Histochemical differences along the intestine of Corydoras paleatus (Siluriformes: Callichthyidae) / Diferencias histoquímicas a lo largo del intestino de Corydoras paleatus (Siluriformes: Callichthyidae)

AbstractThe Neotropical catfish Corydoras paleatus is a facultative air-breather and the caudal half of the intestine is involved in gas exchange. In South America, air-breathing fishes are found in tropical or sub-tropical freshwaters where the probability of hypoxia is high. The aim of this study was to characterize by traditional histochemical and lectinhistochemical methods the pattern of carbohydrate in the intestinal mucosa. Intestine samples were taken from 25 healthy adult specimens collected in Buenos Aires (Argentina). Samples were fixed by immersion in 10 % buffered formalin and routinely processed and embedded in paraffin wax. Subsequently, these sections were incubated in the biotinylated lectins battery. Labeled Streptavidin-Biotin (LSAB) system was used for detection, diaminobenzidine as chromogen and haematoxylin as a contrast. To locate and distinguish glycoconjugates (GCs) of the globet cells, we used the following histochemical methods: PAS; PAS*S; KOH/ PA*S; PA/Bh/KOH/PAS; KOH/PA*/Bh/PAS; Alcian Blue and Toluidine Blue at different pHs. Microscopically, the general structure of vertebrate intestine was observed and showed all the cell types characteristic of the intestinal epithelium. The cranial sector of catfish intestine is a site of digestion and absorption and its structure is similar to other fish groups. In contrast, enterocytes of the caudal portion are low cuboidal cells; and between these, globet cells and capillaries are observed, these latter may reach the mucosal lumen. Underlying the epithelium, observed a well-developed lamina propria-submucosa made of connective tissue; this layer was highly vascularized and did not exhibit glands. According to histochemistry, the diverse GCs elaborated and secreted in the intestine are associated with specific functions in relation to their physiological significance, with special reference to their role in lubrication, buffering effect and prevention of proteolytic damage to the epithelium together with other biological processes, such as osmoregulation and ion exchange. The lectinhistochemical analysis of the intestinal mucosa reveals the presence of terminal residues of glucose, mannose and galactose. In conclusion, this study has shown that GCs synthesized in the intestine of C. paleatus exhibit a high level of histochemical complexity and that the lectin binding pattern of the intestinal mucosa is characteristic of each species and the variations are related with the multiple functions performed by the mucus in the digestive tract. The information generated here may be a relevant biological tool for comparing and analyzing the possible glycosidic changes in the intestinal mucus under different conditions, such as changes in diet or different pathological stages. Rev. Biol. Trop. 64 (1): 327-340. Epub 2016 March 01.

ResumenEl pez neotropical Corydoras paleatus, de respiración aérea de tipo facultativa, utiliza el sector caudal del intestino para el intercambio gaseoso. En América del Sur, los peces con respiración aérea se encuentran en las aguas dulceacuícolas tropicales y subtropicales, donde la probabilidad de hipoxia es alta. El objetivo de este trabajo fue caracterizar mediante técnicas histoquímicas tradicionales y de lectinhistoquímica el patrón de carbohidratos de la mucosa intestinal. Para ello se utilizaron muestras de intestino de 25 ejemplares sanos adultos recolectados en la provincia de Buenos Aires (Argentina). Las muestras fueron fijadas en formol amortiguado al 10 % y se procesaron para su inclusión en parafina. Posteriormente, los cortes fueron incubados con una batería de lectinas biotiniladas. Se utilizó el sistema de marcado con estreptavidina-biotina (LSAB) para su detección, diaminobencidina como cromógeno y hematoxilina como colorante de contraste. Para localizar y diferenciar los glicoconjugados (GCs) de las células caliciformes, se utilizaron las siguientes técnicas histoquímicas: PAS, PAS*S, PAPS, KOH/PA*S, PA/Bh/KOH/PAS, KOH/PA*/Bh/PAS, Azul Alcian y Azul de Toluidina a diferentes pHs. Microscópicamente, se observa la estructura general del intestino de los vertebrados y el epitelio intestinal presenta todos los tipos celulares característicos de esta región. El sector craneal del intestino de este teleósteo, es el sitio de digestión y absorción, y posee una estructura similar a la de otros grupos de peces. En cambio, los enterocitos de la porción caudal, son células cúbicas bajas, entre ellos se observan células caliciformes y capilares sanguíneas que llegan hasta el lumen de la mucosa. Por fuera del epitelio, se observa una lámina propia-submucosa muy desarrollada compuesta por tejido conectivo, altamente vascularizada que no presenta glándulas. De acuerdo con las técnicas histoquímicas, los diversos GCs elaborados y secretados por la mucosa intestinal se encuentran asociados con funciones específicas de importancia fisiológica, como su rol en la lubricación, su efecto amortiguador y la prevención de daños proteolíticos del epitelio junto con otros procesos biológicos, tales como la osmorregulación y el intercambio iónico. El análisis lectinhistoquímico de la mucosa intestinal revela la presencia de residuos terminales de glucosa, manosa y galactosa. En conclusión, en este estudio se demuestra que los GCs sintetizados en el intestino de C. paleatus muestran un alto nivel de complejidad histoquímica y que el patrón de unión de lectina de la mucosa intestinal es característico para cada especie y las variaciones se hallan relacionadas con las múltiples funciones realizadas por el mucus en el tracto digestivo. La información brindada en este trabajo es una herramienta de relevancia biológica para comparar y analizar los posibles cambios glicosídicos del mucus intestinal bajo diferentes condiciones como los cambios en la dieta o diferentes estados patológicos.

