Chromatin condensation,cysteine-rich protamine,and establishment of disulphide interprotamine bonds during spermiogenesis of Eledone cirrhosa (Cephalopoda)

During spermiogenesis in Eledone cirrhosa a single protamine substitutes for histones in nuclei of developing spermatids. This protein displays a peculiar primary structure. It contains 22.6 mol% cysteine residues (19 cysteines in 84 residues). This makes it the most cysteine-rich protamine known. The proportion of basic residues is relatively low (arginine 36.9 mol%, lysine 19.0 mol%). The protamine of E. cirrhosa condenses spermiogenic chromatin in a pattern which comprises fibres with a progressively larger diameter and lamellae that finally undergo definitive coalescence. We have also performed a study that estimates the number of interprotamine disulphide bonds formed during the process of spermiogenic chromatin condensation by means of sequential disappearance of MMNA (monomaleimido-nanogold) labelling. During the first step of spermiogenesis, protamines are found spread over very slightly condensed chromatin with their cysteines in a reactive state (protamine-cys-SH). From this stage the interprotamine disulphide bonds are established in a progressive way. First they are formed inside the chromatin fibres. Subsequently, they participate in the mechanism of fibre coalescence and finally, in the last step of spermiogenesis, the remaining free reactive -SH groups of cysteine form disulphide bonds, thus promoting a definitive stabilization of the nucleoprotein complex in the ripe sperm nucleus.     

TRANSFORMATION OF THE NUCLEUS IN MARCHANTIA SPERMATIDS: MORPHOGENESIS

It is proposed that elongation of the nucleus in spermatids of Marchantia results from interaction between its membranous envelope and microtubules of the spermatid's cytoskeleton. The nucleus may be drawn out in two directions along microtubules until forces attracting the nucleus to them are balanced by forces resisting envelope distortion. Condensation of nuclear chromatin into fibrils of uniform diameter and probable shaping of the nucleus by blebbing of its envelope occur together before elongation is complete. The nucleus becomes crescent shaped and it is prolonged distally into a chromatin-free diverticulum. In accord with their distribution along the axis of the nucleus, chromatin fibrils are compacted together forming a cone-like rod of chromatin which narrows anteriorly and extends distally to the tip of the preexisting diverticulum. Elongation and shaping of the nucleus influence the distribution of its chromatin and thus its ultimate morphology. Coiling of the nucleus is related to a reduction of spermatid cytoplasm during maturation.     

The tensile properties and mode of fracture of elastoidin

The tensile properties and mode of fracture of elastoidin, a collagenous protein fibre from the fins of sharks, were compared with those of rat tail tendon fibres, considered to be a pure form of collagen. Elastoidin fibres were stronger than tendon in the dry state whereas the opposite was observed for fibres tested in the wet state. However, elastoidin was stiffer than tendon whether dry or wet. Scanning electron micrographs of the crosssections and fractured surfaces revealed that elastoidin fibres consisted of fibrils of varying diameter arranged in a lamellar fashion. From the nature of the fractured surfaces, it could be deduced that the primary failure mechanism for elastoidin was probably through a fissuring of the structure.     

草鱼染色体的包装（间期—中期）

以草鱼ＺＣ７９０１细胞株为材料,观察鱼类细胞从间期染色质到中期染色体的包装过程。主要通过（１）分裂期与间期细胞融合,诱导染色体早熟凝集;（２）染色体“伸长”处理;（３）培养细胞的低渗处理;（４）染色质辅展等方法,制作染色体标本,进行扫描和透射电镜观察。观察表明,鱼类染色质的基本结构与哺乳类细胞相同,也是直径约１０ｎｍ的核丝。染色体的色装有两种形式：一种是多级螺旋化形成直径约３００ｎｍ的染色单体,     

Muscle growth in juvenile Atlantic salmon as influenced by temperature in the egg and yolk sac stages and diet protein level

