Histochemistry of the tongue epithelium in four mammals with respect to keratinization

The tongue epithelium was examined in the laboratory rat, guinea pig, rabbit and Domestic cat, using light microscopical, histological fluorescent and histochemical methods. The distributions of the enzymes, acid and alkaline phosphatase were examined. Protein-bound phospholipid and calcium were investigated, together with thiol sulphydryl groups and cysteine disulphide bonds of proteins. A variety of different types of keratinization were shown in the various species, as well as in the same species in different regions of the tongue. The most strongly keratinized structures were the filiform and conical papillae which varied widely from species to species. Those of the rat dorsum were similar to papillae described previously in the House mouse and have strongly keratinized spines. The guinea pig showed some differences but also had keratinized spines. In contrast the rabbit papillae did not have spines but the horny layer over the posterior sides was hardened instead to form pointed edges. Human filiform papillae are similar to the rabbit without spines but the horny layer is less strongly keratinized. In the Domestic cat the conical papillae were also without spines but the horny layer on the anterior and posterior surface was hardened to form claw-like structures.     

A histochemical study of keratinization in the domestic fowl (Gallus gallus)

The microanatomy of the epidermis of the domestic fowl is described and related to the distribution of various histochemical constituents involved in keratinization.
The avian horny layer over the back is composed of a loose network of structurally solid horny cells. This is in contrast to most mammalian epidermal horny cells in which structural keratin is found only in the peripheral cytoplasm, and the interior of the keratinocyte contains soluble products of cytolysis with possibly some free keratin filaments dispersed in the fluid material.
The avian tarsal epidermal horny scales show similarities to both the scales of lizards and snakes and to mammalian tail scales which appear to be homologous structures.
It is suggested that a thin layer of cells containing no detectable disulphide bonds, found in the tarsal scale region of the young chick, is probably mechanically weak and may function as a fission plane for sloughing of the horny layer. A specialized epidermis and thickened horny layer is developed in the fowl on the plantar underside of the toes, but this is quite different in structure from the mammalian plantar epidermis.
The overlapping of zones rich in ribonucleic acid (RNA) and bound cysteine (SH) in the growing feather suggests that protein synthesis and the preparatory stages to keratin disulphide bonding normally occur concurrently in feather formation. This is in contrast to the growing hair which has a region rich in RNA followed immediately before it becomes keratinized by a discrete keratogenous zone weak in RNA but rich in bound cysteine.     

Ultrastructural analysis of the effect of ethane dimethanesulphonate on the testis of the rat,guinea pig,hamster and mouse

Summary The morphological response of the testis of rats, guinea pigs, Syrian hamsters and mice to treatment with the cytotoxin ethane dimethanesulphonate was examined using light and electron microscopy. One to two days after a single administration of ethane dimethanesulphonate to adult rats, guinea pigs, and hamsters, the Leydig cells showed marked ultrastructural alterations suggestive of degeneration and cell death. The former alterations included karyopyknosis, cytoplasmic vesiculation and accumulation of lipid inclusions and large lipofuscin bodies. Fragments of necrotic Leydig cells were often engulfed by the interstitial tissue macrophages. The morphology of the seminiferous epithelium of these three species was unchanged from the morphology observed in vehicle-injected control animals. In contrast, multiple injections of ethane dimethanesulphonate given to mice produced no ultrastructural alterations to Leydig cells yet the seminiferous epithelium exhibited disruption of spermatogenesis. Although the Leydig cells of the mouse appear resistant to ethane dimethanesulphonate, this agent exerts a selective cytotoxic action upon Leydig cells of the rat, guinea pig and hamster thus identifying ethane dimethanesulphonate as a useful chemical for future endocrine and physiological studies of testicular function in three common laboratory species.     

