High-order structure of metaphase chromosomes: evidence for a multiple coiling model

When chromosome preparations made by the conventional air-drying method were processed with the OsO₄/TCH technique and examined by scanning electron microscopy (SEM), spiral structures in chromatids, which have been frequently observed to be present by light microscopy, were found to be composed of 30 nm fibres. In some portions these fibres appeared to be arranged in coils to form thicker fibres. When chromosome preparations were processed for SEM without air drying, chromosomes appeared to consist of fairly homogeneous thick fibrous structures measuring about 200 nm in diameter. In relatively condensed chromosomes, these 200 nm fibres appeared to be arranged perpendicular to the long axis of the chromatid. These findings suggest that chromatid spiral structures represent a regularly loosened state of the compactly spiralized 200 nm fibres which in turn consist of spiralized 30 nm fibres.     

The effect of xylanase on lignocellulosic components during the bleaching of wood pulps

HPLC, SEM and XRD techniques have been proposed as methods for ascertaining the changes occurring in polysaccharides (cellulose and xylans) and fibres during the xylanase bleaching processes. TCF and ECF bleached pulps with and without enzyme pretreatment were analysed. The ratio of carbohydrates present in the pulp, observation of changes occurring in the surface of the fibres and the crystallinity and accessibility of the bleached fibres were determinated. These characteristics have been related with pulp properties. Xylan content decreased when pulp was bleached. Xylanase treatment substantially reduced the xylose content present in pulp, measured by HPLC after the hydrolysis method of the sample. Morphological changes in the fibres occurred when the enzymatic treatment was applied. Bleaching increased the crystallinity of the pulp and enzyme pretreatment also affected the crystallinity of cellulose fibres     

Is mucus involved in biocrystallization?

Summary Freeze-dried intestinal mucus of sea-water-adapted eels was analysed by scanning electron microscopy (SEM) and X-ray microanalysis. Calcite crystals were observed in the mucus fibres; their concentration increased along the hindgut. Random SEM observations made in situ indicated that mucus fibres were involved in the genesis of these crystals. Calcium-rich mucus globules were found fused inside crystal matrices. Single typical rhombohedric crystals of various complexity appeared within the mucus framework. The steps of crystal biogenesis were reconstituted in in-vitro conditions.     

Hypoxia-induced fibre type transformation in rat hindlimb muscles Histochemical and electro-mechanical changes

Twelve male Sprague-Dawley rats (21 days old) were randomly assigned into two experimental groups: sea level control (CONT) and hypobaric hypoxia (HYPO). The HYPO rats were kept in an hypobaric chamber maintaining a simulated altitude of 4000 m (61.1 kPa). After 10 weeks of treatment, the rat hindlimb muscles [soleus (SOL) and extensor digitorum longus (EDL)] were subjected to histochemical and electro-mechanical analyses. Results indicated that compared to CONT the HYPO SOL muscle had a significantly greater relative distribution of fast-twitch-oxidative-glycolytic (FOG) fibres (28.9% SEM 2.0 vs 18.3% SEM 1.8, P less than 0.01) with a significant decrease in slow twitch oxidative fibre distribution (69.5% SEM 2.4 vs 82.9% SEM 3.1, P less than 0.01). Compared to CONT the HYPO EDL muscle also manifested a significant increase in FOG fibre distribution (51.6% SEM 0.8 vs 46.6% SEM 1.1, P less than 0.01), but this was accompanied by a significant decrease in fast twitch glucolytic fibres (44.3% SEM 0.9 vs 49.2% SEM 1.7, P less than 0.05). These histochemical fibre type transformations accompanied significant and expected changes in the electro-mechanical parameters tested in situ, e.g. maximal twitch force, maximal rate of force development, contraction time, half relaxation time, force: frequency curve, and fatigability. It was concluded that chronic hypobaric hypoxia could have a potent influence upon the phenotype expression of muscle fibres.     

The ependymal surface of the fourth ventricle of the rat: a combined scanning and transmission electron microscopic study.

The morphological features of the ependymal surface and supraependymal elements of the fourth ventricle of the rat were examined by scanning electron microscopy (SEM) and by the transmission electron microscopy (TEM). The results confirm the following aspects: 1) The presence of supraependymal elements and microvilli in the ependymal territories, including the sites where the cilia completely cover the ependymal surface; 2) The existence of cilia with oval or spherical thickenings together with supraependymal bulbs similar in size to those of the larger ciliary swellings; 3) Identification of the long supraependymal fibres with intermittent fusiform dilations observed under the SEM with the nerve fibres seen under the TEM; 4) The existence of intraventricular axodendritic synapses.     

