高频电刺激对PC12细胞形态及代谢水平的影响

目的:研究细胞外高频电刺激(HFS)对PC12类交感神经细胞的形态及代谢水平的影响.方法:将HFS作用于传代的PC12类交感神经细胞株,高频电流参数为130 Hz,500μA,60μs;每天固定时间刺激3 h,持续3 d.分别用H-E染色、免疫组织化学、电子显微镜等方法检测经HFS后PC12类交感神经细胞在光镜、电镜下的形态变化及细胞代谢水平的变化.结果:相比对照组,HFS后PC12类神经细胞光镜下突起长度显著缩短,突起个数明显减少;酪氨酸羟化酶免疫组织化学结果显示HFS后PC12细胞胞质灰度降低,提示酪氨酸羟化酶代谢水平下降;电镜下可见胞质中的线粒体、分泌颗粒减少,突起和突触减少.结论:HFS对PC12类交感神经细胞有抑制作用,它抑制PC12类交感神经细胞突起的生长以及颗粒分泌的代谢水平.     

肽类激素分泌机制的研究进展

肽类激素贮存在大型致密核芯分泌颗粒内,通过胞吐分泌,分泌颗粒被膜与细胞膜融合成单层,释放出全部内容物,或通过接触-分离的方式,多次部分释放。对新发现的孔体或融合孔的研究,揭示了分泌颗粒停泊、融合并释放出内容物的机制。排放到细胞外的囊泡内容物的消散速度,取决于融合孔的状态和囊泡内容物的化学性质。由于在细胞外存在完整的膜包分泌颗粒.因此,可能还存在着连同颗粒被膜的整体释放方式。     

胰组织结构提示多肽胰岛素的正常转运经淋巴而非门脉途径

目的　探讨胰岛多肽激素释放形式和细胞外正常转运途径的结构基础。方法　对大鼠和人胰尾部组织进行了光镜 (LM )和透射电镜 (TEM)观察。结果　胰组织主要由腺泡构成的腺小叶和分隔腺小叶的结缔组织所组成。胰腺的导管、血管和淋巴管 ,有髓和无髓神经纤维 ,均穿行于胰结缔组织内。在胰岛周围的结缔组织间隙内也可见到完整的膜包分泌颗粒 (2 0 0～ 5 0 0nm)。胰腺的毛细血管为窗孔 (5 0nm)型。结论　胰组织结构特点提示 :胰岛多肽激素的释放形式 ,可能是连同颗粒膜的整体释放而非传统认为的胞吐分泌 ;释放入胰组织液中的多态胰岛素或分泌颗粒 ,更易进入淋巴而非门脉血液     

神经垂体分泌物释放入脑脊液主要途径结构基础的电镜研究

目的探讨神经垂体分泌物直接释放入脑脊液途径的结构基础。方法透射电镜和扫描电镜观察大鼠神经垂体。结果神经垂体无髓神经纤维终止于毛细血管或垂体细胞周围,垂体细胞突起与神经纤维膨体及终末之间形成细胞间池。有孔型毛细血管内皮外有基膜与血管周隙分隔,血管周隙内可见到完整的膜包分泌颗粒(100~300 nm)。神经垂体表面的囊上皮孔(2~5μm)附近经常可见分泌颗粒。结论细胞间池-血管周隙-囊上皮孔,构成了神经垂体分泌物释放入脑脊液主渠道的结构基础。     

小白鼠腘淋巴结淋巴窦的超微结构

我们观察了幼年小白鼠腘淋巴结淋巴窦的超微结构,窦壁由腔面向外一般出3层构成:(1)一层连续的内皮;(2)由一薄层细胞间质构成的基膜;(3)一层外膜网状细胞及其扁平的突起。内皮细胞核扁圆,异染色质细小,核仁不明显,胞质极薄,含粗面内质网极少,而有大量的吞饮小泡。细胞邻接处相互重叠或嵌合,有20nm宽的间隙相隔。有的地方可见不发达的细胞连接。当巨噬细胞或淋巴细胞穿越内皮时,可出现临时性间隙,内皮边缘与穿越细胞相贴。基膜由电子透明的无定形基质及细的胶原原纤维构成。外膜网状细胞的核异染色质较多,核仁明显,胞质丰富,含许多粗面内质网而很少吞饮小泡。外膜网状细胞之间常见0.5μm宽的间隙。在窦腔内有巨噬细胞、淋巴细胞及星状的网状细胞等。由于小白鼠腘淋巴结的淋巴窦小,故窦内星形的网状细胞很少,其超微结构特点,与外膜网状细胞及淋巴组织内的网状细胞相似,在突起的切面上,常见包有小束的胶原原纤维。巨噬细胞较多,形态不规则,常贴附于内皮细胞表面,核椭圆,常有凹陷,胞质丰富,含许多溶酶体。在取材前1h,于足垫注射中国墨汁的小鼠,巨噬细胞吞噬了大量粗的墨汁颗粒。本文认为,淋巴窦壁可能具有一定的屏障作用。     

