Long bone microstructure gives new insights into the life of pachypleurosaurids from the Middle Triassic of Monte San Giorgio,Switzerland/Italy

The long bone microstructure of four pachypleurosaurid taxa from Monte San Giorgio (Switzerland/Italy) was studied. Pachypleurosaurids are secondarily aquatic reptiles that lived during the Middle Triassic in varying marine environments of the Tethys. All four pachypleurosaurids show high compactness values in their long bones based on a thick cortex and a calcified cartilaginous core, which remains in the medullary region throughout the ontogeny. Parts or even the entire embryonic bone layer composed of a mixture of woven-fibered bone tissue and parallel-fibered bone tissue is preserved in both pachypleurosaurid genera. The rest of the cortex consists of lamellar-zonal bone tissue type. Differences in the microstructure of the bones between the pachypleurosaurids are reflected in the occurrence of remodelling processes, which, if present, affect the innermost growth marks of the cortex or the calcified cartilaginous core. Further variation is present in the spacing pattern of the growth cycles, as well as in the degree of vascularisation of the lamellar-zonal bone tissue type. Our data on the microstructure of the long bones support previous studies on morphology and facies distribution, which indicated different habitats and adaptation to a secondary aquatic lifestyle for each pachypleurosaurid taxon. Life history data furthermore reflect different longevities and ages at sexual maturity. The bone histological data of the stratigraphically youngest and oldest pachypleurosaurid species might indicate possible climate-dependant reproductive seasons similar to Recent lacertilian squamates.     

Ossification of the human fetal ilium.

Ossification of the ilium is similar to that of a long bone. It possesses three cartilaginous epiphyses and one cartilaginous process. Moreover, it undergoes peculiar osteoclastic resorption, comparable with that of the cranium bones. Asymmetrical ossification of the ilium, haversian bone remodelling and apposition of chondroid tissue posterosuperiorly to the acetabulum most probably emphasize the importance of mechanical factors in the morphogenesis of the hip bone during fetal life.     

VALVE MORPHOGENESIS IN THE CENTRIC DIATOM PROBOSCIA ALATA SUNDSTROM1

Valve morphogenesis in Proboscia alata Sundstrom was followed in living cells and during treatment with antiactin and antimicrotubule drugs. Once cleaved, sibling cells rounded up and retracted. Soon, a granular organizing center (OC) appeared adjacent to the stub of the initiated valve. Silicification started within a silicon deposition vesicle (SDV) adjacent to the OC. The elongating valve was initially tubular and sealed at one end, creating the proboscis of the conical valve. The edge of the SDV and thinnest region of the forming valve was lined by a sleeve of bundled microtubules (MTs) that terminated short of the older more rigid part of the valve. The growing proboscis of living cells treated with the anti‐MT drug oryzalin became grossly distorted. EM revealed dense material lining the growing edge of the SDV; immunofluorescence microscopy showed a ring of actin here. Applied to living cells, the antiactin drug cytochalasin D caused the very young proboscis to collapse; in older valves, the base of the proboscis expanded. Thus, valve morphogenesis appeared controlled by the MT cytoskeleton, keeping the proboscis straight while actin molded its conical outline. At the tip of the proboscis was a slit resembling a labiate process. Its morphogenesis involved striated fibers and two MTs, reminiscent of the fibers and MTs associated with raphe morphogenesis. In contrast to spine‐like processes that elongate by tip growth, the tip of the proboscis was formed first, and the consequent “antitip growth” suggests the tip was originally the center of the valve face.     

A microanatomical and histological study of the fin long bones of the Devonian sarcopterygian Eusthenopteron foordi

Meunier F.J. and Laurin M. 2012. A microanatomical and histological study of the fin long bones of the Devonian sarcopterygian Eusthenopteron foordi. —Acta Zoologica (Stockholm) 93 : 88–97. A paleohistological study of the endoskeletal bones of the dorsal and pelvic fins shows that Eusthenopteron foordi had true long bones that grew in length and thickness through endochondral and periosteal ossification, respectively. Endochondral ossification required cartilaginous epiphyses with a growth plate system whose presence is confirmed by both calcified cartilage and thin endochondral bony trabeculae that overlaid the erosive bays located in hypertrophic calcified cartilage. Articulations between axial mesomeres in paired fins were diarthroses. This microanatomical organization, i.e. longitudinal growth of diaphysis and articulations between epiphyses, can be considered an exaptation for terrestrial locomotion as it can produce skeletal elements able to support strong mechanical stress.     

