金鱼精巢支持细胞间连接和血睾屏障

本文主要采用冷冻断裂-蚀刻方法,同时结合超薄切片和镧标记实验,研究金鱼精巢内的支持细胞间连接特点及血-睾屏障的形成。结果发现:1)金鱼精巢内支持细胞间连接呈紧密连接、桥粒和间隙连接同时存在的形式。2)各种连接的数量、面积、分布密度会随着小囊内生精细胞的发育而改变。3)紧密连接在精子发生过程的各个阶段都存在,但在形态上表现为Ⅰ型和Ⅱ型两种。4)血-睾屏障是在粗线期精母细胞期后形成,它是Ⅰ型紧密连接发育为Ⅱ型紧密连接的结果。     

星形胶质细胞可以诱导内皮细胞系紧密连接的形成

为探索星形胶质细胞在血脑屏障内皮细胞紧密连接形成中的重要意义,通过内皮细胞系ECV304与星形胶质细胞体外接触共培养的方法,采用电镜及内皮细胞紧密连接的银染观察星形胶质细胞对内皮细胞系紧密连接的诱导作用。运用Millipore-ERS系统检测紧密连接的功能状况。结果发现,星形胶质细胞可以诱导内皮细胞系形成广泛而连续的紧密连接并产生较高的跨内皮阻抗(transendothelial electrical resistance,TER),于第10d可达321．3Ωcm^2。提示,星形胶质细胞可以诱导ECV304细胞产生紧密连接。同时,ECV304细胞与星形胶质细胞的体外共培养可以作为研究血脑屏障紧密连接结构与功能的一种可靠而简便的体外实验方法。     

ELECTRICAL COUPLING BETWEEN EMBRYONIC CELLS BY WAY OF EXTRACELLULAR SPACE AND SPECIALIZED JUNCTIONS

The meroblastic egg of the teleost, Fundulus heteroclitus, was studied electrophysiologically from cleavage to mid-gastrula stages. The yolk is an intracellular inclusion surrounded by a membrane of high resistivity (50 kΩcm²). This membrane generates a cytoplasm-negative resting potential in later stages. Cells of all stages studied are coupled electrically. In gastrulae, coupling is both by way of specialized junctions between cells and by way of intra-embryonic extracellular space, the segmentation cavity. The latter mode is present because the segmentation cavity is sealed off from the exterior by a high resistance barrier, and the outer membrane of surface cells is of high resistance (50–100 kΩcm²) compared to the inner membrane. It can be inferred that clefts between surface cells are occluded by circumferential junctions. Isolated cells from late cleavage stages develop coupling in vitro, confirming the existence of coupling by way of intercellular junctions. Both modes of coupling could mediate communication between cells that is important in embryonic development.     

CELL CYCLE CHARACTERISTICS OF HUMAN SOLID TUMORS IN VIVO

The duration of the DNA synthetic phase of solid tumors in six patients was determined in vivo by local injections of tritiated thymidine into tumor nodules on the skin and serial biopsies of the lesions. Labeled mitosis curves constructed from the data obtained revealed a distinct wave of labeled mitoses but no evidence of a second wave of DNA synthesis in all six patients. The duration of S phase was 24, 22 and 24 hr in the three patients with malignant melanomas and 19, 22 and 24 hr in the three patients with breast cancer. The method is relatively simple and requires much lower doses of tritiated thymidine; reproducible data are obtainable in patients with solid tumors in a much shorter period of time than with previous experiments.     