大鼠腺垂体促肾上腺皮质激素细胞分泌方式的免疫电镜研究

实验用雄性Wistar大鼠,以在无菌条件下腹腔注射松节油的方法制成急性非特异性腹膜炎,取大鼠腺垂体用免疫电镜方法进行染色.发现促肾上腺皮质激素(ACTH)细胞内分泌颗粒向细胞表面突起,并可见细胞间隙内完整的金标记的内分泌颗粒,实验结果表明,ACTH细胞除传统认为的胞吐分泌外还有其它分泌方式.     

大鼠坐骨神经分支选择性损伤后脊髓背角毛细血管和星形胶质细胞的反应

目的探讨成年大鼠坐骨神经分支选择性结扎切断(SNI)后,腰段脊髓背角内毛细血管和星形胶质细胞的反应。方法成年雄性SD大鼠40只,随机分为光镜组(n=30)和电镜组(n=10)。光镜组又分为对照组、假手术组和SNI手术组。SNI手术为分别结扎切断左侧胫神经和腓总神经,保留腓肠神经,存活20d。部分动物在术后经尾静脉给予30g/L伊文思蓝,切取脊髓L4平面,荧光显微镜观察伊文思蓝的渗出;另一部分在术后进行常规固定取材,切片进行抗鼠IgG和GFAP的免疫组织化学反应。电镜组(对照组和SNI后20d组),脊髓切片进行常规抗大鼠IgG的免疫电镜处理。结果 1.SNI后20d,可见明显的伊文思蓝渗出红色光斑;2.SNI后GFAP阳性细胞变为反应型,数量明显增加,毛细血管壁及其周围的IgG颗粒也显著增加。3.电镜观察:SNI术后20d,IgG阳性颗粒增多,内皮细胞观察到下列变化:(1)内皮细胞向管腔内伸出许多微细的突起,有的含有IgG阳性颗粒。(2)内皮细胞的管腔侧胞膜上有许多"内吞"小凹,凹内可见IgG阳性颗粒,胞质内有含IgG阳性颗粒的小泡,在反管腔侧胞膜上有"胞吐"现象。(3)内皮细胞紧密连接出现间隙,间隙内有IgG颗粒。(4)胞质内出现囊泡,有的泡内显示IgG阳性颗粒。结论 SNI后脊髓背角内毛细血管内皮细胞发生明显的反应,大鼠IgG和伊文思蓝可经过血脊髓屏障渗出,星形胶质细胞被激活。     

衰老大鼠禁水时视上核加压素能神经元超微结构变化

电镜观察不同禁水时间衰老大鼠下丘脑视上核加压素能神经元的超微结构变化。结果发现：禁水６、１２ｈ后，视上核加压素能神经元胞体增大，胞质中粗面内质网排列紧密且增多，高尔基复合体成熟面未成熟分泌颗粒及神经分泌颗粒增多；轴突中神经分泌颗粒增多不明显。而禁水２４ｈ后，神经元胞质内的神经分泌颗粒减少，轴突中的神经分泌颗粒聚集成膨大区域。禁水后，神经胶质细胞减少，突起回缩，使相邻的两神经元直接接触，突触结构重建增多。上述研究结果提示，衰老大鼠下丘脑视上核加压素能神经元在禁水时，其功能活动是增强的，胶质细胞突起回缩，有利于神经元之间的突触结构重建。     

跨膜型TNF-α介导的反向信号对NK细胞杀伤功能的影响

目的：观察跨膜型TNF-α介导的反向信号对NK细胞杀伤功能的影响.方法：用sTNFRI激活NK92细胞的TM-TNF-α反向信号;MTT比色法检测NK92细胞对靶细胞的杀伤作用;β-己糖氨酶释放实验检测NK92细胞在杀伤靶细胞时的胞吐作用;ELISA实验检测NK92细胞sTNF-α的分泌;流式细胞术检测NK92细胞FasL的表达.结果：预先激活TM-TNF-α反向信号,可以促进NK92细胞对靶细胞的杀伤功能;增强NK92细胞的胞吐;促进FasL的表达和sTNF-α的分泌.结论：TM-TNF-α反向信号可能通过促进NK92细胞胞吐、sTNF-α分泌及FasL的表达而促进NK92细胞对靶细胞的杀伤.     