Characterization of chromosome replication during S-phase with bromodeoxyuridine labelling in Chinese hamster ovary and HeLa cells

The same C-banded human polymorphic chromosomes were observed in the light microscope (LM) and then in the scanning electron microscope (SEM) to investigate the structural changes produced by the C-banding technique. C-banded regions, which stained positively in LM, were highly condensed with tightly packed chromatin fibres, resembling non-banded chromosomes. In striking contrast, adjacent non-C-banded regions were represented by loosely arranged fibres, resembling G-banded chromosomes. The significance of these observations in relation to current theories on the effects of C-banding on chromosome structure is discussed.     

Scanning electron microscopy of isolated peripheral nerve fibres

Summary The surface morphology of normal myelinated nerve fibres prepared in different ways for scanning electron microscopy has been studied and compared with the surface features of similar fibres undergoing retrograde changes. Nodes of Ranvier, paranodal specializations, artefactual fractures of the myelin, and the endoneurial collagen sheaths are described. A regular pattern of elevations, usually with a pitted or depressed surface seen on normal myelinated fibres after certain preparative procedures are thought to be artefacts produced during preparation and to be related to the neurokeratin network.Alterations in the surface structure of fibres central to long-standing nerve transections include irregular protuberances, serial surface corrugations and large swellings, all associated with demyelination. Fibres that have undergone retrograde degeneration consist of endoneurial tubes with focal swellings occupied by macrophages or myelin debris, together with fine unmyelinated and small myelinated regenerating axons. Strict centrifugal progression of myelination of regenerating axons was not observed.We thank Mr. R. A. Willis for his collaboration and for taking the SEM photographs of normal nerve fibres, and the Cambridge Scientific Instrument Co. Ltd. for permission to reproduce the SEM photographs of experimental nerve fibres. We also thank Dr. A. Boyde for access to his SEM and for helpful comments on interpretation of the scanning electron micrographs, Prof. J. Z. Young, Dr. P. K. Thomas, and Dr. R. H. M. King for discussion, and Messrs. P. Reynolds and D. Gunn for photography.A grant from the Muscular Dystrophy Group of Great Britain is gratefully acknowledged.     

Identification and Pattern of Primary Sensory Cells in the Body Wall Epithelium of the Tubificid Worm, Limnodrilus hoffmeisteri

This study compares the effectiveness of two examination methods suitable for morphological observations, scanning electron microscopy (SEM) and light microscopic NADH-diaphorase (NADP-d) histochemistry, to identify location and distribution pattern of primary sensory cells situated in the body wall epithelium of Limnodrilus hofmeisteri. SEM observations revealed that grouped sensory cells forming two distinct types of sensillas (or sense organs) are scattered over the body surface. The easily identifiable first type (ciliated sensilla) contains numerous penetrative ciliate sensory cells while no ciliate sensory cells were seen in the second type (sensory bud). NADH-d staining proved to be a suitable method to identify both structures further staining solitary sensory cells, and certain sensory fibres that could not be visualised by SEM preparations. Our results show that NADH-d staining, as a histochemical marker of primary sensory cells, is a suitable and effective method to reveal their morphology and distribution pattern in whole mount preparations in the smallest detail, suggesting that the application of this relatively simple and cheap method may contribute to understand the basic organisation and evolution of the peripheral sensory structures of oligochaetes.     

Biocatalytic surface modification of knitted fabrics made of poly (ethylene terephthalate) with hydrolytic enzymes from Thermobifida fusca KW3b

Knitted fabrics made from poly(ethylene terephthalate) fibres were treated with an enzyme preparation from Thermobifida fusca KW3b showing hydrolytic activity on p-nitrophenyl butyrate. The fabrics were also treated with NaOH and the results were compared. Both enzyme- and alkaline-treated fabrics showed an increase in reactive dye uptake, vertical wicking height and water absorption capacity of the fabrics indicating an increased surface hydrophilicity. However, X-ray photoelectron (XPS) and energy-dispersive spectroscopy (EDS) did not give conclusive results on the presence of newly introduced hydrophilic groups on the surface of the fibres. Analysis of the enzyme-treated poly(ethylene terephthalate) fibres by scanning electron microscopy (SEM) and by atomic force microscopy (AFM) indicated an increase in surface roughness of the fibres, which may contribute to the observed increased hydrophilicity of the PET fabrics. However, much longer treatment times (24 h) were required to obtain these effects with the enzymes compared to the chemical treatment (1 h).     

