Amorphous Ground Substance Research Articles

Contractile Smooth Muscle and Active Stress Generation in Porcine Common Carotids.

The mechanical response of intact blood vessels to applied loads can be delineated into passive and active components using an isometric decomposition approach. Whereas the passive response is due predominantly to the extracellular matrix (ECM) proteins and amorphous ground substance, the active response depends on the presence of smooth muscle cells (SMCs) and the contractile machinery activated within those cells. To better understand determinants of active stress generation within the vascular wall, we subjected porcine common carotid arteries (CCAs) to biaxial inflation-extension testing under maximally contracted or passive SMC conditions and semiquantitatively measured two known markers of the contractile SMC phenotype: smoothelin and smooth muscle-myosin heavy chain (SM-MHC). Using isometric decomposition and established constitutive models, an intuitive but novel correlation between the magnitude of active stress generation and the relative abundance of smoothelin and SM-MHC emerged. Our results reiterate the importance of stretch-dependent active stress generation to the total mechanical response. Overall these findings can be used to decouple the mechanical contribution of SMCs from the ECM and is therefore a powerful tool in the analysis of disease states and potential therapies where both constituent are altered.

Amyloidosis and Anesthesia.

Aim:The aim of this article is to provide a view of amyloidosis and discuss implications for the anesthetic management of patients with this condition.Material and Method:Urine samples from patients with plasma cell dyscrasias were obtained from a urine bank that gathers urine samples from patients who gave research use consent for specimens that would otherwise be considered waste.Results:Patients with amyloidosis may present to the anesthesiologist for procedures relating to diagnosis, surgery relating to the underlying condition (e.g., bronchial laser and organ transplant), or for incidental surgery. The condition carries a significant risk of perioperative morbidity and mortality.Conclusion:The term amyloid was coined by Virchow in the mid 19th century, meaning “starch like.” Amyloidosis is a disease complex, in which there is an abnormal deposition of extracellular hyaline material with particular staining characteristics and which contains protein fibrils embedded in a relatively amorphous ground substance. There are numerous clinical manifestations, the onset is insidious, and the diagnosis may not be made in a patient undergoing anesthesia and surgery for an apparently straightforward problem. Unexpected complications such as heart or kidney failure may arise, either before operation or in the postoperative period. Bullous lesions of the skin or oral mucosa and extensive areas of purpura are but two of the ways, in which amyloidosis may first present. The disease spectrum may be inherited or acquired, localized or systemic, and life threatening or an incidental finding.

Structure of the nucleoli in Angiosperms

The nucleolus structure in the cells of numerous <i>Angiospermae</i> plants was studied with light microscope. The living material was examined under phase and anoptral contrast. Microscopical observations were also carried out on the material fixed in chrome-formalin or in uranyl-formalin. An occurrence of two structural components of nucleolus was demonstrated: an amorphous ground substance and nucleolonema. Cytochemical analysis have shown that considerable amounts of RNA and acid proteins were located in the amorphous substance while nucleolonema was composed of basic proteins and slight quantities of RNA.

Tissue and microstructural deformations in aortic tissue under stretch and after deformation recovery

Elucidating how cardiovascular biomechanics is regulated during health and disease is critical for developing diagnostic and therapeutic methods. The extracellular matrix of cardiovascular tissue is composed of multiple fibrillar networks embedded in an amorphous ground substance and has been found to reveal time-dependent mechanical behavior. Given the multiscale nature of tissue biomechanics, an accurate description of cardiovascular biomechanics can be obtained only when microstructural morphology is characterized and put together in correlation with tissue-scale mechanics. In this study, we sought to determine how the microstructural configuration in aortic tissue changes with tissue-scale loading protocols. At the tissue level, rather intuitively, the width and thickness decrease when specimens undergo elongation and it is found that there is a higher inverse correlation between the elongation of circumferential specimens and the shrinkage along the thickness than along the width. After deformation recovery, the specimens show levels of permanent deformation in both thickness and width as neither recovers the initial values for the unstretched specimen, with higher permanent deformation measured for thickness. At the microstructural level, the networks in the wall inner layer show straighter fibrillar structure under stretch, which partly returns to the crimping structure after deformation recovery. This study indicates a microstructural basis for observations of regional permanent tissue stretch in artery tissues, and furnishes early data for developing multiscale models of cardiovascular viscoelastic/viscoplastic modeling.