Schistosoma manonsi: cercarial penetration of host epidermis at the ultrastructural level

Invasion of the outer layers of the epidermis of mouse ear skin by cercariae of Schistosoma mansoni within 7 min of their application to it has been studied with the optical and the scanning and transmission electron microscopes.Entrance of cercariae was under the edges of the dead flattened keratinized cells of the horny layer (squames), and penetration through this layer was by disarticulation of the stacks of squames at their interdigitations. Mucus from the postacetabular glands was recognized with the light and electron microscopes on the skin surface, especially at squame edges and between layers of squames and along the keratogenous zone. The findings suggested that disarticulation of the squames was not effected solely by the muscular probing and pushing of the parasite, but that it might be aided by swelling of the mucus secretion deposited in the area from the postacetabular glands. Loosening of the interlocked edges of the squames by enzymatic action is also a possibility, but was not evaluated for this report.The migration path along the keratongenous zone was marked by extensive damage to the transitional cells of the granular layer subjacent to the squames. Packets of secretion from the cercarial preacetabular glands were identified below the horny layer in the cytoplasm of these cells. It was considered that the host tissue damage in this area was the result not only of tearing of the tissues by passage of the spiny schistosomules, but also of enzymatic activity, the enzyme source being the granules in the packets of preacetabular gland secretion.     

Ultrastructural characterization of interstitial cells of Cajal associated with the submucosal plexus in the proximal colon of the guinea pig

Interstitial cells of Cajal (ICC) associated with the submucosal (submucous) plexus (ICC-SP) in the proximal colon of the guinea pig were studied by immunohistochemistry and electron microscopy. Whole-mount stretch preparations with c-Kit immunohistochemistry revealed that a number of ICC-SP constituted a dense cellular network around the submucosal plexus. Some of these ICC-SP were observed in the vicinity of the muscularis mucosae in sections immunostained for c-Kit and α-smooth muscle actin. Ultrastructural observation demonstrated, for the first time, that ICC-SP of the proximal colon of the guinea pig retained typical ultrastructural characteristics of ICC repeatedly reported in association with the tunica muscularis of the gastrointestinal tract: a basal lamina, caveolae, many mitochondria, abundant intermediate filaments and the formation of gap junctions with the same type of cells. The most remarkable ultrastructural finding was the presence of thick bundles composed of the processes of ICC-SP connected to each other via large gap junctions. These ICC-SP might be involved in the main mucosal functions of the proximal colon of the guinea pig, namely the transportation of water and electrolytes, possibly via their involvement in the spontaneous contractions of the muscularis mucosae.     

Eosinophils and mast cell homogeneity of the guinea pig eyelid skin, conjunctiva, and ileum

Mast cell heterogeneity has been described on the basis of differential staining reactions, light microscopic morphology, anatomic location, degranulation after polyamines, biochemical contents, growth requirements, and reactions to lymphokines. We have demonstrated typical "connective-tissue mast cells" by using anatomic criteria, histological staining reactions, electron microscopy, and reaction to compound 48/80 in the guinea pig conjunctiva, eyelid skin, and ileum. A second, much larger population of cells in the ileal mucosa and the conjunctiva, and rarely in the eyelid skin stained reddish-blue with acid toluidine blue in tissue fixed in ethanol-acetate-lead subacetate (BLA) and with alkaline Giemsa in formaldehyde-fixed tissue, did not stain with ethanolic or acid toluidine blue in formaldehyde-fixed tissue or with alkaline Giemsa in BLA-fixed tissue, and did not degranulate after 48/80 treatment. These are features of the rat intestinal "mucosal mast cells"; however, ultrastructural and light microscopic studies with the orcein Giemsa stain demonstrated these cells in the guinea pig to be eosinophils. Tissue culture, biochemical, and immunological studies indicate the existence of a second type of mast cell (bone-marrow-derived mast cell), ultrastructurally almost indistinguishable from the connective tissue mast cell. Our studies demonstrate only one mast cell type in the guinea pig and support the contention that other forms of mast cells are immature forms or variants of the connective-tissue mast cell.     

Keratinization of fish skin with special reference to the catfish Bagarius bagarius