Effects of bromodeoxyuridine substitution on metaphase chromosome structures examined by scanning electron microscopy

Chinese hamster chromosomes were differentially substituted with 50 M 5-bromodeoxyuridine (BrdU) to obtain chromosomes with bifilarly and unifilarly substituted (BB-TB) and unifilarly and non-substituted (TB-TT) chromatid constitutions. To avoid the effect of Giemsa staining on the ultrastructure of chromosomes, unstained preparations were exclusively used. When TB-TT chromosomes were prepared with the conventional air-drying method followed by the osmium tetroxide-thiocarbohydrazide (OsO₄-TCH) technique and examined by scanning electron microscopy (SEM), the TB-chromatid appeared somewhat more slender and showed more conspicuous spiral structures, thereby appearing more loosened compared to the TT-chromatid. At higher magnifications, however, 30 nm chromatin fibres which were seen to constitute both chromatids showed no discernible differences in dimension between the TT- and TB-chromatids. On the other hand, TB-TT chromosomes specially prepared for SEM without the process of air-drying appeared in their entirety less extended and no spiral configuration was observed even in the TB-chromatid. The TB-chromatid instead appeared rather less loosened than the TT-chromatid whereas thick fibre-like structures which in turn seemed to consist of 30 nm fibres were more easily discernible in the TT-chromatid compared to the TB. Such seemingly contradictory results obtained from the two different preparatory procedures were tentatively explained on the basis of our multiple coiling model (Taniguchi and Takayama 1986).     

Quantitative assessment of degenerative changes in soleus muscle after hindlimb suspension and recovery

The aim of this study was to quantify the degenerative and regenerative changes in rat soleus muscle resulting from 3-week hindlimb suspension at 45° tilt (HS group, n = 8) and 4-week normal cage recovery (HS-R group, n = 7). Degenerative changes were quantified by microscope examination of muscle cross sections, and the myosin heavy chain (MHC) composition of soleus muscles was studied by sodium dodecyl sulphate polyacrylamide gel electrophoresis. At the end of 3-week hindlimb suspension, histological signs of muscle degenerative changes were detected in soleus muscles. There was a significant variability in the percentage of fibres referred to as degenerating (%dg) in individual animals in the HS group [%dg = 8.41 (SEM 0.5)%, range 4.66%–14.08%]. Moreover, %dg varied significantly along the length of the soleus muscle. The percentage of fibres with internal nuclei was less than %dg in HS-soleus muscles [4.12 (SEM 0.3)%, range 1.24%–8.86%]. In 4-week recovery rats, the greater part of the fibres that were not referred to as normal, retained central nuclei [15.8 (SEM 2.2)%, range 6.2%–21.1%]. A significant increase in the slow isoform of MHC was recorded in the HS-R rats, compared to muscles from age-matched rats (P < 0.01). These results would suggest that a cycle of myofibre degeneration-regeneration occurred during HS and passive recovery, and that the increased accumulation of slow MHC observed in soleus muscles after recovery from HS could be related to the prevalence of newly formed fibres. Accepted: 14 October 1996     

The extracellular matrix of the cuticle of Gordius panigettensis (Gordioiidae,Nematomorpha): observations by TEM,SEM and AFM

The cuticle of Gordius panigettensis (Sciacchitano, 1955) was studied by scanning electron microscopy (SEM), transmission electron microscopy (TEM) and atomic force microscopy (AFM). The cuticle is composed of 30-50 compact layers. The number of the layers is higher in the central part of the animal's body and decreases at the extremities. Each layer is composed of parallel tightly packed fibres approximately 640 nm in diameter and of indefinite length. The fibres run strictly parallel within each layer, while in adjoining layers they run at a variable angle from 45 degrees in the central body to 90 degrees in the extremities. Each fibre shows a barely detectable filamentous inner structure and is enveloped in a thin highly regular net formed by hexagonal meshes. Our results suggested that these fibres should be proteinaceous although non-collagenous. Thinner radial fibres run among the large fibres and across all the layers and span the whole thickness of the cuticle from the epithelial layer located deep underneath the large fibres up to the epicuticle on the external surface of the animal.     

