Monoclonal antibody specific to urochordate Botryllus schlosseri pyloric gland

We describe here the development of a new hybridoma cell line, CF12F6, that produces a specific antibody to Botryllus schlosseri (a colonial tunicate). The monoclonal antibody was isotyped as IgG1 (by enzyme-linked immunosorbent assay), and the cellular localization of the antigenic epitope that reacts specifically with CF12F6 was confined to the cells of the pyloric gland of the zooid (by immunohistochemistry). The pyloric gland participates in osmoregulation, digestion, glycogen storage, and various other secretion functions that will be studied further by the use of monoclonal antibody CF12F6, the first in botryllid ascidians that recognizes cells of a whole, single organ.     

The biochemical composition of the pyloric caeca and its relation to DNA levels in the female starfishAsterias rubens during the reproductive cycle

Summary The biochemical composition of the pyloric caeca of femaleAsterias rubens was studied during the annual reproductive cycle. The amounts of nucleic acids, DNA and RNA were measured in addition to the amounts of lipids, glycogen, other reducing carbohydrates, free amino acids and proteins.A positive correlation of the pyloric caeca index to the biochemical constituents at the cellular level was established. The total DNA content of the pyloric caeca was found to increase with the pyloric caeca index.The relation of the biochemical composition of the pyloric caeca to the size changes of the organ is discussed. The hypothesis that the changes in size of the organ are due to changes both in the number and size of the cells is confirmed. The biochemical composition of the pyloric caeca and its relation to the reproductive cycle is discussed.Abbreviations GI gonad index - CPI pyloric caeca index     

Functions of digestive diverticula in marine invertebrates

Summary The movement of labelled glucose along the digestive tract of two ascidian species, Sidnyum argus and Dendrodoa grossularia is compared. In Dendrodoa grossularia, glucose absorption takes place in all parts of the digestive tract except the oesophagus. Labelled glucose passes through the digestive tract in less than 12 h, and it accumulates in the pyloric ampullae in less than 3 days. In this species, the pyloric gland has more capacity to stock glucidic compounds than other tissues. Under normal feeding conditions, after five days, there is no transfer of labelled glucose to other tissues. Whatever the substances given to Sidnyum argus, its pyloric gland never accumulates labelled products.     

A study of the histology and morphology of the digestive tract of the common eel (Anguilla anguilla)

The anatomy and morphology of the digestive tract of the eel was examined using light and scanning electron microscopy. The oeosphagus showed complex longitudinal folding; stratified epithelium, columnar epithelium and goblet cells striated muscle fibres formed the thick muscular coat. The pneumatic duct entered the oesophagus anterior to the oesophagastric junction. The Y shaped stomach showed large well developed folds which decreased in size and number towards the pyloric sphincter. Columnar epithelium and gastric gland cells were present, the latter being absent from the pyloric region.
The intestine had a heavily thickened anterior region, signs of convulation were noted prior to the ileorectal valve. Intestinal folding showed a complex reticulate pattern with columnar epithelium and goblet cells present. The mucopolysaccharides were studied in the goblet and columnar cells throughout the regions of the gut. Lymphocytes and eosinophilic type cells were found in the connective tissue of the mucosa throughout the gut. The pancreas was a compact organ with few Islets of Langerhans, beta cells were peripherally situated and alpha cells centrally. The unilobular liver acted as a storage organ for oil and glycogen.     

Muscle glycogen synthase isoform is responsible for testicular glycogen synthesis: Glycogen overproduction induces apoptosis in male germ cells

Glycogen is the main source of glucose for many biological events. However, this molecule may have other functions, including those that have deleterious effects on cells. The rate‐limiting enzyme in glycogen synthesis is glycogen synthase (GS). It is encoded by two genes, GYS1, expressed in muscle (muscle glycogen synthase, MGS) and other tissues, and GYS2, primarily expressed in liver (liver glycogen synthase, LGS). Expression of GS and its activity have been widely studied in many tissues. To date, it is not clear which GS isoform is responsible for glycogen synthesis and the role of glycogen in testis. Using RT‐PCR, Western blot and immunofluorescence, we have detected expression of MGS but not LGS in mice testis during development. We have also evaluated GS activity and glycogen storage at different days after birth and we show that both GS activity and levels of glycogen are higher during the first days of development. Using RT‐PCR, we have also shown that malin and laforin are expressed in testis, key enzymes for regulation of GS activity. These proteins form an active complex that regulates MGS by poly‐ubiquitination in both Sertoli cell and male germ cell lines. In addition, PTG overexpression in male germ cell line triggered apoptosis by caspase3 activation, proposing a proapoptotic role of glycogen in testis. These findings suggest that GS activity and glycogen synthesis in testis could be regulated and a disruption of this process may be responsible for the apoptosis and degeneration of seminiferous tubules and possible cause of infertility. J. Cell. Biochem. 114: 1653–1664, 2013. © 2013 Wiley Periodicals, Inc.     

