Endothelial attachment and plasmalemmal apposition in the transcellular movement of intravascular leukemic cells entering the myeloid parenchyma

The plasmalemmal relationship between metastases-forming leukemia cells and myeloid sinus endothelium during the transmural passage of the leukemia cells has been studied in rat bone marrow. After the myeloid vascular system was freed from normal circulating blood cells, the bone marrow was perfused with a suspension of leukemia cells derived from an ascites tumor. The bone marrow was then fixed by perfusion with double aldehyde with and without the addition of tannic acid. Leukemia cells were seen adhering to the adluminal aspect of the sinus endothelium and in all stages of endothelial penetration. The penetration of the sinus wall was independent of endothelial junctions; i.e., the transmural passage into the myeloid parenchyma was transcellular. At these sites, there were restricted areas of close plasmalemmal appositions of the two cell types where the intraplasmalemmal space was reduced to 2.3 nm. This space was interrupted by electron densities of 5 nm diameter and spaced 9 nm center to center. These close plasmalemmal appositions extended over distances ranging from 150 nm to 200 nm. It is suggested on the basis of the structural similarity that these heptalaminar complexes of close plasmalemmal apposition represent the structural equivalent of gap junctions and may be sites of intercellular communication requisite for transmural passage. When tannic acid was added to the fixative, there were extended areas of apparent fusion of the plasmalemmas of the two cell types, at the sites both of adhesion and of endothelial penetration. This fusion was limited to the outer leaflets of the two plasmalemmas, resulting in a single pentalaminar complex. These pentalaminar complexes extended over decidedly longer distances than the presumed gap junctions seen in the nontannic-acid-fixed material. The tannic acid material did not show the heptalaminar gap junction type of plasmalemmal apposition. It is believed likely that the tannic-acid-induced pentalaminar complexes may incorporate the smaller heptalaminar ones. The factors underlying the plasmalemmal configurational differences between the tannic acid and non-tannic-acid material remain undetermined.     

Previously unreported "hemidesmosomal junctions" between folliculo-stellate cells and pituitary adenoma cells

Thirty-eight non-functioning pituitary adenomas were ultrastructurally investigated with particular attention to the Folliculo-Stellate (FS) cells. A large number of FS cells were found in four cases, one of which disclosed a new type of intercellular junction between FS cells and surrounding adenoma cells. These junctions were characterized by 1) the presence of plasmalemmal attachment plaques only in FS cells, 2) the cytoplasmic filaments assembling in parallel to the attachment plaques, 3) the parallel plasma membranes being separated by the intercellular amorphous material and 4) the intercellular space of approximately 25 nm width. They were similar to hemidesmosomes, but were quite different from hemidesmosome-like intercellular specializations which have been described in the normal meninges and human meningiomas. Accordingly, we designated these new junctions as "hemidesmosomal junctions" which appeared to be one of the ultrastructural features characterizing FS cells.     

The association of microtubules with the plasmalemma in epidermal tendon cells of the river crab

The mode of association of microtubules (MTs) with the plasmalemma in epidermal tendon cells of the river crab, Potamon dehaani was studied by thin-section electron microscopy. In the leg muscle, the tendon cells connect striated muscle cells with the cuticle, forming specialized junctions at both ends. At the muscle-tendon cell junction, the apposed plasmalemmas are interdigitated in a zig-zag pattern separated by a uniform space of about 50 nm, where the basal lamina is shared by two cells. At the tendon cell-cuticle junction, the plasmalemma of the tendon cell forms many conical invaginations, into which dense fibrous material extends from the cuticle. Inside the tendon cell, numerous microtubules run parallel to the direction of tension transmission and are arranged into parallel bundles of various sizes. Within such bundles, fine filamentous structures cross-link adjacent MTs. MTs span the entire length of the cell and attach at their both ends to the junctional domains of the plasmalemma. The junctional plasmalemma is characterized by formation of an electron-dense undercoat, through which MTs are connected with the plasmalemma proper. The ultrastructural features of MT association with the plasmalemma are basically the same at both junctions. At the junctions, MTs usually terminate with free ends and are linked laterally to the plasmalemmal undercoat with fine filamentous structures. These observations emphasize the role of the plasmalemmal undercoat as a device of the attachment of MTs to the plasmalemma.     