The influence of enzymatic treatment on wool fibre properties using PEG-modified proteases

The main contribution of the presented work was to introduce the use of proteases modified with the soluble polymer polyethylene glycol (PEG) in the bio-finishing process of wool fibres, to target enzyme action to the outer parts of wool fibres, i.e. to avoid the diffusion and consequent destroying of the inner parts of the wool fibre structure, in the case of native proteases using.

Different proteolytic enzymes from Bacillus lentus and Bacillus subtilis in native and PEG-modified forms were investigated and their influence on the modification of wool fibres morphology surface, chemical structure, as well as the hydrolysis of wool proteins, the physico-mechanical properties, and the sorption properties of 1:2 metal complex dye during dyeing were studied. SEM images of wool fibres confirmed smoother and cleaner fibre surfaces without fibre damages using PEG-modified proteases. Modified enzyme products have a benefit effect on the wool fibres felting behaviours (14%) in the case when PEG-modified B. lentus is used, without markedly fibre damage expressed by tensile strength and weight loss of the fibre. Meanwhile the dye exhaustion showed slower but comparable level of dye uptake at the end of the dyeing.     

Application of calcium alginate immobilized and crude pectin lyase from Bacillus cereus in degumming of plant fibres

Abstract

In this study, the purified pectin lyase was immobilized in calcium alginate beads and compared with crude enzyme for application in degumming of buel and banana plant fibres. From the data of scanning electron microscopy (SEM), it was observed that untreated buel fibres were covered by non-cellulosic materials (pectin, hemicelluloses and waxes) and the surface of enzymatically treated buel fibres looked smoother. Also, the crude alkaline pectin lyase treated buel fibre exhibited a considerably cleaner surface, which suggested that the crude pectin lyase could provide better degumming effects in comparison to the immobilized pectin lyase. In case of banana fibre, the FTIR spectroscopy showed that both crude and immobilized alkaline pectin lyase treatments of banana plant fibres were equally efficient in degumming. The enzymatic degumming of buel and banana with crude pectin lyase resulted in maximum release of galacturonide after 24?h for buel and 15?h for banana fibre. The optimum temperature for degumming of buel and banana fibres with crude pectin lyase was found to be 50?°C and 45?°C, respectively. Also, the maximum galacturonide was released with 200 and 250?U of pectin lyase for buel and banana fibre, respectively.     

Histochemistry and ultrastructure of nerve fibres and contractile cells in the tunica albuginea of the rat testis

Whole-mounted preparations of the tunica albuginea of the rat testis were studied using light microscopy techniques for demonstration of cholinergic nerve fibres (Karnovski-Root method), catecholaminergic nerve fibres (De la Torre's method) and actin filaments (avidin-biotin-peroxidase method). An ultrastructural study of different regions of the albuginea was also performed. Cholinergic fibres were seen to penetrate into the albuginea with the testicular artery to form a broad network in the mediastinum testis, many fibres ending beneath the rete testis epithelium. Catecholaminergic fibres penetrated through the middle part of the cauda epididymis and formed a plexus in the albuginea covering the inferior testicular pole. This plexus gave rise to straight fibres which formed varicosities, some of them appeared related with mast cells. Actin-containing cells were only found beneath the rete testis epithelium. These cells were similar to myofibroblasts. The location of both cholinergic fibres and contractile cells among the rete testis channels suggests that these cells may be involved in the pumping of semen towards the ductuli efferentes and that their contractility may be regulated by cholinergic fibres.     

The lattice arrangement of the collagen fibres in the submucosa of the rat small intestine: scanning electron microscopy

Summary The three-dimensional architecture of the submucosal collagen fibres of the rat (3 weeks old) small intestine was examined by scanning electron microscopy using a selective microdissection method. The main framework of the submucosa was composed of two arrays of collagen fibre bundles running diagonally around the intestinal wall, one set in a clockwise direction, the other counterclockwise. These fibre bundles were about 5 m in diameter and were oriented at a range of angle ± 30°–50° to the longitudinal axis of the intestine. With the advantage of the SEM observation it was demonstrated that these fibres in different arrays did not constitute two separate layers but interwove to form a unified lattice sheet. An irregular network of fine collagen fibrils over the main framework was also seen. The significance of their arrangement is discussed with respect to the skeletal function of the submucosa in the intestine.     