Mechanisms of programmed cell death in the midgut and salivary glands from Bradysia hygida (Diptera: Sciaridae) during pupal–adult metamorphosis

Programmed cell death is involved with the degeneration/remodeling of larval tissues and organs during holometabolous development. The midgut is a model to study the types of programmed cell death associated with metamorphosis because its structure while degenerating is a substrate for the formation of the adult organ. Another model is the salivary glands from dipteran because their elimination involves different cell death modes. This study aimed to investigate the models of programmed cell death operating during midgut replacement and salivary gland histolysis in Bradysia hygida. We carried out experiments of real‐time observations, morphological analysis, glycogen detection, filamentous‐actin localization, and nuclear acridine orange staining. Our findings allow us to establish that an intact actin cytoskeleton is required for midgut replacement in B. hygida and nuclear condensation and acridine orange staining precede the death of the larval cells. Salivary glands in histolysis present cytoplasmic blebbing, nuclear retraction, and acridine orange staining. This process can be partially reproduced in vitro. We propose that the larval midgut death involves autophagic and apoptotic features and apoptosis is a mechanism involved with salivary gland histolysis.     

The annual pyloric cycle of Asterias rubens L. (echinodermata: Asteroidea)

Observations of biochemical changes in the pyloric caeca of Asterias rubens L. in different nutritional states and at different stages of the reproductive cycle show the existence of an annual cycle in the pyloric caeca. This pyloric cycle is closely related to that of the gonads. During the gonadal rest (May to September, when the gonads are barely visible) the pyloric caeca grow and store glycogen. During gonadal activity (October to April), they progressively autolyse and first accumulate, then liberate, lipid reserves. This winter pyloric autolysis provides the gonads with the structural material necessary for their annual growth. The summer pyloric growth must be interpreted as the accumulation of the structural material. Under normal conditions, during gonadal rest, the pyloric glycogen reserves ensure the survival of the animal. Pyloric lipid reserves appear only at the beginning of gonadal activity. Their synthesis is intimately related to the start of the gonadal activity of which they are the principal energy source. We believe that the pyloric cycle is under control of hormones that could be transported through the haemal system.The pyloric caeca are the only reserve organs of starfish. Neither the other digestive organs nor the body wall accumulate energy reserves. During starvation of medium duration (2 to 4 weeks) the pyloric caeca suffice to ensure the survival of the animal by liberating their reserve material and simultaneously starting to autolyse. During prolonged starvation, general autolysis occurs and the size of the starfish diminishes.     

An electron microscopic study of the salivary gland of Rhynchosciara angelae

Summary The structure of the salivary gland of the dipteran insect Rhynchosciara angelae in a defined stage of the larval development, characterized by the synthesis and storage of secretion product, is described. Observations were made with both Nomarski optics and electron microscopy. Filiform projections extending into the lumen of the gland were observed in the apical portion of the cells. At the basal region junctions, characterized as hemidesmosomes, were observed between the membrane of the cell and the basal lamina. The plasma membrane presents numerous infoldings into the cell increasing considerably the surface area at this region. Throughout the cytoplasm of the gland cells numerous mitochondria, Golgi complexes, microtubules, profiles of endoplasmic reticulum, secretion granules and glycogen granules were observed. Carbohydrates were detected on ultrathin sections by using the periodic acid-silver methenamine and the periodic acid-thiosemicarbazide-silver proteinate techniques.     

Helicobacter pylori infection up-regulates gland mucous cell-type mucins in gastric pyloric mucosa

