Spatial organization of the lacunar-canalicular system in the structures of bone lamellae

By means of light microscopical techniques and scanning electron microscopy spatial organization of the lacunar-canalicular system (LCS) has been studied in structures of a mature lamellar bone. A method for making corrosive casts of osseous lacunae and canaliculi is suggested, owing to which their spatial organization can be analysed. Certain data on interconnections of the osseous lacunae with each other and with vascular canals and natural surfaces of the bone are presented. The role of LCS as a component of the microcirculatory bed of the lamellar bone is discussed.     

Morphogenesis of structural elements of the cervical spine in the white rat in prenatal ontogeny

Embryonal development of the spinal column cervical part has been studied in 100 series of sagittal, transversal, frontal sections; time of the main structural elements anlagen (vertebral bodies, arches, joints, ligaments) is noted. The prenatal development of the spinal column cervical part is divided into 3 stages--mesenchymal, cartilagenous, osseous. The first stage lasts up to 16 days of development; during this period anlagen of vertebral bodies, arches, joints, ligaments are formed. The second stage--cartilagenous; mesenchyma is substituted for cartilagenous tissue, cartilagenous cells are differentiated. This stage lasts from the 16th up to the 18th day of embryogenesis. The third stage--osseous--lasts from the 18th up to the 21st day of embryogenesis. During this period structures of the spinal column cervical part acquire a definitive form, the cartilagenous tissue is substituted for the osseous one.     

Ossification of the antler in the Lapland reindeer (Rangifer tarandus tarandus)

A histological and histochemical study of biopsy specimens from the Lapland reindeer antler indicated that the intercellular matrix of the cartilage that forms the partitions of longitudinal channels becomes calcified at virtually the initial stage of formation. The lacunae at the peripheries of the cartilaginous partitions are invaded by osteoblasts in a process comparable to endochondral ossification. The very centres of the partitions evidently become directly converted into bone without the presence of e.g. osteoclasts. The osseous partitions are remodelled by osteoclasts. The antler increases in diameter by periosteal apposition.     

Structure of the erosive lacunae on the periosteal and endosteal surfaces of bone

The periostal and endostal surfaces of the femur, ribs and lumbar vertebrae have been investigated in people at the age of 20-90 years. The erosive lacunae on the surface of the bone are revealed as recesses of various form and size. They are usually encountered as groups forming a resorptive focus, or fuse into large zones of resorption. Comparing the data of the light optic investigation and the results of SAM studies of the bone surface, it is possible to distinguish the erosive lacunae with an active process of resorption from the lacunae, where the process proceeds slowly or is not yet fully completed. On the periostal surface the resorptive foci are revealed more often, while on the endostal surface--large zones of resorption are seen. The erosive lacunae are often situated in the ostial parts of the vascular canals. No age dependence is noted in morphological manifestation of the resorption process on the investigated osseous surfaces of a mature person.     

Developmental reorganization of the skeletal framework and its surface lamina in fusing muscle cells

The skeletal framework of cells, composed of internal structural fibers, microtrabeculae, and the surface lamina, is revealed with great clarity after extraction with detergent. When muscle cells fuse to form a multinucleated myotube, their skeletal framework reorganizes extensively. When myoblasts prepare to fuse, the previously continuous surface lamina develops numerous lacunae unique to this stage. The retention of iodinated surface proteins suggests that the lacunae are not formed by the extraction of lamina proteins. The lacunae appear to correspond to extensive patches that do not bind concanavalin A and are probably regions of lipid bilayer devoid of glycoproteins. The lacunae appear to be related to fusion and disappear rapidly after the multinucleated myotube is formed. When muscle cells fuse, their internal structural networks must interconnect to form the framework of the myotube. Transmission electron microscopy of skeletal framework whole mounts shows that proliferating myoblasts have well developed and highly interconnected internal networks. Immediately before fusion, these networks are extensively reorganized and destabilized. After fusion, a stable, extensively cross-linked internal structure is reformed, but with a morphology characteristic of the myotube. Muscle cells therefore undergo extensive reorganization both on the surface and internally at the time of fusion.     

