ZnT1及游离锌离子在小鼠骺板软骨细胞的定位研究

目的研究锌转运体-1(Zinc transporter 1,ZnT1)和游离锌离子在小鼠骺板软骨细胞的定位分布,探讨ZnT1影响骺板软骨细胞锌离子代谢从而参与骨骼生长的可能机制。方法应用浸入式金属自显影技术(AMG)观察锌离子在小鼠肋骨骺板内的定位分布;应用免疫组织化学SABC法检测ZnT1在小鼠骺板软骨细胞的表达。结果金属自显影技术显示游离锌离子在小鼠肋骨骺板肥大带、增殖带和静止带三层区域结构内有不同程度的表达,其中肥大带软骨细胞层锌离子含量最高;ZnT1免疫阳性反应产物主要定位于软骨细胞膜附近,在小鼠肋骨骺板三层区域结构内有不同程度的表达,从肥大带到静止带,软骨细胞膜上的ZnT1免疫反应逐渐减弱。结论小鼠肋骨骺板内存在大量的游离锌离子和ZnT1蛋白,提示ZnT1可能参与锌离子在软骨细胞的转运和代谢,在骨的形成和发育过程中发挥作用。     

硒酸锌金属自显影技术检测游离锌离子在小鼠肾脏的分布

目的研究游离锌离子在小鼠肾脏的定位分布。方法应用硒酸锌金属自显影技术(ZnSeAMG)检测小鼠肾脏内的游离锌离子分布。结果游离锌离子在肾脏内分布广泛,皮质中有大量AMG反应阳性颗粒,髓质中的AMG阳性颗粒较少。其中,近曲小管、远曲小管、近直小管和远直小管上皮细胞近腔侧均分布有大量的棕黑色AMG阳性颗粒,肾小体、细段和集合管上皮细胞中AMG阳性颗粒较少。结论小鼠肾脏内含有丰富的游离锌离子,锌离子可能参与肾脏的功能。     

ZnT3与Aβ在APP/PS1转基因小鼠脑血管及脉络丛的一致性分布

目的研究锌转运蛋白3(Zinc Transporter 3,ZnT3)与β-淀粉样蛋白(β-amyloid,Aβ)在APP/PS1转基因小鼠大脑血管壁及脉络丛上皮的定位分布,探讨ZnT3影响脑锌平衡从而参与AD发病的可能机制。方法应用免疫荧光技术和共聚焦激光扫描显微镜观察ZnT3和Aβ在APP/PS1转基因小鼠大脑血管壁及脉络丛上皮的共存情况。结果APP/PS1转基因小鼠侧脑室及第三、四脑室的脉络丛上皮细胞均呈ZnT3和Aβ染色阳性,二者共同表达于上皮细胞的胞质内,而细胞核未见任何着色。在APP/PS1转基因小鼠大脑皮层中,几乎所有Aβ阳性的血管壁上均有ZnT3的表达,二者的分布同样具有一致性。结论ZnT3与Aβ在APP/PS1转基因小鼠大脑血管及脉络丛上皮的一致性分布,提示ZnT3可能参与Aβ在大脑血管及脉络丛上皮的沉积。     

游离锌离子在小鼠视网膜的定位研究

目的研究游离锌离子在小鼠视网膜的定位分布。方法应用ZnSe金属自显影技术(AMG)检测硒酸钠注射40 m in后小鼠视网膜内的锌离子。结果注射硒酸钠40 m in后发现游离锌离子主要分布于小鼠视网膜的色素上皮细胞层、光感受器的内节、外核层、外网层、内核层、内网层和神经节细胞层。在色素上皮细胞层、光感受器的内节和内核层与内网层交界处AMG阳性反应最为明显,在光感受器外节和神经纤维层几乎没有AMG阳性反应产物。结论小鼠视网膜内锌离子,在视网膜神经元视觉信息的传导和形成过程中可能起着重要作用。     