烟草类根瘤中ATPase的活性变化及分布特性

采用磷酸铅技术,对烟草类根瘤中ATPase的活性变化及分布特征进行了研究。分生细胞中没有磷酸铅颗粒,非含菌细胞的细胞质和细胞器中有少量的磷酸铅颗粒,但在年轻和成熟根瘤菌中却未见它们。相反,当非含菌细胞和根瘤菌开始衰老后,有大量的磷酸铅颗粒位于细胞的质膜和细胞壁上以及根瘤菌表面的内侧。随着它们进一步衰老,磷酸铅颗粒越来越多,广泛分布在细胞的液泡膜、质膜、细胞壁、胞间层、胞间隙及根瘤菌的表面、细胞质和拟核中。由于细胞的解体,磷酸铅颗粒明显减少,一般只位于质膜和由细胞器解体而来的膜泡状结构上。     

烟草类根瘤中ATPase的活性变化及分布特性

采用磷酸铅技术,对烟草类根瘤中ATPase的活性变化及分布特征进行了研究。分生细胞中没有磷酸铅颗粒,非含菌细胞的细胞质和细胞器中有少量的磷酸铅颗粒,但在年轻和成熟根瘤菌中却未见它们。相反,当非含菌细胞和根瘤菌开始衰老后,有大量的磷酸铅颗粒位于细胞的质膜和细胞壁上以及根瘤菌表面的内侧。随着它们进一步衰老,磷酸铅颗粒越来越多,广泛分布在细胞的液泡膜、质膜、细胞壁、胞间层、胞间隙及根瘤菌的表面、细胞质和拟核中。由于细胞的解体,磷酸铅颗粒明显减少,一般只位于质膜和由细胞器解体而来的膜泡状结构上。     

The fine structure of the gill epithelium of a fresh-water flea,Daphnia magna (Crustacea: Phyllopoda) and changes associated with acclimation to various salinities

Two kinds of epithelial cells, dark and light types, are alternately arranged in the gill of Daphnia magna. The dark cell has numerous mitochondria and an elaborate tubular system containing two kinds of cytoplasmic tubules, small about 70 nm in diameter, and large about 130 nm in diameter. The former occur in bundles and seem to be smooth-surfaced endoplasmic reticulum. The latter, lined with a ridged surface coat and frequently open at the lateral and basal cell membrane, are regarded as extensions of the cell membrane. The atypical cell membrane of the dark cell is modified by repeated subunits of a cytoplasmic coat on the inner leaflet of the unit membrane. The light cell exhibits a high degree of basal infoldings of the cell membrane, which represent a magnification of the surface area of the cell. Large mitochondria between the infoldings often come into intimate association with the infolded cell membrane to form a regular array of parallel mitochondria interposed with the double cell membranes. The results suggest that at least the dark epithelial cells play an important role in the osmoregulation of this animal.     

Stress fibers in the splenic sinus endothelium in situ: molecular structure, relationship to the extracellular matrix, and contractility

In the present study, we investigated structural and functional aspects of stress fibers in a cell type in situ, i.e., the sinus endothelium of the human spleen. In this cell type, stress fibers extend underneath the basal plasma membrane and are arranged parallel to the cellular long axis. Ultrastructurally, the stress fibers were found to be composed of thin actin-like filaments (5-8 nm) and thick myosin-like filaments (10-15 nm X 300 nm). Actin filaments displayed changes in polarity (determined by S-1-myosin subfragment decoration), which may allow a sliding filament mechanism. At their plasmalemmal attachment sites, actin filaments exhibited uniform polarity with the S-1-arrowhead complexes pointing away from the plasma membrane. Fluorescence microscopy showed that the stress fibers have a high affinity for phalloidin and antibodies to actin, myosin, tropomyosin, and alpha-actinin. Vinculin was confined to the cytoplasmic aspect of the plasmalemmal termination sites of stress fibers, while laminin, fibronectin, and collagens were located at the extracellular aspect of these stress fiber-membrane associations. Western blot analysis revealed polypeptide bands that contained actin, myosin, and alpha-actinin to be major components of isolated cells. Exposure of permeabilized cells to MgATP results in prominent changes in cellular shape caused by stress fiber contraction. It is concluded that the stress fibers in situ anchored to cell-to-extracellular matrix contacts can create tension that might allow the endothelium to resist the fluid shear forces of blood flow.     

Unusual mitochondrial ultrastructure in the pig adrenal cortex

Unusually large mitochondria with few cristae were observed in the cells of the boundary layer between the zonae fasciculata and reticularis of the pig adrenal. These mitochondria occasionally contained parallel arrays of beaded filaments which appeared to be composed of repetitive electron opaque particles, measuring 10 to 11 nm in diameter. The possibility that these filaments are arranged in closely packed arrays of tubular structures with a central filament is discussed.     