Initial stages of tunic morphogenesis in the ascidian Halocynthia: a fine structural study

Tunic morphogenesis in embryos of the ascidian Halocynthia roretzi was examined by scanning and transmission electron microscopy. For this purpose it was necessary to modify the classical embedding procedure. Soon after reaching the initial tail-bud stage, tunic deposition is initiated on the dorsal side of the embryo. As soon as the embryo is completely covered by the tunic, larval fins are formed. The test cells settle onto the embryo. At this stage only the outer cuticle and the outer tunic compartment have appeared. Tunic morphogenesis is accompanied by ultrastructural modifications of the epidermis characteristic of secreting cells. Cytochemical investigations reveal polysaccharide glycogen-like material in the lumen of epidermal lacunae and in the outer compartment of the tunic. Our observations strongly suggest that this material is stored in the lacunae and discharged into the outer compartment. The significance of fluffy osmiophilic material that appears at the early tail-bud stage and enlaces the whole embryo is discussed.     

Osteocyte lacuna size and shape in women with and without osteoporotic fracture

Osteocytes have been hypothesized to control the amount and location of bone tissue which is resorbed or formed, based on the strain magnitude they perceive, and therefore may play a role in the bone loss of osteoporosis. The shape of osteocyte lacunae influences the mechanical strain applied to the osteocyte; thus, it is important to quantify their shape to further understand the mechanical environment of this cell. Previous studies of the size and shape of lacunae have been contradictory and limited to two-dimensional measurements on iliac crest biopsies. This investigation measured the size and shape of osteocyte lacunae in trabecular bone near a typical fracture site from three-dimensional image sets obtained by confocal microscopy. Bone tissue specimens were obtained from individuals undergoing hip replacement subsequent to fracture, and matched cadaveric specimens without fracture. After extensive image processing to differentiate the lacunae from the matrix, the volume and anisotropy of the lacuna were determined. No significant difference was found in the size (volume) or shape (anisotropy) of the lacunae between women with and without osteoporotic fracture, although there was a large range of sizes and shapes in both groups. These results suggest that the size or shape of the lacunae, which influences the strain in osteocytes, does not play a role in osteoporotic fracture. In addition, this study provides geometric measures of lacunae that are important in computational modeling of the mechanical environment of osteocytes.     

Organization and differentiation of the segmentary plate in common toad embryo Bufo bufo L., following heat shock (author's transl)

A heat shock greatly disturbs the shaping and the growth of embryos with 4 or 5 somites. One part of a segmentary plate with a size including four control somites stays inorganized, whereas posterior segmentation gets on normally. The differentiation of the unsegmented somitic block (fusing myoblasts, myogenesis) is the same as the one of the previous somites; the dermatome and the sclerotome issued from the unsegmented block are not cut up. Ultrastructural observations of treated segmentary plates show that the somitic block undergoes important changes. The non-segmented somitic mesoderm of the control animals obtains a loose aspect; its constitutive cells separated by lacunae are brought into contact by the means of filopods and junctions. 18 hours after the heat shock, the cells of the unsegmented block keep close to each other and are tightened together by many "pseudo-desmosomes", the intercellular spaces are reduced to small meatus. The nuclei of the somitic cells are clear and provided with a restricted amount of chromatine spots; dark granular nuclear like chromatin material is thrown out of the nucleus. In embryos fixed 40 hours after heat shock, several large plurinucleated cells, are formed in the non-segmented somitic block. Inside these plurinucleated cells, there is synthesis of myofilaments and differentiation of myofibrilles. Numerous intercellular lacunae are formed, some yet apparent "pseudo-desmosomes" form "pinchings" between the membranes of muscular cells and therefore confine series of intercellular lacunae.     

Morpho-functional correlations of the structure of bone cells and adjoining bone matrix in the developing bone

By means of scanning and transmissive electron microscopy methods structure of the developing bone has been studied. Interconnection of the cell structure and spatial organization of the adjoining matrix has been demonstrated. On the surface of the growing bone not only forming areas have been revealed, where under osteoblasts at various functional states, osteoid layer is determined, but also areas of resorption and completed osteogenesis. This demonstrates an interrupted character of osteogenesis at modelling. At the same time for the remodelling process presence of erosive lacunae is specific; they are filled with a newly deposited collagenous matrix. Therefore, it is possible to suppose that formation of the bone as an organ during the postnatal development includes in itself both mechanisms supporting its form at outgrowth of the osseous matrix volume (modeling) and its continuous rearrangement and adaptation to real conditions of functioning (remodelling).     