锌转运蛋白9在实验性脑疟小鼠脑内的表达改变

目的 探讨锌离子转运蛋白9在实验性脑疟小鼠大脑皮层和海马中的表达改变。方法 6周龄C57BL/6小鼠随机分为2组,对照组给予正常饮食,感染组（PbA组）经腹腔注射1×10 6个伯氏疟原虫（P.bergheiANKA）寄生的红细胞,采用吉姆萨染色检测红细胞感染率,并监测小鼠生存率;采用Western blot和免疫组织化学染色检测ZnT9在大脑皮层和海马中的表达改变;激光共聚焦扫描显微镜检测ZnT9在小鼠大脑中的共定位表达。结果 PbA感染组在感染后5-7天逐渐出现脑疟症状,外周血红细胞内有大量感染的疟原虫存在。ZnT9主要定位于神经元中,ZnT9在实验性脑疟小鼠大脑皮层和海马中表达降低。结论 ZnT9在实验性脑疟小鼠神经元细胞中表达降低,可能参与调控锌离子在神经元内稳态的维持。     

锌缺乏对神经病理性疼痛模型小鼠行为学的影响

目的研究饮食锌缺乏对神经病理性疼痛模型小鼠痛域的影响。方法制备饮食锌缺乏小鼠,采用小鼠坐骨神经分支选择损伤模型,金属自显影和原子吸收光谱检测小鼠脊髓后角锌稳态,检测小鼠痛域。结果 AMG结果证实,与假手术组相比,模型组小鼠和锌缺乏组小鼠脊髓后角游离锌离子减少,与模型组小鼠相比,锌缺乏小鼠脊髓后角游离锌离子减少。原子吸收光谱结果表明,锌缺乏组小鼠总锌含量较假手术组减少。痛域检测结果表明,锌缺乏小鼠痛域下降。结论锌离子可能参与小鼠脊髓后角痛觉专递。     

硒酸锌金属自显影技术检测游离锌离子在小鼠卵巢的分布

目的研究游离锌离子在小鼠卵巢的定位分布。方法应用硒酸锌金属自显影技术(ZnSeAMG)检测小鼠卵巢内的游离锌离子分布。结果游离锌离子在卵巢内分布广泛,在原始卵泡、初级卵泡、次级卵泡、闭锁卵泡以及间质细胞中都有大量棕黑色的AMG染色颗粒。结论小鼠卵巢内含有丰富的游离锌离子,锌离子可能参与了卵泡的发育以及性激素的合成。     

大鼠脑脊髓不同部位室管膜上皮细胞表面的超微结构特征--扫描电镜观察

目的 研究大鼠脑脊髓室管膜上皮细胞的区域性结构特征，并初步探讨其功能。方法 采用扫描电子显微镜观察法。结果 侧脑室壁室管膜上皮细胞游离端呈不规则多边形，可见纤毛及微绒毛，偶见分泌泡。侧脑室脉络丛上皮细胞游离端呈不规则多角形、梭形或三角形，微绒毛、分泌泡丰富，纤毛少见，可见“丛上细胞”。第Ⅲ脑室室管膜上皮细胞具明显区域性，脑室底部上皮细胞微绒毛短小、散在，有少量分泌粒，无纤毛；侧壁分泌泡较多，纤毛丰富；均可见“丛上细胞”。第Ⅲ脑室脉络丛上皮细胞可见大量细长微绒毛，分泌泡及“丛上细胞”较少。室问孔室管膜上皮呈梭形，长轴与室间孔平行，游离面罕见微绒毛，“丛上细胞”较多。穹窿下器及终板处室管膜细胞纤毛少见，而正中隆起处可见大量纤毛及微绒毛，并可见单个散在的纤毛上皮细胞。中脑导水管壁具有平行走向的纵嵴，游离面可见大量纤毛、微绒毛及分泌泡。第Ⅳ脑室室管膜上皮具大量长纤毛及微绒毛，分泌泡罕见。第Ⅳ脑室脉络丛上皮游离面呈多型性，可见短小密集的微绒毛及处于不同分泌周期的分泌泡，以及形态各异的“丛上细胞”，偶见纤毛上皮细胞及特殊的单鞭毛上皮细胞。脊髓中央管结构较简单，游离面可见大量长纤毛、微绒毛及分泌泡。结论 大鼠脑脊髓室管膜不同部位上皮细胞游离面超微结构差异，可能与其功能不同有关。分泌泡、纤毛及微绒毛的大小及多寡，可能是脑脊液的分泌、流动及代谢活性中分别有区域性不同的形态学基础。     