Mechanics of growth pulsations as the basis of growth and morphogenesis in colonial hydroids

Growth and shaping in colonial hydroids (Hydrozoa, Cnidaria) are realized due to the functioning of special colony elements, growing tips located at the terminuses of branched colony body. Unlike in plants, the growing tips of colonial hydroids are sites of active cell movements related to morphogenesis and lacking proliferation. The activity of hydroid growing tips is expressed as growth pulsations: cyclic repetitions of their apex extensions and retractions. The parameters of growth pulsations are species specific and related to the shape of a forming element. Here, the succession of cell movements and changes in mutual arrangement within the growing tip are described in detail at all pulsation phases. The role of the inner cell layer in the tip activity was demonstrated for the first time. Relationships between the growing tip parameters, length and diameter, and pulsations are discussed. A scheme is proposed for cyclic processes in both epithelial layers. An explanation is provided for the two-step mode of growth pulsations with relative independence of the main phases. It was proposed that successive activities of the tip ecto- and endoderm serve as driving forces provided there is a hard outer skeleton. This scheme makes it possible to explain some patterns of growth and morphogenesis in colonial hydroids, such as gradually increasing growth rate of a new tip and its maximum growth rate, differences in the parameters of growth pulsations between shoot and stolon tips, shoot base inclination towards the stolon tip, etc., and provides a basis for further improvement of the model of morphogenesis in hydroids.     

Mechanics of growth pulsations as the basis of growth and morphogenesis in colonial hydroids

Growth and shaping in colonial hydroids (Hydrozoa, Cnidaria) are realized due to the functioning of special colony elements, growing tips located at the terminuses of branched colony body. Unlike in plants, the growing tips of colonial hydroids are sites of active cell movements related to morphogenesis and lacking proliferation. The activity of hydroid growing tips is expressed as growth pulsations: cyclic repetitions of their apex extensions and retractions. The parameters of growth pulsations are species specific and related to the shape of a forming element. Here, the succession of cell movements and changes in mutual arrangement within the growing tip are described in detail at all pulsation phases. The role of the inner cell layer in the tip activity was demonstrated for the first time. Relationships between the growing tip parameters, length and diameter, and pulsations are discussed. A scheme is proposed for cyclic processes in both epithelial layers. An explanation is provided for the two-step mode of growth pulsations with relative independence of the main phases. It was proposed that successive activities of the tip ecto-and endoderm serve as driving forces provided there is a hard outer skeleton. This scheme makes it possible to explain some patterns of growth and morphogenesis in colonial hydroids, such as gradually increasing growth rate of a new tip and its maximum growth rate, differences in the parameters of growth pulsations between shoot and stolon tips, shoot base inclination towards the stolon tip, etc., and provides a basis for further improvement of the model of morphogenesis in hydroids.     

The endocytic network in plants

Endocytosis and vesicle recycling via secretory endosomes are essential for many processes in multicellular organisms. Recently, higher plants have provided useful experimental model systems to study these processes. Endocytosis and secretory endosomes in plants play crucial roles in polar tip growth, a process in which secretory and endocytic pathways are integrated closely. Plant endocytosis and endosomes are important for auxin-mediated cell-cell communication, gravitropic responses, stomatal movements, cytokinesis and cell wall morphogenesis. There is also evidence that F-actin is essential for endocytosis and that plant-specific myosin VIII is an endocytic motor in plants. Last, recent results indicate that the trans Golgi network in plants should be considered an integral part of the endocytic network.     