Fine structure of the spermatozoa of Chiton marginatus (mollusca: Amphineura), with special reference to nucleus maturation

The spermatozoon of Chiton marginatus is a long uniflagellate cell displaying structural features of “modified sperm.” The nucleus presents a conical shape with a long apical cylindrical extension. The chromatin is homogeneously dense. Scattered inside the condensed nucleus, a few nuclear lacunae are visible. The acrosomal complex is lacking. Some mitochondria are located in a laterofrontal structure side by side with the nucleus. The typical midpiece is absent. The cytoplasm forms a thin layer around the nucleus and the mitochondria. The proximal centriole is in a basal nuclear indent. The distal centriole serves to form the axoneme tail with the usual microtubular pattern. During nuclear maturation, the early spermatid nucleus is spherical and contains fine granular chromatin patches. The nuclear envelope shows a deposit of dense material at the base of the nucleus, forming a semicircular invagination occupied by a flocculent mass. In middle spermatid stage, the chromatin gets organized in filaments, coiled as a hank, attached over the inner surface of the basal thickening of the nuclear envelope. The nucleus starts to elongate anteroposteriorly. At the pointed apical portion of the spermatid, a group of microtubules is observed seeming to impose external pressure to the nucleus giving rise to the long apical nuclear point. The mitochondria have a basal position. Late spermatids have an elongated conical nucleus. The chromatin filaments are further condensed, and lacunae appear inside the nucleus. Some mitochondria migrate to a lateral position.     

Structure of the osseous callus formed under electric stimulation. An experimental histological study

The study tried to evidence histologically, histometrically and mechanically the comparative qualities of the osseous callus experimentally formed in dogs and rabbits under the influence of an electric stimulation. The materials were decalcified or polished, and the collagen network and the Haversian systems analysed in the callus. The tearing strength in kg-power on the surface unity of the osseous tissue was measured using the EDZ 20 apparatus. The results indicated that the osseous tissue formed under electric stimulation had a structure and a tearing strength similar to that of controls, and that the consolidation time of fractures is reducing.     

The ultrafine structure of the body wall of sexually mature thorny-headed worms Echinorhynchus gadi (Acanthocephala)]

Studies of the fine structure of the adult acanthocephalan Echinorhynchus gadi have given a new information on the structure and organization of the body wall of these parasitic helminths. Their body surface is covered by glycocalyx of mucopolysaccharide nature. Just under it there is the surface membrane which has numerous invaginations forming a network of branching canals from which membrane vesicles are isolating. In their turn these canals pass through "the cytoplasmic canals" of the cortical matrix. Between the surface membrane and cortical matrix there is the base plate. These three structures form the striped layer underlain by the felt layer. It is formed by three layers of fibrous strands (one circular and two longitudinal), which are parallel to the body surface. These strands consist of loosely laid fibrils. The lowest layer is a radial one which occupies 2/3 of the body wall. It consists of the radial strands beginning from the cortical matrix and ending at the basement membrane. Numerous lipid droplets and glycogen granules are formed here. Two types of fibrils with 0.26 and 0.05 diameter have been detected for the first time. The radial layer in the cytoplasm was found to have crystalline structures and polymembrane bodies, numerous nuclei with light karyoplasm and distinct nucleoli. The location of the nuclei is of two types: either in the cytoplasm or in the "lacunae". We have shown that the "lacunae" are specialized sites of the cytoplasm whose boundaries are marked by the fibres of two types. Besides, this type of the acanthocephalan was found to have two "giant lacunae" extending along the body.(ABSTRACT TRUNCATED AT 250 WORDS)     

A histochemical localization on Maclura pomifera lectin during osteogenesis

Summary Mandibular condyles of 4-week-old Wistar strain rats and mandibles of ICR strain mice from 14 days gestation stage to 2 days postnatal stage were used to investigate the localization of Maclura pomifera lectin (MPA) during two modes of osteogenesis. During endochondral ossification of the mandibular condyle, MPA was only localized at the peripheral regions of calcified cartilage after the destruction of chondrocyte lacunae. Bone extracellular matrix (ECM) was not reacted with MPA. In intramembranous ossification of mice mandibles, MPA was stained intensively in the early bone ECM. The intensity of the MPA reaction decreased during bone development. In both cases of osteogenesis, chondroclasts and osteoclasts showed the strong affinity to MPA. These results indicated that the time- and position-specific changes within ECM proceeded during osteogenesis and that MPA was the useful probe to detect chondroclasts and osteoclasts.     