Endothelial plasmalemmal vesicles have a characteristic striped bipolar surface structure

Capillary endothelial cells have a large population of small (65-80 nm diameter in transmission electron microscopy) vesicles of which a large fraction is associated with the plasmalemma of the luminal and abluminal side. We studied the fine structure and distribution of these plasmalemmal vesicles by high resolution scanning electron microscopy in cultured endothelial cells obtained from bovine adrenal cortical capillaries. Cell monolayers were covered with polylysine-coated silicon chips, split in high potassium buffer, fixed in aldehyde mixtures, and then treated with OsO4 and thiocarbohydrazide. After critical point drying, the specimens were coated with a thin (less than 2 nm) continuous film of chromium. On the cytoplasmic aspect of the dorsal plasmalemmal fragments seen in such specimens, plasmalemmal vesicles appear as uniform vesicular protrusions approximately 70-90 nm in diameter, preferentially concentrated in distinct large fields in which they occur primarily as single units. Individual plasmalemmal vesicles exhibit a striped surface fine structure which consists of ridges approximately 10 nm in diameter, separated by furrows and oriented as meridians, often ending at two poles on opposite sides of the vesicles in a plane parallel to the plasmalemma. This striped surface structure is clearly distinct from the cage structure of coated pits found, at low surface density, on the same specimens. The cytoplasmic aspect of the plasmalemma proper is covered by a fibrillar infrastructure which does not extend over plasmalemmal vesicles but on which the latter appear to be anchored by fine filaments.     

Crystalline fibrils and complexes of membranes in the parietal layer in sieve elements

Summary Fibrils are described which were found in the parietal layer in well differentiated sieve elements under the electron microscope. The fibrils were usually about 0.1 micrometers wide and appeared, in longitudinal section, to be composed of closely parallel filaments. In transverse section they appeared as a lattice, possibly of closely packed tubules. Some of the fibrils were joined to complexes of membranes in the parietal layer and to plasmatic filaments. Their relationship to these structures and to fibrils which may cause movements in other kinds of cells is discussed.     

Substructure of sidearms on squid axoplasmic vesicles and microtubules visualized by negative contrast electron microscopy

We present a high-resolution electron microscopic study of the sidearms on microtubules and vesicles that are suggested to form the crossbridges which produce the microtubule-based vesicle transport in squid axoplasm. The sidearms were found attached to the surfaces of the anterogradely transported vesicles in the presence of ATP. These sidearms were made of one to three filaments of uniform diameter. Each filament measured 5-6 nm in width and 30-35 nm in length. The filaments in some of the sidearms had splayed apart by pivoting at their base, thereby assuming a "V" shape. The spread configuration illustrated the independence of the individual filaments. The filaments in other sidearms were closely spaced and oriented parallel to each other, a pattern called the compact configuration. In axoplasmic buffer containing AMP-PNP, structures indistinguishable from the filaments of the sidearms on the vesicles were observed attached to microtubules. Pairs of filaments, thought to represent the basic functional unit, were observed attached to adjacent protofilaments of the microtubules by their distal tips. These data support a model of vesicle movement in which a pair of filaments within a sidearm forms two crossbridges and moves a vesicle by "walking" along the protofilaments of the microtubule.     

F-actin distribution during the cellularization of the Drosophila embryo visualized with FL-phalloidin

The changing distribution of polymerized actin during the cellularization of the Drosophila blastoderm was investigated in fixed whole embryos using FL-phalloidin as a specific stain. Prior incubation of FL-phalloidin with F-actin from both rabbit and locust muscle blocked the staining action, whereas G-actin at the same concentration had no effect. At the initiation of cellularization bands of F-actin filaments, shaped into rough hexagons, were found around each forming cell close to the surface bulges. These bands interlinked across the whole embryo. Above the level of the hexagons was a fine meshwork of F-actin associated with many folds of the plasmalemma. Below the hexagons was a layer of small irregular actin aggregates. During the process of cellularization the hexagonal actin network was associated with the tips of the extending plasmalemmas until the cells reached their full length. It is suggested that this actin network acts as a contractile ring system which cleaves the embryo into cells. The network was then found to rapidly break down. Microfilament bundles formed rings associated with the bases of the cells. These are presumed to cleave off the fully formed cells from the underlying yolk sac. During the first phase of cell membrane growth the fine F-actin meshwork remained associated with the apical plasmalemmas. However, the mesh rapidly disappeared during the second period of extension. After this, actin aggregates were visible close to the apical surfaces of the cells. F-actin was also observed to be associated with the newly formed plasmalemmas along their length during the whole of the process of cleavage.     

