Three-dimensional high-resolution reconstruction of the human gastro-oesophageal junction.

The aim of this study was to obtain detailed information regarding the three-dimensional structure of the gastro-oesophageal region, and, in particular, the fiber orientation of the different muscle layers of the junction. This was achieved by a study of an en bloc resection of the gastro-oesophageal junction (GOJ) harvested from a human cadaver. The excised tissue block was suspended in a cage to preserve anatomical relationships, fixed in formalin and embedded in wax. The tissue block was then processed by a custom-built extended-volume imaging system to obtain the microstructural information using a digital camera which acquires images at a resolution of 8.2 microm/pixel. The top surface of the tissue block was sequentially stained and imaged. At each step, the imaged surface was milled off at a depth of 50 microm. The processing of the tissue block resulted in 650 images covering a length of 32.25 mm of the GOJ. Structures, including the different muscle and fascial layers, were then traced out from the cross-sectional images using color thresholding. The traced regions were then aligned and assembled to provide a three-dimensional representation of the GOJ. The result is the detailed three-dimensional microstructural anatomy of the GOJ represented in a new way. The next stage will be to integrate key physiological events, including peristalsis and relaxation, into this model using mathematical modeling to allow accurate visual tools for training health professionals and patients.

Crura ultrastructural alterations in patients with hiatal hernia: a pilot study.

BACKGROUND: Laparoscopic fundoplication for gastroesophageal reflux disease (GERD) and hiatal hernia has been validated worldwide in the past decade. However, hiatal hernia recurrence still represents the most frequent long-term complication after primary repair. Different techniques for hiatal closure have been recommended, but the problem remains unsolved. The authors theorized that ultrastructural alterations may be implicated in hiatal hernia. Thus, this study was undertaken to investigate the presence of these alterations in patients with or without hiatal hernia. METHODS: Samples from Laimer-Bertelli connective membrane and muscular crura at the esophageal hiatus were collected from 19 patients with GERD and hiatal hernia (HH group), and from 7 patients without hiatal hernia enrolled as the control group (NHH group). Specimens were processed and analyzed by transmission electron microscopy. RESULTS: Muscle and connective samples from the NHH group did not present any ultrastructural alteration that could be detected by transmission electron microscopy. Similarly, connective samples from the HH group showed no ultrastructural alterations. In contrast, all muscle samples from the HH group exhibited sarcolemmal alterations, subsarcolemmal vacuolar degeneration, extended disruption of sarcotubular complexes, increased intermyofibrillar spaces, and sarcomere splitting. CONCLUSION: The evidence of ultrastructural alterations in all the patients in the HH group raises the suspicion that the long-term outcomes of antireflux surgery depend not only on the surgical technique, but also on the underlying muscular diaphragmatic illness.

Ciliated surface in the esophagogastric junction zone: a precursor of Barrett's mucosa or ciliated pseudostratified metaplasia?

We have previously reported squamous metaplasia-like change at the esophagogastric junction (EGJ). In the present study, we examined these lesions histologically and by immunohistochemistry and electron microscopy. Samples of EGJ mucosa, 3 cm long and comprising 1.5-cm long portions of both columnar and squamous mucosa, were obtained from 43 esophagectomy resection specimens. Squamous metaplasia-like change was observed in 21 (49%) of the 43 cases. The squamous metaplasia-like change was generally positive with immunohistochemical stains for tubulin and cytokeratins (CKs) 4, 7, 8, 13, and 18, and was generally negative with stains for CKs 10, 14, and 20. This pattern of immunoreactivity is very similar to that of bronchial mucosa. Also, many cilia were detected at the apices of the cells by electron microscopy in 5 (31%) of the 16 cases that were able to be examined. Therefore, squamous metaplasia-like change at the EGJ has both a similar appearance and a similar immunohistochemical profile to respiratory bronchial epithelium. These findings may suggest that squamous metaplasia-like change at the EGJ is not a precursor of Barrett's mucosa but rather is a form of pseudostratified metaplasia.

Murine peptidoglycan recognition proteins PglyrpIalpha and PglyrpIbeta are encoded in the epidermal differentiation complex and are expressed in epidermal and hematopoietic tissues.

