糖尿病大鼠穹窿下器室管膜细胞的形态学变化

目的:观察糖尿病大鼠不同时期穹窿下器的室管膜细胞的形态学变化.方法:雄性Wistar大鼠,每只大鼠给予链脲佐菌素60 mg/kg,腹腔一次性注射,建立Ⅰ型糖尿病大鼠模型.造模成功后分别于2、4及8周,扫描电镜和透射电镜观察穹窿下器不同时期室管膜细胞的形态学变化.结果:扫描电镜观察,对照组细胞膨隆,表面光滑,有清晰的微绒毛.2周组细胞扁平、塌陷,细胞表面可见到圆形小破孔.4周组室管膜细胞表面凹凸不平,出现皱褶,细胞膜可见大小不一的多个小破孔,并可见细胞膜缺失、细胞器裸露的细胞.8周组室管膜细胞表面皱褶明显,可见大量的球形分泌颗粒.透射电镜观察显示,对照组穹窿下器室管膜细胞脑室面完整,有少量的微绒毛和乳头状微突起,模型组室管膜面可见大量分泌颗粒.结论:糖尿病可导致大鼠穹窿下器室管膜细胞破损及分泌大量的分泌颗粒.     

大鼠中幼、晚幼红细胞与小鼠浆细胞瘤SP 2/0细胞融合后的光镜和电镜观察

本文报道大鼠中幼或晚幼红细胞与小鼠浆细胞瘤sP2/0细胞异种杂交后的光镜和电镜观察结果。融合后杂交细胞体积变大,微绒毛、指状突和皱褶突起减少。细胞的核质比明显变小,核内异染色质增多,核仁数目减少或消失。少数杂交细胞出现线粒体肿胀,核固缩甚至排核。长期传代的杂交细胞保持较少的表面微绒毛及皱褶突起、核质比例变小等现象,某些细胞的胞质中可见有空泡或致密颗粒。上述现象为杂交细胞去恶性提供了细胞形态以及超微结构变化的根据。本文并对杂交后细胞核、胞质及细胞表面的形态变化与去恶性之间的可能关系进行了讨论。     

cAMP对人胃腺癌细胞系SGC-7901的影响——微绒毛和Na~+—K~+—ATP酶活性变化的电镜观察

本实验通过外源性 cAMP 和茶碱对人胃腺癌细胞系 SGC-7901的连续作用,观察到外源性 cAMP 和茶碱对该细胞株的增殖,具有明显的抑制作用。在透射电镜和扫描电镜下,均可见到细胞表面较光滑,微绒毛减少或消失。同时见到细胞膜表面上的 Na~+—K~+—ATP 酶活性受到抑制。本文讨论了外源性 cAMP 对人胃腺癌 SGC-7901细胞系增殖的抑制作用,以及膜表面微绒毛和Na~+—K~+—ATP 酶活性变化的关系。     

高、低转移的二株亚克隆细胞系的超微结构比较研究

对来自同一祖先的两株具有不同转移能力的亚克隆细胞系Ｈｃａ－１６Ａ３－Ｆ（Ｆ）和Ｈｃａ－Ａ２－Ｐ（Ｐ）进行了形态学观察，并应用体视学方法进行定量比较分析。结果表明，Ｆ细胞的剖面平均直径、核质比、核仁核比均大于Ｐ（Ｐ＜０．００１）。Ｆ的核畸形、核仁边集多见，胞质中粗面内质网、游离核糖体丰富，细胞表面微绒毛较多。Ｐ中有较多的脂滴。Ｆ和Ｐ中均没有发现Ａ型病毒颗粒。表明同源而具有不同转移能力的瘤细胞在形态上也有一定的差异。     