The angular process of Monodelphis domestica (Didelphidae, Marsupialia) and its relation to the middle ear: an ontogenetic and evolutionary morphologic study

In most marsupials, the angular process is inflected medially. By using an ontogenetic series of Monodelphis domestica, the development of this characteristic structure has been described. In contrast with the eutherian mammals, in marsupials there is retained a close connection between the dentale and the tympanicum and goniale; it is well known that these 2 elements of the middle ear are derived from the angulare and prearticulare of the reptilian lower jaw. At the neonatal stage, the dentale and tympanicum are both relatively vertically orientated; during the following 2 weeks, they take an increasingly oblique position, which is primarily caused by the rapid growth of the braincase. Only after the eruption of the first teeth, the ascending ramus of the dentale takes a more and more vertical position, whereas the angular process remains with its tip near the medioventral floor of the tympanic bulla. The bulla shows at this place a rectangular fenestra which is covered by a membrane of loose connective tissue; the tip of the angular process, which is always free of muscular insertions, maintains contacts with this fenestra throughout life. During juvenile and adult life stages, the process becomes somewhat removed from the fenestra for obvious reasons, but at a gape of about 40 to 50 degrees it inevitably must touch the "inferior tympanic membrane" and possibly also the tympanic ring. It is speculated that the relationship between the angular process and the tympanic bulla represents a specific form-function complex for sound transmission, which may be a modified retention from archaic mammalian conditions. Further details of the ontogenetic development of the tympanic region have been described which may be of some relevance for the evolutionary morphology of mammals: The tympanic process of the petrosal, which fixes the posterior end of the tympanic ring, is formed by 'Zuwachsknochen' (additional bone) but not by cartilage. The styloid process remains cartilaginous throughout life: its free tip ends in the lateral wall of the tympanic cavity and it is closely connected with the collum mallei and the posterior end of the tympanicum; it guides the chorda tympani and may therefore be homologous with the cartilage of Spence. The cartilage of Paauw is interpreted in terms of functional morphology. A model of evolutionary transformation of the dentale-tympanicum complex in mesozoic mammals in outlined on the basis of the ontogenetic findings in Monodelphis and other didelphid and dasyurid marsupials.     

Cellular interactions during cartilage and bone development.

Both interactions between like cells, as between chondrogenic cells in a developing cartilaginous rudiment, and between unlike cells, as in epithelial-mesenchymal interactions, are dealt with in this review. Such interactions may involve direct apposition of cell membranes or may be mediated via interaction with peri- or extracellular matrices. An ontogenetic approach is taken in which cellular interactions involved in five processes of the development of cartilage and bone are discussed, the five being (1) origin of the cells, (2) migration of the cells within the embryo, (3) localization of the cells at their final embryonic site, (4) differentiation, and (5) morphogenesis. Some emphasis is placed on interactions affecting neural crest-derived cells both before and during their migration and on interactions, especially epithelial-mesenchymal interactions, that precede cytodifferentiation of chondroblasts or osteoblasts. Whether epithelial or mesenchymal specificity is required for such interactions to occur is discussed with reference to the otic vesicle-otic mesenchyme interaction that leads to differentiation and morphogenesis of the cartilaginous otic capsule.     

Specific effects of retinoic acid on the skeletal morphogenesis of the 11-day mouse embryo forelimb bud in vitro

Using our improved method for culturing 11-day mouse forelimb buds in vitro, we have investigated the effects of a local application of all-trans-retinoic acid (RA) on growth, cartilaginous differentiation and skeletal patterning in the mammalian limb bud. Carrier implants of catgut impregnated with DMSO or various doses of RA in DMSO were inserted at the apex of the buds in the proximo-distal axis just beneath the apical ectodermal ridge. After 6 days of culture, cartilaginous skeletons were stained and explants were processed for morphological analysis and quantitative study using computerized optical image analysis. Buds treated with low doses of RA exhibited stimulated growth and chondrogenesis. Moreover, hypertrophied and fused metacarpals were seen within explants treated with the lowest dose. High doses strongly inhibited growth and skeletal morphogenesis. An intermediate dose sustained cartilaginous differentiation at the same level as low doses, but concomitantly disturbed the skeletal pattern. These results are discussed considering reported RA effects on other experimental systems including avian limb bud as an in vivo model or cell cultures as an in vitro simplified model.     