Peptidoglycan recognition proteins (PGRPs or PGLYRPs) are pattern recognition molecules that are found in insects and mammals and are critical for innate immune responses. PGRPs bind peptidoglycan, a ubiquitous component of bacterial cell walls, and are involved in killing bacteria, degrading peptidoglycan, and initiating host defense reactions. Relatively little is known about the four mammalian PGRPs. In this article, we report the sequences of mouse PglyrpIalpha and PglyrpIbeta and provide details of their expression in wild-type mouse tissues. PglyrpIalpha and PglyrpIbeta are encoded within the epidermal differentiation complex on mouse chromosome 3F. Both genes are expressed in epidermal and hematopoietic tissues. PglyrpIbeta is expressed in each of 16 tissues tested, while PglyrpIalpha expression is limited to fewer tissues, including the lung and spleen as well as several tissues of the digestive system. Both proteins are expressed in epithelial cells throughout the gut, and immunohistochemical staining shows expression in salivary glands, the squamous epithelium of the stomach, and the villi of the jejunum. Immunohistochemical staining further shows expression of both PglyrpIalpha and PglyrpIbeta in macrophages in the spleen. PglyrpIalpha is not expressed in resting RAW264.7 macrophage-like cells, but is induced by stimulation with lipopolysaccharide. PglyrpIbeta is constitutively expressed in RAW264.7 cells and is unaffected by lipopolysaccharide or peptidoglycan stimulation. Computational and experimental data suggest that these proteins are secreted. This work provides a step toward understanding the roles of PglyrpIalpha and PglyrpIbeta in host defense and chronic inflammatory conditions induced by bacteria or their components.

Ultrastructural analysis of spinal primary afferent fibers within the circular muscle of the cat lower esophageal sphincter.

We have analyzed the ultrastructural characteristics and environment of spinal primary afferent fibers that run within the circular muscle of the cat lower esophageal sphincter. These were selectively labeled by anterogradely transported cholera toxin B subunit conjugated with horseradish peroxidase. Most of the labeled fibers were perpendicular to the muscle cells but some ran sinuously or parallel to the muscle cells. All the labeled fibers were unmyelinated and exhibited relatively rare varicosities. Most of the fibers were in large nerve fiber bundles surrounded by perineurium and probably project to the mucosa. Only some fibers that were in small nerve fiber bundles with no perineurium ran parallel to the musculature and established close relationships with smooth muscle cells. They might be a small subpopulation of the spinal tension receptors, most of the other spinal tension receptors being located in the myenteric plexus area, between the circular and longitudinal muscle.

Neural inhibition in the rat lower esophageal sphincter: role of beta 3-adrenoceptor activation.

1. Electrical field stimulation (EFS) of the rat lower esophageal sphincter produced tetrodotoxin-sensitive relaxant responses. 2. The relaxant responses were not significantly affected by propranolol but were attenuated by 6-hydroxydopamine and cyanopindolol, indicating beta 3-adrenoceptor activation. 3. The relaxant responses were significantly reduced in tissues previously exposed to BRL 37344, a beta 3-adrenoceptor agonist. 4. It was concluded that EFS-induced relaxation was adrenergic in origin and was mediated by beta 3-adrenoceptors.

Beta(3)-adrenoceptors mediate smooth muscle relaxation in the rat lower oesophageal sphincter.