人表皮的冷冻复型电镜观察

本文应用冷冻复型法,对5例人小腿部脾经与胃经低电阻线下的表皮,进行了电镜观察。发现棘细胞具有粗大突起和微绒毛。颗粒细胞呈梭形,细胞表面呈波浪状;其胞膜的劈裂面上膜内微粒较棘细胞者少;但膜被颗粒的外排小孔则较之为多,其大小为50～75nm,平均数为47.5/μm~2;胞质内除个别线粒体外,主要为呈各种走向的张力细丝束;其间散布着膜被颗粒,接近角化细胞者多位于胞膜下方,直径为60～88nm。颗粒细胞核膜的膜内微粒及按孔均很稀少。角化细胞呈扁平状,胞膜明显增厚,其劈裂面上除桥粒区外,无膜内微粒和外排小孔;深层细胞的胞质可见大量细丝,而浅层者胞质呈均质状。桥粒在深层细胞多,至角化层渐减少,且呈退化状态。在颗粒层细胞间隙内,可见由膜被颗粒外排物质先附于细胞外表面,继而出现板层状膜及原纤维状结构,后者直径为70～100nm,具有间距为60～70nm的横纹。     

不同基因型别乙型脑炎病毒在不同细胞系上的生长特点比较

目的研究不同基因型别乙型脑炎病毒在不同细胞系上的生长特点, 为乙脑病毒的研究中细胞系的选择提供科学依据。方法选择BHK-21、Vero、C6/36、PK-15、DF-1、N2a、SH-sy5y和MDCK细胞系, 通过空斑测定法和RT-qPCR法评价基因1型(G1, NX1889株)、基因3型(G3, P3株)和基因5型(G5, XZ0934株)乙脑病毒在上述细胞系中的增殖能力。结果 3种基因型别乙脑病毒在BHK-21、Vero、C6/36、DF-1、N2a和PK-15细胞系中存在明显的细胞病变效应(CPE), 在同一种细胞系中引起CPE的特点并未观察到明显不同。病毒感染SH-sy5y和MDCK细胞系后没有明显的CPE出现, 但是在SH-sy5y细胞系中存在病毒的增殖, 而MDCK细胞系为非敏感细胞系。G1、G3和G5型乙脑病毒在BHK-21、Vero和SH-sy5y细胞系中的增殖曲线没有明显差异;在C6/36和PK-15细胞系中, G1型乙脑病毒的滴度高于其他两型乙脑病毒;在DF-1细胞系中, G5型高于其他两者, 在N2a细胞系中, G5型低于其他两者。结论 3种不同基因型别的乙脑病...     

小白鼠腹膜间皮的扫描电镜观察及机能探讨

我们用扫描电镜观察了5只小白鼠的脏、壁层腹膜间皮及肠系膜腹膜间皮,发现腹膜间皮细胞表面有排列形式不一,形态各异的微绒毛、火山口样凹陷、细胞表面的圆形缺损区及颗粒等。在肠系膜腹膜间皮还发现了细胞间孔。我们还观察到间皮细胞具有六种不同的形态。     

醋酸棉酚对HeLa细胞作用的透射电镜和扫描电镜观察

本文报道人体子宫颈癌HeLa细胞系细胞的超微结构,及其在培养液中加入10,20μg/ml醋酸棉酚,作用24,48h后的变化状态。HeLa细胞在常态生长情况下,具有一般癌细胞的超微结构特点。在醋酸棉酚的作用下,这些细胞间的紧密连接未见发生改变。而在细胞内,线粒体首先发生变化,由肿胀、嵴断裂、空泡化,直至溶解成颗粒状物。棉酚对HeLa细胞的损害作用,具有剂量相关性和作用时间相关性。在棉酚浓度加大和作用时间增长时,可见除线粒体以外的其他细胞器的改变,包括溶酶体大量增多、核膜受损及核内异染色质改变等。扫描电镜下,可见微绒毛断落,细胞表面出现极不正常的泡状突起。本文还对棉酚损伤HeLa细胞的性质及其意义进行了讨论。     