Studies on formation and resorption of fish scales

Summary Scale formation in Cyprinodon variegatus was found to be initiated at about 26 to 30 days after hatching. Ultrastructural investigation revealed that within 4 to 6 h in the first-formed scales the marginal cells begin to flatten and differentiate into osteogenic cells, which later change to osteoblasts and fibroblasts. These cells are separated from the surrounding epithelial cells by a basal lamina. The osteoid is formed by the marginal and osteogenic cells; the osseous layer by the osteoblasts; and the fibrillary plate by the fibroblasts.The osteoid is formed within 2 to 3 h after the initiation of the scale, and within 20 to 24 h the osseous layer is formed. Hydroxyapatite crystals are deposited in the matrix of the osseous layer without apparent association with collagen fibers. No matrix vesicles or dense bodies are evident at the sites of calcification. The fibrillary plate arises 18 to 20 h after the initiation of the scale. It is also partially calcified, but not before the third week of scale formation. The crystals develop almost exclusively between the collagen fibers at the extreme edge of the calcifying front, but solid calcification of the fibers results with further growth of the crystals. The fibroblasts appear to participate in calcification of the fibrillary plate.Contribution No. 332, Belle W. Baruch Institute for Marine Biology and Coastal Research, University of South Carolina, Columbia, South Carolina, 29208, USA     

Lacunal gas discharge as a measure of productivity in the seagrasses Zostera capricorni,Cymodocea serrulata and Syringodium isoetifolium

A new method for estimating the rate of photosynthetic fixation of carbon in seagrasses is described. This method, which is sensitive and simple to apply, is based on the close relationship between photosynthetic rate and the volume of gas discharged through the lacunae.During photosynthesis, the gas discharged from the lacunae was composed primarily of oxygen (32.5%) and nitrogen (67.5%). The rate of discharge was proportional to the rate of photosynthesis and hence was a function of light intensity. There was a linear relationship between gases discharged from the lacunae and oxygen released into the water column. Calibration curves were derived relating volume of gases (oxygen and nitrogen) released from the lacunae to total oxygen produced during photosynthesis for three species of seagrasses (Zostera capricorni Aschers., Cymodocea serrulata (R. Br.) Aschers. & Magnus and Syringodium isoetifolium (Aschers.) Dandy). Molar ratios of ¹⁴C fixed to oxygen produced were close to unity (1.008±0.016; n=8) indicating that measurements of lacunal gas released may be used to measure productivity.     

A histochemical localization on Maclura pomifera lectin during osteogenesis

Mandibular condyles of 4-week-old Wistar strain rats and mandibles of ICR strain mice from 14 days gestation stage to 2 days postnatal stage were used to investigate the localization of Maclura pomifera lectin (MPA) during two modes of osteogenesis. During endochondral ossification of the mandibular condyle, MPA was only localized at the peripheral regions of calcified cartilage after the destruction of chondrocyte lacunae. Bone extracellular matrix (ECM) was not reacted with MPA. In intramembranous ossification of mice mandibles, MPA was stained intensively in the early bone ECM. The intensity of the MPA reaction decreased during bone development. In both cases of osteogenesis, chondroclasts and osteoclasts showed the strong affinity to MPA. These results indicated that the time- and position-specific changes within ECM proceeded during osteogenesis and that MPA was the useful probe to detect chondroclasts and osteoclasts.     

Influence of metal ion solutions on rabbit osteoclast activities in vitro

The purpose of the present study was to compare the effects of various metal ions (aluminium, chromium, cobalt, gold, iron, strontium, titanium and vanadium) on rabbit osteoclast activities, with respect to their number, size, resorptive capacity and their capacity to release proteinases. Marked heterogeneous osteoclastic behaviour was observed early in culture with metal ions (24 h) in term of resorption parameters. In contrast, protease activities (cysteine-proteinase and metalloproteinase activities) were not modulated in our culture conditions. Aluminium, iron, gold and titanium reduced the number of osteoclasts significantly. Aluminium and gold had no effect on osteoclast-mediated resorption on dentin-slices, although aluminium induced a greater number of very small lacunae. Titanium reduced only the mean surface area per lacunae, cobalt reduced the mean surface area of lacunae and increased their number, and iron reduced both parameters. Strontium had no effect on osteoclast formation and on total dentin slice surface resorbed. However, strontium increased the number of small lacunae formed on dentin-slices by osteoclasts. Chromium had no effect on osteoclast activities. These findings indicate that metal ions induce very early effects on osteoclasts, which can contribute to periprosthetic pathologies via different cellular mechanisms.     