Ultrastructural observations of astrocyte end-feet in the rat central nervous system

Astrocytic end-feet in the rat CNS were studied by thin-section electron microscopy. Astrocyte processes that enclose neuronal elements extended to blood vessels and the pia mater, where the processes expanded to form end-feet or glial limiting membranes. At the end-feet, cell junctions such as gap junctions and desmosome-like junctions were formed between the astrocyte processes. The end-foot plasma membrane facing the basal lamina was undercoated with electron-dense, layered materials, with an internal substructure of filamentous networks, with which bundles of glial filaments (GFs) appeared to be closely associated via fine filamentous structures, often showing a hemi-desmosome-like appearance. In specimens treated with Triton X-100, the internal substructure of the undercoat was better visualized and the association with GFs was well preserved. At the end-feet, some unique tubular structures were found in spatial relationship to the plasmalemmal undercoat. Plectin visualized by immunofluorescence was localized to astrocytes and their processes, especially at the end-feet facing the pia mater. Immunoelectron microscopy located plectin on fine filamentous structures lying between GFs and the plasmalemmal undercoat. These observations suggest that plasmalemmal undercoats at the astrocyte end-feet may serve as attachment sites of GFs to the plasma membrane and that plectin may be involved in such attachment.     

The renal chloride cell of the fresh-water catfish,Parasilurus asotus,with special reference to the tubular membrane system

The kidney of the fresh-water catfish, Parasilurus asotus, was examined by electron microscopy. A special type of cell, very similar in appearance to the chloride cell of the teleostean gill filaments, is found in the kidney and is referred to as a renal chloride cell. This cell is characterized by an extensive tubular membrane system with a rather constant diameter of approximately 600 A. A number of mitochondria are closely associated with this system. Application of ruthenium red as an extra-cellular space marker revealed that the tubular system is a highly organized derivative of the cell membrane, infolded from the basal and lateral surfaces of the cell. The fine structural resemblance to other types of cells known to possess active transport of electrolytes suggests that these cells are involved in intrarenal osmoregulation.     

Evolution of metaplastic squamous cells of alveolar walls in pulmonary fibrosis produced by paraquat

Sequential histologic, ultrastructural, immunohistochemical and morphometric studies were made of the evolutional changes of metaplastic and regenerating alveolar epithelial cells in monkeys from 3 days to 8 weeks after paraquat administration. In the early proliferative phase, many alveoli were lined by single-layered and stratified squamous epithelium and bronchiolized epithelium (i.e., presumably derived from bronchi and bronchioles). The regenerating epithelial cells had well developed bundles of actin-like filaments, which were arranged parallel to the basal surfaces of the cells and were associated with zonulae adherentes; these cells also had intermediate filaments and some desmosomes, but lacked basement membranes, hemidesmosomes and anchoring fibrils. They covered either denuded, wavy and disrupted original epithelial basement membranes or areas of developing intraalveolar fibrosis. In zones of squamous epithelial cell metaplasia associated with intraalveolar fibrosis, fibronexus-like structures appeared to be responsible for the initial adhesion of the cells to the underlying connective tissue. In later phases, single-layered and stratified squamous epithelial cells disappeared, and only bronchiolized epithelial cells, with hemidesmosomes and anchoring fibrils on their basal surfaces, were found in fibrotic alveoli. Although bronchiolized and squamous metaplastic epithelial cells are generally thought to be formed as late events in pulmonary damage, such cells play an important role in early, temporary repair of damaged alveoli.     

Connections of intermediate filaments with the nuclear lamina and the cell periphery

We investigated the relationship between intermediate filaments (IFs) and other detergent- and nuclease-resistant filamentous structures of cultured liver epithelial cells (T51B cell line) using whole mount unembedded preparations which were sequentially extracted with Triton X-100 and nucleases. Immunogold labelling and stereoscopic observation facilitated the examination of each filamentous structure and their three-dimensional relationships to each other. After solubilizing phospholipid, nucleic acid and soluble cellular protein, the resulting cytoskeleton preparation consisted of a network of cytokeratin and vimentin IFs linked by 3 nm filaments. The IFs were anchored to and determined the position of the nuclear lamina filaments (NLF) network and the centrioles. The NLF was composed of the nuclear lamina filaments measuring 3-6 nm in diameter which radiated from and anchored to the skeleton of the nuclear pores. The IFs located in the nuclear region appeared to be interwoven with the NLF. At the cell surface, the IFs seemed to be attached to the putative actin filament network. They formed a focally interrupted plexus-like structure at the cell periphery. Fragments of vimentin filaments were found among the filamentous network located at the cell surface, and some filaments terminated blindly there.     