Background. Two types of mucous cell are present in gastric mucosa: surface mucous cells (SMCs) and gland mucous cells (GMCs), which consist of cardiac gland cells, mucous neck cells, and pyloric gland cells. We have previously reported that the patterns of glycosylation of SMC mucins are reversibly altered by Helicobacter pylori infection. In this study, we evaluated the effects of H. pylori infection on the expression of GMC mucins in pyloric gland cells. Methods. Gastric biopsy specimens from the antrums of 30 H. pylori‐infected patients before and after eradication of H. pylori and 10 normal uninfected volunteers were examined by immunostaining for MUC6 (a core protein of GMC mucins), α1,4‐N‐acetyl‐glucosaminyl transferase (α4GnT) (the glycosyltransferase which forms GlcNAcα1‐4Galβ‐R), and GlcNAcα1‐4Galβ‐R (a GMC mucin‐specific glycan). Results. MUC6, α4GnT, and HIK1083‐reactive glycan were expressed in the cytoplasm, supranuclear region, and secretory granules in pyloric gland cells, respectively. The immunoreactivity of MUC6 and α4GnT, but not of GlcNAcα1‐4Galβ‐R, in the pyloric gland increased in H. pylori‐associated gastritis, and after the eradication of H. pylori, the increased expression of MUC6 and α4GnT in the gastric mucosa of H. pylori‐infected patients decreased to almost normal levels. This up‐regulation was correlated with the degree of inflammation. Conclusions. In addition to the synthesis of GMC mucins increasing reversibly, their metabolism or release may also increase reversibly in H. pylori‐associated gastritis. The up‐regulation of the expression of gastric GMC mucins may be involved in defense against H. pylori infection in the gastric surface mucous gel layer and on the gastric mucosa.     

Amplification and invasiveness of epithelial progenitors during gastric carcinogenesis in trefoil factor 1 knockout mice

Abstract. Objective: It is not known whether or not epithelial progenitors of the pyloric antrum are involved in gastric carcinogenesis. Normally, these progenitors give rise to two main cell lineages: pit and gland mucous cells. This study was designed to examine the changes that occur in pyloric antral mucous cell lineages and their progenitors during development of gastric adenoma and carcinoma in trefoil factor 1 (TFF1) knockout mice. Materials and methods: Pyloric antral mucosal tissues of TFF1 knockout mice at ages from 3 days to 17 months were processed for histochemical analysis using Ulex europaeus and Grifforia simplifolica lectins as markers for pit and gland mucous cells, respectively. The dividing epithelial progenitors were identified by using immunohistochemical and electron microscopy techniques. Results: TFF1 loss was associated with amplification of both mucus‐secreting pit and gland cells. Both lectins examined bound not only to mature mucous cells, but also to most of epithelial progenitors which gradually amplified with age and frequently were seen in mitosis. Analysis of 12‐ to 17‐month‐old TFF1‐deficient stomachs revealed occasional groups of poorly differentiated mucosal cells with features similar to those of epithelial progenitors (or stem cells), in the basal portion of the antral mucosa. These cells eventually invaded the muscularis mucosa while maintaining some capacity to differentiate. Conclusion: This study shows that the progenitors of pit and gland mucous cells contribute to gastric carcinogenesis in the pyloric antrum of TFF1 knockout mice, strongly supporting the concept of stem cell origin of cancer.     

Storage metabolism in the Pacific oyster (Crassostrea gigas) in relation to summer mortalities and reproductive cycle (west coast of France)

We describe seasonal changes in the biochemical composition of digestive gland, adductor muscle and gonad and surrounding mantle area in Crassostrea gigas from the Western Atlantic coast of France. Seasonality in histology of storage tissues and glycogen storage capacity in isolated vesicular cells were also studied. Proteins, the main muscle components did not contribute to the gametogenetic effort. Glycogen and lipids were stored in the digestive gland, gonad and surrounding mantle area during the wintering period and the gonad and surrounding mantle area represented the main storage compartment supplying the reproductive effort. Gametogenesis in spring and summer was associated with an increase in lipid and protein contents and took place at the expense of glycogen reserves. Histological study of storage tissue in the gonad led us to define four seasonal stages of storage tissue development. In vitro, glycogen storage capacity in isolated vesicular cells was high from November to March and markedly reduced during gametogenesis, decreasing below detectable levels after spawning. This physiological state should be taken into account with relation to summer mortalities occurring in commercial growing areas.     

Role of eyestalk hormone in the carbohydrate metabolism of a marine penaeid prawn,Parapenaeopsis hardwickii (MIERS) (Crustacea,Decapoda, Penaeidae)

Data furnished here concern with the role of eyestalk hormone in the regulation of carbohydrate metabolism in Parapenaeopsis hardwickii. Bilateral eyestalk ablation has brought about a significant (P < 0.01) fall and rise in the glycogen content in the midgut gland and abdominal muscle respectively. Although eyestalk ablation resulted in a significant (P < 0.01) depletion of fat in midgut gland, n0 significant (P > 0.05) change was observed in the abdominal muscle. Eyestalk extract administration in eyestalk-less prawns has significantly (P < 0.05) restored the glycogen and fat metabolites in the midgut gland. There was an obvious change in the glycogen content of the midgut gland and abdominal muscle of normal prawns when injected with eyestalk extracts from prawns in different molting stages. Eyestalk extract from intermolt prawns caused a significant (P < 0.05) decrease and increase in the glycogen quantity in the midgut gland and abdominal muscle respectively. Eyestalk extract from premolt and postmolt prawns has, although not significantly (P > 0.05), decreased and increased the utilization of glycogen respectively in the midgut gland. The physiological significance of these findings are discussed briefly.Paper forms part IV of the series