Atlantic salmon Salmo salar eggs derived from a single family were incubated at two different water temperature regimes, with a mean temperature between fertilization and first feeding differing between 6 and 10° C (HT) and 2–6° C (LT). From first feed the fry were kept under the same rearing conditions and fed either high (50%) or low (45%) protein diet level of equivalent energy content until smoltification. All treatments were carried out in duplicate tanks. At first feeding the groups were similar in mass, but thereafter the HT‐fish were heavier and longer compared to the LT‐fish throughout the experiment. The groups fed the high protein diet were significantly heavier and longer compared with the corresponding low protein diet. A strong positive relationship was observed between L_F and total white muscle cross‐sectional area (CSA), white muscle fibre diameter and fibre number. There were also equivalent relationships with body mass. There were no significant differences in CSA, the mean diameter or the number of white muscle fibres per CSA between groups at first feed. Muscle fibre number and CSA increased in all groups during the experiment, whereas fibre diameter reached a plateau when the fish reached > 9 cm L_F. There were only minor effects of pre‐hatch and yolk sac stage temperature on CSA and fibre number per CSA during the juvenile stage. In short periods the LT‐group had larger CSA and higher fibre number than the HT‐groups, but this differences had disappeared by the end of the juvenile stage. No differences in mean fibre diameter were found between groups, except at the time of smoltification. When the fish approached smoltification a decrease in mean fibre diameter and an increase in muscle fibres <25 µm was seen and taken as an indication of recruitment of new fibres (hyperplasia). Only minor differences in CSA, fibre number or fibre diameter was observed between high and low protein diet groups.     

Turbidimetric and morphological studies of type I collagen fibre self assembly in vitro and the influence of fibronectin

Collagen undergoes several stages of self assembly including turbidimetric lag, growth and plateau steps. The later stages of type I collagen self assembly were studied by turbidity—time measurements, low angle laser light scattering and by determination of the birefringence retardation of collagen fibres formed in vitro. These studies were conducted in the presence and absence of fibronectin to evaluate the effect of fibronectin on the kinetics and extent of type I collagen fibrillogenesis. The results of these studies indicate that the collagen fibres observed at the end of the lag phase appear to be identical to fibres seen in the growth phase of turbidity—time curves based on fibre diameter and birefringence retardation measurements. Birefringence retardation measurements suggest that the diffracting unit may be the collagen fibril and that the volume fraction of fibrils in fibres is about 0.95 using a model developed for a series of parallel ellipsoids. Morphological observations suggest that the distribution of fibre diameters formed in vitro during the growth phase is narrow and appears to be independent of time with only the mass of collagen in fibres increasing during the growth phase. During the growth phase, layers of parallel fibres are formed with alternating layers appearing almost orthogonal. In the presence of fibronectin the mechanism of fibre formation appeared unchanged. It was concluded that fibronectin appeared to modify the kinetics of self assembly by preventing collisions between collagen molecules.     

Open and closed domains in the mouse genome are configured as 10‐nm chromatin fibres

The mammalian genome is compacted to fit within the confines of the cell nucleus. DNA is wrapped around nucleosomes, forming the classic ‘beads‐on‐a‐string’ 10‐nm chromatin fibre. Ten‐nanometre chromatin fibres are thought to condense into 30‐nm fibres. This structural reorganization is widely assumed to correspond to transitions between active and repressed chromatin, thereby representing a chief regulatory event. Here, by combining electron spectroscopic imaging with tomography, three‐dimensional images are generated, revealing that both open and closed chromatin domains in mouse somatic cells comprise 10‐nm fibres. These findings indicate that the 30‐nm chromatin model does not reflect the true regulatory structure in vivo.     