锌转运体ZNT7在小鼠视网膜的定位分布

目的研究锌转运体-7(zinc transporter 7,ZNT7)在小鼠视网膜的定位和分布。方法应用免疫组织化学技术检测CD-1小鼠视网膜内的ZNT7免疫反应产物的表达。结果ZNT7在小鼠视网膜内分布广泛,在神经节细胞和色素上皮细胞内ZNT7免疫阳性反应产物的表达最丰富,在无长突细胞和视神经纤维层中ZNT7免疫阳性反应产物为中等程度的表达,在内网层、外网层和光感受器外节中ZNT7免疫阳性染色较淡,在外核层和光感受器内节中ZNT7几乎没有表达。结论ZNT7可能在维持视网膜锌稳态过程中起到重要的作用。     

锌转运体8与小鼠睾丸间质细胞睾酮生成相关

目的锌转运体8(ZnT8)与胰岛素等激素的分泌有关,但其在睾丸的功能尚不清楚,本实验旨在探讨ZnT8在小鼠睾丸的分布与功能。方法通过免疫组织化学技术检测成年雄性小鼠睾丸ZnT8的分布;乳鼠腹腔注射hCG(human chorionic gonadotrophin,人绒毛膜促性腺激素),注射5d后,检测乳鼠睾丸ZnT8的分布;化学发光法检测乳鼠血清睾酮水平。结果 ZNT8主要分布在成年雄鼠睾丸间质细胞,hCG处理乳鼠在血清睾酮升高的同时伴有睾丸间质细胞ZnT8表达增加。结论 ZnT8的功能可能与小鼠睾丸间质细胞睾酮合成有关。     

游离锌离子在小鼠颈上神经节的分布

目的研究游离锌离子在小鼠交感节后神经元中的分布。方法应用ZnSe金属自显影技术和透射电镜检测硒酸钠注射1.5和24 hr后小鼠颈上神经节内的游离锌离子。结果注射硒酸钠1.5 hr后的小鼠颈上神经节中几乎所有的神经元均有锌阳性反应产物分布,透射电镜下观察锌离子主要分布于神经元细胞核周围的高尔基复合体和小泡状结构内。注射硒酸钠24 hr后有锌阳性反应的神经元约为10%,提示锌离子在交感节后神经轴突内进行轴突运输。结论小鼠颈上神经节神经元胞体内含有丰富的游离锌离子,游离锌离子在轴突内进行轴突运输。     

Zinc transporter 3 and zinc ions in the rodent superior cervical ganglion neurons

Previous studies have revealed that zinc-enriched (ZEN) terminals are present in all parts of the CNS though with great differences in intensity. The densest populations of both ZEN terminals and ZEN somata are found in telencephalic structures, but also structures like the spinal cord demonstrate impressive ZEN systems spreading terminals several segments around the respective ZEN somata. The present study evaluates whether sympathetic neurons in the superior cervical ganglia (SCG) are ZEN neurons, i.e. contain vesicles that have zinc transporter 3 (ZnT3) proteins in their membranes and contain zinc ions. ZnT3 immunoreactivity (IR) was found in the somata and processes in the postganglionic neurons of mouse SCG. Only a small fraction of neurons (less than 5%), expressed varying degrees of ZnT3. Colchicine treatment, however, increased the number of ZnT3-positive neurons three-fold, suggesting an accumulation of ZnT3 protein in the somata. A small proportion of the postganglionic axons revealed dotted accumulations of ZnT3 IR along their courses. Double labeling showed that all ZnT3-positive neurons and axons were also tyrosine hydroxylase-positive with strong immunofluorescence, while no colocalization was found between ZnT3 and the vesicular acetylcholine transporter (VAChT) or neuropeptide Y IR. VAChT-positive preganglionic neurons were found to terminate on ZnT3 neuronal somata. 6-Methoxy 8-para toluene sulfonamide quinoline fluorescence and zinc selenium autometallography (ZnSe(AMG)) revealed that a subgroup of SCG cells contained free or loosely bound zinc ions. It is therefore concluded that ZnT3 and zinc ions are present in a subpopulation of TH-positive, NPY-negative neurons in the rodent SCG, supporting the notion that vesicular zinc ions may play a special role in the peripheral sympathetic adrenergic system.     

