The Effects of Deoxynivalenol on the Ultrastructure of the Sacculus Rotundus and Vermiform Appendix, as Well as the Intestinal Microbiota of Weaned Rabbits.

Deoxynivalenol (DON) is a mycotoxin found in grains that poses a potential threat to human and animal health, and the gastrointestinal tract is the primary target organ. There are few studies focused on the toxicology of DON to rabbits, especially on the relation among DON, microbiota, and the gut-associated lymphoid tissue. A total of 30 weaned rabbits (35 d) were evenly divided into the control group and DON group (1.5 mg/kg bodyweight (BW)) based on their body weight. After a 24-day trial, the ultrastructures of the sacculus rotundus and vermiform appendix were observed using a scanning electron microscope and transmission electron microscopy. The morphology and microflora in the ileum, caecum, and colon were also examined. The results proved that the ultrastructure of the sacculus rotundus and vermiform appendix, as well as the integrity of the intestinal barrier (especially for the ileum), were impaired after DON was administrated to the rabbits. Compared to the control group, the relative abundance and diversity of the microflora decreased in all three intestinal segments in the DON group, particularly in the ileum and caecum. In conclusion, the toxic effect of DON on weaned rabbits may be performed by destroying the structure of the sacculus rotundus and vermiform appendix, as well as affecting the structure and diversity of the intestinal flora.

Romanowsky-Giemsa as a counterstain for immunohistochemistry: optimizing a traditional reagent.

We describe a detailed protocol for using Romanowsky-Giemsa (RG) counterstaining on formalin fixed, paraffin embedded tissue sections that are stained immunohistochemically (IHC) after antigen retrieval using hot acidic citrate buffer. RG staining is easy to perform and provides consistent results that are similar to hematoxylin and eosin (HE) staining. The counterstaining was applied after a variety of antibodies that used the DAB chromogen and the intensity of IHC stained structures was preserved. Moreover, RG counterstaining provided finer cell detail than HE, methyl green or nuclear fast red. A detailed troubleshooting guide is provided for the RG staining protocol.

Distribution of zinc iodide-osmium positive dendritic cells in the human appendix

Dendritic cells are antigen-presenting cellsfound in almost every type of tissue, includinglymphatic tissue, blood and skin. In the intestinaltract, these cells are likely to play a pivotalrole in the initiation and regulation of immuneresponses. Our earlier study of the human colonand ileum revealed the presence of zinc iodideosmium-positive dendritic cells. In the presentstudy we demonstrate the presence of ZIO-positivedendritic cells in the human appendix.ZIO-positive cells were seen in the region ofcrypts of Lieberkühn as well as in the surfaceepithelium. The cells showed a single longprocess directed towards the lumen. They werelong, slender, and triangular. In the region ofthe lymphoid follicle, two different types ofdendritic cells were noted. The follicular dendriticcells present in the germinal center werefew in number, larger in size and with thickdendritic processes. However, in the mantlezone typical dendritic cells were seen. Theywere smaller in size and had many thin processes.The distribution of dendritic cells in thehuman appendix confirms the role of theappendix in the immune response (AU)

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A new simplified catalyzed signal amplification system for minimizing non-specific staining in tissues with supersensitive immunohistochemistry.

