Principles of spatial organization of cellular sheets]

The principles of the spatial organisation of epithelial sheets are described, and a new approach to studying their histoarchitecture is proposed. The approach is based on the concept of modular structure of tissues and consists in constructing a family of topological and geometrical models of tissue structure and their experimental verification. Theoretical evidence was obtained that at least 11 topological variants of cell mosaic patterns can be realized in simple epithelia but not one as has been suggested so far. Nine variants of epithelial mosaics predicted have been found experimentally in various animals, and the discovery of the other two is expected in the future. New thus far unknown properties of tissues such as translation symmetry and stoichiometry of the composition were described. By using this approach, it is possible to predict and find new variants of topology of multirow and multilayer epithelia and prognose their changes during normal and pathological morphogenesis.     

Topological Patterns in Metazoan Evolution and Development

Topological patterns in the development and evolution of metazoa, from sponges to chordates, are considered by means of previously elaborated methodology, with the genus of the surface used as a topological invariant. By this means metazoan morphogenesis may be represented as topological modification(s) of the epithelial surfaces of an animal body. The animal body surface is an interface between an organism and its environment, and topological transformations of the body surface during metazoan development and evolution results in better distribution of flows to and from the external medium, regarded as the source of nutrients and oxygen and the sink of excreta, so ensuring greater metabolic intensity. In sponges and some Cnidaria, the increase of this genus up to high values and the shaping of topologically complicated fractal-like systems are evident. In most Bilateria, a stable topological pattern with a through digestive tube is formed, and the subsequent topological complications of other systems can also appear. The present paper provides a topological interpretation of some developmental events through the use of well-known mathematical concepts and theorems; the relationship between local and global orders in metazoan development, i.e., between local morphogenetic processes and integral developmental patterns, is established. Thus, this methodology reveals a “topological imperative”: A certain set of topological rules that constrains and directs biological morphogenesis.     

The role of Hippo signaling pathway and mechanotransduction in tuning embryoid body formation and differentiation

Embryoid bodies (EBs) are the three-dimensional aggregates of pluripotent stem cells that are used as a model system for the in vitro differentiation. EBs mimic the early stages of embryogenesis and are considered as a potential biomimetic body in tuning the stem cell fate. Although EBs have a spheroid shape, they are not formed accidentally by the agglomeration of cells; they are formed by the deliberate and programmed aggregation of stem cells in a complex topological and biophysical microstructure instead. EBs could be programmed to promisingly differentiate into the desired germ layers with specific cell lineages, in response to intra- and extra-biochemical and biomechanical signals. Hippo signaling and mechanotransduction are the key pathways in controlling the formation and differentiation of EBs. The activity of the Hippo pathway strongly relies on cell–cell junctions, cell polarity, cellular architecture, cellular metabolism, and mechanical cues in the surrounding microenvironment. Although the Hippo pathway was initially thought to limit the size of the organ by inhibiting the proliferation and the promotion of apoptosis, the evidence suggests that this pathway even regulates stem cell self-renewal and differentiation. Considering the abovementioned explanations, the present study investigated the interplay of the Hippo signaling pathway, mechanotransduction, differentiation, and proliferation pathways to draw the molecular network involved in the control of EBs fate. In addition, this study highlighted several neglected critical parameters regarding EB formation, in the interplay with the Hippo core component involved in the promising differentiation.     

Effect of flow regime on the architecture of a Pseudomonas fluorescens biofilm

A comparison of the effects of laminar versus turbulent flow regime on the characteristics of a single-species biofilm is presented. The study was carried out by growing Pseudomonas fluorescens biofilms in a flow cell and studying the different layers of the biological matrix with a confocal laser-scanning microscope. The following conclusions were obtained: i) a higher concentration of cells was found in the upper layers of the microbial films than in their inner layers, regardless of the flow regime; ii) the fraction of cells in the overall biofilm mass decreased with time as the film grew; and iii) under laminar flow the total number of cells was higher than in biofilms formed under turbulent flow, but the latter had a higher number of cells per unit volume. Such conclusions, together with the fact that the biofilms were more dense and stable when formed in contact with turbulent flows, favor the design of more compact and efficient biofilm reactors operating in turbulent conditions.     