The articular disc of the hand.

The carpal articular disc have been studied in serial sectioned embryonal and fetal hands. It can be concluded that the articular disc is an extensive fibrous system that comes from the ulnar edge of the distal end of the radius and reaches, bordered by the deep layer of the antebrachial fascia, the base of metacarpal V. In this course, insertions take place into the ulnar aspect of the head of the ulna, into the ulnar styloid process and into the ulnar carpals. Emphasis has been laid upon the fact that the sheath of the m. extensor carpi ulnaris tendon is part of the fibrous system of the disc. The positions of the dorsal tendons seen with respect to the fascial implicate a mutual shift during pronation and supination of the hand. The relations between the so-called ligamentum subcruentum and prestyloid recess have been established, the presence of cartilaginous primordia in the developing dis has been discussed. We come to the conclusion that the evidence for drawing a parallel between phylogenetic and human embryological phenomena is still insufficient. In this connection we have stressed the modus of development of the pisotriquetral joint space.     

Chemically mediated mechanical expansion of the pollen tube cell wall

Morphogenesis of plant cells is tantamount to the shaping of the stiff cell wall that surrounds them. To this end, these cells integrate two concomitant processes: 1), deposition of new material into the existing wall, and 2), mechanical deformation of this material by the turgor pressure. However, due to uncertainty regarding the mechanisms that coordinate these processes, existing models typically adopt a limiting case in which either one or the other dictates morphogenesis. In this report, we formulate a simple mechanism in pollen tubes by which deposition causes turnover of cell wall cross-links, thereby facilitating mechanical deformation. Accordingly, deposition and mechanics are coupled and are both integral aspects of the morphogenetic process. Among the key experimental qualifications of this model are: its ability to precisely reproduce the morphologies of pollen tubes; its prediction of the growth oscillations exhibited by rapidly growing pollen tubes; and its prediction of the observed phase relationships between variables such as wall thickness, cell morphology, and growth rate within oscillatory cells. In short, the model captures the rich phenomenology of pollen tube morphogenesis and has implications for other plant cell types.     

Stereom morphogenesis and differentiation during regeneration of adambulacral spines of Asterias rubens (Echinodermata,Asteroida)

Summary The very first mineral deposits appearing in regenerating fractured adambulacral spines of Asterias rubens are minute polyhedrons that cover the surface of fractured trabeculae. Polyhedrons fuse together forming a fold from which a microspine differentiates. Microspines develop into long linear trabeculae which send out lateral processes at regular length intervals. Lateral processes from adjacent trabeculae fuse together, bridging the trabeculae and giving the regenerate the typical meshwork structure of stereom. Most of the regenerate is built up according to this growth pattern which ensures its longitudinal growth. Simultaneously, the initial fascicular stereom of the stub sends out short radial processes which branch into upward and downward directed subprocesses. The latter fuse with their equivalents located above or below, building up longitudinal rows of stereom meshes. These rows then bridge together by additional branched or unbranched lateral processes, so forming a new stereom layer which progressively covers the whole stub. Up to three new layers of stereom are formed in this way at the stub periphery. These become continuous with the stereom layers of the regenerate by fusion of reciprocal subprocesses, so ensuring the continuity between the stub and the regenerate. In both structures the first stage of mineralization results in an open stereom. Stereom thickening occurs in a second stage of mineralization (that is chronologically separated from the formation of the open stereom) and results in the differentiation of the original stereom fabrics (i.e. fascicular stereom). Regeneration of removed spines starts with the formation of a new spine base made of labyrinthic stereom. The development of the latter mostly relies on short branched and unbranched processes which fuse with each other or with predifferentiated meshes. After completion of its base, the regenerating spine lengthens and thickens similarly to the regenerating fractured spines. The diversity of the stereom growth processes observed in the present work may be reduced to the combination of one to three elementary events, viz. the development of long linear processes, of short unbranched processes and of short branched processes. A survey of the literature allows the suggestion that the implementation of these elementary events is sufficient to describe most types of stereom morphogenesis.Senior research assistant NFSR (Belgium)     