Histologie de la peau et des follicules pileux

The skin consists of an outer epidermis, the dermis, and the hypodermis. It includes nerves, blood vessels, glands and hair follicles. Epidermis is a continually renewing, stratified squamous epithelium. It is populated by keratinocytes (80 %) and dendritic cells (20 %) : melanocytes, Langerhans and Merkel cells. In standard histology, keratinocytes are arranged in layers that represent different stages of their differentiation while melanocytes and Langerhans cells appear as clear cells respectively between the basal and the supra-basal cells of epidermis. The Merkel cells cannot be clearly identified. Dendritic processes of the dendritic cells can only be recognized by immunocytochemistry. At the dermal-epidermal junction, a PAS reactive basement membrane follows the contour of the basal cells. Dermis consists of collagenous and elastic fibers embedded into an amorphous ground substance. Fibroblasts, macrophages, mast cells and lymphocytes are its resident cells. Hypodermis is composed of adipocyte lobules defined by fibrous connective tissue septa. Hair follicle consists of 3 parts : the lower portion, from the base of the follicle including hair bulb to the insertion of the arrector pili muscle or buldge ; the isthmus, from the insertion of the arrector pili to the entrance of the sebaceous duct, and the infundibulum, from the entrance of the sebaceous duct to the follicular orifice. The lower portion is composed of the dermal hair papilla, the hair matrix, the hair, and the inner and the outer root sheaths. The hair matrix cells within hair bulb give rise to the hair and to the inner root sheath. With the electron microscope, one can obtain a more detailed view of the characteristic skin structures. Much of them can now be explained in terms of function and in many instances, in correlation with its biochemical composition. An attempt has been made in this paper to precisely give the location of molecules that are relevant in basic skin functions and understanding of auto-immune and genetic diseases.

Comparative Anatomy of Laboratory Animal Corneas with a New-Generation High-Resolution In Vivo Confocal Microscope

Purpose: The aim of the current study was to compare the corneas of three commonly used laboratory animals with a new in vivo confocal microscope. Methods: Six eyes of three adult male New Zealand albino rabbits, six eyes of three adult male Lewis rats, and six eyes of three adult male Swiss mice were used in this study. Corneas were analyzed in vivo using the Rostock Cornea Module of the Heidelberg Retina Tomograph (HRT)-II. For all eyes, 20 confocal microscopic images of each layer, that is, the superficial and basal corneal epithelia, the Bowman layer, the anterior and posterior stroma, and the endothelium, were recorded. The images were then analyzed qualitatively and compared among animals. Cellular densities of anterior and posterior stroma keratocytes of rabbits and endothelium density of the three different animals were also measured and compared. Results: The Rostock Cornea Module of the HRT II was successfully used to analyze all corneal layers of these three commonly used laboratory animals. Although the cellular patterns of the corneal layers of these three animals, as observed with in vivo confocal microscopy, were quite similar, some differences were seen in terms of endothelial cell density and stroma appearance. Superficial cells were seen as hyper- and hyporeflective polygonal cells. Basal cells had dark cytoplasm without visible nuclei and were closely organized. A Bowman layer was observed in all three animals as an amorphous tissue containing fine subepithelial nerve plexus. In rabbits, the stroma consisted of an amorphous ground substance with hyper-reflective structures corresponding with keratocyte nuclei. In rats and mice, numerous reflective stellate structures with no clearly visible nuclei were observed within the stroma. Besides endothelial cell density, the endothelium was similar among the three animals and was seen as hyper-reflective cells with dark limits organized in a honeycomb pattern. Conclusions: The Rostock Cornea Module of the HRT II can provide high-resolution images of all corneal layers of rabbits, rats, and mice without sacrificing animals or preparing tissue. This new device may be useful for evaluating the cornea during experimental animal studies.

Defective bone remodelling in osteoprotegerin-deficient mice.