Structure,composition and mechanical properties of the silk fibres of the egg case of the Joro spider, <Emphasis Type="Italic">Nephila clavata</Emphasis> (Araneae,Nephilidae)

The silk egg case and orb web of spiders are elaborate structures that are assembled from a number of components. We analysed the structure, the amino acid and fibre compositions, and the tensile properties of the silk fibres of the egg case of Nephila clavata. SEM shows that the outer and inner covers of the egg case consist of thick, medium and thin silk fibres. The silk fibres of the outer cover of the egg case are probably produced by the major and minor ampullate glands. The silk fibres of the inner cover of the egg case from cylindrical glands appears to be distinct from the silk fibres of the major ampullate glands based on their micro-morphology, mole percent amino acid composition and types, and tensile behaviour and properties. Collectively, our investigations show that N. clavata uses silk fibres from relatively few glands in varying combinations to achieve different physical and chemical properties (e.g., color, diameter, morphology and amino acid composition) and functional and mechanical properties in the different layers of the egg case.     

Scanning electron microscopic observations of the ependymal cell and the subependymal layer of the third brain ventricle in the rat

This investigation was undertaken to clarify the three dimensional ultrastructure of the subependymal layer in relation with the ependymal cell layer in rat brain using the scanning electron microscope (SEM). The subependymal layer existing below the ependyma of the third ventricle in the brain of mature albino rats was examined with S E M. The hypothalamus freshly excised after median sagittal section was treated by collagenase with or without trypsin for a short while to remove the ependymal cells at the ventricular wall. After the enzymatic pretreatment of the specimen, many ependymal cells were removed and the subependymal layer was partially exposed. Most of the ciliated ependymal cells remaining at the ventricular wall extended long, single basal processes which then penetrated into the subependymal layer. The subependymal layer was composed of a delicate framework of thin processes of glial cells, ependymal cells and, in addition nerve cells. Scattered among the neuropil just beneath the ependymal cell layer, there were relatively small, globular subependymal cells. Occasionally, there were large bundles of unmyelinated nerve fibres in the subependymal layer. The individual nerve fibres distinctly showed many axonal varicosities within the fibres. Intermingled with the nerve fibres, glial processes of various forms were present. The structure of the ependymal cells and the subependymal layer was compared with the findings already reported in the studies using light and transmission electron microscope.     

The spatial relationship between the musculature and the NADPH-diaphorase activity, 5-HT and FMRFamide immunoreactivities in redia, cercaria and adult Echinoparyphium aconiatum (Digenea)

The spatial relationship between the musculature and the NADPH-diaphorase (NADPH-d) activity, 5-HT and FMRFamide immunoreactivities in redia, cercaria and adult Echinoparyphium aconiatum was studied using scanning electron microscopy (SEM), NADPH-d histochemistry, immunocytochemistry, and confocal scanning laser microscopy (CSLM). TRITC-conjugated phalloidin was used to stain the musculature. Staining for NADPH-d was observed in the central (CNS) and peripheral nervous system (PNS) of all three stages. NADPH-d positive nerves occurred very close to muscle fibres. 5-HT-immunoreactive (5-HT-IR) nerve cells and fibres occurred in the CNS and PNS and close to muscle fibres. FMRFamide-IR nerve fibres were observed in the CNS and PNS of adult worms. This is the first time, the presence of the NADPH-d has been demonstrated in the larval as well as the adult stages of a fluke.     

Monoaminergic and cholinergic nerve fibres in the human dental pulp

Summary Distribution of cholinesterase-containing nerve fibres was studied in 15 human dental pulps by the thiocholine method. Falk's fluorescent method was used to demonstrate catecholamines (8 dental pulps).Cholinesterases were localized partly in the subodontoblastic plexus sending out fine branches towards odontoblasts, and partly in the nerve fibres attached to the blood vessel walls. These fibres in contrast to those of the subodontoblastic plexus were finer and showed fine varicosities.Monoaminergic terminals were localized mainly along blood vessel walls, however, some fibres having no relation to the blood vessels were also found.Cholinesterase-containing nerve fibres in the periphery of the pulp are considered to be sensitive nerve fibres originating from n.V. Distribution of cholinesterase-containing nerve fibres and monoaminergic terminals along the blood vessel walls indicates that the blood vessels in the human dental pulp might be under both parasympathetic and sympathetic control.