We investigated non-specific staining in a catalyzed reporter deposition (CARD) reaction and improved its blocking methods in supersensitive immunohistochemistry, based on our simplified catalyzed signal amplification (CSA) system (Hasui et al. 2002). In the CARD reaction using biotinyl tyramide, non-specific staining could be reduced by pretreatment with a casein solution or 3% bovine serum albumin (BSA)-phosphate buffer saline (PBS) with 0.1% Tween 20. In the CARD reaction using FITC-labeled tyramide, non-specific staining could be blocked by pretreatment with 0.3% BSA-PBS with 0.1% Tween 20 or 3% polyethylene glycol-PBS with 01% Tween 20. Thus, our new simplified CSA system features: 1) destruction of the endogenous peroxidase activity; 2) blocking of the nonspecific reaction of the primary antibody; 3) a primary antibody reaction; 4) blocking of the non-specific reaction of the polymer reagent by casein treatment; 5) a polymer reaction; 6) blocking of the non-specific reaction of CARD reaction by casein treatment; 7) a CARD reaction; and 8) detection of deposited tyramide. This new system proved useful for detecting an extremely low amount of antigen in the endogenous biotin-rich tissues such as the gastrointestinal tract and liver. By this method, the Ki67 antigen in the G1 phase cell cycle could be detected and a metabolic disorder of the Ki67 antigen was implicated in a carcinoid tumor in the stomach. We believe that this new simplified CSA system represents a new standard of supersensitive immunohistochemistry for use in light-microscopic investigation.

Bacterial translocation in the normal human appendix parallels the development of the local immune system.

Experimental modes and pathological conditions may result in bacterial translocation (BT), that is, the passage of indigenous bacteria colonizing the intestine through the intestinal mucosa to mesenteric lymph nodes. Yet no data are available on BT in the normal human gut. We determined the occurrence of BT and its extent in histologically normal, incidentally removed human vermiform appendices (VA) from individuals of different ages and correlated the findings with the development with age of associated lymphatic tissue. BT appears to pertain to normal antigen-sampling processes of the GALT in the VA. It also parallels the development of the GALT and its maintenance during adulthood. In the first two weeks after birth, when bacterial colonization of the gut evolves and when the VA lacks the protection of secretory IgA, BT was not detected. Thereafter, BT occurs along with development of the local GALT, which is fully built up after the first year. A physiological uptake of, or invasion by, bacteria may be instrumental (1) for tolerance induction against the indigenous flora and (2) for the stimulation and normal development of the GALT.

Linfoma de apêndice cecal: relato de caso / Appendicecal lymphoma cecal: case story

Introdução - O linfoma primário de apêndice é uma patologia incomum. Sua apresentação, assim como em outros tumores deste órgão, geralmente simula quadro de apendicite aguda, sendo difícil seu diagnóstico clinico. Objetivo _ Relato de caso de paciente submetido a apendicectomia, cujo diagnóstico patológico foi de linfoma primário de apêndice cecal. Material e Método _ Paciente masculino, 41 anos com sintomas de abdome agudo inflamatório. Submetido à investigação clinica e exames complementares, sendo encaminhado à cirurgia com suspeita de apendicite aguda. Resultados _ O exame patológico diagnosticou linfoma de apêndice cecal. Após a alta hospitalar o paciente foi encaminhado para tratamento oncológico complementar. Conclusão _ Sendo um tumor de localização rara neste local, sua descrição na literatura é infreqüente, com os índices de sobrevida pouco discutidos. Porém, quando o tumor é restrito ao apêndice, sem comprometimento ganglionar e com tratamento complementar adequado, o prognóstico parece ser promissor.

[Participation of a distant cholinergic mechanism in the response of the vascular bed to intoxication by organophosphorus inhibitors of cholinesterase]. / Uchastie distantnogo kholinergicheskogo mekhanizma v reaktsii sosudistogo rusla na intoksikatsiiu fosfororganicheskimi ingibitorami kholinésterazy.

Poisoning with phosphacol causes dose--dependent decrease of blood flow speed in small vessels of mesoappendix. Administration of LD50 of phosphacole results in blood stagnation, simultaneous blood pressure fall, which leads to death of part of the animals. Electron microscopic study revealed the presence of acetyl and buthyryl cholinesterase in endotheliocytes of mesoappendicular capillaries, the activity of which was completely suppressed by administration of LD50 of phosphacol. 0,0-dimethyl-0 (2,2-dichlorvinyl) phosphate LD10 caused the damage of endotheliocyte surface. It was suggested that endothelial cholino-receptors that are activated through the rise of redundant acetyl-choline level in blood on the background of cholinesterase inhibition participate in the mechanism of pathological reactions described. Such variant of toxic effect was characterized as distant.