小白鼠胸膜间皮的扫描电镜观察

目的　观察小白鼠脏、壁胸膜间皮细胞在扫描电镜下的结构以及两者在结构上的异同。方法　用扫描电镜对比观察 5只小白鼠的脏胸膜及各部壁胸膜的结构。结果　脏、壁胸膜间皮细胞表面有大量粗、细微绒毛 ,较多的火山口样凹陷及一些细胞间孔 ;脏胸膜间皮表面粗微绒毛密集且分支多 ,分支内有大量串珠样颗粒 ,多见具有芽状或颗粒状微绒毛的幼稚间皮细胞 ;壁胸膜间皮表面粗微绒毛稀疏 ,细微绒毛密集 ,多见微绒毛退化消失且有即将与间皮分离的衰老细胞、火山口样凹陷及细胞间孔。结论　脏胸膜间皮可能具有较强分泌功能 ,而壁胸膜间皮吸收功能较强     

在扫描电镜下观察培养不同时间的人鼻咽癌细胞株(CNE)表面特殊结构变化过程

在常规培养瓶中附贴小块盖玻片,使CNE细胞株在玻片上生长。培养6、24小时和2、3、4天后,各取出附在玻片的瘤细胞进行扫描电镜观察。检查结果,泡状突起和微绒毛是瘤细胞表面结构变化的主要特征。培养后一天,瘤细胞表面出现大量分布均匀的泡状突起,第二天开始分布于细胞周边,第三天后则几乎完全消失。而在培养第一天时,细胞边缘仅有少量微绒毛存在;到第二天时,微绒毛在细胞表面呈稀疏排列;第三天后,可见瘤细胞表面布满致密的微绒毛。此外,培养一天时,在细胞表面还查见叶状伪足和折叠皱纹状结构,后者于第二天消失。丝状伪足在不同培养时间内均可查见,但多分布于细胞一侧,也有的沿细胞周边分布。丝状伪足可见有2级分叉,其末端呈吸盘样结构。本实验表明,CNE细胞株的表面结构不是恒定不变的,而是具有明显的易变性,并可因培养时间的长短,而发生不同的改变。     

小鼠类巨噬细胞系超微结构的观察

本文报道了小鼠类巨噬细胞系(MMC-1)的超微结构。在透射电镜下,可见暗细胞、亮细胞及空细胞。这三种细胞有一定的比例。在其他培养细胞中也见到了类似的三种细胞。暗细胞电子致密度最高,呈圆形、椭圆形或梭形。表面有许多突起,有的呈丘状。胞质可分内、外两部分。外胞质无细胞器;内胞质中有丰富的线粒体、粗面内质网和聚核蛋白体。溶酶体易见,高尔基复合器发达,有成堆的微纤维。胞核不规则,多呈肾形,位于细胞的一侧。核膜清楚,常染色质较多。有的胞核中可见核仁。亮细胞突起少,线粒体肿胀,溶酶体与脂滴易见,核肿胀。空细胞的突起更少,内质网明显减少或消失,溶酶体多,次级溶酶体易见胞核大,染色质稀疏。本文根据此细胞的超微结构特点,证实其为巨噬细胞,并根据其来源称之为类巨噬细胞。     