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Microanatomy of the digestive tract and accessory organs of the Japanese flathead (Inegocia japonica Cuvier, 1829) (Scorpaeniformes,Platycephalidae)

The Japanese flathead, Inegocia japonica Cuvier, 1829 is a commercially important fish in small-scale coastal fisheries in Thailand; however, an explanation of its digestive biology is missing. This study describes the digestive tract and accessory organs of I. japonica, using morphological and histological methods. The fish (10 individual fish, 24.5 ± 0.98 cm in total length) were obtained from Libong Island, Thailand. Integrated morphological and histological data showed that the digestive tract was composed of oesophagus, stomach, pyloric caeca and intestine, with accessory organs. All digestive tracts consisted of four layers, including mucosa, submucosa, muscularis and serosa. Two stomach regions were identified (cardiac and pyloric stomachs). Several clusters of gastric glands were identified in the cardiac stomach. Each gland was a unicellular structure. The apical surface of this gland contained the vacuolar cell. The intestine was lined with a simple columnar structure with goblet cells that was similar to pyloric caecum. Goblet cells were rare in the anterior intestine, in contrast to the posterior intestine where goblet cells were abundant. The numerous of hepatocyte was mostly observed in the liver, whereas an exocrine acinar cell of pancreas was also identified. The results of our observations provided the first information of the digestive tract of I. japonica and can be applied to advanced study, such as physiology and histopathology.     

Ultrastructure of the cardiac and pyloric glands of the gastric mucosa of the South African hedgehog,Atelerix frontalis

The cardiac and pyloric glands in the gastric mucosa of the South African hedgehog, Atelerix frontalis, are described. The cardiac area of the stomach contains proper cardiac glands and lacks undifferentiated fundic glands. The cardiac glands are simple tubular, coiled, and lined with columnar cells ultrastructurally similar to those of the gastric surface epithelium. Secretory granules with varying electron densities fill the apical cytoplasm of these cells. In contrast to other mammals, these glands lack mucous neck cells. The neck of the pyloric glands contains only a single cell type, whereas the basal regions of these glands contain “light” and “dark” cells. The secretory granules in the “dark” cells and the pyloric neck cells have a moderate electron density and often contain an electron dense core. An electron-lucent cytoplasm with numerous polysomes is characteristic of the “light” cells. Some “light” cells contain electron-dense granules in the apical cytoplasm. The presence of only neutral mucins in the cardiac gland cells denotes the absence of mucous neck cells. The acidic mucins within the pyloric neck cells seem to indicate that these cells are mucous neck cells, whereas the neutral mucins within the basally located pyloric gland cells show at least a partial functional difference from the pyloric neck cells. © 1993 Wiley-Liss, Inc.     

Peculiarities of the male urogenital apparatus of two grunt species (Teleostei, Haemulidae)

An unusual testicular structure in the white grunt, Haemulon plumieri , and the French grunt, H. flavolineatum , and a urinary exocrine gland in H. flavolineatum are described. The testes of both species are comprised predominantly of efferent ducts, with spermatogenetic tissue restricted to the gonadal periphery. The epithelial cells of the efferent ducts produce glycogen which may serve a role in the storage and nutrition of sperm. The urinary gland is a male accessory gland thought to be associated with reproduction. The gland is mesonephric in origin and the epithelial cells of the gland produce mucins which may be involved in intraspecific chemical communication.     