The stele – a developmental perspective on the diversity and evolution of primary vascular architecture

The stele concept is one of the oldest enduring concepts in plant biology. Here, I review the history of the concept and build an argument for an updated view of steles and their evolution. Studies of stelar organization have generated a widely ranging array of definitions that determine the way we classify steles and construct scenarios about the evolution of stelar architecture. Because at the organismal level biological evolution proceeds by changes in development, concepts of structure need to be grounded in development to be relevant in an evolutionary perspective. For the stele, most traditional definitions that incorporate development have viewed it as the totality of tissues that either originate from procambium – currently the prevailing view – or are bordered by a boundary layer (e.g. endodermis). Consensus between these two perspectives can be reached by recasting the stele as a structural entity of dual nature. Following a brief review of the history of the stele concept, basic terminology related to stelar organization, and traditional classifications of the steles, I revisit boundary layers from the perspective of histogenesis as a dynamic mosaic of developmental domains. I review anatomical and molecular data to explore and reaffirm the importance of boundary layers for stelar organization. Drawing on information from comparative anatomy, developmental regulation, and the fossil record, I propose a stele concept that integrates both the boundary layer and the procambial perspectives, consistent with a dual nature of the stele. This dual stele model posits that stelar architecture is determined at the apical meristem by two major cell fate specification events: a first one that specifies a provascular domain and its boundaries, and a second event that specifies a procambial domain (which will mature into conducting tissues) from cell subpopulations of the provascular domain. If the position and extent of the developmental domains defined by the two events are determined by different concentrations of the same morphogen (most likely auxin), then the distribution of this organizer factor in the shoot apical meristem, as modulated by changes in axis size and the effect of lateral organs, can explain the different stelar configurations documented among tracheophytes. This model provides working hypotheses that incorporate assumptions and generate implications that can be tested empirically. The model also offers criteria for an updated classification of steles in line with current understanding of plant development. In this classification, steles fall into two major categories determined by the configuration of boundary layers: boundary protosteles and boundary siphonosteles, each with subtypes defined by the architecture of the vascular tissues. Validation of the dual stele model and, more generally, in-depth understanding of the regulation of stelar architecture, will necessitate targeted efforts in two areas: (i) the regulation of procambium, vascular tissue, and boundary layer specification in all extant vascular plants, considering that most of the diversity in stelar architecture is hosted by seed-free plants, which are the least explored in terms of developmental regulation; (ii) the configuration of vascular tissues and, especially, boundary layers, in as many extinct lineages as possible.     

The tensor-based model for growth and cell divisions of the root apex. II. Lateral root formation

In this work, the formation of the virtual lateral root (VLR) is shown. The VLR is formed using the 2D simulation model of growth and cell divisions based on the concept of growth tensor, specified for radish. Growth is generated by the field of growth rates of an unsteady type (GT field). Principal directions of growth (PDGs) are assumed to define the orientation of cell divisions. Temporal sequences of the VLR formation are a result of an application of the GT field to the polygon meshwork representing cell pattern of already initiated primordium. The computer-generated lateral root (LR) develops realistically, and its cell pattern is vivid and similar to that observed in anatomical sections. The real and virtual LRs show similar cellular organization, both originate from a small group of cells situated in two-cell layers of the pericycle and both layers are engaged in the LR development. The LR formation seems to be controlled at the tensor level and individual cells presumably detect PDGs and obey them in the course of the cell divisions. PDGs are postulated to affect the cellular organization of the LR. Using the method of computer simulations, cellular aspects of the LR morphogenesis are discussed.     