Morphological changes in BHK-21 cells infected with S/N (H2N1) influenza virus

Summary Morphological changes developing in BHK-21 cells infected with the S/N virus (a recombinant of A₂/Singapore and A/NWS viruses), at the approximate m.o.i. of 1 TCD₅₀ per cell, were studied by electron microscopy. Substantial alterations were observed in both the nucleus and cytoplasm. Changes in the nuclei were first detected eight hours after infection. The nucleoli substance was reduced and dense inclusions consisting of fibrillar structures 40–60 Å in diameter were formed. The greatest number of inclusions and their distribution over the whole nuclear area were found 24 hours after infection. Electron-dense inclusions were also detected in the cytoplasm. They were identical in shape and size with those observed in the nuclei but they consisted of fibrils 35–40 Å in diameter. These inclusions were rare eight hours after infection. Their number increased with the incubation period, reaching a maximum 24 hours after infection. Six hours after infection filamentous structures 80–90 Å in diameter were observed in the cytoplasm. It is assumed that they represent viral ribonucleoprotein. They were at first (hour 6) distributed diffusely through the whole cytoplasm and were later (hour 8–10) found mainly near the cytoplasmic membrane. Twenty four hours after the infection the filamentous structures were abundant both in the central part of the cytoplasm and in the proximity of the cellular membrane. Sometimes they formed dense agglomerations 2000–4200 Å in width. Maturation of the virion took place at the plasma membrane and in cytoplasmic vacuoles in whose proximity the filamentous structures were found. The particles either budded directly from the smooth cell membranes or from processes formed on the cell surface. Numerous rod-like particles were observed.     

Morphological observations on the replication of herpesvirus saimiri in monkey kidney cell cultures.

Owl and African green monkey kidney cell cultures have been infected with 1 p.f.u./cell of herpesvirus saimiri and sample cultures have been taken for examination by electron microscopy at 3 to 6 hourly intervals over a period of 7 days; the experiments were repeated several times. The peculiarly slow replication cycle of Herpesvirus saimiri has enabled distinct cytoplasmic and nuclear phases in virus maturation to be clearly distinguished; the overall fine structural features were similar in both cell types. Immature particles were first detected in the nucleus and cytoplasm 63 h after infection. Thereafter, abundant cytoplasmic immature particles matured by budding through cytoplasmic membranes until about 100 h, whereas nuclear immature particles budded through the inner nuclear membrane or intranuclear invaginations of it later, from about 100 h until cytolysis was complete at 160 h. Morphological differences were also observed between particles budding at cytoplasmic membranes and the nuclear envelope. At the former site the membrane overlying the bud showed an electron opaque thickening which imparted to the mature particle an asymmetrical appearance. Such thickenings of the envelope were not observed in mature particles of nuclear origin. Unusual tubular and laminated nuclear structures were seen towards the end of the replicative cycle corresponding with the phase of nuclear virus maturation by budding; the morphology of the latter structures is described.     

Frog virus 3 replication: electron microscope observations on the terminal stages of infection in chronically infected cell cultures.

An examination of BHK, CEF, and FHM cells chronically infected with frog virus 3 has been made by scanning and transmission (thin section, freeze fracture, and surface replica) electron microscopy. With minor differences the pattern of virus development is similar in all three cell line. Virus particles were detected in cell nuclei which subsequently became degenerate very late in infection. Three inclusions were associated with frog virus 3 cytoplasmic foci of infection; lamella structures, extensive microtubule formation (in BHK and FHM cells), and linear crystalline structures. The last two structures may play a role in creating or maintaining the cell rounding c.p.e. revealed by scanning electron microscopy. Very late in infection most BHK and FHM, but not CEF, cells are stripped of the plasma membrane. Replicas of frozen fractured BHK cells featured cytoplasmic foci of infection, budding at the plasma membrane, and showed that at early times when virus is detected in the nucleus, the nuclear membranes are intact and morphologically unaltered. Budding at the plasma membrane was better resolved by scanning and as surface replicas. This demonstrated that sparse to profuse localized budding occurred. Frequently virus particles were located singly, or as multiples, at the end of, or along, cytoplasmic protrusions which occur both on the body of the cells and at the cytoplasmic/coverslip 'interface'.     