POST-MITOTIC AGE OF MONONUCLEAR CELLS MIGRATING INTO TA-3(St) SOLID TUMORS

Subcutaneous transplantation of 5 × 10⁶ TA-3(St) mammary carcinoma cells into A/J mice produced rapidly growing tumors that showed a substantial accumulation of lymphocytes, monocytes and macrophages. This must have resulted primarily from an influx of circulating cells, since none of the cell types exhibited any significant local proliferation as indicated by 1 hr ³H-TdR uptake. The post mitotic age of the various leukocyte types in circulation of normal and tumor bearing animals, and those appearing within the tumors, was evaluated by a repeated ³H-TdR labeling protocol designed to label newly formed leukocytes. Tumor transplantation (or injection of an equivalent volume of saline in control animals) was preceded by thirteen ³H-TdR injections (12.5 μCi every 8 hr) and followed by eight more injections at equivalent intervals. Animals were serially sacrificed at 5-14 days after tumor transplantation or saline injection. Total lymphocyte labeling in the blood during these intervals was higher in tumor bearing mice (approximately 25% between 5 and 12 days) than in the saline injected controls (approximately 20%), indicating that tumor transplantation resulted in an increase in the proportion of young lymphocytes in circulation. More significantly, a much higher labeling (57-60% at 5-12 days) was exhibited by lymphocytes isolated from the tumors, indicating selective migration (and/or retention) of newly formed lymphocytes within the tumor. This finding applied to lymphocytes in all size categories. Although blood monocyte labeling in the control and experimental animals was similar (e.g. 83% at day 5 and 52% at day 14), significantly higher labeling (e.g. 93% and 70% at the respective intervals) was exhibited by monocytes isolated from tumors. This suggested a preferential accumulation of young monocytes at the tumor site. An identical macrophage labeling pattern compared to that shown by monocytes within the tumor indicated a rapid monocyte-macrophage transition. Since these findings in the present strain-specific solid tumor are similar to those previously obtained in this laboratory in the strain-nonspecific Ehrlich ascites tumor, the phenomenon of selective localization of newly formed mono-nuclear leukocytes appears to be a general occurrence in transplanted murine tumors.     

SPECIALIZED MEMBRANE JUNCTIONS BETWEEN NEURONS IN THE VERTEBRATE CEREBELLAR CORTEX

"Gap" junctions, the morphological correlate for low-resistance junctions, are demonstrated between some mossy fiber terminals and granule cell dendrites in some lower vertebrate cerebella (gymnotid and frog). Most of the gap junctions (GJs) seen in the gymnotid-fish cerebellum exhibit an asymmetrical configuration, the electron-opaque cytoplasmic material underlying the junction being more extensive in the dendritic than in the axonal side. In the frog cerebellum, the GJs have a symmetrical distribution of such electron-opaque material. In both species the GJs are encountered at the same synaptic interface as the conventional synaptic zone (CSZ), constituting "mixed synapses" in a morphological sense. The axonal surface covered by CSZs is larger than that covered by GJs. In mammalian cerebellum, GJs are observed only in the molecular layer, between perikarya, dendrites, or perikarya and dendrites of the inhibitory interneurons. These GJs are intermixed with attachment plates and intermediary junctions interpreted as simply adhesive. In the mammalian cerebellum, a new type of junction which resembles the septate junctions (SJs) of invertebrate epithelia is observed between axonal branches forming the tip of the brush of basket fibers around the initial segment of the Purkinje cell axon. It is suggested that such junctions may be modified forms of septate junctions. The physiological implications of the possible existence of high-resistance cross-bridges between basket cell terminals, which may compartmentalize the extracellular space and thus regulate extracellular current flow, must be considered.     

JUNCTIONS BETWEEN INTIMATELY APPOSED CELL MEMBRANES IN THE VERTEBRATE BRAIN

Certain junctions between ependymal cells, between astrocytes, and between some electrically coupled neurons have heretofore been regarded as tight, pentalaminar occlusions of the intercellular cleft. These junctions are now redefined in terms of their configuration after treatment of brain tissue in uranyl acetate before dehydration. Instead of a median dense lamina, they are bisected by a median gap 20–30 A wide which is continuous with the rest of the interspace. The patency of these "gap junctions" is further demonstrated by the penetration of horseradish peroxidase or lanthanum into the median gap, the latter tracer delineating there a polygonal substructure. However, either tracer can circumvent gap junctions because they are plaque-shaped rather than complete, circumferential belts. Tight junctions, which retain a pentalaminar appearance after uranyl acetate block treatment, are restricted primarily to the endothelium of parenchymal capillaries and the epithelium of the choroid plexus. They form rows of extensive, overlapping occlusions of the interspace and are neither circumvented nor penetrated by peroxidase and lanthanum. These junctions are morphologically distinguishable from the "labile" pentalaminar appositions which appear or disappear according to the preparative method and which do not interfere with the intercellular movement of tracers. Therefore, the interspaces of the brain are generally patent, allowing intercellular movement of colloidal materials. Endothelial and epithelial tight junctions occlude the interspaces between blood and parenchyma or cerebral ventricles, thereby constituting a structural basis for the blood-brain and blood-cerebrospinal fluid barriers.     

A SURVEY OF DEHYDROGENASES IN VARIOUS EPITHELIAL CELLS IN THE RAT

Several different epithelial elements that have intense active transport or protein secretory functions were histochemically assayed in several dehydrogenase media by a recently perfected method. The mitochondria represented the only site of activity, not only when tested in the succinate and D-β-hydroxybutyrate media, but also when tested in the lactate, malate, and isocitrate media. The reaction for D-β-hydroxybutyric dehydrogenase in the mouse kidney was curiously limited to the mitochondria of the distal segment of the proximal convoluted tubule, a finding that most convincingly shows that dehydrogenase activity may be differentiated in certain instances from diaphorase activity by the ditetrazole methods and that D-β-hydroxybutyric dehydrogenase is not present in all mitochondria. Tetranitro-BT is favored over nitro-BT in studies conducted on most organs prepared without fixation and on formalin-fixed tissues that consist of lipid-containing or active transport cells.     