Acetylation of histone H4 in complex structural transitions of spermiogenic chromatin

In spermiogenic nuclei of the cephalopod mollusc Sepia officinalis histones are replaced by a precursor-protamine molecule, which is later converted into protamine. Simultaneously, spermiogenic chromatin undergoes a complex structural change. Somatic-like chromatin belonging to the earliest spermatid is progressively reorganized into: (a) granules of 20 nm diameter, (b) fibres of 30-35 nm, and (c) fibres of 40-50 nm. In the final phases of spermiogenesis these fibres of 40-50 nm join to form larger structures of condensed chromatin, and lastly, the uniformly packed chromatin in the sperm nucleus. Using specific antibodies for mono- and hyperacetylated forms of histone H4, in this work we show that the first structural remodelling of chromatin (from somatic-like organization into 20 nm granules) is given concomitantly with a massive mono-acetylation of H4 (acetylation in lysine 12), whereas the structural remodelling from 30-35 to 40-50 nm fibres is produced simultaneously with hyperacetylation of H4 and the nuclear removal of histones.     

Helical Structure Revealed upon the Condensation and Decondensation of Chromosomes of Vicia faba

By using conventional and stereo electron microscopy, helical structures were revealed in prophase and telophase chromosomes of root tip cells of Vicia faba. Longitudinal and transverse sections of the chromosomes showed that both prophase and telophase chromosomes were composed of chromatin fibres about 0.5μm in diameter and among the 0.5μm chromatin fibre thinner chromatin fibres with a diameter of about 0.2μm were found. In transverse sections, prophase chromosomes appeared to be a circular structure which contained a low electron density centre encircled by the 0.5 μm fibre. In longitudinal section of the chromosomes, the 0.5 μm fibres were seen to be orientated parallel to each other while constituted roughly a right angle to the long axis of the chromosome. Helical coils consisting of the 0.5μm fibre were identified easily by stereo electron microscopy. In transverse sections of telophase chromosomes, both the circular structure similar to that of the prophase chromosomes and the hoof-shaped structure composed of the 0.5μm fibre were observed, demonstrating4 the de-spiralization of the helical coils in the decondensation of the chromosomes. Based on these observations, the radial loop. model and the multiple coiling model are discussed.     

SPERMIOGENESIS IN THE PULMONATE SNAIL, EUHADRA HICKONIS II. STRUCTURAL CHANGES OF THE NUCLEUS

In accordance with the characteristic shape of the nucleus and degree of condensation of the nuclear substance, spermiogenesis in Euhadra hickonis can be roughly divided into four stages. The chromatin in the highly polymorphic nucleus of the first stage, early spermatid, forms relatively thick (ca. 50 nm) fibrils which associate here and there into irregular clumps. In the next stage, the spermatid nucleus becomes conspicuously spherical, its contents appear more finely homogeneous and the irregular clumps of chromatin are few. In the third stage, the nucleus gradually takes on an ellipsoidal shape as the antero-posterior axis shortens. The anterior part of its envelope becomes structurally modified in preparation for the adherence to it of the developing acrosome, and an implantation fossa forms posteriorly at the center of a second area where the nuclear envelope has been modified. The diameter of the chromatin fibrils again increases and those near the implantation fossa become oriented perpendicular to the nuclear envelope.
As the nucleus elongates in the fourth stage, a concentric sheath of microtubules closely surrounds it. These appear to depolymerize as the nuclear elongation proceeds, so that they are no longer present in the head region of the mature spermatozoon. The diameter of the chromatin fibrils increases to about 10 nm and they become oriented parallel to the long axis of the cell. With the decrease in the nuclear volume the fibrils unite laterally to form longitudinal sheets, and these finally merge in the mature spermatozoon into a mass of very dense chromatin without perceptible internal structure.     