THE FINE STRUCTURE OF CHONDROCOCCUS COLUMNARIS : I. Structure and Formation of Mesosomes

Cells of Chondrococcus columnaris were sectioned and examined in the electron microscope after fixation by two different methods. After fixation with osmium tetroxide alone, the surface layers of the cells consisted of a plasma membrane, a dense layer (mucopeptide layer), and an outer unit membrane. The outer membrane appeared distorted and was widely separated from the rest of the cell. The intracytoplasmic membranes (mesosomes) appeared as convoluted tubules packaged up within the cytoplasm by a unit membrane. The unit membrane surrounding the tubules was continuous with the plasma membrane. When the cells were fixed with glutaraldehyde prior to fixation with osmium tetroxide, the outer membrane was not distorted and separated from the rest of the cell, structural elements (peripheral fibrils) were seen situated between the outer membrane and dense layer, and the mesosomes appeared as highly organized structures produced by the invagination and proliferation of the plasma membrane. The mesosomes were made up of a series of compound membranes bounded by unit membranes. The compound membranes were formed by the union of two unit membranes along their cytoplasmic surfaces.     

Structure of oxidized thioredoxin to 4 with 5 A resolution

The structure of the oxidized form of Escherichia coli thioredoxin, space group C2, has been determined from X-ray crystallographic data, to a resolution of 4.5 Å using two heavy-atom derivatives, platinum diaminedichloride and 3-pyridyl mercuric chloride. The electron density maps show the molecular shape and the packing of the thioredoxin molecules as well as the positions of the cupric ions necessary for crystallization of thioredoxin. The shape of the thioredoxin molecule is ellipsoidal with approximate dimensions 25 Å × 34 Å × 35 Å. The two thioredoxin molecules in the asymmetric unit appear very similar. They are related by a translation vector with components (0, 0.1, 0.5) along the axis of the unit cell and not by a 2-fold rotation axis. Each of the two molecules in the asymmetric unit belongs to separate infinite layers of molecules parallel to the xy plane. The basic unit in these layers is a dimer formed by interaction of two thioredoxin molecules across the crystallographic 2-fold axis. The structural role of the cupric ions in the crystal lattice is to bridge these dimers within the layers.     

Topological determination of early morphogenesis in Metazoa

This paper presents a topological interpretation of some developmental events through the use of well-known concepts and theorems of combinatorial geometry. The organization of early embryo using a simulation of cleavage considering only blastomere contacts is examined. Each blastomere is modeled as a topological cell and whole embryo—as cell packing. The egg cleavage results in a pattern of cellular contacts on the surface of each blastomere and whole embryo, a discrete morphogenetic field. We find topological distinctions between different types of early egg cleavage and suggest a topological classification of cleavage. Blastulation and gastrulation may be related to an inevitable emergence of discrete curvature that directs development in three-dimensional space. The relationship between local and global orders in metazoan development, i.e., between local morphogenetic processes and integral developmental patterns, is established. Thus, this methodology reveals a topological imperative: a certain set of topological rules that constrains and directs biological morphogenesis.     

Rearrangement of cells in storeyed cambium of<Emphasis Type="Italic"> Lonchocarpus sericeus</Emphasis> (Poir.) DC connected with formation of interlocked grain in the xylem

The history of cellular events in the storeyed cambium of Lonchocarpus sericeus (Poir.) DC was analysed on the basis of changes in the cell arrangement in successive layers and strata of axial parenchyma in the xylem. The mechanism of formation of the regular interlocked grain was investigated. Inclination of fusiform cells changes intensively whereas height and position of storeys in the successive layers of axial parenchyma are constant. As a result, new contacts between cells are formed by means of the intrusive growth of ends of cells belonging to one storey between the tangential walls of cells of the neighbouring storey and unequal periclinal divisions, which give a new shape to the initials. The concept of intrusive growth between the radial walls of the fusiform initials in the formation of xylem with interlocked grain should be revised on this basis.     