Previous studies have reported enhanced osteoclastogenesis, increased bone resorption and osteoporosis in osteoprotegerin (OPG)-deficient mice. In the present study, we show that the tibial epiphyses contain abundant, thin trabeculae lined with numerous osteoclasts and cuboidal osteoblasts. The increase in osteoblasts and osteoclasts was associated with a dramatic increase in calcein labelling of the mineralization fronts and replacement of much of the intertrabecular marrow with numerous alkaline phosphatase-positive preosteoblasts. Furthermore, the discrete, linear cement lines seen in wild-type mice were replaced by a randomly oriented meshwork of cement lines that were stained intensely for tartrate-resistant acid phosphatase and osteopontin in the OPG-/- mice. These indices of accelerated bone remodelling in mutant bone were associated with irregular trabecular surfaces, a disorganized collagen matrix interspersed with amorphous ground substance and numerous fissures between old and new bone. In total, these observations indicate that enhanced osteoclastic activity in OPG-/- epiphyses led to a coupled increase in osteoblast differentiation and activity and an increase in bone remodelling. The high bone turnover, disorganized matrix and impaired attachment of new to old bone in the cement lines in OPG-/- mice appear to cause bone fragility.

Age changes in the cells of the intra-articular disc of the temporomandibular joints of rats and marmosets

Cells in the intra-articular disc of the temporomanadibular joint were studied ultrastructurally at three different ages to investigate any age changes. Rats aged 2, 15.5 months, and 2.5 years, and marmosets aged 21 months, 7 years, and between 10.5 and 14 years were studied. In the first two age groups of the rat and the first of the marmoset, the cells were generally rounded and had moderate amounts of rough endoplasmic reticulum and other organelles associated with protein synthesis and secretion. Many cells had conspicuous amounts of microfilamentous material and cell membranes were closely applied to the collagen fibrils of the extracellular matrix. Occasionally, a narrow, irregular space containing microfilamentous material lay adjacent to the cell membrane. In the 2.5-year-old rats and the two older age groups of marmosets, cells with chondrocyte-like morphology were present. These cells were surrounded by a conspicuous pericellular matrix devoid of collagen fibrils and composed of microfilamentous material embedded in an amorphous ground substance. They resembled cells described in fibrocartilage from other sites, but differed from chondrocytes in hyaline cartilage by lacking a pericellular capsule. Thus, rats and marmosets both show cellular age changes in the intra-articular disc of the mandibular joint, which can be considered as changing from fibrous to fibrocartilaginous with age, a condition similar to that reported in humans.

Anorexia Nervosa with Pancytopenia

Anorexia Nervosa with Pancytopenia

Ultrastructural and histochemical features of the ground substance in cyclosporin A-induced gingival overgrowth.

The overgrowth-affected gingiva of patients treated with cyclosporin A after kidney transplant was examined with ultrastructural and histochemical methods to evaluate the involvement of connective tissue. Gingival overgrowth has the same clinical signs as local edema. The ultrastructural study showed that the dimensional increase was largely due to increased production of amorphous ground substance by fibroblasts, possibly resulting from an increased release of histamine by mast cells. The histochemical data revealed that the affected tissues contained higher levels of glycosaminoglycans and that cyclosporin A induced comparably high levels of glycosaminoglycans in in vitro cultures of fibroblasts obtained from normal gingiva. The combination of ultrastructural and histochemical data, therefore, strongly suggests that the response of the connective tissue in gingival overgrowth cannot be ignored and may be the main cause of the observed pathological condition.

Tensile strength and histological changes of abdominal aorta of malnourished rats

The resistance of the abdominal aorta of rats after 6, 7 and 8 weeks of malnutrition, compared with control animals, was evaluated by longitudinal tensiometry. Weakness of this vessel in malnourished rats was demonstrated; microscopic examination of the aorta stained by Masson, Calleja and hematoxylin-eosin methods showed a decrease in amorphous ground substance and an increase in the width of elastic laminae. There was no visible alteration either in the endothelial lining layer or in the smooth muscle fibers. Such alterations of the aorta are, to the authors' knowledge, the first reported modifications in the peripheral vasculature after malnutrition.