[The distant effects of acetylcholine--links in the pathogenesis of poisoning by cholinesterase inhibitors]. / Distantnye éffekty atsetilkholina--zven'ia v patogeneze intoksikatsii ingibitorami kholinésterazy.

Scanning electron microscopy showed that the capillary endothelial cells of rats poisoned by O,O-dimethyl-O(2,2-dichlorvinyl) phosphate swell and become wrinkled, while some of the cells acquire fenestrae. In 24 h. these changes become weaker. Mass deformity of erythrocytes was seen at the same time and lasted less than 24 h. Since the capillary endothelium and the erythrocytes are devoid of cholinergic innervation but possess cholinoreceptors, the occurring effects may be explained by the distant action of acetylcholine accumulating in the blood in poisoning by cholinestarase inhibitors.

Mucin-related epitopes distinguish M cells and enterocytes in rabbit appendix and Peyer's patches.

The biochemical composition of the apical membranes of epithelial M cells overlying the gut-associated lymphoid tissues (GALT) is still largely unknown. We have prepared monoclonal antibodies (MAbs) directed against carbonate-washed plasma membranes from epithelial cells detached with EDTA from rabbit appendix, a tissue particularly rich in GALT. As determined by immunofluorescence microscopy, several MAbs specifically recognized either M cells or enterocyte-like cells of the domes from rabbit appendix, sacculus rotundus, and Peyer's patches. M cells were identified by their large ventral pocket containing lymphoid cells and by specific labeling with antivimentin. Among various characterized MAbs, MAb 104 recognized rabbit immunoglobulins and was used as an apical marker for M cells in the rabbit appendix, MAb 58 selectively stained an integral membrane glycoprotein of greater than 205 kDa located at the apex of M cells, and MAb 214 stained a smaller soluble glycoprotein associated with the apical surfaces from neighboring enterocytes. In addition, both MAbs 58 and 214 also labeled luminal mucus and secretory granules in goblet cells. The selective association of mucin-related molecules at the surfaces of either M cells or enterocyte-like cells of the follicle-associated epithelium suggests that specific carbohydrate antigens are differentially expressed by epithelial cells and could account for the differential binding properties of pathogens.

[The absorbing peripheral lymphatic apparatus and blood vessels of the vermiform appendix: the 3-dimensional and ultrastructural aspects]. / Appareil lymphatique périphérique absorbant (ALPA) et vaisseaux sanguins de l'appendice vermiforme: aspects tridimensionnels et ultrastructuraux.

We studied the lymphatic angiotectonics of the vermiform appendix of the rabbit, using direct injection of Neoprene latex, electron microscopy and three-dimensional models. We observed that the vermiform appendix is made of absorbing lymphatic vessels distributed among the tubular glands of the tunica mucosa. Moreover, these absorbing lymphatic vessels continue through the straight vessels into a vast, basket-like lymphatic network, which envelopes the medium and basal portions of each single lymphoid follicle. In addition, the vessels of this network reach the submucosal network, whose vessels are peculiar in shape and size. The submucosal network, in turn, drains the lymph through the small caliber intramuscular vessels in the valved subserous precollector lymphatic vessels. We also highlighted the ultrastructural characteristics of the absorbing lymphatic vessels, which completely lack openings at the contacts between the adjacent endothelial cells, which were always joined by specialized junction complexes. Moreover, we reported the presence of intraendothelial channels which we affirm are dynamic morphologic entities, through which lymphocyte migration from the peri-interfollicular lymphoid tissue inside the absorbing lymphatic vessels takes place.

Mucocele do apêndice - relato de um caso / Mucocele of the appendix - a case report

Os autores apresentam um caso de mucocele do apêndice cecal, sendo tal relato justificado pelo caráter incomum da patologia, bem como pela apresentação clínica pouco usual. É ainda digno de nota a dimensão excessiva atingida pelo órgão acometido.