1. The role of beta(3)-adrenoceptors in isoprenaline-induced relaxation of carbachol-precontracted ring segments of the rat lower oesophageal sphincter (LOS) was examined. 2. Isoprenaline (10(-8)M-10(-5)M) relaxed ring segments of the LOS in a concentration-dependent manner. Propranolol (10(-7)M) had very little antagonist effect on isoprenaline-induced relaxation. 3. Dobutamine (10(-7)M-10(-4)M), salbutamol (10(-7)-10(-4)M) and BRL 37344 (10(-8)-10(-5)M) also relaxed carbachol-contracted ring segments of the LOS in a concentration-dependent manner. The relaxant responses to these agonists were similarly not antagonized by propranolol (10(-7)M). 4. Cyanopindolol (10(-6)M), produced a parallel rightward displacement of isoprenaline, dobutamine, salbutamol and BRL 37344 concentration-response curves with similar potencies. The pKB values range from 7.3 +/- 0.1 to 7.7 +/- 0.2. 5. The relaxant effect of isoprenaline in the rat LOS was not inhibited by NG-nitro-L-arginine (L-NOARG; 3 x 10(-5)M), glibenclamide (10(-5)M) or tetraethylammonium (1 mM). 6. It was concluded that beta(3)-adrenoceptors mediate isoprenaline-induced relaxation in rat lower oesophageal sphincter and that activation of these receptors was not linked to either ATPase-, Ca(2+)-dependent K+ channels or to NO release.

Morphological characterization of the squamocolumnar junction of the esophagus in patients with and without Barrett's epithelium.

Barrett's esophagus is a metaplastic condition in which the normal stratified squamous epithelium of the distal esophagus is replaced by columnar epithelium. Our group has previously characterized a unique surface cell (the "distinctive cell") at the junction of squamous and Barrett's epithelium. This cell is notable for the simultaneous presence on its surface of both squamous and columnar cell features. The aims of our present study were, first, to evaluate prospectively the frequency with which Barrett's patients have the distinctive cell at the squamo-Barrett's junction; second, to further elucidate the characteristics of the distinctive cell; and third, to perform a combined morphological study of the squamo-Barrett's junction using scanning electron microscopy followed by transmission and light microscopy. We divided study patients into two groups: Group I consisted of Barrett's patients and group II of non-Barrett's control patients. Of eight group I Barrett's patients with junctional biopsies, three were noted to have the distinctive cell (37.5%). In contrast, this cell was not observed in any of the group II control patients. Biopsies in control patients as well as Barrett's patients without the distinctive cell revealed abrupt squamogastric or squamo-Barrett's junctions by scanning electron microscopy and light microscopy. In contrast, biopsies from the Barrett's patients with the distinctive cell revealed junctions that were not abrupt and had the distinctive cells overlying normal squamous epithelium. By scanning electron microscopy, the distinctive cells were flattened, polygonal cells with surface microvilli (a columnar cell feature) and were demarcated from one another by shallow depressions, or by intercellular ridges (a squamous cell feature). By transmission electron microscopy, the distinctive cells were cuboidal in shape with abundant apical microvilli and secretory vesicles. We have confirmed that distinctive cells are present in some Barrett's patients. This cell is a morphologic hybrid, sharing features of both squamous and columnar cells, and may be analogous to hybrid cells identified in other locations that undergo metaplasia (eg, the human cervix). Its origin may be the result of transformation of multipotential basal cells of squamous epithelial origin. We hypothesize that the distinctive cells may represent an intermediate stage in the development of Barrett's epithelium.

Histology of the mucosa of the oesophagogastric junction and the stomach in adult Rana perezi.

The histological structure of the frog digestive mucosa changes at the oesophagogastric junction. The pseudostratified ciliated mucosal epithelium of oesophageal type changes to a simple mucus-secreting epithelium of gastric type. The glands straighten and the muscularis mucosae develops as a complete layer. The muscularis increases in thickness. Unlike the mammalian stomach, in the frog the surface of the plicae forms convoluted ridges that delimit furrow-shaped pits. Two types of gastric glands are distinguished, fundal and pyloric. The former consist of mucous, oxynticopeptic and endocrine cells. The pyloric glandular cells are mainly of mucus-secreting type with scattered endocrine cells. Scattered endocrine cells of P, D, G, A, EC, and EC-L-like types are found in the glands along the stomach. It is concluded that the mucosal structure of the anuran oesophagogastric junction and stomach is less complicated than that of mammals, including man.

Morphological studies of the developing human esophageal epithelium.