The study of chromatin and chromosome structure on preparations of interphase nucleus derivatives resulting from nuclear wall removal. IV. Structural heterogeneity of stretched chromatin in membrane-free cell nuclei from human peripheral lymphocytes

Densely aggregated chromatin of mature human or animal peripheral lymphocytes is inaccessible for structural investigation on preparations of both intact cell and conventionally spread chromatin. Giemsa- and DAPI-positive "free chromatin" structures, in addition to amembraneous nuclei, were isolated from intact lymphocytes gently treated with Triton-X-100. Surface stretching of both these nuclei and structures, shortly fixed in methanol-glacial acetic acid (3:1), revealed three main types of these "free chromatin" structures: dense chromatin structures (DCS), loose chromatin structures (LCS) and nuclear spreads (NS). The share of each nuclear derivative may be shifted by changing either detergent concentration and(or) the time of incubation in detergent solution. Each DSC consists of condensed "residual" nucleus, similar in from and size with an intact lymphocyte nucleus, and involves 1-15 uni- or olygonemic chromatin sprouts of different length. LSC contain heterogeneously loosened spindle-shape or drop-like nuclei, being several times longer and wider than DCS-nuclei, and 1-3 long uni- or olygonemic chromatin tail-pieces and incidentally observed lateral chromatin sprouts. The majority of LCS contain either a chromocenter of different number of end-to-end associated spindle-shape domains of condensed chromatin. The latter reached 2-5 x 1.5 microns being cross-striated or spiral in structure. NS represent spread chromatin fibrillar structures varying from 150 to 500 microns in length and from 1.5 to almost 50 microns in width. NS consist of 0.3-0.4 micron smooth and 0.4-0.8 micron beaded chromatin fibres. Thin fibres produce web-like domains of NS. and thick fibres form olygonemic bundles or end-to-end association of unit chromatin fibres within NS. Some portion of thick unit fibres of NS gave rise to local splitting into two thin fibres with a similar bead patterns. Thick argyrophilic fibers of the nucleolus also displayed a beaded structure and commonly spread hand-in-hand with the basic chromatin fibre aggregations.     

STRUCTURAL BIOLOGY OF THE FIBRES OF THE COLLAGENOUS AND ELASTIC SYSTEMS

The different types of fibres of the collagenous and elastic systems can be demonstrated specifically in tissue sections by comparing the typical ultrastructural picture of each of the fibre types with studies using selective staining techniques for light microscopy. A practicalmodus operandi, which includes the recommended staining procedures and interpretation of the results, is presented. Micrographs and tables are provided to summarize the differential procedures. Reticulin fibres display a distinct argyrophilia when studied by means of silver impregnation techniques, and show up as a thin meshwork of weakly birefringent, greenish fibres when examined with the aid of the Picrosirius-polarization method. In addition, electron-microscopic studies showed that reticulin fibres are composed of a small number of thin collagen fibrils, contrasting with the very many thicker fibrils that could be localized ultrastructurally to the sites where non-argyrophilic, coarse collagen fibres had been characterized by the histochemical methods used. The three different fibre types of the elastic system belong to a continuous series: oxytalan—elaunin—elastic (all of the fibre types comprising collections of microfibrils with, in the given sequence, increasing amounts of elastin). The three distinct types of elastic system fibres have different staining characteristics and ultrastructural patterns. Ultrastructurally, a characteristic elastic fibre consists of two morphologically different components: a centrally located solid cylinder of amorphous and homogeneous elastin surrounded by tubular microfibrils. An oxytalan fibre is composed of a bundle of microfibrils, identical to the elastic fibre microfibrils, without amorphous material. In elaunin fibres, dispersed amorphous material (elastin) is intermingled among the microfibrils.     

The lateral musculature of the common bully, Gobiomorphus cotidianus, a freshwater fish from New Zealand

A histochemical and ultrastructural study has shown that the myotome of the common bully, Gobiomorphus cotidianus , is composed of three muscle fibre types: white, pink and small diameter fibres. There are no red fibres. Both white and pink fibres have characteristics similar to these fibres found in other teleosts. The small diameter fibres are located in the position usually occupied by red fibres and are identified by their small size and poor staining characteristics. At the ultrastructural level these small fibres are seen to have few mitochondria and a poorly developed sarcoplasmic reticulum. It is suggested that the small diameter fibres are a type of tonic muscle used for positioning the trunk.     