Characterization of bovine oviduct epithelial cell monolayers cultured under serum-free conditions

Summary We have developed a culture system for early bovine embryos in serum-free media conditioned by oviduct cell monolayers. A gentle mechanical procedure for oviduct cell isolation has been applied for this purpose avoiding the use of proteolytic enzymes. The aim of the present study was to identify the cell types present in the monolayers and to examine their fate in primary culture in serum-free or in serum-containing media by means of electronmicroscopical, immunocytochemical, and biochemical analyses. The cell dissociation procedure yielded two cell populations: ciliary cells and secretory cells that gradually dedifferentiate during culture. These cells formed a confluent monolayer after 6 d of culture in Tissue Culture Medium 199 medium supplemented with 10% fetal calf serum. Confluent cells displayed a typical epithelial cell morphology as assessed by phase contrast and electron microscopy and all the cells contained cytokeratin filaments as determined by immunocytochemistry. The overall histoarchitecture of the monolayer was preserved after washing and further culture for 7 d in serum-free medium. However, some degenerative signs indicate that the serum-free culture should not be extended for more than 7 d. Confluent oviduct cells also maintained their metabolic and protein secretory activity when deprived of serum. Total protein content in the culture supernatant linearly increased as a function of time and numerous peaks were detected after separation of proteins by high performance ion exchange chromatography. Protein elution patterns were reproducible and most of the proteins present in the culture medium were neosynthesized as determined by the incorporation of radiolabeled amino acids into nondialyzable proteins.     

Basic Types of Structural Changes in the Leaf Mesophyll during Adaptation of Eastern Pamir Plants to Mountain Conditions

Quantitative characteristics of mesophyll structure were compared in leaves of eleven alpine plant species grown under natural conditions in the Eastern Pamirs at various altitudes, from 3800 to 4750 m. Basic types of changes in mesophyll structure, associated with plant adaptation to mountain conditions, were characterized. These changes manifested themselves in different numbers of cell layers and cell sizes in the palisade tissue and, as a consequence, in changed leaf thickness and cell number per unit of leaf area. Three plant groups were identified by the changes in the leaf structural characteristics depending on the type of mesophyll structure, ecological group of plant species, and altitude of plant habitat. The first group comprised alpine xerophytes with an isopalisade structure, in which the volume of palisade cells decreased and their number per unit of leaf area increased with the altitude of plant habitat. The number of mesophyll layers and leaf thickness decreased or did not change with altitude. The second group comprised subalpine plant species with a dorsoventral structure of mesophyll; these species occur below the line of continuous night frost. In these plant species, the number of mesophyll layers, leaf thickness, and cell number per unit of leaf area increased with altitude. The third group comprised mesophyte plants with a dorsoventral and homogenous mesophyll structure, which are encountered in a wide range of habitats, including the nival belt (from 4700 to 5000 m). In this group, cell volume increased and cell number per unit of leaf area decreased with altitude. We present a general scheme of leaf structural changes, which explains the changes in the quantitative characteristics of mesophyll as a function of altitude and highland environmental conditions.     

凤仙花花药发育中多糖和脂滴组织化学研究

以不同发育时期的凤仙花花药为实验材料,采用组织化学方法,对花药发育中的结构变化及多糖和脂滴物质分布进行观察。结果表明:(1)凤仙花的花药壁由6层细胞组成,包括1层表皮细胞,2层药室内壁细胞,2层中层细胞和1层绒毡层细胞。其中绒毡层细胞的形态不明显,很难与造孢细胞区分,且在小孢子母细胞时期退化。(2)在小孢子母细胞中出现了一些淀粉粒,但减数分裂后,早期小孢子中的淀粉粒消失,又出现了一些小的脂滴;随着花粉的发育,小孢子形成大液泡,晚期小孢子中的脂滴也消失;小孢子分裂形成二胞花粉后,营养细胞中的大液泡降解、消失,二胞花粉中又开始积累淀粉;接近开花时,成熟花粉中充满细胞质,其中包含了较多的淀粉粒和脂滴。(3)在凤仙花的花药发育中,绒毡层细胞很早退化,为小孢子母细胞和四分体小孢子提供了营养物质;其后的中层细胞退化则为后期花粉发育提供了营养物质。     

The fine structure of the compound sense organs on the cirri of Nereis diversicolor