This article focusses on the structural development of human esophageal ciliated epithelium. A combination of transmission electron microscopic (TEM), scanning electron microscopic (SEM), radioautographic, and light microscopic (LM) analyses were carried out using intact fetal tissues between 8 and 20 weeks of gestation as well as cultured esophageal explants. Up to the age of 10 weeks, the stratified esophageal epithelium consisted of two longitudinal primary folds. The surface cells were undifferentiated and contained large glycogen aggregates. Between 11 and 16 weeks, the primary folds (now up to four) had developed secondary folds. The thickness of the epithelium drastically increased (123%) in concomittance with a differentiation of surface columnar ciliated cells. These highly specialized surface cells exhibited junctional complexes and well-developed organelles with numerous microvilli interspersed among the cilia. Transverse sections revealed the internal structure of the cilia with a consistent pattern of nine doublet microtubules surrounding a central pair of single microtubules. Freeze-fracture studies illustrated the presence of a ciliary necklace composed of 6 ring-like rows of intramembranous particles. They also revealed the structure of ciliary cell tight junctions consisting of up to nine anastomosing strands (P-face) or complementary grooves (E-face). Ultrastructural studies (LM, TEM, SEM) of the esophageal squamous epithelium obtained after 15 days of culture showed that the newly formed epithelium was similar to adult human epithelium. Finally LM and SEM observations established that the esophagogastric junction was not yet well delineated, consisting of a transitional area composed of a mixture of esophageal ciliated cells and gastric columnar mucous cells.

Scaning electron microscope study of the veins at the human esofago-gastric junction

Veias da mucosa e submucosa do segmento de juncao esofagogastrica foram estudadas ao microscopio eletronico da varredura, em cinco individuos necropsiados, apos injecoes pela via gastrica esquerda, de material acrilico polimerizavel, utilizado no preparo de proteses dentarias, seguidas da metalizacao pelo metodo de "sputtering". Foram descritos dois tipos morfologicos fundamentais de veias: 1)veias com dilatacoes fusiformes ou ampulares ircunscritas e de superficies relativamente lisas, nas zonas 1, 2 e 3 (submucosa do estomago e esofago); 2) veias com numerosas pequenas dilatacoes e microdilatacoes vesiculares, na zona 3, (mucosa do esofago. O papel dessas caracteristicas e considerado na formacao de varizes do esofago.

Coexistence of vasoactive intestinal polypeptide and neuropeptide Y immunoreactivity within axon terminals in the canine and human lower esophageal sphincter: electron microscopy by a double immunogold labeling procedure.

Subcellular localization of vasoactive intestinal polypeptide (VIP) and neuropeptide Y (NPY) immunoreactivity within nerves was investigated in the lower esophageal sphincter of dogs and humans by double immunogold staining of sections prepared for electron microscopy. Coexistence of VIP and NPY immunoreactivity was clearly demonstrated in the large granular vesicles (LGVs) in axon terminals that were closely associated with the smooth muscle cells, as well as in the LGVs within the perikarya of neurons located in the myenteric plexus. Some LGVs appeared immunopositive only for VIP or NPY. This phenomenon might have been partly due to the fact that the double-labeling procedure with immunogold particles of different sizes was performed on both faces of each section. The results obtained in this study suggest that VIP and NPY are synthesized in the same neuron, stored in the same axon terminal, and released together to act on sphincter muscle cells.

Distinct muscarinic receptors, G proteins and phospholipases in esophageal and lower esophageal sphincter circular muscle.