Immunohistochemical localization of keratan sulfate and chondroitin 4- and 6- sulfate proteoglycans in subregions of the tectorial and basilar membranes

Proteoglycans containing keratan sulfate (KSPG) and 4- and 6-sulfated epitopes of chondroitin sulfate (CSPG) were identified in distinct domains of the tectorial and basilar membranes by ultrastructural immunogold labeling with monoclonal antibodies. In the tectorial membrane (TM), the highest concentration of gold particles was present in the upper fibrous layers of the limbal, middle and marginal zones with all three antibodies. Reactivity with anti-KSPG exceeded that with anti-4S and anti-6S CSPG, especially in the marginal zone. The cover net showed no reactivity for any antibody. Labeling density of gold particles with all three antibodies increased markedly from base to apex. In the basilar membrane (BM), all three PGs were most highly concentrated in regions of amorphous ground substance bordering the upper and lower filamentous bands. As in the TM, reactivity for anti-KSPG in the BM exceeded that for either CSPG antibody and staining with all three antibodies was stronger and more widespread in the apical as compared to the basal turns. These results provide the first ultrastructural demonstration of KSPG and CSPG in distinct subregions of the TM and BM. The preferential distribution and marked increase in PGs from base to apex in both TM and BM supports a role for these macromolecules in regulating structural and mechanical properties of these highly specialized extracellular membranes.

Semi-Serial Electron-Micrographic Reconstruction of Putative Transducer Sites in Pacinian Corpuscles

The Pacinian corpuscle (PC) is composed of an afferent neurite surrounded by an accessory capsule formed by concentric layers of lamellae. Projecting from the neurite, which is elliptical in cross-section, are "filopodia" or axonal, spike-like extensions. These filopodia are the putative sites of transduction. It has been proposed that two populations of filopodia organized in morphofunctional opposition exist, and that this arrangement is responsible for the bidirectional sensitivity of PCs as seen in receptor potential recordings. In order to explore this possibility, PCs obtained from cat mesentery were processed for electron microscopy, and semiserial reconstructions were made. We evaluated the extensions' (n > 110) locations, inclusions, shapes, and sizes. The filopodia were found to project along the major elliptical axis of the neurite, their density being approximately 2.8 per micron. The filopodia were found to contain filaments, vesicles, and amorphous ground substance, and dense accumulations of mitochondria were found at their bases. Measurements of their size (i.e., length, width, and height) suggest that there are two different types of filopodia. No other obvious relations among filopodial type, location along the neurite, and landmarks for transduction were found. The presence of the two filopodial types may be the basis for the bidirectional sensitivity of the PC.

Dentine proteoglycans: composition, ultrastructure and functions

Proteoglycans (PGs) have been visualized in the predentine and dentine with cationic dyes by staining thin sections with Alcian Blue, bismuth nitrate, or using Spicer's high-iron diamine (HID) method. The precise location may be obtained by adding cationic dyes such as Cuprolinic Blue, ruthenium hexamine trichloride or cationic detergent (cetylpyridinium chloride) to the fixative. These methods induced the formation of aggregates which varied in shape and number according to the method used. Rapid freezing followed by freeze-substitution revealed an amorphous ground substance, homogeneously stained with Alcian Blue, located in the predentine between the collagen fibres. These PGs may be involved in transport and diffusion in predentine. In dentine, small granules and needle-like structures were observed along the collagen fibres. This second group of PGs differs in composition, distribution and functions from the predentine PGs. The same distribution was seen when hyaluronidase-gold labelling was used. Labelling with antibodies and autoradiography also gave evidence of two distinct groups of PGs. In predentine, as an hydrated gel, PGs seems to act as mineral inhibitors, whereas immobilized on a surface, as seen at the dentine edge, they act as nucleating agents. The interaction between PGs and phospholipids seems also to play a role in the mineralization process.

Hormonal control of the cervix in pregnant gilts. IV. Relaxin promotes changes in the histological characteristics of the cervix that are associated with cervical softening during late pregnancy in gilts.