Summary A study of the fine structure of the sense organs on the prostomial cirri and palps of Nereis diversicolor shows them to consist of two types of cell. There are between 7 and 15 sensory cells and a similar number of associated cells which contain many osmiophilic granules. The cell bodies of both are sub-epidermal, having a long distal process which reaches the surface in a raised sensory hillock. The sensory cells carry a cilium, which passes through the cuticle and emerges surrounded by a sheath formed from the outer layers of the epicuticle. Scanning electron micrographs show the surface of the cirrus to be covered by hair-like epicuticular microvilli, through which the sheathed cilia protrude. There is also a second type of sensory cell which occurs singly between the epithelial cells. The distal membrane of this cell is formed into a tuft of approximately 55 large microvilli which open through a pore in the epicuticle. It is suggested by their position and structure, that both these receptors resemble chemoreceptors.We should like to acknowledge the advice and technical help of Dr. J. A. Nott of the N.E.R.C. unit of Electron Microscopy, Menai Bridge, and Dr. P. E. Secker of the School of Electronic Engineering for use of the Cambridge Stereoscan. The work is supported by a grant from the Science Research Council to D.A.D.     

MOLECULAR STRUCTURE OF PLANT FIBERS DETERMINED BY X-RAYS

It has been shown that the wall of the plant fiber is probably built up of unit groups of atoms which have assumed the form of a space lattice. The elementary cell of the lattice is an orthorhombic structure with the dimensions 6.10 x 5.40 x 10.30 Å.u., and contains two unit groups equal in size to two C₆H₁₀O₅ groups. The crystallographic unit cell would contain 4 of these elementary cells and would be represented by Fig. 9 rather than by Fig. 3. The groups of atoms, C₆H₁₀O₅, are arranged in parallel chains running lengthwise of the fiber. In each chain the odd numbered groups have a different orientation from the even numbered. The chains, parallel to one another are spaced 6.10 Å.u. in one direction and 5.40 Å.u. at right angles to that. In these two directions the odd numbered chains also would have a different orientation from the even numbered. On account of the cylindrical shape of the fiber, the elementary cells are arranged in the form of concentric cylinders or layers. The dimensions of the fibers are such that the fiber wall is about 40,000 elementary cells in thickness, or in other words, the fiber is composed of that many concentric layers. If it could be magnified sufficiently, a cross-section of a fiber would show the end view of each cylinder as a dotted circle. The dots, representing the unit groups of atoms, would have considerable uniformity of spacing in both the tangential and the radial directions, 6.10 Å.u. in one and 5.40 Å.u. in the other. The structure could not be as rigidly exact as might be inferred, since the wall is deposited more or less rhythmically during a period of several days or weeks* in which adjustments in the arrangement of the unit groups undoubtedly occur. It is common knowledge that the fibers, under the microscope, rarely appear as true circles on cross-section; usually they appear as irregular, many-sided polygons and the wall thickness is normally uneven. For our purpose it is simpler to think of the fiber as composed of concentric cylinders with diameters so large in proportion to the size of the unit groups that in relatively large segments they closely approach the parallelism of the planes of a rectangular lattice, sufficiently close to be capable of producing diffraction patterns. Although these conclusions seem to be in agreement with the diffraction patterns obtained from various positions of a bundle of approximately parallel fibers, the fact must not be overlooked that the structure cannot be proved with as great certainty as can the structure of a well formed crystal. The very nature of the fiber, its cylindrical shape, and the many internal adjustments which must take place, militate against a clean-cut demonstration. Models, made more or less to scale, were used in working out this structure. The unit group was constructed according to Irvine''s suggestion that all the groups are glucose residues. An intensive study is now under way in which an attempt is being made to bring the models into agreement with the chemical and physical properties of the cellulose fibers and with the diffraction patterns. A report on that part of the work will soon be submitted for publication.     

Differentiation During Encystment and Excystment in Oxytricha fallax*

During encystment of Oxytricha fallax, a wall composed of 4 distinct layers, each derived from a different kind of endoplasmic vesicle, is formed between the 2 unit membranes that cover the vegetative cell. Numerous autophagic vacuoles arise in the endoplasm and later (during excystment undergo internal changes comparable to those characteristic of food vacuoles. Mitochondria aggregate into a band. The 2 macronuclei fuse and their nucleoli become homogeneous. Except for the 2 cell membranes, all visible cortical structures, including cilia, kinetosomes, and microtubules, disappear. Despite the absence of visible ciliature in the mature cyst, the various primordia of the normal vegetative ciliature arise during excystment in the same positional relations to one another as is characteristic of developments during cell division.     

