免疫电镜技术运用过程中电子染色方法的选用

目的探究免疫电镜不同染色方法对免疫组化阳性实验结果影响的关系。方法组织切片经常规免疫组化（雌激素受体GPR30）并行DAB-硫酸镍铵显色后进行电镜切片，然后分为双氧铀-柠檬酸铅双染、双氧铀单染与未染色3组，以便对不同电子染色结果进行比较。结果GPR30免疫阳性产物位于细胞核外的膜性结构上，在铀-铅双染组显示很高的电子密度，但是背景染色也很深，在铀单染组的反差比较好，而未染色组的反差更好。结论免疫电镜技术中针对不同的免疫阳性反应选用不同的电子染色方法，有利于阳性结果的判断与鉴别。     

苯丙氨酸缓解高血压大鼠血管重塑

目的：应用激光共聚焦显微镜观察高血压时血管重塑，以及苯丙氨酸治疗后对血管重塑的影响。方法： 以Wistar-Kyoto 大鼠 (WKY)和自发性高血压大鼠(SHR)为研究对象，采用固定压力灌注和荧光染色激光共聚焦显微镜观察分析肠系膜小动脉的管腔、厚度和血管平滑肌的排列方向。结果： SHR较WKY 管腔缩小，管壁增厚，同时血管平滑肌排列方向紊乱。应用苯丙氨酸治疗后，管腔扩大，管壁变薄，血管平滑肌排列方向改善。结论： 高血压大鼠肠系膜小动脉存在血管重塑现象，苯丙氨酸可以改善血管重塑。     

Separate and variably shaped chromosome arm domains are disclosed by chromosome arm painting in human cell nuclei

Fluorescence in situ hybridization (FISH) with microdissection probes from human chromosomes 3 and 6 was applied to visualize arm and subregional band domains in human amniotic fluid cell nuclei. Confocal laser scanning microscopy and quantitative three-dimensional image analysis showed a pronounced variability of p- and q-arm domain arrangements and shapes. Apparent intermingling of neighbouring arm domains was limited to the domain surface. Three-dimensional distance measurements with pter and qter probes supported a high variability of chromosome territory folding.     

Tissue-specific expression of the tight junction proteins claudins and occludin in the rat salivary glands

Tight junctions (TJs) are essential features of endothelial barrier membranes and of fluid-secreting epithelial cells, such as in the salivary glands. Novel integral membrane proteins have been identified as components of TJs, namely claudins and occludin. The aim of the present study was to determine the distribution of occludin and claudins in the large salivary glands of the rat. The parotid, submandibular and sublingual salivary glands were harvested from adult Sprague-Dawley rats and cryostat sections were stained using immunoperoxidase and immunofluorescence methods. Claudin-1 was expressed in endothelial cells of microvessels and in short selected segments of the duct system. Claudin-3 was expressed principally in the acinar cells and intercalated ducts, while claudin-4 was principally expressed by the striated and interlobular ducts. Claudin-5 was specific to endothelial cells of microvessels. Occludin was ubiquitously detected in the duct system. Double labelling and confocal microscopy showed some co-localization of claudin-3 with claudin-4, and minimal co-localization of occludin with claudin-4, in the striated ducts. Claudin 2 was not detected in any of the salivary glands. The results indicate specificity of the chemical composition of tight junctions in the rat salivary glands, and may reflect different physiological roles for TJs in the glandular and duct epithelial cells, and in endothelial cells of salivary gland microvessels.     

T cell receptor induced intracellular redistribution of type I protein kinase A

The productive activation of CD4(+) T lymphocytes, leading to proliferation and cytokine secretion, requires precise temporal regulation of intracellular cyclic AMP concentrations. The major effector molecule activated by cyclic AMP in mammalian cells is the cyclic AMP-dependent protein kinase A (PKA). The type I PKA isozyme mediates the inhibitory effects of cyclic AMP on T-cell activation. Using laser scanning confocal microscopy, we demonstrated that the regulation of PKA type I activity involves spatial redistribution of PKA type I molecules following T-cell receptor (TCR) stimulation. In resting T cells, PKA type I was located in membrane proximal regions and distributed equally across the cell. Shortly after antigen engagement, T cells and antigen-presenting cells formed an area of intense contact, known as the immunological synapse. TCR concentrated at the synapse, whereas PKA type I molecules redistributed to the opposite cell pole within 10 min after T-cell stimulation. Type I PKA redistribution was solely dependent on TCR signalling, because we observed the same temporal and spatial distribution after antibody-mediated cross-linking of the TCR-associated CD3 complex. Segregation of TCR and PKA type I molecules was maintained for at least 20 min. Thirty minutes after stimulation, PKA type I partially colocalized with the TCR. After 60 min, PKA type I distribution again approached the resting state. Considering that initial TCR signals lead to increases in intracellular cyclic AMP, PKA type I molecules may be targeted towards localized cyclic AMP accumulations or transported away from these areas, depending on the requirements of the cellular response.     