Intermediate filament proteins in choroid plexus and ependyma and their tumors.

The intermediate filament protein types of normal choroid plexus and ependymal tissue and their putative tumors were investigated. In normal human choroid plexus tissue, but not in ependyma, keratin could be demonstrated immunohistochemically. By immunoblotting, keratins 8, 18, and 19 were found, but glial fibrillary acidic protein (GFAP) was absent. In mouse and rat, choroid plexus epithelium and ependymal lining cells were keratin-positive. In addition, many ependymal cells were vimentin-positive. Keratin was immunohistochemically found in three of four choroid plexus papillomas, two of two choroid plexus carcinomas, and the lining cells of three neuroepithelial cysts. GFAP-positive cells were present in some choroid plexus tumors. In contrast, none of the eight ependymomas contained keratin, but all were strongly positive for GFAP. The results show that choroid plexus lining cells and choroid plexus tumors have true epithelial characteristics in their cytoskeleton, in contrast to ependymomas, which do not show keratin positivity but show glial filaments, as would be seen in astrocytic tumors.     

Changes in the vesicular zinc pattern following traumatic brain injury

The present study aims at evaluating the significance of zinc ions on the development of brain damage in a model of traumatic brain injury (TBI). The zinc ion specific autometallographic technique, the ZnSe(AMG) method, using silver enhancement of in vivo-captured zinc ions bound in zinc-selenium nanocrystals was applied to follow changes in the vesicular zinc pattern. Balb/c mice, ZnT3 knockout (ZnT3-Ko) mice, a mouse genetically knocked out for the protein ZnT3 responsible for sequestering zinc into synaptic vesicles, and littermates from the genetically un-manipulated mother type mice, wild type (Wt), were used. The Wt and the Balb/c mice exhibited instantaneously a boost in the zinc staining adjacent to the lesion involving all six neocortical layers. Ultra-structural analyses revealed that the in vivo created ZnSe nanocrystals were still confined to the vesicles of the zinc-enriched (ZEN) neurons in the neuropil. No differences between the Balb/c and Wt mice were seen at any time points. In the ZnT3-Ko mice the ZEN terminals stayed void of AMG grains, but a number of neuronal somata around the lesion became loaded with ZnSe nanocrystals. These silver-enhanced ZnSe nanocrystals were confined to the cytoplasm of the somata and their proximal dendrites. No such soma staining was seen in the Wt or Balb/c mice. We speculate that vesicular zinc may not contribute to neuronal damage following TBI.     

游离锌离子在APP/PS1转基因小鼠嗅球内的定位

目的研究锌离子在APP/PSI转基因小鼠嗅球内的定位,为锌离子参与阿尔茨海默病发病的相关性研究提供重要的形态学依据。方法应用浸入式金属自显影技术(AMG)检测锌离子在野生型小鼠和APP/PSI转基因小鼠嗅球内的分布。结果在野生型小鼠和APP/PSI转基因小鼠嗅球内,AMG反应产物呈棕黑色,嗅球的5层结构清晰可见,其中颗粒层锌离子含量最高。在APP/PSI转基因小鼠嗅球内,还可见锌离子聚集在老年斑内,AMG阳性的老年斑主要分布于含锌量较高的颗粒层,其它各层仅见散在分布。野生型小鼠嗅球内未见AMG阳性的老年斑。结论APP/PSI转基因小鼠嗅球的老年斑内含有大量的锌离子,提示锌离子在嗅球Aβ老年斑的形成过程中发挥重要作用,参与AD的发病和进展。