Acetylcholine (ACh)-induced contraction of esophageal circular smooth muscle cells was inhibited by the M2 muscarinic antagonist methoctramine. In lower esophageal sphincter (LES) cells contraction was inhibited by the M3 antagonist p-fluoro-hexa-hydro-sila-difenidol (pF-HSD). Pertussis toxin (PTX) reduced ACh-induced contraction of esophageal but not of LES cells, which suggested that different receptor-linked G proteins are involved. Antibodies against G13 antagonized contraction of esophageal cells and G9-G11 antibodies antagonized contraction of LES cells. The phosphatidylinositol-specific phospholipase C (PLC) inhibitors, U-73122 and neomycin, reduced ACh-induced contraction of LES but not of esophageal cells. Conversely, propranolol and p-chloromercuribenzoic acid (pCMB), which inhibit a phosphatidylcholine-specific phospholipase D (PLD)-dependent pathway, reduced contraction of esophageal but not of LES muscle cells. At 1 and 5 sec after the administration of ACh (10(-5) M), inositol 1,4,5-trisphosphate (IP3) increased only in LES muscle, which suggested that contraction results from PLC-induced IP3 production in the LES but not in the esophagus. The IP3 receptor antagonist heparin, and depletion of intracellular Ca++ stores by thapsigargin or A23187, inhibited ACh-induced contraction of LES but not of esophageal muscle. It was concluded that ACh-induced esophageal contraction depends preferentially on M2 receptors, a PTX-sensitive G13 protein, phosphatidylcholine-specific PLD and production of diacylglycerol (DAG) and is independent of IP3 formation and the release of intracellular Ca++. Conversely, LES contraction is mediated through M3 receptors, a PTX-insensitive G9-G11 protein, activation of PLC, IP3 formation and the release of intracellular Ca++.

Detection by scanning electron microscopy of a distinctive esophageal surface cell at the junction of squamous and Barrett's epithelium.

Metaplastic columnar epithelium replaces the normal squamous epithelium in Barrett's esophagus. We characterized the surface epithelial cells of the junction between squamous and Barrett's epithelium using scanning electron microscopy and light microscopy. In four biopsy specimens from the squamous-Barrett's junction in three patients, we found a distinctive cell type having features intermediate between those of squamous and columnar epithelium. Its distinguishing characteristic is the presence on its surface of two disparate structures not normally present on the same cell in the gastrointestinal tract: microvilli (a scanning electron microscopy feature of glandular epithelium) and intercellular ridges (a scanning electron microscopy feature of squamous mucosa). The surface characteristics of this newly recognized cell were strikingly similar to those of cells found in the transformation zone of the uterine cervix, an area in which squamous epithelium physiologically replaces columnar epithelium. We also examined 28 biopsies of the gastroesophageal junction area from 14 patients with and without a history of heartburn but with no evidence of Barrett's esophagus. None of these biopsies showed the distinctive cell. We hypothesize that this distinctive cell represents an intermediate step in either the development or the healing of Barrett's epithelium, during which surface characteristics of two different cell types, columnar and squamous, coexist on the same cell.

[SEM study on the mucosa of the pre-stomach in Bubalus buffalus]. / Studio al SEM sulla mucosa dei prestomaci in Bubalus buffalus.

Morpho-structural features of pre-stomach mucosa in the first period of post-natal life in Bubalus buffalus have been studied by SEM. The rumen presents a well defined morpho-structural architecture from 10 to 100 days of life, while in reticulum and in omasum numerous morphological variations have been noticed. During the development, in fact, in these organs the establishment of a typical morphological pattern has been observed.

The actions of some beta-receptor agonists and xanthines on isolated muscle strips from the human oesophago-gastric junction.

Isolated preparations from the circular muscle layer of the human oesophago-gastric junction were mounted in organ baths and isometric tension recorded. During an equilibration period, active resting tension developed suggesting that the preparations were representing the lower oesophageal sphincter. Active tension was abolished by exposing the preparations to Ca(++)-free medium. The two xanthines theophylline and enprofylline almost equipotently relaxed the preparations in a concentration-dependent manner (10(-7)-10(-3) M). Within therapeutic concentrations, theophylline inhibited active resting tension by 30-60%, while enprofylline lowered tension by less than 20%. Inhibitory actions of adenosine were demonstrated, and this suggests that adenosine antagonism is not the mechanism of action for xanthines in the oesophagus. Non-selective beta-receptor stimulation with isoprenaline inhibited active tension by 70% (10(-7) M), while beta 2-receptor stimulation with terbutaline inhibited tension by 47% (10(-5) M). Dobutamine, believed to preferentially stimulate beta 1-receptors, inhibited active tension in a concentration-dependent manner (10(-7)-10(-4) M). Metoprolol (10(-6) M), a selective beta 1-receptor antagonist, shifted the concentration-response curve for isoprenaline to the right, but left the maximal response unchanged. It is concluded that xanthines and beta-receptor agonists have inhibitory actions on circular muscle from the human oesophagogastric junction. The experimental data suggest the presence of beta 1- as well as beta 2-receptors, both mediating inhibition of active resting tension.