Confocal observation of senile plaques in Alzheimer's disease: Senile plaque morphology and relationship between senile plaques and astrocytes

Senile plaques In the brains of Alzheimer's disease (AD) were examined by confocal laser scanning microscopy (CLSM) with the following three findings. First, in sections stained with Congo red, the serial CLSM images of optical sections clearly revealed that a classic plaque is composed of a plaque core and a corona. Radially arranged process-like structures, corresponding to bundles of amyloid fibrils, formed amyloid cores and stronger signals were detected in the center of some cores. Second, in sections stained with Congo red and anti-gllal fibrillary acidic protein (GFAP), reactive astrocytes were found around the senile plaques and many astrocytlc processes surrounded the plaque cores and some processes had penetrated into them. Third, three-dimensional reconstruction on classic plaque revealed that the surface of classic plaque showed a 'coral-like' appearance.     

A micro cell culture method utilizing modified microscope slides for use in fluorescent antibody and enzyme immunoassays

Summary A technique for culturing small quantities of mammalian cells on modified microscope slides is described. The modified microscope slides were Bellco Glass, Inc., toxoplasmosis slides and the cell cultures used were early passage bovine embryonic lung cells and continuous cell lines of porcine and canine origins. The slide cell cultures were either uninfected or infected with selected viruses or the obligate intracellular protozoanEncephalitozoon caniculi for utilization in direct and indirect fluorescent antibody testing or in peroxidase antiperoxidase immunosorbant assays.     

大鼠三叉神经脊束间质核内内脏神经初级传入终末与NOS阳性投射神经元的联系

目的　探查间质核 (INV)内内脏传入终末与向臂旁核 (PBN)投射的NOS阳性神经元之间的联系。　方法　逆行、跨神经节追踪以及免疫荧光组织化学方法 ,结合激光共聚焦扫描显微镜观察。　结果　PBN内注射四甲基罗丹明 (TMR)后 ,逆行标记细胞主要位于注射侧的INV ,大多属 2 0 μm以下的中、小型细胞。NOS阳性细胞与TMR逆行标记细胞分布区域重叠。NOS TMR双标记细胞分别占NOS阳性细胞总数的 5 4 8% (17 31)和TMR逆行标记细胞总数的 34% (17 4 9)。舌咽和迷走神经内注射生物素化葡聚糖胺 (BDA)跨神经节标记的内脏神经初级传入终末点状膨体贴近双标记细胞胞体 ,呈紧密接触状。　结论　可能存在经INV向PBN投射的内脏伤害性信息传导通路 ,作为神经递质和神经信息分子的NO可能参与其内脏伤害性信息的传递和调控     

Blood Flow and Leukocyte Adhesiveness Are Reduced in the Microcirculation of a Peritoneal Disseminated Colon Carcinoma

Dynamic behavior of leukocytes in the microcirculation of solid tumor tissue was visualized using a fluorescent labeling technique combined with the use of a real-time confocal laser-scanning microscope (CLSM) system. Colon tumor cells (RCN-9) were inoculated into the peritoneal cavity of male Fischer 344 rats. Tumor-free rats were similarly injected with physiological saline (intraperitoneally). Ten days after tumor inoculation, the mesentery was exteriorized and subjected to vital microscopic observation under the CLSM system. Leukocytes were labeled with rhodamine 6G (100 g kg^–1, intravenously), and their behavior within the microvessels (10–30 m in diameter) was analyzed both in the solid tumor tissues and the normal mesentery. Wall shear rate was calculated from the measured values of vessel diameter and erythrocyte flow velocity. In tumor microvasculature of tumor-bearing rats, the centerline erythrocyte velocity (0.73 ± 0.58 mm s^–1, mean±standard deviation) and wall shear rate (210 ± 151 s^–1 were significantly lower than those of the tumor-free rats (1.27 ± 0.83 mm s^–, 344 ± 236 s^–1, respectively). Despite such reduced flow conditions, flux of the rolling leukocytes as well as density of the adhered leukocytes both decreased significantly in tumor microvasculature as compared with normal controls. The methods developed in this work show promise in improving our understanding of tumor biology and pathophysiology. © 1998 Biomedical Engineering Society. PAC98: 8722Fy, 8745Hw, 8745Ft, 8764-t, 4262Be     

鸡胚心脏外形变化及心外膜的发生

本研究用扫描电镜、透射电镜和光镜,观察了早期鸡胚心脏发生的外形变化及心外膜的发生。1.心脏发生经历2个阶段,首先是心脏的成襻过程,心管表面的一定部位出现沟,沟相应部位内面出现嵴。心襻的完成,标志着心脏各部原基的确立。第2个阶段是心脏各部原基的进一步分化,包括心腔内部的分隔及心脏各部位置的变化。心房与心锥的移位,可能是受左右心室移位的影响。2.心外膜片并非由心肌外层原位分化,而是由静脉窦处的间充质细胞增殖,并迁移覆盖在心肌层表面形成。