Deletion of the gene encoding c-Cbl alters the ability of osteoclasts to migrate, delaying resorption and ossification of cartilage during the development of long bones

During development of the skeleton, osteoclast (OC) recruitment and migration are required for the vascular invasion of the cartilaginous anlage and the ossification of long bones. c-Cbl lies downstream of the vitronectin receptor and forms a complex with c-Src and Pyk2 in a signaling pathway that is required for normal osteoclast motility. To determine whether the decreased motility we observed in vitro in c-Cbl(-/-) OCs translated into decreased cell migration in vivo, we analyzed the long bones of c-Cbl(-/-) mice during development. Initiation of vascularization and replacement of cartilage by bone were delayed in c-Cbl(-/-) mice, due to decreased osteoclast invasion of the hypertrophic cartilage through the bone collar. Furthermore, c-Cbl(-/-) mice show a delay in the formation of secondary centers of ossification, a thicker hypertrophic zone of the growth plate, and a prolonged presence of cartilaginous remnants in the spongiosa, confirming a decrease in resorption of the calcified cartilage. Thus, the decrease in motility of c-Cbl(-/-) osteoclasts observed in vitro results in a decreased ability of osteoclasts to invade and resorb bone and mineralized cartilage in vivo. These results confirm that c-Cbl plays an important role in osteoclast motility and resorbing activity.     

Morphogenesis of the highly specialized nasal skull in the sperm whale (Physeter macrocephalus). I

Investigated the morphogenesis of the nasal structures of the chondrocranium and determatocranium in 15 embryos and foetuses of the sperm whale (Physeter macrocephalus). In the very early stages of the morphogenesis, the nasal capsule of Physeter shows a conspicuous similarity with that of all other odontocetes. In the following stages there are some important differences. Most peculiar is the occurrence of the cartilaginous tectum nasi with the cupulae nasi anteriores, elements which are reduced in all other odontocetes. This cartilaginous complex as a slender band projects obliquely forward from the upper edge of the anterior septal margin. It is free, i.e. not accompanied by membraneous bones. The complex represents the most important factor in the morphogenesis and growth of the characteristic big forehead in Physeter, which contains the spermaceti organ unique within the odontocetes. Other important differences concern the changes in the orientation of the nasal passages and the adjacent skeletal structures. Nevertheless, these differences have to be taken as specializations related to the development of the far-advanced spermaceti organ. As a whole, the embryonic nasal structures in Physeter belong to the same general type of nasal capsule which is common to all odontocetes. The results presented here suggest a close phylogenetic relationship between Physeter and the other Odontoceti.     

Growth and proteoglycan metabolism of chick embryonic cartilaginous long bone rudiments and of isolated epiphyses

Summary Turnover of extracellular matrix (ECM) proteoglycans was studied in chick cartilaginous femur rudiments grown in organ culture. Femora from six-day-old embryos showed nearly normal growth rates during the first few days in culture. By labeling the rudiment with ³⁵S-sulfate or ¹⁴C-glucosamine, it was demonstrated that the cartilaginous ECM undergoes rapid turnover. It was also found that the metabolic fate of the proteoglycans is to be released as macromolecules into the culture medium. When a rudiment was cut to obtain two epiphyses it was observed that each part grows and synthesizes proteoglycans at nearly normal rates, which indicates that the isolated epiphyses, like the whole rudiment, behave as autonomous systems. We suggest that the turnover of ECM components is part of the continuous remodelling process rudiments undergo during their growth and development. In order to study cell-ECM interaction in morphogenesis, we made an attempt to prepare an intact cell-free ECM. Epiphyses were heated at 45.2° C for 1 h. The treatment caused complete cessation of growth and biosynthesis. When the cut surface of a live epiphysis was brought into apposition to a heat-treated epiphysis and the attached pair placed in organ culture, it was found that the heat-treated epiphysis begins to grow and reaches almost the same size as its live counterpart. We discuss the possible advantage of this new experimental system for studies on the role of ECM in morphogenesis.     