Ultrastructure of interstitial cells of Cajal at the gastro-oesophageal junction of the monkey (Macaca fascicularis).

The ultrastructure of the interstitial cells of Cajal (ICC) in the oesophagus of the monkey resembled that described in the oesophagus of other mammalian species but differed in their paucity and almost lack of smooth endoplasmic reticulum, caveolae and filaments. The plasmalemma of the ICC was in close contact (20- to 30-nm gaps) with that of smooth muscle cells. This may occasionally take the form of a desmosome, but gap junctions have not been observed. Vesiculated axon profiles, containing large granular or agranular vesicles were in close contact (20- to 30-nm gaps) with the plasmalemma of ICC. In a few vesiculated profiles a presynaptic density could be recognized. The intercalation of the ICC between the vesiculated axon profiles and the smooth muscle cells suggest a role in oesophageal motility. Between 3 and 21 days following bilateral vagotomy some ICC showed regressive changes such as increased electron density and shrinkage of the cytoplasm, crowding of the organelles and dissolution of the nuclear chromatin material. Axon profiles in the vicinity of the affected ICC contained glycogen granules suggesting injury. In late stages, the number of ICC and smooth muscle contacts was reduced. The results suggest that the vagus nerves exert a trophic influence on the ICC and that the intercellular relationships between ICC and smooth muscle cells possess a degree of plasticity. It is tentatively suggested that these vagal effects may be mediated via the oesophageal myenteric ganglia.

Scanning electron microscope study of the esophagogastric junction in the tufted capuchin monkey, Cebus apella.

A scanning electron microscopic study of the esophagogastric junction in the Cebus apella monkey showed the following characteristics: Numerous irregular folds with epithelial cells polyhedral in shape and in longitudinal extension; the luminal surface of the glandular epithelium presents a dome-shaped mucous cells with a cobblestone-like topography.

Estrutura microscópica da junçäo esofagogástrica e do esfíncter esofagiano inferior na cobaia e no rato albino / Microscopical structure of the esophagogastric junction and the lower esophageal sphincter in the guinea pig and albine rat

A estruturaçäo geral do esôfago abdominal e da junçäo esofagogástrica do rato albino e da cobaia é apresentada neste trabalho. Diferenciaçöes histológicas foram observadas aos níveis das junçöes esofagogástricas em ambas as espécies as quais provavelmente estariam relacionadas à oclusäo tônica observável no esfíncter esofagiano inferior. O papel de outros fatores subsidiários, à oclusäo bem como dos componentes relaxadores ao nível da junçäo foram apresentados e discutidos, comparativamente, com o de outros mamíferos

The muscle coat of the lower esophageal sphincter in patients with achalasia and hypertensive sphincter. An electron microscopic study.

The muscle coat of the lower esophageal sphincter (LES) of seven patients with achalasia and three patients with a hypertensive sphincter has been studied with the electron microscope. In these pathological conditions the ultrastructural pictures differ both from normal and from one another. In achalasia, the LES muscle wall components (nerve endings, smooth muscle cells, interstitial cells of Cajal and connective tissue) are altered, but, while all the nerve endings and interstitial cells are affected, only a few smooth muscle cells are damaged. The severity of the alterations is more pronounced in the older patients. On the contrary, there is no damage of the muscle wall components in the hypertensive sphincter, whereas an increase in the cytoplasmic organelles (smooth endoplasmic reticulum and mitochondria) has been found in all interstitial cells and in some smooth muscle cells. Moreover, the ultrastructural picture of the hypertensive sphincter does not seem to change with patients' age. Since the LES components specifically altered in achalasia are the nerve endings and the interstitial cells of Cajal, they are regarded as principally responsible for the altered motility. On the contrary, the ultrastructural picture of the hypertensive sphincter suggests an enhancement of the activity of the activity of all the interstitial cells and of some smooth muscle cell; therefore, we consider the hyperfunction of these cells as the cause of this esophageal motor disorder.