Summary Histochemical reactions indicating keratinization have previously been demonstrated in parts of the epidermis of Bagarius bagarius. Fluorescence histochemistry and electron microscopy have now confirmed these results. Elevated areas of the epidermis are capped by a layer of dead cells with altered contents. On the outer aspect of these cells a dense layer, 18 nm thick, beneath the plasma membrane corresponds to the resistant envelope found in keratinized cells in tetrapod vertebrates. In Bagarius this layer does not extend to all faces of the keratinized cells, but a similar envelope has been detected in two other sites of piscine keratinized epidermis investigated, namely in the breeding tubercles of Phoxinus phoxinus and in the teeth of Lampetra fluviatilis. In the elevated areas of Bagarius-epidermis, the epithelial cells undergo progressive changes in cytoplasmic organization as they become more superficial. The second tier from the surface is sealed by tight junctions and is separated from the overlying keratinized cells by a sub-corneal space resembling that found in keratinized amphibian epidermis. Histochemical evidence of a high lipid content in the outer layers of the epidermis correlates with the presence of lipid inclusions and lamellated membranous profiles in the material studied by electron microscopy. Histochemical results show that the fin skin of Blennius pholis is not keratinized, but secretes a cuticle, histochemically reactive for both proteins and glycoproteins.     

Fine structure of the horny teeth of the lamprey,Entosphenus japonicus

The fine structure of the horny teeth of the lamprey, Entosphenus japonicus, was examined by light- and electron-microscopy. Most of the horny teeth consisted of two horny and two nonhorny layers. The primary horny layer was well keratinized, and the cells were closely packed and intensely interdigitated, being joined together by many modified desmosomes. The plasma membrane of the horny cell, unlike the membranes of other vertebrates, was not thickened. The intercellular spaces were filled with electron-dense material. Microridges were seen on the free surface. Structures resembling microridges were found on the underside of the primary horny layer. The secondary horny layer displayed various stages of keratinization. The keratinization started at the apex and developed toward the base. In the early stage of keratinization, the superficial cells became cylindrical and were arranged in a row forming a dome-shaped line. Their nuclei were situated in the basal part of the cells. The appearance of the nonhorny layers varied according to the degree of keratinization of the horny layers beneath them. The nonhorny cells were joined together by many desmosomes and possessed many tonofilament bundles. The replacement and keratinization of the horny teeth are discussed in the light of these results.     

Structure of the sensory innervation of the anal canal in the pig. A light- and electron-microscopical study

The sensory innervation of the anal canal of the pig was investigated by light and electron microscopy. The distribution of the different types of sensory nerve endings correlates with the histology of different zones: (1) After the rectal mucosa there was a zone lined with nonkeratinized stratified squamous epithelium. (2) A middle zone was lined with keratinized stratified squamous epithelium. Here the dermis already showed a papillary and reticular layer. (3) The last zone showed hairy skin with a high hair density. The following nerve endings were found: Free nerve endings reached the stratum superficiale in nonkeratinized squamous epithelium and the stratum granulosum in the keratinized squamous epithelium. Dermal free nerve endings were found in all zones near the epithelium and two different types were identified as those derived from C-fibers and those from A-delta-fibers. Merkel nerve endings showed different features depending on their location. Few Merkel-like cells were found in the epithelium of the anal crypts. Typical Merkel Tastscheiben were located at the base of epithelial ridges or pegs in zones 2 and 3. The number of Merkel cells varied up to 200. The myelinated afferent fiber supplied 10-15 Merkel cells. Merkel cells were also found regularly in the outermost layer of the external rooth sheath of hair follicles at about the same level as perifollicular nerve endings. Lamellated corpuscles were found in the dermis of all zones except the cranial part of zone 1, where the anal crypts are located. Generally they consisted of a central nerve terminal which may be branched. Each terminal was surrounded by an inner core of concentrically arranged lamellae of the terminal Schwann cell and one or several inner cores were included in a capsule of perineural cells. The size of the corpuscle, the regularity of the inner core and the number of capsular layers depended on the location of the corpuscle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Light- and electron-microscopic study of substance P-immunoreactive neurons in the guinea pig retina

Substance P (SP) immunoreactivity in the guinea pig retina was studied by light and electron microscopy. The morphology and distribution of SP-immunoreactive neurons was defined by light microscopy. The SP-immunoreactive neurons formed one population of amacrine cells whose cell bodies were located in the proximal row of the inner nuclear layer. A single dendrite emerged from each soma and descended through the inner plexiform layer toward the ganglion cell layer. SP-immunoreactive processes ramified mainly in strata 4 and 5 of the inner plexiform layer. SP-immunoreactive amacrine cells were present at a higher density in the central region around the optic nerve head and at a lower density in the peripheral region of the retina. The synaptic connectivity of SP-immunoreactive amacrine cells was identified by electron microscopy. SP-labeled amacrine cell processes received synaptic inputs from other amacrine cell processes in all strata of the inner plexiform layer and from bipolar cell axon terminals in sublamina b of the same layer. The most frequent postsynaptic targets of SP-immunoreactive amacrine cells were the somata of ganglion cells and their dendrites in sublamina b of the inner plexiform layer. Amacrine cell processes were also postsynaptic to SP-immunoreactive neurons in this sublamina. No synaptic outputs onto the bipolar cells were observed.     