Specific effects of retinoic acid on the skeletal morphogenesis of the 11-day mouse embryo forelimb bud in vitro

Using our improved method for culturing 11-day mouse forelimb buds in vitro, we have investigated the effects of a local application of all-trans-retinoic acid (RA) on growth, cartilaginous differentiation and skeletal patterning in the mammalian limb bud. Carrier implants of catgut impregnated with DMSO or various doses of RA in DMSO were inserted at the apex of the buds in the proximo-distal axis just beneath the apical ectodermal ridge. After 6 days of culture, cartilaginous skeletons were stained and explants were processed for morphological analysis and quantitative study using computerized optical image analysis. Buds treated with low doses of RA exhibited stimulated growth and chondrogenesis. Moreover, hypertrophied and fused metacarpals were seen within explants treated with the lowest dose. High doses strongly inhibited growth and skeletal morphogenesis. An intermediate dose sustained cartilaginous differentiation at the same level as low doses, but concomitantly disturbed the skeletal pattern. These results are discussed considering reported RA effects on other experimental systems including avian limb bud as an in vivo model or cell cultures as an in vitro simplified model.     

Plasma membrane-adjacent actin filaments, but not microtubules, are essential for both polarization and hyphal tip morphogenesis in Saprolegnia ferax and Neurospora crassa

The organization and roles of F-actin and microtubules in the maintenance and initiation of hyphal tip growth have been analyzed in Saprolegnia ferax and Neurospora crassa. In hyphae of both species, the apex is depleted of microtubules relative to subapical regions and near-normal morphogenesis occurs in concentrations of nocodazole or MBC which remove microtubules, slow growth, and disrupt nuclear positioning. In contrast, each species contains characteristic tip-high arrays of plasma membrane-adjacent F-actin, whose organization is largely unaltered by the loss of microtubules but disruption of which by latrunculin B disrupts tip morphology. Hyphal initiation and subsequent normal morphogenesis from protoplasts of both species and spores of S. ferax are independent of microtubules, but at least in S. ferax obligatorily involve the formation of F-actin caps adjacent to the hyphal tip plasma membrane. These observations indicate an obligatory role for F-actin in hyphal polarization and tip morphogenesis and only an indirect role for microtubules.     

dmd-3, a doublesex-related gene regulated by tra-1, governs sex-specific morphogenesis in C. elegans

Although sexual dimorphism is ubiquitous in animals, the means by which sex determination mechanisms trigger specific modifications to shared structures is not well understood. In C. elegans, tail tip morphology is highly dimorphic: whereas hermaphrodites have a whip-like, tapered tail tip, the male tail is blunt-ended and round. Here we show that the male-specific cell fusion and retraction that generate the adult tail are controlled by the previously undescribed doublesex-related DM gene dmd-3, with a secondary contribution from the paralogous gene mab-3. In dmd-3 mutants, cell fusion and retraction in the male tail tip are severely defective, while in mab-3; dmd-3 double mutants, these processes are completely absent. Conversely, expression of dmd-3 in the hermaphrodite tail tip is sufficient to trigger fusion and retraction. The master sexual regulator tra-1 normally represses dmd-3 expression in the hermaphrodite tail tip, accounting for the sexual specificity of tail tip morphogenesis. Temporal cues control the timing of tail remodeling in males by regulating dmd-3 expression, and Wnt signaling promotes this process by maintaining and enhancing dmd-3 expression in the tail tip. Downstream, dmd-3 and mab-3 regulate effectors of morphogenesis including the cell fusion gene eff-1. Together, our results reveal a regulatory network for male tail morphogenesis in which dmd-3 and mab-3 together occupy the central node. These findings indicate that an important conserved function of DM genes is to link the general sex determination hierarchy to specific effectors of differentiation and morphogenesis.