大鼠视上核神经分泌细胞的衰老形态变化

本文选用青年(3个月)、成年(10—12个月)和老年(28—30个月)雄性Wistar大鼠,对其下丘脑视上核神经分泌细胞分别作了光镜和电镜观察与测定。结果表明:老年大鼠视上核前部的神经分泌细胞密度与细胞核体积明显低于青年和成年大鼠,提示老年大鼠视上核有神经元消失和细胞活性降低的表现;神经分泌细胞核周质内的醛复红阳性颗粒未测出三个年龄组大鼠间的差异,含醛复红阳性颗粒的胞突在老年时却显示增粗和膨大;老年动物神经分泌细胞超微结构的主要变化为神经分泌颗粒和脂褐素增加,尤以后者为甚。老年大鼠少数神经分泌细胞还表现有粗面内质网和核蛋白体减少,排列紊乱,池囊扩张和脱颗粒,高尔基复合体池囊变窄或扩张,胞质和胞突内有自噬体样物质出现等衰老改变;多数细胞胞质内的其他结构同青年和成年大鼠者相似。此外。老年大鼠视上核神经毡结构中有多层膜包绕胞突,终末和突触等的鞘样结构出现增多。本研究表明大鼠衰老时视上核神经分泌细胞活性降低。     

The fine structure of rat liver sinusoids,space of Dissé and associated tissue space

Three structurally distinct zones are present in the liver sinusoid. The endothelium and basement (boundary) membrane of the portal vein extend uninterruptedly into the peripheral zone. The intermediate zone, comprising 90% or more of the length of the sinusoid, possesses a fenestrated lining and no basement membrane. The short central zone has unfenestrated endothelium and a basement membrane. Both are continuous with those of the central vein. The space of Dissé encircles all three zones of the sinusoid. It contains fat storage cells, perisinusoidal cells, numerous microvillae of liver cells and reticular fibers. These fibers are bundles of unit collagen fibers enclosed by cytoplasm of nearby cells. The space of Dissé is continuous with the tissue space at both ends of the sinusoid. The liver cells lack a basement membrane whether they abut on the space of Dissé or on the tissue space proper. The unique feature of liver fine structure is the combination of fenestrations and lack of basement membranes in the intermediate zone of the sinusoid. Blood plasma is thereby afforded intimate contact with the parenchymal cells and has access to the tissue space at both ends of the sinusoid. This structural situation fits the known facts of liver function.     

大白鼠心脏小强荧光细胞超微结构研究

本文采用醛诱发荧光的荧光组织化学方法,首先定位小强荧光细胞,在电子显微镜下观察它们的超微结构。心脏的小强荧光细胞体积小,胞质含大量两种不同电子密度的颗粒泡。它们的线粒体丰富,每个沿核水平切面有20个线粒体;高尔基复合体发达,每组为4～7个扁囊构成;粗面内质网一般呈单独扁囊,散布在胞质。小强荧光细胞与胆碱能神经末梢形成传入性突触,相邻小强荧光细胞常形成粘合斑结构和相嵌联结。它们相邻的毛细血管,多为有孔型毛细细管。它们在相邻细胞之间、邻近血管和细胞间隙部位有胞吐现象。结果提示,这种细胞可能以内分泌或旁分泌方式,起局部调节作用。     

HISTOLOGICAL AND FINE STRUCTURAL STUDIES ON PIGMENTED NEUROECTODERMAL TUMOR OF INFANCY

Pigmented neuroectodermal tumor of infancy originating from the anterior maxilla in a two-month-old male has been studied by light and electron microscopy. The tumor is characterized histologically by two types of neoplastic cells embedded in considerable amounts of fibrous stromal tissue. The first type of cell is cuboidal to columnar in shape with an epithelial appearance having abundant cytoplasm; either scanty or a heavy accumulation of melanin pigment is observed in the cytoplasm. These cells are aligned along the cleft-like space or arranged in small ductal structures. Electron microscopy shows the characteristic features of melanocytic cells having a varying degree of asynchronous maturation of melanosomes. The second type of cells is small and round in shape and has a hyperchromatic nucleus and scanty cytoplasm, resembling neuroblastic cells or lymphocytes. Electron microscopy reveals cytoplasmic processes resembling immature neurits which protrude into the intracellular space; a small number of secretory granules having a central core surrounded by a single limiting membrane are observed in the cytoplasm and cytoplasmic processes.     