[The ultrastructure of the epithelial endocrinocytes of the appendiceal mucosa in appendicitis patients]. / Ul'trastruktura éndokrinotsitov épiteliia slizistoi obolochki cherveobraznogo otrostka bol'nykh appenditsitom.

The electron microscopic study of the vermiform process epithelium under inflammation showed five types of endocrinocytes, based on the ultrastructure of secretory granules: EC, DI, L, I, P. The overwhelming majority of endocrinocytes form EC-cells. Besides, cells of a "mixed" type, involving exo- and endo-endocrinic cells, were revealed. Morphologic and functional alteration of the human vermiform process epithelium endocrinocytes under inflammation have been noticed.

[The endocrinocyte differentiation of the mucosal epithelium of the human appendix]. / Differentsirovka éndokrinotsitov épiteliia slizistoi obolochka cherveobraznogo otrostka cheloveka.

By electron microscopy, the cambial elements of intestinal epithelium (band cells) are revealed to be sources of formation of the vermiform process epithelium endocrinocytes. By a successive differentiation the band cells are transformed into either secretory cells with monohormonal contents or "mixed" cells. The latter in their turn can be differentiated into mucous cells or lose their mucous component by transforming into endocrinocytes. In addition, mucous cells can be transformed into endocrinocytes by a direct differentiation.

[The ultrastructure of the epithelial endocrinocytes of the appendiceal mucosa in the human fetus]. / Ul'trastruktura éndokrinotsitov épiteliia slizistoi obolochki cherveobraznogo otrostka ploda cheloveka.

An electron microscope study of the epithelium of vermiform processes of 11-26 week old human fetuses showed seven types of endocrinocytes differing in ultrastructure and dimensions of secretory granules. In addition to the six known types of endocrinocytes (EC, D, D1, I, L and P), endocrinocytes of a 7th type were discovered which are beyond the International classification of endocrinocytes of the gastroenteropancreatic system. The overwhelming majority of endocrinocytes of the vermiform process epithelium form EC-cells. Besides, cells of a "mixed" type, both exo-endocrine and endo-endocrine cells, were revealed.

Siblings with microvillous inclusion disease.

Two male siblings from a consanguinous Pakistani family had fatal diarrhoea with an onset at 24 and 48 hours after birth. A diagnosis of microvillous inclusion disease (MVID) was established by showing characteristic light and electron microscopic features in the small intestinal biopsy specimen on day 6 of life in case 1. The typical abnormalities of MVID were also demonstrated retrospectively in case 2 by examining archival appendicular tissue from 10 years previously. These cases are consistent with an autosomal recessive inheritance for MVID. Retrospective diagnosis of MVID is possible by examining appropriate archival material, which may aid genetic counselling and future research.

Differential binding of lectins to M cells and enterocytes in the rabbit cecum.

BACKGROUND: The afferent limb of the intestinal immune system is represented by the gut-associated lymphoid tissue, in which antigenic material, including complete bacteria, is taken up from the lumen by specialized epithelial cells (M cells). Because the adherence of micro-organisms to epithelial can be mediated by lectin-sugar bindings, the glycoconjugates of the surfaces of M cells and enterocytes were compared. METHODS: A set of 28 lectins and corresponding sugars was used for light and electron microscopy of fixed and unfixed sections. M cells were identified by anti-vimentin antibodies. RESULTS: M cells of the cecal lymphoid patches selectively bound lectins specific for fucose or N-acetylgalactosamine. The labeled glycoconjugates were located in the apical membrane and in the membrane of vesicles in the apical cytoplasm. Enterocytes were selectively labeled by galactose-specific lectins. In contrast, the lectin-reactivity of M cells and enterocytes did not differ in the jejunal Peyer's patches. CONCLUSIONS: The results suggest that there may be selectivity mediated by glycoconjugates in the uptake of antigenic material by cecal M cells but not by jejunal M cells.