Developmental changes in cell wall structure of phloem fibres of the bamboo Dendrocalamus asper

BACKGROUND AND AIMS: Bamboo culms have excellent physical and mechanical properties, which mainly depend on their fibre content and anatomical structure. One of the features which is known to contribute to the high tensile strength in bamboo is the multilayered structure of the fibre cell wall. The aim of this study was to characterize the development of the layered structure in fibre cell walls of developing and maturing culms of Dendrocalamus asper. METHODS: Cell wall development patterns were investigated in phloem fibre caps of vascular bundles in the inner culm wall areas of Dendrocalamus asper of three different age classes (<6 months old, 1 year old, 3 years old). A combination of light microscopy and image analysis techniques were employed to measure cell wall thickness and to determine number of cell wall layers, as well as to describe the layering structure of fibre walls. Two-dimensional maps showing the distribution pattern of fibres according to the number of cell wall layers were produced. KEY RESULTS: The cell walls of fibres in phloem fibre caps located in the inner part of the culm wall of D. asper developed rapidly during the first year of growth. Six different fibre types could be distinguished based upon their cell wall layering and all were already present in the young, 1-year-old culm. In the mature stage (3 years of age) the multilayering was independent of the cell wall thickness and even the thinner-walled fibres could have a large number of wall layers. The multilayered nature of cell wall structure varied considerably between individual cells and was not exclusively related to the cell wall thickness. Nevertheless, fibres at the periphery of the fibre bundles and immediately adjacent to the phloem elements exhibited a consistent and high degree of layering in their cell walls. CONCLUSIONS: The multilayered structure of fibre cell walls was formed mainly during the first year of growth by the deposition of new wall layers of variable thickness, resulting in a high degree of heterogeneity in the layering patterns amongst individual fibres. A degree of 'order' in the distribution of multilayered fibres within the caps does exist, however, with multilayered cell walls common in fibres adjacent to phloem elements and around the edge of the fibre cap. These findings confirm the observations, primarily in Phyllostachys viridi-glaucescens. The layering structure was not found to be specifically related to the thickness of the cell wall.     

Interaction of collagen with hydrophobic protein granules in the egg capsule of the dogfish scyliorhinus canicula

The egg capsule of the dogfish is a composite material containing collagenous fibrils and 2 mum spherical hydrophobic protein granules. The latter appear to owe much of their hydrophobicity to an exceptionally high tyrosine content (approximately 20% of total amino acid residues). The hydrophobic component appears to form as an emulsion in the secretory granules of the D and E zone gland cells of the nidamental gland. Droplets of the hydrophobic material appear to become coated with remarkably regular layers of radially-arranged collagen molecules which form a series of concentric, evenly spaced layers around each hydrophobic granule. Numerous disclinations were seen where the layers around adjacent granules interfered with one another. The layers are thought to represent a lamellar liquid crystalline phase previously described for this collagen (Knight et al., 1993). The fine structural appearance of the concentric layers and evidence for radial arrangement of collagen molecules within them is compatible with the suggestion that the layers are built from a dumbbell-shaped unit approximately 35 nm long with hydrophobic groups concentrated at the ends. This unit may represent a dumbbell-shaped molecule or an oligomer of two or more molecules lying parallel with one another in a head-to-tail arrangement. Such a unit can be readily incorporated into models for the micellar, hexagonal columnar and final fibrillar phases previously described for this collagen (Knight et al., 1993). Evidence from the TEM study of stretched egg capsule wall suggests that there is a mechanical interaction between the hydrophobic granules and the collagen fibrils in the fully formed material. We suggest that the radial, concentric layered arrangement of collagen molecules is established by hydrophobic interactions within the liquid crystalline material and locked into place by oxidative covalent cross-linking to give a 3-dimensional cross-linked meshwork of collagen fibrils and hydrophobic granules. The latter arrangement helps to account for the high tensilestrength and toughness of this material.