The structure of the smooth muscle fibres in the body wall of the earth worm

1. The structure of the smooth muscle fibres in the longitudinal muscle coat of the body wall of Lumbricus terrestris has been investigated by phase contrast light microscopy and electron microscopy. 2. The muscle fibre is ribbon-shaped, and attached to each of its two surfaces is a set of myofibrils. These are also ribbon-shaped, and they lie with their surfaces perpendicular to the surfaces of the fibre, and their inner edges nearly meeting in the middle of the fibre. These fibrils are oriented at an angle to the fibre axis, and diminish greatly in width as they approach the edge of the fibre. The orientation of the set of fibrils belonging to one surface of the fibre is the mirror image of that of the set belonging to the other surface; thus, when both sets are in view in a fibre lying flat on one face, the fibre exhibits double oblique striation. A comparison of extended and contracted fibres indicates that as the fibre contracts, the angle made between fibre and fibril axes increases (e.g. from 5 to 30 degrees ) and so does the angle made between the two sets of fibrils (e.g. from 10 to 60 degrees ). 3. The myofibril, throughout its length, contains irregularly packed filaments, commonly 250 A in diameter, which are parallel to its long axis and remain straight in contracted muscles. Between them is material which probably consists of much finer filaments. Thus A and I bands are absent. 4. Bound to one face of each fibril, but not penetrating inside it, is a regularly spaced series of transverse stripes. They are of two kinds, alternating along the length of the fibril, and it is suggested that they are comparable to the Z and M lines of a cross-striated fibril. The spacing of these stripes is about 0.5 micro ("Z" to "Z") in extended muscles, and 0.25 micro in contracted muscles. A bridge extends from each stripe across to the stripeless surface of the next fibril.     

A histochemical study of the lateral muscles of five teleost species

A qualitative histochemical study has been made of the myotomal muscles of five teleost fish (glass fish, Chanda ranga; carp, Carassius carassius; coalfish, Gadus virens; black mollie, Molliensia sp. and grey mullet, Mugil cephalus ) . Three or four main fibre types were distinguished in these species on the basis of the distribution and relative activities of glycogen, lipid, aglycerophosphate dehydrogenase, phosphorylase, and succinic dehydrogenase. The so-called red and white fibre types were found to have similar histochemical properties to previously investigated species. All the species studied, with the exception of the glass fish, Chanda ranga , were found to have one or two types of pink fibre situated between the red and white fibre regions. In the carp, coalfish and mullet, the pink fibres were found to be composed of small and large diameter fibres which were similar to red and white fibres respectively, except for their staining for succinic dehydrogenase. Considerable differences were found in the relative amounts of pink muscles between species. Minor fibre components were found in several species. These consisted of very small diameter fibres which did not stain well with any of the histochemical procedures used. It is suggested that these fibres represent areas of continuing muscle growth. The results obtained are discussed in relation to the division of labour between myotomal muscles during swimming.     

Changes in muscle structure associated with somatic growth in Idiosepius pygmaeus, a small tropical cephalopod

Mantle muscle tissue of Idiosepius pygmaeus was examined to describe changes in structure and organization associated with growth. Growth in I. pygmaeus|DD was a function of both an increase in muscle fibre number and fibre size within muscle blocks. Continuous fibre production over the observed life span of I. pygmaeus was indicated by the presence of very small muscle fibres (< 1.0 μm in diameter) in substantial proportions in all sizes of individuals. Muscle blocks became larger as animals increased in size, although new muscle blocks were generated in all sizes of individuals. Mantle muscle fibres had a maximum size of 11 μm. Therefore, for an individual to continue increasing muscle block sizes, new fibres must be produced. This is further evidence of continuous fibre production throughout the size range of I. pygmaeus examined. The relative rates of muscle fibre generation and fibre growth depended on the size of the animal and position along the mantle (anterior, mid or posterior mantle). The predominance of small fibres and blocks at the anterior end of the mantle suggested that this was the primary growth region. Mitochondriapoor and mitochondria-rich muscle fibres from small individuals had much larger mitochondrial cores than muscle fibres from larger animals. Changes in the muscle structure are discussed with respect to the metabolic and energetic requirements of I. pygmaeus , and how these may change with growth.     