Occluding junctions in the epithelia of the gut-associated lymphoid tissue (GALT) of the rabbit ileum and caecum.

The zonulae occludentes of the dome epithelia and adjacent non-dome epithelia in four locations of the gut-associated lymphoid tissue (GALT) in the rabbit ileum and caecum (Peyer's patches, sacculus rotundus, caecal lymphoid patches, appendix) were studied in freeze-fracture replicas. In all locations the zonulae occludentes of the dome epithelium are composed of more junctional strands than in the corresponding non-dome epithelium. In the dome epithelia of Peyer's and caecal lymphoid patches the zonulae occludentes show considerable structural variation; the number of superimposed strands is approximately 10 (range 5-18). In the dome epithelia of sacculus rotundus and appendix, in addition to zonulae occludentes, extended networks of junctional strands (fasciae occludentes) are present particularly between M-cells and enterocytes. The zonulae occludentes consist of approximately 8 to 9 (range 5-15) superimposed strands; the fasciae occludentes extend up to a depth of 20 microns on the lateral membranes. The presence of the fasciae occludentes correlates with the appearance of regularly shaped clusters of lymphocytes, which are most developed in the dome epithelia of sacculus rotundus and appendix. These results suggest (1) that in contrast to the dome epithelia of Peyer's and caecal lymphoid patches those of sacculus rotundus and appendix are compartmentalized, and (2) that the mobility of lymphocytes and diffusion of antigens in the dome epithelia of sacculus rotundus and appendix is restricted.

[A scanning electron microscopy study of the normal rabbit appendix]. / Stúdia normálneho králicieho apendixu v rastrovacom elektrónovom mikroskope.

The surface relief of the mucous membrane of the normal rabbit appendix was studied under scanning electron microscopy. Two different types of surface relief were identified. One consisted of relatively regular crests and grooves of the mucosa, the other one exhibited scattered crater-like openings into which "heads of lymphatic nodes" were inserted to varying extent and which were separated from the surface mucosa by broad alveolar cavities. Since adequate terminology has not yet been coined for the description of the observed structures, the authors use the terminology of classical histology and from analogous descriptions of other reliefs.

Immunohistochemical and ultrastructural studies of twelve argentaffin and six argyrophil carcinoids of the appendix vermiformis.

Endocrine cell types in 12 argentaffin and six argyrophil carcinoids and in nonneoplastic epithelia of the appendix vermiformis were investigated histochemically, immunohistochemically, and ultrastructurally. The nonneoplastic epithelia contained serotonin (Ser), peptide YY (PYY), glicentin (Gli), neurotensin (Neu), and somatostatin (So) cells in decreasing frequency. Out of 30 nonneoplastic Ser cells examined ultrastructurally, 28 cells were EC1 cells and two were non-EC cells. Eleven of 12 argentaffin carcinoids could be immunostained with anti-Ser serum and all of those 11 were composed almost totally of Ser cells. One of the 11 contained a small number of Neu cells. Ultrastructurally, 11 argentaffin carcinoids were composed predominantly of EC1 and/or ECn cells, and one was composed primarily of non-EC cells. Out of the six argyrophil carcinoids, five were argyrophil, non-argentaffin carcinoids; three consisted almost totally of PYY cells; one consisted of 60% PYY cells, 40% So cells and a few Gli cells; and one consisted of Ser cells alone. Ultrastructurally, the first four of those tumors were composed of D1 and/or L cells and the latter tumor was composed of ECn cells. The remaining one argyrophil carcinoid contained a few Ser-positive argentaffin cells and consisted almost totally of ECn cells which were found in both parts, with and without argentaffinity. It is concluded that the appendiceal carcinoids comprise two distinct groups on the basis of the main constituting cell type: Ser-positive, argentaffin carcinoids, composed of EC cells and peptide (especially of PYY)-positive, and Ser-negative, argyrophil non-argentaffin carcinoids of D1 and/or L cells.