大鼠松果体分泌物释放到蛛网膜下隙脑脊液途径的电镜研究

为探讨松果体分泌物在细胞外转运到蛛网膜下隙脑脊液途径的结构基础,本研究运用透射电镜技术对成年大鼠松果体浅部进行了观察。结果显示:(1)大鼠松果体毛细血管为有孔型(孔径50nm),借基膜与宽阔的血管周隙分隔;(2)松果体细胞借基膜与结缔组织间隙分隔,松果体细胞之间形成与松果体囊结缔组织间隙及血管周隙相通的细胞间小管;(3)松果体囊表面为扁平上皮,上皮下为疏松结缔组织间隙,其内可见松果体细胞的胞体和突起。囊上皮细胞间存在瓣膜样连接(叶瓣间宽处可达1μm)和囊上皮孔(2.5μm)。以上结果提示,松果体细胞表面可分为结缔组织间隙面、连接面和细胞间小管面三个功能面,松果体分泌物可能通过血管周隙-细胞间小管-松果体囊结缔组织间隙-囊上皮瓣膜样连接或囊上皮孔,释放入蛛网膜下隙脑脊液中转运。     

Eosinophilic leucocytes in nasal allergy—movement of enzymes

The changes in peroxidase of the granules of eosinophils were examined in cells in the nasal mucous membranes of persons allergic to house dust. There was a slight leakage of peroxidase from eosinophil granules immediately after placing an antigen-treated disc (house dust, Torii, 2560 μg/disc) on the nasal mucous membrane. Leakage of peroxidase from the eosinophil granules into the cytoplasm, and extracellular release from some eosinophils, was greater 20 min after provocation, than at 30 sec. There were two forms of peroxidase release from the plasma membrane. In one, the granule membrane fused with the plasma membrane and the peroxidase was released from the cell through destruction of the two fused membranes. This was considered to be active secretion. In the other form, the plasma membrane was damaged by peroxidase released from the granules into the cytoplasm. This was considered to be passive release following degeneration of the cell.     

Ultrastructural changes in guinea-pig bone marrow basophils during anaphylaxis

A sequence of ultrastructural changes related to the degrees of severity of the anaphylactic reaction was observed in the bone marrow basophils of guinea-pigs. This sequence of changes might represent the sequential stages of a process of degranulation which consisted of (1) formation of haloes of low electron-density around the granules, (2) development of vacuoles by intercommunicating the adjacent haloes and of confluent vacuoles by joining the small vacuoles, and (3) disruption of cell membrane with concomitant expulsion of granules into the intercellular space or sinusoid. The vacuoles might communicate with the cisternae of the endoplasmic reticulum. Degranulation was found to be a result of an antigen—antibody reaction in which the reaginic antibodies produced had a selective affinity for the cell membrane of the basophils. The mechanism of degranulation might involve two factors: a physicochemical factor of changing the nature of the cell membrane resulting from the antigen—antibody interaction, and a mechanical factor of pressure exerted on the cell membrane by the confluent vacuoles. The alterations in the granules themselves, such as relatively larger size, lighter electron-opacity and looser texture, appeared to be associated with the histamine release from the granules. The structural changes of the granule contents encountered in anaphylaxis suggest that the ultrastructure of the basophil granules may be composed of an orderly interlacing framework of microfilaments onto which other constituent substances are bound.     