Electron microscopy of porcine cytomegalovirus

Summary Porcine cytomegalovirus was examined by electron microscopy in the nasal mucosa of piglets from a natural epizootic of the disease with the object to determine the localization of nucleocapsids arranged in crystalline arrays in the cell and to investigate the structure of most common viral forms. The study revealed the following. 1. The morphogenesis of porcine cytomegalovirus is marked by intranuclear formation of nucleocapsids arranged in crystalline arrays. Large quantities of crystals found in the cytoplasm were apparently extruded from the nucleus at late stages of infection; 2. nucleocapsids in the cytoplasm were coated with an electron dense amorphous material. This coat was also seen at the surface of capsids in cytoplasmic and extracellular enveloped particles, but was absent from the surface of capsids located in the nucleus; 3. the majority of enveloped virus particles showed an electron translucent zone separating the capsid from the envelope, and 4. the outer envelope of cytoplasmic and extracellular particles had projections and showed a clear unit membrane structure, whereas the outer envelope of intranuclear particles and those located in the perinuclear cisterna had the appearance of an electron dense single membrane.     

Cultured Reed-Sternberg cells HDLM-1 and KM-H2 can be induced to become histiocytelike cells. H-RS cells are not derived from lymphocytes.

Two Hodgkin's Reed-Sternberg cell (H-RS) lines, HDLM-1 and KM-H2, have phenotypes and functional properties very similar to those of H-RS cells in tissues. These two types of cells were induced to differentiate with a combination of phorbol ester, retinoic acid, and extracellular matrix. The induced cells displayed the morphology of histiocytes or histiocytelike cells, with a small, round or oval, eccentric nucleus and abundant cytoplasm. In ultrastructural studies, many cytoplasmic projections and rugae were observed. These induced cells exhibited abundant cytoplasmic lysosomal enzymes, such as esterase, acid phosphatase, alpha 1-antitrypsin, or lysozyme. The histiocytic nature of these induced cells was further confirmed by the increased expression of many monocyte/histiocyte markers, including CD11b, CD11c, CD13, CD14, CD15, CD33, CD68, Mac387, and 1E9. In functional tests, the induced cells were shown to produce interleukin-1, tumor necrosis factor, macrophage colony-stimulating factor, and/or prostaglandin E2. Phagocytosis was detected in less than 5% to 10% of the cells when Candida albicans was added to cultures. The results strongly suggest that H-RS cells are related to cells of histiocyte lineage.     

N-methyl-D-aspartate receptors are present in vagal afferents and their dendritic targets in the nucleus tractus solitarius.

N-Methyl-D-aspartate receptors are present in the nodose ganglion, which contains the cell bodies of vagal afferents, and in the nucleus tractus solitarius, where these afferent fibers terminate. This suggests that N-methyl-D-aspartate receptors are located presynaptically on visceral vagal afferents and/or their target neurons in the nucleus tractus solitarius. To test this hypothesis, we combined anterograde transport of biotinylated dextran amine, following injections into the left nodose ganglion, with electron microscopic immunogold labeling of antipeptide antiserum against the R1 subunit of the N-methyl-D-aspartate receptor in the nucleus tractus solitarius of rat brain. Within the medial nucleus tractus solitarius, the N-methyl-D-aspartate receptor R1 immunoreactivity was seen in dendrites (39% of 639 profiles), axons and axon terminals (41%), and a few neuronal perikarya and glia. Many vagal afferent axons and terminals (40% of 468 profiles) contained N-methyl-D-aspartate receptor R1 immunogold labeling. In addition, 42% of the dendrites contacted by vagal afferent terminals (n = 206) contained N-methyl-D-aspartate receptor R1 immunoreactivity. In axons and dendrites, the gold particles were occasionally seen within asymmetric postsynaptic junctions or at non-synaptic sites on the plasma membrane. More commonly, however, N-methyl-D-aspartate receptor R1 labeling was seen on membranes of vesicular cytoplasmic organelles, suggesting that there is abundant N-methyl-D-aspartate receptor protein available for activity-dependent mobilization to the plasmalemma. Since many vagal afferents are glutamatergic, our results implicate N-methyl-D-aspartate receptors in autoregulation of the presynaptic release and postsynaptic responses to glutamate at the level of the first central synapse in the nucleus tractus solitarius.     

