Isolation of villous microvessels from the human placenta

A method for isolating the microvessels of the human placental villi has been developed in order to culture perivascular cells. It consists of an initial selection of the villi by serial sieving. The villi retained by the 75 μm sieve were digested by collagenase-dispase. A Percoll gradient permitted the isolation of microvessels still surrounded by stromal fibres and cells. Another digestion by collagenase-dispase eliminated the contaminant elements and allowed, after a new Percoll gradient, microvessels with endothelium, basement membrane and a few perivascular cells to be obtained. Each step of the isolation of microvessels was monitored by light or electron microscopy. Our study confirms the isolation of microvessels embedded in their basement membrane and the preservation of endothelial and perivascular cells after digestion. This method, which has permitted the culture of placental endothelial cells and pericytes, appears of interest for studying microvascular angiogenesis and permeability.     

Ultrastructure of the kidney of a South American caecilian,Typhlonectes compressicaudus (Amphibia,Gymnophiona)

The ultrastructure of the renal corpuscle, the neck segment, the proximal tubule and the intermediate segment of the kidney of a South American caecilian, Typhlonectes compressicaudus (Amphibia, Gymnophiona) was examined by means of transmission electron microscopy (TEM), scanning electron microscopy (SEM) and freeze-fracture technique. The glomerular filter apparatus consists of the podocyte epithelium, a distinct basement membrane, a subendothelial space and the capillary endothelium. Emanating from the podocyte cell body, several long primary processes encircle neighboring capillaries. The short slender foot processes originating from the primary processes interdigitate with those from other primary processes, thereby forming the meandering filtration slit. Thick bundles of microfilaments are found in the primary processes, but absent in the foot processes. The basement membrane consists of a lamina rara externa and a rather thin lamina densa (50 nm thickness). The wide subendothelial space contains abundant microfibrils, a few collagen fibrils and many thin processes of mesangial cells. The endothelium is flat and fenestrated (compared to mammals displaying relatively few fenestrations); some of the fenestrations are bridged by a diaphragm. The glomerular mesangium is made up of the mesangial cells and a prominent mesangial matrix containing microfibrils and collagen fibrils. The cells of the neck and intermediate segments display numerous cilia with their microtubules arranged in the typical 9 + 2 pattern. The basal bodies of the cilia are attached to thick filaments with a clear crossbanding pattern of 65 nm periodicity. The proximal tubule is composed of cells typical for this segment (PT cells) and light cells lacking a brush border (bald-headed cells). The PT cells measure 10-25 micron in height and 15-30 micron in width and do not interdigitate at their lateral borders with each other. Their basolateral cell membrane is amplified by many folds projecting into lateral intercellular spaces and into basal recesses. The brush border is scarce and composed of loosely arranged short microvilli.     

Reticular meshwork of the spleen in rats studied by electron microscopy

The reticular meshwork of the rat spleen, which consists of both fibrous and cellular reticula, was investigated by transmission electron microscopy. The fibrous reticulum of the splenic pulp is composed of reticular fibers and basement membranes of the sinuses. These reticular fibers and basement membranes are continuous with each other. The reticular fibers are enfolded by reticular cells and are composed of two basic elements: 1) peripheral basal laminae of the reticular cells, and 2) central connective tissue spaces in which microfibrils, collagenous fibrils, elastic fibers, and unmyelinated adrenergic nerve fibers are present. The basement membranes of the sinuses are sandwiched between reticular cells and sinus endothelial cells and are composed of lamina-densalike material, microfibrils, collagenous fibrils, and elastic fibers. The presence of these connective tissue fibrous components indicates that there are connective tissue spaces in these basement membranes. The basement membrane is divided into three parts: the basal lamina of the reticular cell, the connective tissue space, and the basal lamina of the sinus endothelial cell. When the connective tissue space is very small or absent, the two basal laminae may fuse to form a single, thick basement membrane of the splenic sinus wall. The fibrous reticulum having these structures is responsible for support (collagenous fibrils) and rebounding (elastic fibers). The cells of the cellular reticulum--reticular cells and their cytoplasmic processes, which possess abundant contractile microfilaments, dense bodies, hemidesmosomes, basal laminae, and a well-developed, rough-surfaced endoplasmic reticulum, and Golgi complexes, which are characteristic of both fibroblasts and smooth muscle cells--are considered to be myofibroblasts. They may play roles in splenic contraction and in fibrogenesis of the fibrous reticulum. The contractile ability may be influenced by the unmyelinated adrenergic nerve fibers that pass through the reticular fibers. The three-dimensional reticular meshwork of the spleen consists of sustentacular fibrous reticulum and contractile myofibroblastic cellular reticulum. This meshwork not only supports the organ but also contributes to a contractile mechanism in circulation regulation, in collaboration with major contractile elements in the capsulo-trabecular system.     