We recently described relaxin-induced changes that occur in both the physical properties and the biochemical composition of the uterine and vaginal portions of the cervix during the last third of gestation in the gilt. This study employed morphometric analysis to examine both the changes that occur in the histological characteristics of the uterine and vaginal portions of the cervix between days 80 and 110 of pregnancy in intact gilts and the effects of relaxin on these changes in ovariectomized gilts given progesterone to maintain pregnancy. There were four treatment groups: intact day 80 control gilts, sham-ovariectomized day 110 control gilts, ovariectomized progesterone-treated day 110 gilts, and ovariectomized progesterone- plus relaxin-treated day 110 gilts. The histological characteristics of the uterine portion of cervices obtained from intact controls on day 110 differed markedly from those of intact controls on day 80; there was a reduction in the density and organization of collagen fiber bundles, a reduction in the density of smooth muscle fiber bundles, and an increase in amorphous ground substance. The histological characteristics of cervices removed on day 110 from ovariectomized gilts given progesterone only did not differ from those of day 80 controls. After replacement therapy with progesterone plus relaxin, however, the histological characteristics of the cervix on day 110 did not differ from those in intact day 110 controls. Additionally, this study demonstrates that the relaxin-dependent changes are more dramatic in the uterine than in the vaginal portion of the cervix. These findings are consistent with and extend our earlier findings that relaxin-dependent changes in the physical properties and biochemical composition of the uterine portion of the cervix far exceed those in the vaginal portion of the cervix during late pregnancy in the gilt. We conclude that the relaxin-dependent changes in the histological characteristics of the cervix described in this report may contribute at least in part to the cervical softening that occurs during late pregnancy in gilts.

Hormonal control of the cervix in pregnant gilts. IV. Relaxin promotes changes in the histological characteristics of the cervix that are associated with cervical softening during late pregnancy in gilts

We recently described relaxin-induced changes that occur in both the physical properties and the biochemical composition of the uterine and vaginal portions of the cervix during the last third of gestation in the gilt. This study employed morphometric analysis to examine both the changes that occur in the histological characteristics of the uterine and vaginal portions of the cervix between days 80 and 110 of pregnancy in intact gilts and the effects of relaxin on these changes in ovariectomized gilts given progesterone to maintain pregnancy. There were four treatment groups: intact day 80 control gilts, sham-ovariectomized day 110 control gilts, ovariectomized progesterone-treated day 110 gilts, and ovariectomized progesterone- plus relaxin-treated day 110 gilts. The histological characteristics of the uterine portion of cervices obtained from intact controls on day 110 differed markedly from those of intact controls on day 80; there was a reduction in the density and organization of collagen fiber bundles, a reduction in the density of smooth muscle fiber bundles, and an increase in amorphous ground substance. The histological characteristics of cervices removed on day 110 from ovariectomized gilts given progesterone only did not differ from those of day 80 controls. After replacement therapy with progesterone plus relaxin, however, the histological characteristics of the cervix on day 110 did not differ from those in intact day 110 controls. Additionally, this study demonstrates that the relaxin-dependent changes are more dramatic in the uterine than in the vaginal portion of the cervix. These findings are consistent with and extend our earlier findings that relaxin-dependent changes in the physical properties and biochemical composition of the uterine portion of the cervix far exceed those in the vaginal portion of the cervix during late pregnancy in the gilt. We conclude that the relaxin-dependent changes in the histological characteristics of the cervix described in this report may contribute at least in part to the cervical softening that occurs during late pregnancy in gilts.

Electron microscopic studies on reticular fibers in the pig sheathed artery and splenic cords.

The reticular fibers of the capillary sheath and splenic cord were studied in the pig spleen by transmission electron microscopy to reveal their components and the presence of sialic acid in the amorphous ground substance. Collagen fibrils, elastic fibers, microfibrils, nerve fibers and smooth muscle cells were observed in the reticular fibers of the splenic cord, while only microfibrils were recognized in the reticular fibers of the capillary sheath. The binding of LFA lectin to the splenic cord was stronger than the capillary sheath. These findings may suggest that the reticular fibers of the splenic cord include numerous functional elements and perform an important role during contraction or dilation of the spleen. On the other hand, it seems to be reasonable to term the capillary sheath as "capillary basement membranous reticular tissue", since the reticular fiber of the capillary sheath resembles the basement membrane of the capillary in its components.

Monoclonal antibodies specific for rat relaxin. VII. Passive immunization with monoclonal antibodies throughout the second half of pregnancy prevents development of normal mammary nipple morphology and function in rats.