PROTEIN METABOLISM IN TUMOR CELLS AT VARIOUS STAGES OF GROWTH IN VIVO

Protein metabolism of Yoshida ascites hepatoma cells was studied in the early phase of logarithmic proliferation and in the following stage in which cell mass remains constant (resting phase). The rate of protein synthesis was measured by a short-time incorporation of [⁸H]lysine, while degradation was concurrently assessed by following the decrease of specific activity of [¹⁴C]lysine-labeled proteins. Most of the labeled amino acid injected intraperitoneally into the animal was immediately available for the tumor cells, with only a minor loss towards the extra-ascitic compartment. It was thus possible to calculate the dilution of the isotope in the ascitic pool of the lysine, which increased concurrently with the ascitic plasma volume. Amino acid transport capacity did not change in the log vs. the resting cells. This fact permitted the correction of the specific activity of the proteins synthesized by tumors in the two phases, taking into account the dilution effect. Protein synthesis was found to proceed at a constant rate throughout each of the two phases, although it was 30% lower during the resting as compared to the log phase. When cell mass attained the steady-state, protein degradation occurred at such a level as to balance the synthesis. Throughout the resting phase the amount of lysine taken up by the cells and renewed from the blood remained unchanged. Protein turnover, as studied in subcellular fractions, exhibited a similar rate in nuclei and microsomes, where it proceeded at a higher level than in mitochondria. On the whole, the results encourage the use of the Yoshida ascites hepatoma as a suitable model for studying protein turnover in relation to cell growth in vivo.     

HPD在胃癌细胞各时相中的转运分布和损伤部位的关系

本文探讨了HPD衍生物加红光对人胃低分化腺癌MGC 80-3细胞不同周期的生物学效应。我们观察到HPD的转运与分布决定于细胞周期。G_1期在30分至60分钟内HPD从膜转运至胞质;S、G_2期则直接进入胞质的不同部位;而M期在核部位弥散分布。同步化细胞经HPD加红光处理后,引起细胞大量光敏杀伤,S与G_1期较明显,而M期光敏性最小。我们还观察到:不同周期细胞HPD的分布和HPD的光敏损伤部位密切相关。核仁对HPD的选择性结合也很明显。     

乳腺癌中Vimentin与P-糖蛋白表达和其它预后指标的相关性研究

检测乳腺癌 V im entin与 P-糖蛋白的表达 ,对其相关性进行研究 ,同时与其它病理指标进行比较分析。应用免疫组织化学技术 (S- P法 )对 6 1例浸润性乳腺癌石蜡包埋组织进行检测。Vim entin阳性 2 2例 (36 .1% ) ,P-糖蛋白阳性 18例(2 9.5 % ) ,在 P-糖蛋白阳性或阴性表达组中 ,Vimentin的阳性表达率分别为 6 6 .7% (12 / 18)和 2 3.3% (10 / 43) ,两者呈显著的正相关 (P<0 .0 5 ) ;同时 ,Vimentin的表达在高病理分级病例中为 5 7.1% ,亦呈显著的正相关 (P<0 .0 5 ) ,而与淋巴结转移和 PCNA指数均无明显的相关性 (P>0 .0 5 )。在乳腺癌中 V im entin的表达可能是多药耐药性的一个表型 ,且表明肿瘤细胞分化程度较低 ;所以 Vimentin的表达提示病人预后较差     

大肠癌细胞GST-Pi免疫细胞化学的超微结构定位

探讨GST－Pi在大肠癌细胞内亚微结构定位及其与大肠癌发生的关系。应用免疫电镜技术（免疫胶体金法）对6例大肠腺癌及3例正常大肠粘膜细胞进行GST－Pi的定位观察。6例大肠癌细胞内均出现GST－Pi胶体金阳性颗粒,主要分布于胞浆的线粒体、溶酶体、近胞膜部位的胞头中,也分布在核内和核膜上。金颗粒呈团状、点灶状分布,图象清晰。3例正常粘膜细胞内未见GST－Pi的金颗粒阳性表达,结果表明,GST－Pi在癌细胞内的特异性表达可作为大肠癌的诊断指标之一。