Morphological Evidence for the Existence of Two Distinct Types of Mitochondria Rich Cells in the Gill of the Crayfish Astacus leptodactylusEschscholtz

An ultrastructural study was performed on gill epithelia of the crayfish Astacus leptodactylus . The filament and the lamina of the podobranch were examined. In both structures a vascular system is lined by a specialized epithelium covered by a cuticle. Different types of epithelium were described in the gill filaments: either a thin epithelium with few organelles and probably involved in gas exchange or, in other filaments, a thick epithelium with characteristics of a transporting epithelium (abundant mitochondria in close association with plasma membrane infoldings). The epithelium of the lamina is also of a transporting type but in this case the complex of membrane infoldings and mitochondria is in an opposite orientation to that of the filament epithelium suggesting a different role in osmoregulation. © 1997 Published by Elsevier Science Ltd on behalf of The Royal Swedish Academy of Sciences.     

Coil-1 of rod domain of NF-L is essential for its assembly in vivo

Neurofilaments take highly ordered structures composed of parallel mays of 10 nm filaments linked to each other with frequent cross-bridge. It is composed of three components named NF-L, NF-M and NF-H. NF-L is able to form filamentous network alone in Sf9 cells, while M could not. To identify which domain is essential for the assembly of NF-L, two chimera proteins named ML and MML were constructed: ML was composed of the head domain of NF-M and other domains of NF-L; MML was composed of the head and Coil-1 domains of NF-M and Coil-2 and tail domains of NF-L. ML was not only able to form filaments in Sf9 cells, but also co-assemble with NF-M into parallel filamentous bundles. MML could not assemble into filaments. Thus the Coil-1 domain of NF-L was essential for its assembly. Project supported by the National Grant for Outstanding Young Scientists.     

Coil-1 of rod domain of NF-L is essential for its assembly in vivo

Neurofilaments take highly ordered structures composed of parallel arrays of 10 nm filaments linked to each other with frequent cross-bridge. It is composed of three components named NF-L, NF-M and NF-H. NF-L is able to form filamentous network alone in Sf9 cells, while M could not. To identify which domain is essential for the assembly of NF-L, two chimera proteins named ML and MML were constructed: ML was composed of the head domain of NF-M and other domains of NF-L; MML was composed of the head and Coil-1 domains of NF-M and Coil-2 and tail domains of NFL. ML was not only able to form filaments in Sf9 cells, but also co-assemble with NF-M into parallel filamentous bundies. MML could not assemble into filaments. Thus the Coil-1 domain of NF-L was essential for its assembly.     

Ultrastructural neuronal pathology in transgenic mice expressing mutant (P301L) human tau

Transgenic mice expressing mutant (P301L) human tau develop neurofibrillary tangles, amyotrophy and progressive motor disturbance. We present ultrastructural features of neuronal degeneration in this model that suggests involvement of both neurofibrillary and autophagic processes in neurodegeneration. Neurons undergoing neurofibrillary degeneration contain tau-immunoreactive, 15–20 nm-wide straight or wavy filaments with no periodic twists. Tau filaments were found in two types of affected neurons. One type resembled neurons with neurofibrillary tangles (NFT) that were filled with numerous filaments that displaced sparse cytoplasmic organelles to the periphery. Microtubules were almost completely absent. The nucleus remained centrally located, but showed lobulations due to deep infoldings. The other type resembled ballooned neurons seen in some human tauopathies. The nucleus was peripherally placed, but normal appearing. The cytoplasmic organelles were dispersed throughout the swollen perikarya, the Golgi complex was fragmented and duplicated, while mitochondria and other organelles appeared normal. Tau filaments similar to those in NFT were sparse and not tightly packed. Microtubules were also sparse. Many autophagic vacuoles were present in these cells. Heterogeneous appearing axonal swellings resembling spheroids in human tauopathies were present in gray and white matter. Unlike normal appearing axons, axonal spheroids were filled with tau-immunoreactive filaments and autophagic vacuoles, in addition to normal appearing neurofilaments and microtubules. These P301L transgenic mice exhibit many features common to human tauopathies, making them a valuable model to study the pathogenesis of these uncommon disorders.