We recently demonstrated that relaxin-dependent development of the mammary nipples during the second half of pregnancy is required for pup survival during lactation in the rat. The two related objectives of this investigation were to 1) characterize the effects of endogenous relaxin on the histological modifications that normally occur in the mammary nipples, and 2) test the hypothesis that the cause of lactational failure in relaxin-deficient rats is attributable to failure of the nipples to grow and develop during the second half of pregnancy. Endogenous relaxin was neutralized by daily iv injection of a highly purified monoclonal antibody specific for rat relaxin (MCA1) to intact rats from days 12-22 of pregnancy. Mammary nipples were collected on day 22 of pregnancy and routinely prepared for light microscopy. Tissue cross-sections (6 microns) obtained from the midpoint of mammary nipples were stained with either Gomori's trichrome stain (to visualize collagen) or orcein (to visualize elastin). Nipple size as well as histological characteristics of nipple cross sections were determined by morphometric analysis. MCA1-treated rats were significantly different from controls with the following parameters: shorter length of the nipples; smaller cross-sectional areas of the entire nipple, lactiferous duct lumen, and blood vessels; greater percentage of the analysis field composed of collagen; lower percentage of the analysis field composed of amorphous ground substance; and longer elastin fibers. To test the hypothesis that the cause of lactational failure in relaxin-deficient rats is attributable to the failure of nipples to grow and develop, MCA1 and control rats were cesarean sectioned between 2100-2400 h on day 22 of pregnancy, and lactation was examined using normal foster pups from intact donor females. Unlike pups fostered to controls, pups fostered to MCA1-treated dams failed to grasp the nipples, stimulate PRL release, or have milk in their abdomens. This study demonstrates that endogenous relaxin promotes not only growth, but also modifications of the histological characteristics of the nipple that are consistent with relaxin's effects on the cervix and mammary glands. Additionally, this study provides evidence that lactational failure in relaxin-deficient rats is attributable to the small size and different histology of the mammary nipples, which results in the inability of the pups to attach to the nipple, stimulate PRL release, and obtain milk from the dams.

Supramolecular organization of extracellular matrix glycosaminoglycans, in vitro and in the tissues

Connective tissues maintain shape against external and internal stress. They are molecular hierarchies in which fundamental building units come together in tiers of increasing complexity and mutual interactions, based on information carried in the precursor molecules secreted by cells. The collagen fibril is the end product of well-understood self-aggregation controlled by its amino acid sequences, but the interfibrillar amorphous ground substance has not hitherto been seen as structured by analogous aggregations prescribed by the primary structures of the characteristic glycosaminoglycans dissolved therein. Transmission electron microscopy with morphometry and stereology has demonstrated their existence in tissues. Nuclear magnetic resonance defined their secondary structures, rotary shadowing electron microscopy delineated their aggregates in vitro, and molecular dynamics stimulations showed how the latter can spring from the former. The driving forces to aggregation are hydrophobic and hydrogen bonding, offset by electrostatic repulsion between polyanionic charges. The relative stabilities of the aggregates are determined by this balance, and hence by the position and number of their charges, particularly the sulfate ester groups. Corneal stroma is a system of collagen fibrils, highly ordered to ensure transparency, in which glycosaminoglycan aggregates are suggested to determine the ordered spacing as yardsticks in a way that has parallels in all connective tissues.

Visualization of the Lytechinus pictus egg jelly coat in platinum replicas

The jelly coat of Lytechinus pictus is composed of a three-dimensional network of interconnected fibers. Both the jelly ultrastructure and its individual molecular components were visualized using rotary-shadowed platinum/carbon replicas. Bulk egg jelly, shed dry onto mica chips, displays large rope-like strands composed of bundles of smaller fibers associated with an amorphous ground substance. Additional fibers extend laterally from the rope-like strands forming branching networks. Isolation of the major fucose sulfate-rich polysaccharide complex showed that its overall structure is similar to that of whole egg jelly. Individual molecular components of jelly, separated by acid extraction and sprayed onto mica chips in 50% glycerol, exhibit repeating topological features including branching and large numbers of internal and terminal loops. Size and complexity of individual components range from small circular molecules of less than 50 nm in diameter to highly branched networks exceeding 9.5 μm in total length.