三尖杉酯碱诱导HL-60细胞程序性死亡过程中的细胞质和细胞核出泡与染色质凝集

本文利用视频显微影像反差增强技术(VideoEnhancement Contrast,VEC)对三尖杉酯碱诱导的单个HL-60活细胞程序死亡(Apo-ptosis,Apo)全过程进行了观察,结果表明每个Apo细胞在染色质凝集前都要发生细胞核的出泡,而每一个核出泡又都是由相应的质出泡所诱导的,但并不是每个质出泡都能诱导核出泡,质出泡的次数远远高于核出泡,提示核、质出泡可能与染色质凝集有关,并且核、质出泡是程序死亡细胞形成Apo小体所必需的。进一步研究则说明核、质出泡与微丝解聚和重组有关。核、质出泡虽可加速细胞程序死亡过程中的染色质凝集,但并不是程序死亡细胞染色质凝集所必需的,提示HL-60细胞程序死亡过程中的核变化和质变化可能是相对独立的。     

Sperm penetration into immature mouse oocytes and nuclear changes during maturation: an EM study

The ultrastructure of oocyte and sperm nuclei was studied in mouse ovarian oocytes inseminated in vitro and cultured for 1 1/2 and 3 h in a medium containing dbcAMP or lacking the maturation inhibitor. In oocytes blocked at the germinal vesicle (GV) stage, certain maturation-linked changes were noted. Sperm apposition and sperm-oocyte fusion were similar to that during fertilization of ovulated oocytes. The sperm nucleus and its nuclear envelope remained intact after penetrating into the ovarian oocyte. One and a half h after removal of the drug (time 0 of maturation) the germinal vesicle (GV) and sperm nucleus remained intact. In oocytes maturing for 3 h, the nuclear envelopes of the GV and sperm nucleus had fragmented. The NE of the oocyte formed quadruple membranes while the NE of the sperm remained as flat vesicles. Oocyte chromatin condensed to form chromosomes, whereas at the same time the sperm chromatin was in the process of decondensation and was surrounded by fragments of the sperm NE. The sperm chromatin, composed of DNA complexed with protamines, consisted of thin fibrils; the individual fibrils measured 3.8 nm in diameter. Near the penetrated spermatozoa only occasional Mts were detected which were not related to the proximal centriole which was recognizable in the neck-piece of the flagellum. Thus in mouse oocytes the introduced sperm centriole is not capable of behaving as a centrosome and organizing microtubules in the form of an aster.     

Striae albae: a morphological study on the human skin

Specimens of abdomen skin, comprising alternate areas of striae albae and healthy skin, were removed during surgical lipectomy from multiparous and obese women between the ages of 24 and 53 years. A flattening and thinning of the striae albae surface and the almost complete disappearance of dermal papillae was observed in paraffin and thin sections. The papillary dermis was found to be almost completely replaced by straight bundles of collagen fibres running parallel to the skin surface. Immunofluorescence data revealed in these bundles high positivity for type I collagen. The underlying reticular dermis was also found to contain large densely packed bundles of collagen fibres running parallel to the skin surface. Both papillary and reticular dermis collagen fibres were mainly arranged orthogonally to the main axis of the stria. Furthermore, the density of the collagen fibre bundles and the diameter of the collagen fibrils was found to be greater than that of the clinically healthy skin. A larger number of elastic fibres, which presented an abnormal ultrastructural appearance, were visible in pathological papillary and reticular dermis.