The structure of the proboscis musculature in <Emphasis Type="Italic">Baseodiscus delineatus</Emphasis> (Delle Chiaje, 1825) (Heteronemertea) and the comparative analysis of the proboscis musculature in heteronemerteans

The proboscis musculature was studied in the nemertean Baseodiscus delineatus using confocal laser scanning and electron transmission microscopy. Three muscle layers were differentiated in the proboscis wall: the outer-longitudinal, the diagonal, and the inner-circular layer. The endothelium consists of two cell types: apical supportive cells with rudimentary cilia and subapical myocytes making up the inner-circular musculature of the proboscis. The supportive cells have thin processes attached to the basal extracellular matrix and their perikarya are spread over the apical surfaces of myocytes. The endothelium of B. delineatus is characterized by a folded basal layer of the extracellular matrix and by different heights of myocyte processes, giving an impression that the inner-circular musculature is multilayered. Comparative analysis shows that the diagonal musculature of Baseodiscus is not homologous to that of other heteronemerteans. An assumption is made that the inner-circular muscles have endothelial origin in all heteronemerteans.     

An Electron Microscopic Study of the Intestinal Villus : I. The Fasting Animal

The structure of the intestinal villus of the rat was studied in thin sections of tissue fixed in buffered osmium tetroxide and embedded in methacrylate. The simple columnar epithelium investing the villus is surmounted by a striated border consisting of slender projections of the cell surface. These microvilli are arranged in almost crystalline, hexagonal array, and increase the apical surface area of the cell by a factor of 24. The core of each microvillus is filled with fine fibrils which arise from the filamentous substance of the terminal web underlying the striated border. Each microvillus is covered by a tubular extension of the plasma membrane of the epithelial cell. Pinocytotic vesicles originating from the plasma membrane occur at the bases of the intermicrovillous spaces. The nucleus, mitochondria, and the endoplasmic reticulum of the epithelial cell display no unusual features. Small bits of ergastoplasm occur in the apical cytoplasm. A thin basement membrane separates the epithelium from the lamina propria which consists of vessels, nerves, and numerous lymphocytes, eosinophiles, mast cells, plasma cells, smooth muscle fibers, and macrophages suspended in a delicate stroma of fibroblasts and collagen fibers. Intercellular fat droplets often occur in this stroma, even in animals fasted for 40 hours. The blood capillaries are distinguished by their extremely attenuated, fenestrated endothelial cells. The lacteal has a thicker endothelium which, although not fenestrated, appears to have significant interruptions, especially at the margins between neighboring lining cells. Strands of smooth muscle always accompany the lacteal but do not form an integral part of its wall. Unmyelinated nerves, many of which are too small to be distinguished with the light microscope, course through the lamina propria in association with the vessels. The nerve fibers evidently do not cross the basement membrane into the epithelium. Neuromuscular junctions or other terminal apparatus were not found.     