Fine structure of atrial natriuretic peptide (ANP)-granules in the atrial cardiocytes in the hamster, guinea pig, rabbit, cat and dog.

In the hamster, guinea pig, rabbit, dog and cat, the right and left atria and ventricles were examined by immunohistochemistry, and the right auricular cardiocytes were studied by transmission electron microscopy. Moreover, ANP-granules in the cardiocytes were analyzed by ultrastructural morphometry. Immunohistochemically, the most intensely ANP-reactive cardiocytes were localized in the right auricle, particularly more prominent in the hamster and guinea pig than in the rabbit, dog and cat. The immunoreaction in the dog and cat was weaker than that in the rabbit. ANP-immunoreactivity was not detected in the ventricular myocardium of any of all species examined, but was occasionally observed in the subendocardium of the ventricular septum. Ultrastructurally, ANP-granules were localized principally in the perinuclear region associated with the Golgi apparatus and scattered throughout the sarcoplasmic layers. The Golgi apparatus of the cardiocytes was better developed in the hamster and guinea pig than in the rabbit, dog and cat. It was poorly-developed in the dog and cat. By ultrastructural morphometry, the number of granules was greatest in the hamster followed by the guinea pig, rabbit and dog or cat, in this order. On the other hand, the diameter of granules was largest in the guinea pig and reduced via the hamster to the rabbit. The diameter was significantly smaller in the dog than in the rabbit. The diameter of granules of the cat was lay between the rabbit and dog.     

Some ultrastructural observations on human palmar stratum corneum.

Ultrastructure of human palmar stratum corneum was examined for cell junctions and cytoplasmic material. Interdigitated microvilli are shorter and less uniform in man. Most of these processes are either conical or truncated in shape, as distinct from the relatively smooth cell surfaces except for sigma-shaped junctions in hairy sites. It is suggested that this provides strength against shearing force in the palm and sole. There is less cytoplasmic breakdown in palmar horny cells than in hairy cells.     

Morphological studies of fiber types of striated muscle fibers of the cremaster in the guinea pig

THe fine structure of the striated muscle fibers of the cremaster of the guinea pig was studied using the cholinesterase technique and light and electron microscopy. Under light microscopy, isolated single muscle fibers showed two types of nerve endings: the first one presented elliptic or oval areas having digit-like structures inside, some of the borders of which were heavily stained. These fibers had only one end-plate. The second type presented elongated clear areas with most of the density located on the borders. Several nerve endings were apparent in these fibers. By electron microscopy, the former had large and numerous sarcolemmal foldings and these characteristics were also observed in unstained fibers. In the latter, the foldings were scanty or absent. At the ultrastructural level, the fibers having only one end-plate presented a regular array of fibrils with an abundant sarcoplasmic reticulum ('Fibrillenstruktur' type) in contrast to the multi-innervated fiber with an irregular distribution pattern of fibrils and a scarce sarcoplasmic reticulum ('Felderstruktur' type). The striated muscle fiber layer of the cremaster probably contains both fast and slow fibers. The possible functional role for the slow striated muscle fibers is discussed.     

Metabolism of arachidonic acid by guinea pig Clara cells.