Distribution of elements in the pancreatic exocrine cells determined by electron probe X-ray microanalysis

We measured the intracellular electrolytes of acinar cells by making electron probe X-ray microanalysis of hydrated and dehydrated sections of freshly frozen dog pancreas.The concentrations of electrolytes in the cytoplasm were: Na 4.8±2.1, K 132±15, Cl 14±4.7, P 165±36, S 19±2.8 and in zymogen granules: Na 6±5, K 60±16, Cl 31±20, P 36±8, S 172±25, Ca 7±5 (mean±S.D. mmol/kg wet weight).The cytoplasm, which is rich in rough endoplasmic reticulum, had low Na and high K concentrations, as compared with levels in the acinar cells of other exocrine glands such as the submandibular gland, the bulk of which is occupied by secretory granules. Though the representative feature of secretory granules was a high S content, occasionally low S peaks of spectra from secretory granules were obtained. These findings may reflect the content of mature zymogen granules and immature condensing vacuoles.Pilocarpine stimulation increased cytoplasmic Na, Cl and Ca and decreased K levels in the pancreatic acinar cells. This indicates that secretory stimulation increases the permeability of the cell membrane to Na, Cl and K ions and that there is a simultaneous Ca release from the intracellular Ca stores such as zymogen granules and endoplasmic reticulum, and/or Ca influx from the extracellular space.     

Dynamics of epithelial cells in the corpus of the mouse stomach. II. Outward migration of pit cells

The pit cells (or surface mucous cells) present along pit walls and gastric surface have been investigated by electron microscopy and radioautography after a pulse or continuous infusion of ³H-thymidine. For these studies, the pit region has been subdivided into four segments: three of equal length along the pit wall, respectively named low pit, mid pit and high pit, and a last one at the surface named pit top. The pit region includes an average of 37 pit cells, characterized by dense mucous granules accumulated along the apical membrane in an organelle-free zone referred to as ectoplasm. Continuous ³H-thymidine infusion reveals that pit cells come from pre-pit cells, which are believed to arise in the isthmus region from the undifferentiated granule-free cells through a pre-pit cell precursor stage. The pre-pit cells, characterized by the presence of a few mucous secretory granules scattered in the cytoplasm, migrate outward (i.e., in the direction of the gastric lumen). When the secretory granules line up along the apical membrane in the ectoplasm, the pre-pit cell becomes pit cell. It is estimated that 87% of pit cells differentiate from pre-pit cells, while the remaining 13% come from their own mitoses. Observations at successive times after a ³H-thymidine pulse demonstrate that pit cells, like pre-pit cells, migrate toward the gastric surface where they are eventually lost. The continuous ³H-thymidine infusion results indicate that this migration takes 3.1 days on the average. Cells spend almost a day in each pit wall segment. In the low pit segment, cells produce more and larger mucous secretory granules than do pre-pit cells. In the mid and high pit segments, the number and size of the granules generally keeps on increasing, thus indicating that mucous differentiation is progressing. The secretory granules arising in the Golgi apparatus of pit wall cells are mostly spherical; they retain this shape during the few minutes taken to cross the cytoplasm and enter the apical ectoplasm. They spend about an hour in the ectoplasm, where they change to an ovoid shape as they approach the apical membrane to finally release their content by exocytosis. The mucous differentiation along the pit wall is associated with a progressive decline in the organelles: nucleoli and mitochondria decrease in size while the amount of free ribosomes diminishes. When pit cells reach the free surface, they produce fewer, smaller secretory granules and at a lower rate than in mid and high pit. Meanwhile, organelles decline further, while mitochondria tend to swell and disintegrate. Clearcut signs of degeneration appear in some of the cells. These cells find their way into the gastric lumen either by direct extrusion or indirectly after being phagocytosed by a neighbor cell which is itself eventually extruded. Thus a sequence of cells—the pit cell lineage—derived from the stationary undifferentiated granule-free cells, includes pre-pit cell precursors, pre-pit cells, and pit cells, which all migrate in the direction of the gastric lumen, where pit cells are eventually lost. © 1993 Wiley-Liss, Inc.     

Identification and localization of adrenomedullin-storing cardiac mast cells

Adrenomedullin (AM), a potent vasodilatory hypotensive peptide, is expressed in the heart, where it is known to play a protective action. Light-microscopy immunocytochemistry (ICC) demonstrated the presence of AM immunoreactivity not only in the coronary-vessel wall and ventricular myocytes of the human and rat heart, but also in sparse voluminous cells located in the perivascular space. These cells displayed the same location of toluidine blue-positive mast cells, and electron microscopy ICC showed AM-immunogold staining over the granules of rat cardiac mast cells. The incubation of rat left ventricle fragments with the mast-cell histamine releaser compound 48/80 evidenced groups of AM-positive cells undergoing degranulation and caused an increase of approximately 50% in the AM concentration in the incubation medium. Collectively, our findings provide evidence that at least a subset of cardiac mast cells are able to synthesize and store AM, and upon stimulation to release it near coronary arterioles and venules.