An electron microscopic study of the intestinal villus. I. The fasting animal

The structure of the intestinal villus of the rat was studied in thin sections of tissue fixed in buffered osmium tetroxide and embedded in methacrylate. The simple columnar epithelium investing the villus is surmounted by a striated border consisting of slender projections of the cell surface. These microvilli are arranged in almost crystalline, hexagonal array, and increase the apical surface area of the cell by a factor of 24. The core of each microvillus is filled with fine fibrils which arise from the filamentous substance of the terminal web underlying the striated border. Each microvillus is covered by a tubular extension of the plasma membrane of the epithelial cell. Pinocytotic vesicles originating from the plasma membrane occur at the bases of the intermicrovillous spaces. The nucleus, mitochondria, and the endoplasmic reticulum of the epithelial cell display no unusual features. Small bits of ergastoplasm occur in the apical cytoplasm. A thin basement membrane separates the epithelium from the lamina propria which consists of vessels, nerves, and numerous lymphocytes, eosinophiles, mast cells, plasma cells, smooth muscle fibers, and macrophages suspended in a delicate stroma of fibroblasts and collagen fibers. Intercellular fat droplets often occur in this stroma, even in animals fasted for 40 hours. The blood capillaries are distinguished by their extremely attenuated, fenestrated endothelial cells. The lacteal has a thicker endothelium which, although not fenestrated, appears to have significant interruptions, especially at the margins between neighboring lining cells. Strands of smooth muscle always accompany the lacteal but do not form an integral part of its wall. Unmyelinated nerves, many of which are too small to be distinguished with the light microscope, course through the lamina propria in association with the vessels. The nerve fibers evidently do not cross the basement membrane into the epithelium. Neuromuscular junctions or other terminal apparatus were not found.     

Fine Structure of the Tentacles of Phoronis australis Haswell (Phoronida,Lophophorata)

The epidermis of the tentacles of Phoronis australis consists of six cell types: supporting cells, choanocyte-like sensory cells, both types monociliated, secretory A-cells with a mucous secretion, and three kinds of B-cells with mucoprotein secretions. On cross-sections of the tentacle, one can distinguish four faces: the frontal one, heavily ciliated and located between the two frontolateral rows of sensory cells, the lateral and the abfrontal ones. The orientation of the basal structures of the cilia is related to the direction of their beat. The basiepidermal nervous system is grouped mainly at the frontal and abfrontal faces. The basement membrane is thickest on the frontal face and consists of circular collagen fibrils near the epidermis and longitudinal ones near the peritoneum. All peritoneal cells surrounding the mesocoel are provided with smooth longitudinal myofibrils, and isolated axons are situated between these cells and the basement membrane. The wall of the single blood capillary in each tentacle consists of epitheliomuscular cells with circular myofilaments, lying on a thin internal basal lamina; there is no endothelium.     

No direct contact between the excretory system and the circulatory system in Prostomatella arenicola Friedrich (Hoplonemertini)

Excretory and circulatory systems in Prostomatella arenicola are examined at the ultrastructural level. Interdigitating cells, which rest on a thin fibrillar basal lamina, line the lumina of the lateral vessels. A layer of muscle cells and an underlying sheath of fibrillar extracellular material surround each vessel.The excretory system consists of one pair of laterally situated branched protonephridia. Each protonephridium is composed of several terminal cells, an efferent duct and a nephridiopore. The terminal parts of the protonephridia are not restricted to the vicinity of the circulatory system; they can also be found dorsally or laterally to the nerve cords between muscle cells. The presumed filtration area arises as a hollow cylinder from the terminal cell. This cylinder is perforated by numerous clefts which are never bridged by a filter diaphragm. Instead, each terminal cell cylinder is surrounded by an extracellular matrix. The terminal cells neither extend into the lumen of the lateral vessel nor contact the vessel lining cells.Phylogenetic implications of the results are discussed.     

Histological and ultrastructural specialization of the digestive tract of the intestinal air breather hoplosternum thoracatum (teleost)

The digestive tract of Hoplosternum thoracatum consists of an esophagus, gastric area, anterior digestive intestine with elaborate folds, digestive intestine with decreasing folds and thin, smooth-surfaced respiratory intestine. The upper tract has a mucoid columnar lining which is gently folded, whereas the gastric area has numerous pits opening into the tubular secretory glands. Striated muscle comprises the anterior muscularis but is replaced by inner circular and outer longitudinal smooth muscle layers in the gastric region. The digestive intestinal mucosa is elaborately folded, consisting of columnar cells with prominent brush borders. Mucosa, submucosa, circular and longitudinal muscularis and serosa layers are present throughout the tract. Goblet cells occur in both the digestive and respiratory intestine. Major changes that appear in the respiratory intestine are a drastic reduction in mucosa epithelial thickness and the penetration of an elaborate capillary bed into the epithelium. The other basic layers are not significantly reduced in thickness. The air-blood barrier consists of the thin epithelium, basement lamina and very thin capillary endothelium. Regional cellular composition and ultrastructural features are correlated with respective digestive and respiratory functions.     