A method for the isolation of non-ciliated bronchiolar epithelial (Clara) cells from the guinea pig is described. Following digestion of the lung tissue with Type XXIV protease, the isolated lung cells showed a viability greater than 90% and contained 3% of Clara cells. Several cell populations were then separated on the basis of size using 2 centrifugal elutriations. The macrophages and endothelial cells were removed from the Clara cells enriched fractions by differential adherence on Petri dishes. The Clara cell-rich suspension was then further purified by centrifugation on Percoll non-continuous density gradients consisting of 48-52-55% Percoll solution. The lower interface and the pellet of the non-continuous gradient consisted of approximately 80% Clara cells. Identification of isolated Clara cells was confirmed by light microscopic observations after nitroblue tetrazolium staining and by ultrastructural characteristic features as observed by electron microscopy. The metabolism of arachidonic acid into prostaglandins and TxB2 by purified Clara cells was examined by enzyme immunoassay (EIA) and leukotriene formation was investigated by reverse phase high performance liquid chromatography (RP-HPLC). Enriched guinea pig Clara cells incubated with arachidonic acid released TxB2, PGE2 and 6-keto PGF1 alpha, but did not produce leukotrienes. These cells could however transform exogenous leukotriene A4 into leukotriene B4. These results suggest that guinea pig Clara cells possess the enzymes of the cyclooxygenase pathway required for TxB2, PGE2 and 6-keto-PGF1 alpha synthesis. Clara cells do not possess the 5-lipoxygenase enzyme but show some leukotriene A4 hydrolase activity since they can produce leukotriene B4 upon incubation with leukotriene A4.     

Fucoidin inhibits attachment of guinea pig spermatozoa to the zona pellucida through binding to the inner acrosomal membrane and equatorial domains

Our previous study has shown that fucoidin, an algal heteropolysaccharide, is a potent inhibitor of sperm-zona binding in the guinea pig, hamster and human. To visualize the surface site of fucoidin binding, a biotinated derivative (B-Fuc) of the native fucoidin was prepared. B-Fuc retained the inhibitory activity and was used in conjunction with FITC-avidin to localize its binding sites on guinea pig spermatozoa using fluorescence microscopy. In living acrosome-reacted spermatozoa, B-Fuc bound predominantly to the inner acrosomal membrane and equatorial segment domains. The binding was effectively competed by a 10-fold excess of native fucoidin, but not by a 10-fold excess of heparin or a 20-fold excess of biotinated normal rabbit serum IgG. B-Fuc binding patterns on dead spermatozoa were quite different from that of living spermatozoa. The post-acrosomal region, rather than the inner acrosomal membrane and equatorial domains, was intensely labeled. This indicates the importance of using living cells in assessing true surface binding sites whenever possible. We conclude that the inner acrosomal membrane and/or equatorial domains are critical for zona binding in the guinea pig.     

Epidermal keratinization in the salamander and a comparison with other amphibia

In a variety of amphibians examined the stratum corneum was one cell in depth, although in Xenopus it was up to three cells deep. The flattened horny cells were closely fused together along their lateral membranes to form a continuous sheet. Disulphide bonds of keratin were most concentrated in the peripheral cytoplasm, but the interiors of the cornified cells were sufficiently well keratinized to prevent more than slight enzymatic cytolysis of the normal cell components. Characteristically large, weakly stainable, non-shrunken nuclear remnants were found in the salamander and frog horny layers, but the clawed toad had small pyknotic (parakeratotic) nuclei. The mature amphibian keratinocytes contained free fats, bound phospholipids, calcium and sulphydryl groups, together with acid phosphatase and non-specific esterase. Cornification appears to begin by a process of separate individual cell keratinization and lateral membranes of neighbouring cells only later become fused together. This differs from the process in higher vertebrates in which the cells undergoing keratinization form a uniform transitional layer in the epidermis. In the amphibian epidermis neighbouring cells occur in different stages of keratinization.     

Leu-enkephalin-like material in nerves and enterochromaffin cells in the gut

Summary The distribution and cellular localization of leu-enkephalin in the gut and pancreas was studied by immunohistochemistry using two different antisera, one specifically directed against leu-enkephalin and the other cross reacting with met-enkephalin. The results were identical with both antisera. In all species examined, enkephalin-immunoreactive material was found in nerves of the smooth muscle, particularly numerous in the myenteric plexus. Here, immunoreactive nerve cell bodies were observed occasionally. In addition, enkephalin-immunoreactive material was demonstrated in gut endocrine cells of chicken, mouse, rat, pig and monkey but not of guinea pig, cat and man. Enkephalin cells were detected also in the exocrine parenchyma of the porcine pancreas. They were rare in the gut of mouse, rat and monkey but numerous in the antrum and duodenum of pig where they were identified as 5-hydroxytryptamine-storing enterochromaffin cells. The enkephalin-containing cells of the porcine antrum and duodenum were defined ultrastructurally by the consecutive semithin/ultrathin section technique. The ultrastructural features were typical of enterochromaffin cells, the most characteristic ones being the irregular shape and high electron density of the cytoplasmic granules. The immunoreactive material was confined to the cytoplasmic granules.     