Post-embedding staining of rat gastric mucous cells with lectins

Summary The mucous cells of the rat stomach were stained with lectins by two post-embedding staining methods for electron microscopy. The mucous granules of surface mucous cells and foveolar mucous cells were stained weakly by Ricinus communis agglutinin-ferritin and wheat germ agglutinin-ferritin. The mucous granules of mucous neck cells were stained by concanavalin A-ferritin, Ricinus communis agglutinin-ferritin and wheat germ agglutinin-ferritin. The mucous granules of pyloric gland cells showed an affinity for wheat germ agglutinin-ferritin and concanavalin A-ferritin, while Ricinuscommunis agglutinin-ferritin only slightly stained the granules. The granules of mucous neck cells and pyloric gland cells were also stained by the concanavalin A-horseradish peroxidase-colloidal gold method, but the granules of surface and foveolar mucous cells were not stained by this method. Periodic acid oxidation of the sections before the standard concanavalin A-ferritin procedure enhanced the staining of the granules of mucous neck cells and pyloric gland cells slightly. Reduction of the sections after the periodic acid oxidation weakened the staining. Similar results were obtained using the concanavalin A-horseradish peroxidase-colloidal gold method. Though the staining with Ricinus communis agglutinin-ferritin was inhibited by periodic acid oxidation of the sections before staining, the staining with wheat germ agglutinin-ferritin was not inhibited by the oxidation. It is suggested that the paradoxical staining is closely related to the position of the concanavalin A-binding sugar residues in the carbohydrate chains.This work was supported in part by a grant-in-aid (No. 457008) from the Ministry of Education, Science and Culture, Japan and a grant-in-aid for cancer research (55-21) from the Ministry of Health and Welfare, Japan     

Morphological changes in external and internal feeding structures during the transition phyllosoma-puerulus-juvenile in the Western Rock Lobster (Panulirus cygnus,Decapoda: Palinuridae)

External and internal feeding structures of the pelagic final phyllosoma, the transitional puerulus, and the benthic juvenile Western Rock Lobster, Panulirus cygnus, were studied by means of scanning electron microscopy. The study revealed that the external feeding structures of phyllosomata are well equipped for capture and mastication of food. The foregut, however, is not clearly divided into pyloric and cardiac regions and a gastric mill is absent, although a comb row and gland filter are present. Juveniles, on the other hand, have a well-developed gastric mill and gastric teeth, and a cardiopyloric valve separates the foregut into cardiac and pyloric regions. External mouthparts of juveniles are suitable for mastication of solid food particles and bear numerous setae. In contrast, external mouthparts of pueruli are largely non-setose. Furthermore, although the foregut is differentiated into pyloric and gastric regions and a gland filter and comb row are present, a functional gastric mill is absent during the puerulus stage. Absence of such structures indicates that the puerulus may be a non-feeding stage. It is postulated that absence of (or reduced) feeding may be a response to an increased risk of predation rather than a result of the considerable morphological changes taking place during the transition from a planktonic to a benthic lifestyle, as has been previously proposed. © 1994 Wiley-Liss, Inc.     

An autoradiographic investigation of the proliferative activity of pyloric caeca and gondas of Asterias rubens

The proliferative activity of the pyloric caeca of Asterias rubens was investigated. Autoradiographic experiments using intracoelomically injected (methyl-³H)-thymidine were performed throughout the year and incorporation into pyloric caeca and into gonads was studied. Tritiated thymidine was found to be incorporated mainly in the coelomic lining of both organs. Cell divisions in the coelomic lining may be necessary for the growth of these organs, for the production of coelomocytes or, in the case of the pyloric caeca, for growth of the digestive epithelium. Proliferative activity of the digestive epithelium of the pyloric caeca was only observed in the median duct. It is hypothesized that new cells, arising from mitosis, grow from the median duct to the side lobes and differentiate into storage cells, for example. The existence of a mitosis-inducing or mitosis-stimulating substance is discussed. In the ovaries follicle cells were found to incorporate (methyl-³H)-thymidine; in the testis, proliferation of the germinal epithelium occurred simultaneously in all spermatogenic columns. First, the spermatogonia and then later the spermatocytes became labeled. Absorption of substances from the coelomic fluid is discussed.     

Seasonal variations in biochemical composition during the reproductive cycle of the veined rapa whelk Rapana venosa (Valenciennes, 1846) from the northern coast of China

Seasonal variations in the biochemical composition of Rapana venosa in relation to reproductive cycle and environment on the northern coast of China were investigated from March 2012 to February 2013. The results indicated that R. venosa has an annual reproductive cycle with synchronized gonad development in both females and males. Gametogenesis was initiated in September and gametes developed slowly during the winter, followed by rapid gonad development during spring and summer. Most individuals from this study were sexually mature between May and June, and gamete release occurred mainly between May and August. The peak of spawning was found in July and the recovery of the gonad was observed between August and November. The key biochemical components including glycogen, protein and lipid were analysed in four tissues, specifically the gonad, digestive gland, mantle and foot. The declining glycogen content in the gonad, digestive gland and mantle during maturation suggested that glycogen was consumed during the development of the gonad. Lipids and protein can be stored in the digestive gland and used during the winter in a period of food shortage. The protein and lipid contents in the ovaries increased during gonad development, which suggested that the protein and lipid had been accumulated as vitellin in oocytes.