Lungs of Polypterus and Erpetoichthys

The wall of the asymmetrical saclike lungs of the fishes Polypterus and Erpetoichthys consists of several functionally different tissue layers. Their lumen is lined by a surface epithelium composed of (1) highly attenuated cells, termed pneumocytes I; (2) pneumocytes II with lamellar bodies, presumably indicating surfactant production; (3) mucous cells; and (4) ciliated cells. Underlying the pneumocytes I is a dense capillary net. The thin continuous endothelium of this net, together with the pneumocytes I, constitute the very thin blood-air barrier. The basement membrane of epithelium and endothelium fuse in the area of the blood-air barrier (thickness 210 m?m). Secretory and ciliary cells form longitudinal rows in the epithelium. Below the zone with a gas-exchanging tissue, a layer of connective tissue containing collagen and special elastic fibers occurs. The blood vessels that give rise to or drain the superficial capillary plexus are located in this connective tissue. The outermost layer of the lung consists of muscle cells, a narrow inner zone with smooth muscle cells, and an outer, broader zone with cross-striated muscle cells. The lung is innervated by myelinated and nonmyelinated nerve fibers. The morphology of the gas-exchange tissue in the lungs of these primitive bony fish is fundamentally very similar to that of the lungs of tetrapod vertebrates. The morphologic observations are in close agreement with physiologic data, disclosing well-developed respiratory capacities. Structural simplicity can be regarded as a model from which the lungs of the higher vertebrates derived. In addition to respiratory function, the lungs seem also to have hydrostatic tasks.     

Fine structure of the respiratory organs of the Climbing perch, Anabas testudineus (Pisces: Anabantidae)

An electron microscopic study has been made of the three respiratory organs of climbing perch. The gill structure is similar to that of the other telcosts but the thickness of the water/blood barrier is much greater, being as great as 20 μm in some specimens. The increased thickness is due to a multilayered epithelium which is thinner (3.5–7 μm) over the marginal channel of the secondary lamellae. The other two main layers, basement membrane and pillar cell flange, are relatively thin (about 1 μm).
The pillar cells have a typical structure, but in certain regions they are contiguous with one another and line well-defined blood channels. Some of the columns of basement membrane material in such regions may be common to adjacent pillar cells.
The air-breathing organs are (a) the lining of the suprabranchial chambers , and (b) the labyrinthine plates attached to the dorsal region of branchial arches. Electron microscopy showed that their structure is well adapted for gas exchange, the air/blood barriers being only 0.12–0.3 μm, comprising an epithelial layer, basement membrane, and thin capillary endothelium. The many parallel blood channels of the respiratory islets of both organs are separated by pillar-like structures which differ from the pillar cells of the secondary lamellae. Thus the hypothesis that the air-breathing organs represent modified gills is not supported by this study.
The fine structure of the non-respiratory region of the air-breathing organs is similar to that of the skin, and includes chemoreceptor-like cells. Evidence concerning the possible homology of pillar cells with plain muscle cells is discussed.     

Muscle cell attachment in Caenorhabditis elegans

In the nematode Caenorhabditis elegans, the body wall muscles exert their force on the cuticle to generate locomotion. Interposed between the muscle cells and the cuticle are a basement membrane and a thin hypodermal cell. The latter contains bundles of filaments attached to dense plaques in the hypodermal cell membranes, which together we have called a fibrous organelle. In an effort to define the chain of molecules that anchor the muscle cells to the cuticle we have isolated five mAbs using preparations enriched in these components. Two antibodies define a 200-kD muscle antigen likely to be part of the basement membrane at the muscle/hypodermal interface. Three other antibodies probably identify elements of the fibrous organelles in the adjacent hypodermis. The mAb IFA, which reacts with mammalian intermediate filaments, also recognizes these structures. We suggest that the components recognized by these antibodies are likely to be involved in the transmission of tension from the muscle cell to the cuticle.     