Leu-enkephalin-like material in nerves and enterochromaffin cells in the gut. An immunohistochemical study.

The distribution and cellular localization of leu-enkephalin in the gut and pancreas was studied by immunohistochemistry using two different antisera, one specifically directed against leu-enkephalin and the other cross reacting with met-enkephalin. The results were identical with both antisera. In all species examined, enkephalin-immunoreactive material was found in nerves of the smooth muscle, particularly numerous in the myenteric plexus. Here, immunoreactive nerve cell bodies were observed occasionally. In addition, enkephalin-immunoreactive material was demonstrated in gut endocrine cells of chicken, mouse, rat, pig and monkey but not of guinea pig, cat and man. Enkephalin cells were detected also in the exocrine parenchyma of the porcine pancreas. They were rare in the gut of mouse, rat and monkey but numerous in the antrum and duodenum of pig where they were identified as 5-hydroxytryptamine-storing enterochromaffin cells. The enkephalin-containing cells of the porcine antrum and duodenum were defined ultrastructurally by the consecutive semithin/ultrathin section technique. The ultrastructural features were typical of enterochromaffin cells, the most characteristic ones being the irregular shape and high electron density of the cytoplasmic granules. The immunoreactive material was confined to the cytoplasmic granules.     

Activity of specific and non-specific cholinesterases in the subcommissural organ of guinea pig and albino rat

Summary The localizations of specific and non-specific cholinesterases were demonstrated by light and electron microscopical methods in the secretory cells of the subcommissural organ of the guinea pig and albino rat.The activity of non-specific cholinesterase at light microscopical level appeared slightly stronger compared to the activity of the specific cholinesterase. No differences in the localizations of the both enzymes could be noticed.In electron microscopic specimens no differences could be observed between the localization or intensity of the specific and non-specific cholinesterase reactions except some nerve fibres round the secretory hypendymal cells in which only a specific cholinesterase reaction product was noticed. The reaction product was found mainly in the cytoplasmic and nuclear membranes, in the rough and smooth surfaced endoplasmic reticulum and around some secretory granules in the ependymal and hypendymal secretory cells of the subcommissural organ in guinea pig and albino rat.The possible role of cholinesterases in the secretory cells of the subcommissural organ is further discussed. Their participation in the metabolism and/or secretion of the hormonal end products is suggested.     

Essence of keratin in lips and associated structures of a freshwater fish Puntius sophore in relation to its feeding ecology: Histochemistry and scanning electron microscope investigation

Morphological specializations in the lips and associated structures of Puntius sophore were examined by scanning electron microscopy and histochemically. The upper lip (UL), in P. sophore, is associated with the horny upper jaw sheath (HUJS) on its ventral side and with the rostral cap (RC) on its dorsal side through a thin and extensive fold of skin (FSUR). The lower lip (LL) is greatly enlarged, conspicuous and associated with horny lower jaw sheath (HLJS) on the dorsal side and ventrally continues with ventral head skin (VHS). On the lateral sides there is a thin and extensive fold of skin (FSLS) between the lower lip and VHS. In contrast to the mucogenic epithelia of the UL, LL, the RC and fold of skins, the horny jaw sheaths are keratinized in nature and surface epithelial cells are characteristically modified into unculi. The UL and the LL are equipped with epithelial cells (EC), mucous cells (MC) and taste buds (TB) while in addition to these cells club cells (CC) are also present in the RC. Keratin found in unculi is an extremely strong protein which is tough and insoluble, they form the hard but un-mineralized structures. Keratin in unculi could be regarded as an adaptation for browsing or scraping food materials from the substrate as the fish grubs about the bottom. The elaboration of mucus is considered to lubricate the surface and protect the epithelia from abrasions. Taste buds are associated to locate and select palatable food and to trigger a ‘pick-up’ reflex.