Broccoli necrotic yellows virus in cauliflower and in the aphid, Brevicoryne brassicae L.

A rhabdovirus resembling broccoli necrotic yellows virus was found in brassica crops in Victoria. In plants, the virus was restricted to the phloem parenchyma cells. In aphid vectors (Breuicoryne brassicae), a virus of similar (but not identical) form was found in most types of cells except those of the gut and developing embryos. It was shown by transmission experiments that these particles are a host-specific variant of the virus found in plants. In cauliflower, the virus matured and accumulated in cytoplasmic vesicles, and did not accumulate in the nucleus or the perinuclear space, nor at the cell surface. In aphids, the virus matured and accumulated mainly in the nucleus; particles were less frequently found in the cytoplasm and never in large regular arrays comparable with those found in the plant. The virus therefore probably develops in different ways in the two hosts studied.     

Structural characterization of bioengineered human corneal endothelial cell sheets fabricated on temperature-responsive culture dishes

For the purpose of corneal regenerative medicine, we fabricated human corneal endothelial cell sheets on temperature-responsive dishes, which could be non-invasively harvested as intact, transplantable sheets by simply reducing the culture temperature. Cells demonstrated hexagonal cell shape with numerous microvilli and cilia, and also exhibited abundant cytoplasmic organelles similar to these cells in vivo. Immunofluorescence for type IV collagen and fibronectin revealed that abundant extracellular matrix (ECM) was deposited on the basal surface throughout culture, and the deposited ECM was harvested along with the cell sheets by reducing culture temperature to 20 degrees C. Faint ECM remnants were observed on the dish surfaces after cell sheet detachment. Immunofluorescence for ZO-1 showed that tight junctions were established between cells, and immunoblotting indicated that intact ZO-1 was maintained during cell sheet harvest, while conventional proteolytic cell harvest methods resulted in the degradation of ZO-1. These results suggest that these transplantable corneal endothelial cell sheets can be applied to treat patients with damaged corneas.     

Cytoplasmic and nuclear distribution of the protein complexes mTORC1 and mTORC2: rapamycin triggers dephosphorylation and delocalization of the mTORC2 components rictor and sin1

The mammalian target of rapamycin (mTOR) is part of two distinct complexes, mTORC1, containing raptor and mLST8, and mTORC2, containing rictor, mLST8 and sin1. Although great endeavors have already been made to elucidate the function and regulation of mTOR, the cytoplasmic nuclear distribution of the mTOR complexes is unknown. Upon establishment of the proper experimental conditions, we found mTOR, mLST8, rictor and sin1 to be less abundant in the nucleus than in the cytoplasm of non-transformed, non-immortalized, diploid human primary fibroblasts. Although raptor is also high abundant in the nucleus, the mTOR/raptor complex is predominantly cytoplasmic, whereas the mTOR/rictor complex is abundant in both compartments. Rapamycin negatively regulates the formation of both mTOR complexes, but the molecular mechanism of its effects on mTORC2 remained elusive. We describe that in primary cells short-term treatment with rapamycin triggers dephosphorylation of rictor and sin1 exclusively in the cytoplasm, but does not affect mTORC2 assembly. Prolonged drug treatment leads to complete dephosphorylation and cytoplasmic translocation of nuclear rictor and sin1 accompanied by inhibition of mTORC2 assembly. The distinct cytoplasmic and nuclear upstream and downstream effectors of mTOR are involved in many cancers and human genetic diseases, such as tuberous sclerosis, Peutz-Jeghers syndrome, von Hippel-Lindau disease, neurofibromatosis type 1, polycystic kidney disease, Alzheimer's disease, cardiac hypertrophy, obesity and diabetes. Accordingly, analogs of rapamycin are currently tested in many different clinical trials. Our data allow new insights into the molecular consequences of mTOR dysregulation under pathophysiological conditions and should help to optimize rapamycin treatment of human diseases.