Electron microscopy of the esophageal ganglion complex of the gastropod pulmonate Triodopsis divesta. I. Ultrastructure of the epineurium

The epineurium of the esophageal complex of the gastropod pulmonate Triodopsis divesta was examined by electron microscopy. The epineurium consists of two main regions: an inner dense fibrous region adjacent to the avascular neural tissue of the ganglion and an outer cellular region comprised of a variety of cell types embedded in a connective tissue matrix. The dense fibrous region contains smooth muscle cells and associated nerve processes and is invested on the neural side by thin processes of glial cells. The outer highly cellular region contains smooth muscle cells, nerve processes, wandering cells (amebocytes), globular cells, and myoepithelial cells comprising the walls of the vascular system. In addition, a cell type not previously identified in other gastropod epineuria is present. These cells resemble neurosecretory cells. The morphology and structural interrelationships of these various constituents are presented and the possible functions of individual cell types and the epineurium in general are discussed in relation to information available on other molluscs.     

The endothelial microenvironment in the venous valvular sinus: thromboresistance trends and inter-individual variation

The valve sinuses of the deep venous system are frequent sites of venous thrombus initiation. We previously reported that, in comparison with the non-valvular lumenal endothelium, the valve sinus endothelium had decreased expression of von Willebrand factor (vWF) and increased expression of endothelial protein C receptor (EPCR) and thrombomodulin (TM), suggesting alteration in the procoagulant/anticoagulant balance. We hypothesized that increased stasis in the deeper recesses of the venous valves would be associated with a gradient of increased thromboresistance. Expression of EPCR, TM, and vWF was analyzed via quantitative confocal immunofluorescence in residual saphenous veins collected following coronary artery bypass procedures. In agreement with our hypothesis, endothelial expression of vWF in the valve sinus decreased from the uppermost to the deepest region of the valve sinus. In contrast to our hypothesis, EPCR expression decreased from the uppermost to the deepest region of the valve sinus (p < 0.001) and TM expression remained unchanged throughout the valve sinus. Comparison of the non-valvular lumenal endothelium with the valve sinus endothelium demonstrated significantly decreased vWF expression (p < 0.001) in the valvular sinus consistent with our previous report; however, we did not observe statistically significant differences in EPCR or TM expression in this comparison. In addition, remarkable inter-individual variation in expression of these three proteins was also observed. These findings suggest that the genesis of these observations is more complex than predicted by our initial hypothesis, likely due, at least in part, to the complex rheology of the valvular sinus microenvironment.     

Heterogeneous histochemical reaction pattern of the lectin Bandeiraea (Griffonia) simplicifolia with blood vessels of human full-term placenta

Bandeiraea simplicifolia lectin (BS-I) stains vascular endothelium in various species. In humans, less than 10% of the specimens studied exhibit a reaction with BS-I. In the present histochemical study, the reactivity of BS-I with placental blood vessels and its correlation with the blood group from mother and newborn child was investigated. Acetone-fixed cryosections of representative tissue segments of human full-term placenta and umbilical cord were stained with BS-I. The staining pattern of tissues from patients with different blood groups was identical, although the reaction of BS-I in the placenta was heterogeneous. BS-I did not react with the umbilical cord. Vascular smooth muscle cells at the insertion site of the umbilical cord into the chorionic plate, and endothelium deeper in the chorionic plate, became progressively stained. The endothelial cells and tunica muscularis of smaller arteries and veins in stem villi lost their reactivity in parallel with decreasing vessel size. Arterioles and venules reacted heterogeneously. Capillaries, trophoblastic basement membranes, especially epithelial plates, and sometimes the syncytiotrophoblast were labelled in several terminal villi. The data indicate that 1) the placenta binds BS-I to fetal endothelium independent of the blood group, 2) cell-surface antigens on placental endothelial cells are expressed heterogeneously and 3) cell-surface glycans are constituted in an organ-specific manner on human endothelial cells.     

The structure of the rectal pads of Periplaneta americana L. With regard to fluid transport

