore, pre-treatment with mST2 protein significantly inhibited the production of IL-4 and IL-5, but not IFN-gamma, from OVA-stimulated splenocytes in vitro, and intravenous mST2 gene transfer resulted in a drastic reduction in the number of eosinophils and in the levels of IL-4 and IL-5 in bronchoalveolar lavage fluid, compared with those in response to transfer of non-coding plasmid vector or of lipid alone. CONCLUSION: These results suggest that increases in endogenous mST2 protein after allergen exposure may modulate Th2-mediated airway inflammation, and that in vivo gene transfer of mST2 can be applicable to use in a novel immunotherapy for allergic diseases.     

转录因子Pitx3和Nurr1在中脑多巴胺能神经元终末分化中的表达特点

目的:探讨转录因子垂体同源盒家族因子3(Pitx3)和孤儿核受体相关因子1(Nurr1)在多巴胺(DA)能神经元终末分化中的表达特点。方法:采用免疫荧光方法检测体外培养的中脑源性神经干细胞(mNSCs)诱导分化后和大鼠胚胎发育至出生腹侧中脑Pitx3、Nurr1和酪氨酸羟化酶(TH)的表达。结果:(1)体外培养的mNSCs诱导分化48 h的Map-2阳性细胞不表达Pitx3、Nurr1和TH;分化7 d的TH阳性细胞均表达转录因子Pitx3和Nurr1;(2)在大鼠腹侧中脑黑质(SN)、腹侧被盖区(VTA)和中缝背核(DRN)可见大量TH与Pitx3(或Nurr1)共表达的神经元;(3)Pitx3和Nurr1阳性细胞主要分布于E16.5、P0大鼠SN、VTA和DRN区,其中Pitx3阳性细胞还少量分布于腹侧中脑非DA能神经元区域,Nurr1阳性细胞在腹侧中脑分布范围较Pitx3更广泛。结论:转录因子Pitx3、Nurr1在中脑DA能神经元的终末分化和生存维持中起重要作用。     

Two types of acid-sensing ion channel currents in rat hippocampal neurons

Two types of acid-sensing ion channel (ASIC)-like currents in cultured rat hippocampal neurons were recorded and their characteristics were studied by using a whole-cell recording technique. The results revealed that the ASIC-like currents, induced by a quick drop of the extracellular pH, decayed with different time constants (τ) of 229 ± 16 (Type I) and 1209 ± 56 ms (Type II). The ASIC-like currents displayed different sensitivities to extracellular proton (pH_0.5 was 6.17 ± 0.04 for Type I and 5.70 ± 0.07 for Type II) and amiloride, a specific ASIC channel blocker (IC₅₀ was 1.19 ± 0.37 μM for Type I and 0.14 ± 0.02 μM for Type II). Among all the 360 recorded neurons, ASIC-like currents were induced in 314 neurons (87.2%). In the neurons expressing ASICs, Type I currents were evoked from 269 neurons (85.7%) and Type II currents were induced only from 45 neurons (14.3%). As these ASIC-like currents presented various electrophysiological and pharmacological properties, further experiments should be conducted to decipher the complex subunit composition of ASICs in the hippocampus.     

Role of pituitary adenylate cyclase-activating polypeptide (PACAP) in the action of ginsenoside Rh2 against beta-amyloid-induced inhibition of rat brain astrocytes

Ginsenoside, the active principles in Panax ginseng root, has been demonstrated to show neurotrophic and neuroprotective actions for prevention of neuron degeneration. Deposition of β-amyloid peptide (Aβ) causes neurotoxicity through the formation of plaques in brains with Alzheimer's disease. Pituitary adenylate cyclase-activating polypeptide (PACAP) is introduced as a neurotrophic factor to promote cell survival. However, effect of Rh2, one of ginsenosides, on PACAP expression induced by Aβ remains unclear. In the present study, we found that Rh2 stimulates PACAP gene expression and cell proliferation in type I rat brain astrocytes (RBA1) cells and both effects were not modified by the estrogen antagonists (MPP or ICI 182780). Also, Rh2 ameliorates the RBA1 growth inhibition of Aβ. Moreover, blockade of PACAP receptor PAC1 using PACAP (6-38) inhibits all the actions of Rh2. These results suggest that Rh2 can induce an increase of PACAP to activate PAC1, but not estrogen receptor, and thereby leads to attenuate Aβ-induced toxicity. Thus, ginseng seems useful in the prevention of dementia.