Fine structure of the respiratory lamellae of teleostean gills

Summary The blood-water pathway in respiratory lamellae of teleostean gills consists of an epithelial layer one or two cells thick, a basal lamina and a thin layer of cytoplasm which lines the blood lacunae. This layer of cytoplasm is formed by flange-like extensions of the pillar cells. The resulting location of the pillar cell perikarya between the surfaces of the blood lacunae is probably of paramount importance for maintenance of the flattened form of the lamellae.Collagenous bundles traverse the pillar cells within tubes formed by infolding of the cellular surface. These bundles, which are oriented normal to the flattened aspect of the lamellae, no doubt provide further protection against distension or collapse of the blood spaces. A compartment filled with collagenous tissue is interposed between the basal lamina and the lining layer of the lacunae in some of the species studied.Regulation of blood flow to the respiratory surfaces is thought to result in part from contraction of the pillar cells. This contractility presumably resides in tracts of filaments which course through the cytoplasm of the pillar cells parallel to the collagenous bundles. Since nervous tissue has not been demonstrated within the gill lamellae it is possible that contraction of the pillar cells is under some form of hormonal control, although existence of local control mechanisms (e.g. self-stimulation of the cells as a result of anoxia) is not excluded.Within the limited number of species studied, the structure of the blood-water pathway does not appear to be correlated with the characteristics of the normal habitat of a particular species.This work was performed during the tenure of a post doctoral traineeship under USPHS Grant 5 T 1 GM-136 to the Department of Biological Structure, University of Washington.Particular thanks are due Dr. John H. Luft of the University of Washington for his advice and criticism while this work was in progress and to Drs. Douglas Kelly, James Koehler and Daniel Szollosi for critical assistance with the manuscipt.     

Developmental morphology of the neonatal alligator (Alligator mississippiensis) ovary

American alligator (Alligator mississippiensis) ovary development is incomplete at hatching. During the months following hatching, the cortical processes of oogenesis started in ovo continues and folliculogenesis is initiated. Additionally, the medullary region of the gonad undergoes dramatic restructuring. We describe alligator ovarian histology at hatching, 1 week, 1 month, and 3 months of age in order to characterize the timing of morphological development and compare these findings to chicken ovary development. At hatching, the ovarian cortex presents a germinal epithelium containing oogonia and a few primary oocytes irregularly scattered between somatic epithelial cells. The hatchling medulla shows fragmentation indicative of the formation of lacunae. By 1 week of age, oocytes form growing nests and show increased interactions with somatic cells, indicative of the initiation of folliculogenesis. Medullary lacunae increase in diameter and contain secretory material in their lumen. At 1 month, nest sizes and lacunar diameters continue to enlarge. Pachytene oocytes surrounded by somatic cells are more frequent. Trabeculae composed of dense irregular connective tissue divide cortical nests. Three months after hatching oocytes in meiotic stages of prophase I up to diplotene are present. The ovary displays many enlarged follicles with oocytes in diplotene arrest, thecal layers, lampbrush chromosomes, and complete layers of follicular cells. The medulla is an elaborated complex of vascularized lacunae underlying the cortex and often containing discrete lymphoid aggregates. While the general morphology of the alligator ovary is similar to that of the chicken ovary, the progression of oogenesis and folliculogenesis around hatching is notably slower in alligators. Diplotene oocytes are observed at hatching in chickens, but not until 3 months in alligators. Folliculogenesis is completed at 3 weeks in chickens whereas it is still progressing at 3 months in alligators.     

Localization of osteopontin in resorption lacunae formed by osteoclast-like cells: a study by a novel monoclonal antibody which recognizes rat osteopontin

The characteristics of a monoclonal antibody produced against osteoclast-like multinucleated cells (MNCs) formed in rat bone marrow cultures were examined immunohistochemically and biochemically. The in vitro immunization was performed using as immunogen the MNCs from rat bone marrow cell culture, which revealed many characteristics of osteoclasts. After screening and cloning of hybridomas, the monoclonal antibody HOK 1 was obtained. This antibody reacted weakly with stromal cells and intensely with both MNCs and their putative migratory traces on culture dishes. Immunofluorescent examination of paraffin sections revealed intense reactivity on the epithelium of the choroid plexus, the ileum and the proximal-convoluted tubules of the kidney, and also on bone cells such as osteocytes, osteoblasts, and osteoclasts. Western blotting using purified rat osteopontin verified that the antigen recognized by HOK 1 was osteopontin. Positive HOK 1 immunoreactivity was further observed in the resorption lacunae formed by a culture of MNCs on human tooth slices and on the surface of osteoclasts. The present data suggested that osteopontin is preferentially present on the resorption lacunae in resorbing calcified matrices and that osteoclasts under a specific state might trap this protein on their cell surface.