The rectum of Periplaneta americana L. is lined with cuticle and has six radially arranged cushion-shaped thickenings, the rectal pads, composed of columnar cells. Narrow strips of simple rectal cells lie between the pads. Tall junctional cells form a thin but continuous collar around the pads where they join the rectal cells. The epithelium is surrounded by a layer composed of circular and longitudinal muscles and connective tissue. This layer of muscle and connective tissue is innervated and tracheated, and is separated from the pad surface by a subepithelial sinus. Fluid flowing through the sinus enters the haemolymph through openings in the muscle layer whre large tracheae penetrate. These openings can be sealed by muscle contractions that appress the muscle around the openings against the pad surface. The tracheae pass on into the pads, following basement membrne-lined indentations of the pad surface. Within the pad tracheolar cells send fine branches between the cells. Near the apical and basal surfaces the lateral membranes of pad cells are bridged by septate desmosomes that form a continuous band around the cells. Between apical and basal septate desmosomes is an interconnected labyrinthine system of intercellular spaces. There are three kinds of space, dilations and apical sinuses, both of variable size, and narrow communicating channels about 200 Å wide. The membranes of the latter have mitochondria closely associated with them. Continuity between the system of spaces and the subepithelial sinus is established by the basement membrane-lined invaginations of the basal surface where tracheae penetrate between pad cells. Apical surfaces of the pad cells are highly infolded and are also associated with mitochondria. However, unlike the lateral membranes facing the narrow channels, the apical membranes have a cytoplasmic coating of particles. Both associations of mitochondria with membranes constitute discrete structural entities that are found in many transporting epithelia, and we have termed them “plasmalemma-mitochondrial complexes.” As the rectal pads are organized into systems of spaces that ultimately open in the direction of fluid movement, existing models of solute-coupled water transport can be applied. However, the rectal pads are structurally more complex than fluid-transporting tissues of vertebrates. This complexity may be related to the ability of the rectum to withdraw water from ion-free solutions in the lumen. We present a structural model involving solute recycling to explain the physiological characteristics of rectal reabsorption.     

Ultrastructural localization of lectin-binding sites in different basement membranes

Summary In the present work we localized binding sites for the lectins WGA, RCA I, con A and SBA at the ultrastructural levels in morphologically different basement membranes. These different basement membranes included (a) thin ones, for example, tubular basement membrane of the mouse kidney which separates epithelial cell layers from mesenchymal cells and glomerular basement membrane which separates epithelial cells from other epithelial cells, (b) thick multilayered ones, for example, Reichert's membrane which is built up during the embryonic development of rodents and as an example of a pathologically thickened basement membrane, the basement membrane of the Engelbreth-Holm-Swarm (EHS) sarcoma. We were able to show that, in contrast to the thick multilayered basement membranes, the thin ones showed a strong positive SBA-binding pattern. Thick basement membranes otherwise revealed very strong labelling with the lectins WGA and RCA I. Our findings lead us to conclude that thin and thick basement membranes differ markedly in the quality and quantity of the carbohydrates which they contain.     

The accessory reproductive structure in the simultaneous hermaphrodite Diplectrum bivittatum

The general gonad morphology in the simultaneous hermaphrodite, Diplectrum bivittatum from the Gulf of Mexico, agrees with previously published accounts on the gonad morphology of other Diplectrum spp. In D. bivittatum there is a well developed accessory reproductive structure (ARS) which consists of a post-ovarian sinus and numerous well developed villi which project into the sinus from the sinus wall. The ARS varies seasonally: the post-ovarian sinus is reduced in February and enlarged during June, September and October when the species is reproductively active. The villi are better developed in September specimens. Two pillar-like structures within the ARS are described for the first time. Their columnar and smooth muscle cell composition suggests several possible reproductive functions.     

Ultrastructure of the pronephric kidney in upstream migrant sea lamprey, Petromyzon marinus L

The pronephric kidneys were examined in upstream migrant sea lampreys, Petromyzon marinus L., by transmission and scanning electron microscopy. Each pronephros consists of an enlarged renal corpuscle (glomus) and ciliated nephrostomes, but there are no renal tubules. The renal corpuscle contains an extensive mesangium, which consists of a highly fibrous extracellular matrix, numerous mesangial cells, granulocytes, and macrophages. The extracellular matrix contains microfibrils with a morphology similar to amyloid P microfibrils, fibrils with a periodicity similar to fibrin, and abundant collagen. Often these fibrillar components are aggregated in the region of the basement membrane, giving it a thickened appearance. Some podocytes of the visceral epithelium appear swollen, and their cytoplasm contains numerous vacuolar inclusions, and many have only primary major processes with only a few or no foot processes. The morphological features of the pronephric kidney of the lamprey at this time in the life cycle reflect the regression of this organ, but some features also resemble those seen in renal pathologies of higher vertebrates.