Effects of the 2, 4-D herbicide on gills epithelia and liver of the fish Poecilia vivipara / Efeitos do herbicida 2, 4-D no epitélio das brânquias e no fígado do peixe Poecilia vivípara

The 2,4-dichlorophenoxyacetic acid, usually named 2,4-D is one of the most widely used herbicides in the world. Acute toxicity of 2,4-D herbicide was investigated through its effects on guppies (Poecilia vivipara Bloch et Schneider 1801). Fish were exposed to the herbicide at concentrations of 10, 20 and 40µl per liter of water for 24 hours to determine its effects on gills and liver epithelia. The estimated LC50 was 34.64µl of 2,4-D per liter of water. Histochemical analyses and Feulgen's reaction were conducted to detect glycoconjugates and DNA, respectively, in gills and liver epithelia. Histochemistry revealed qualitative variations of glycoconjugates present on mucous cells and granules. The four types of mucous cells contained neutral granules, acids, or both. Increasing amounts of syalomucins were observed from the control group to the group exposed to the highest concentration of 2,4-D, suggesting increased mucous viscosity and the formation of plaques that could inhibit gas exchange and osmoregulation. Lamellar fusion observed in the group exposed to 40µl of 2,4-D suggests a defense mechanism. Hepatocytes showed vacuolization in the 10 and 20µl/L groups. The 40 µl/L group showed normal hepatocytes as well as changed ones, many Ito cells, micronuclei, and nuclear swelling. These effects may be associated with toxicity or adaptative processes to cellular stress. The data from this study indicates the importance of assessing similar risks to aquatic species and suggests that Poecilia vivipara is an adequate biological model for analysis of environmental contamination.

A toxicidade aguda do herbicida 2,4-D foi investigada através dos efeitos no peixe Poecilia vivípara (Bloch et Schneider, 1801). Grupos de peixes foram expostos ao herbicida nas concentrações de 10, 20 e 40µl por litro de água, durante 24 horas. As brânquias e o fígado foram estudados. A concentração letal média (CL50) do herbicida para a espécie em questão foi de 34,64µl/l. Foram realizadas colorações histoquímicas e coloração de Feulgen para identificar glicoconjugados e DNA, respectivamente, nos tecidos acima citados. Os métodos histoquímicos revelaram os tipos de glicoconjugados presentes nas células mucosas e nos grânulos. Os quatro tipos de células mucosas apresentaram glicoconjugados neutros, ácidos, ou ambos em um mesmo tipo celular. Observou-se a presença crescente de sialomucinas do grupo controle até o grupo exposto a maior concentração de 2,4-D, sugerindo aumento da viscosidade do muco e, consequentemente, formação de placas que impedem as trocas gasosas e a osmorregulação. A fusão lamelar observada no grupo exposto a 40µl de 2,4-D sugere ser um mecanismo de defesa. Os hepatócitos apresentaram processo de vacuolização nos grupos 10 e 20µl/l. No grupo de 40µl/l, observou-se a presença de células de Ito, micronúcleos e hepatócitos normais e outros com edema nuclear. Este estudo indica a importância da avaliação de riscos semelhantes a espécies aquáticas e sugere a espécie Poecilia vivipara como modelo biológico adequado para análises de contaminação ambiental.

Lectin histochemical aspects of the ovarian lamellae epithelium of Genypterus blacodes (Schneider, 1801) / Padrão de reação lectino-histoquímico do epitélio laminar ovariano em Genypterus blacodes (Schneider, 1801)

The composition and distribution of the glycoconjugates (GCs) secreted by the epithelium of ovarian lamellae with reference to the reproductive biology of Genypterus blacodes (Schneider, 1801) through lectin hi stochemistry is here discussed. In this species, the epithelial cells that line the ovarian cavity presented sharp morphological variations along the reproductive cycle related to the mucus secretion that accompanies oocyte ma turation. During sp awning season, residues of mannose and N-acetylglucosamine were detected in the glycocalyx of those cells using lectinhistochemistry. N- acetylgalactosamine and fucose were also observed in the same zone. The greatest variations in the lectinhistochemical pattern were found in the apical cytoplasm composition in comparison to the basal zone of the cells. The results of the present study were discussed by comparing their possible functional implications.

A composição e distribuição dos glicoconjugados (GCs) secretado pelo epitélio do ovário de lamelas com referência à biologia reprodutiva de Genypterus blacodes (Schneider, 1801) através da histoquímica com lectinas é aqui discutida. Nesta espécie, as células epiteliais que revestem a cavidade do ovário apresentou acentuada variação morfológica ao longo do ciclo reprodutivo relacionados com a secreção de muco que acompanha a maturação do oócito. Durante a época de desova, de resíduos de manose e N-acetilglicosamina foram detectados no glicocálix dessas células usando histoquímica de lectinas. N-acetilgalactosamina e fucose também foram observados na mesma zona. As maiores variações no padrão de lectinas foram encontradas na composição do citoplasma apical, em comparação com a zona basal das células. Os resultados do presente estudo foram discutidos, comparando as suas possíveis implicações funcionais.

Componentes glicosilados de la envoltura de Mycobacterium tuberculosis que intervienen en la patogénesis de la tuberculosis / Structural components of the envelope of Mycobacterium tuberculosis that intervene in the pathogenesis of tuberculosis

Los componentes glicosilados de la envoltura de Mycobacterium tuberculosis tienen un papel importante en la inmunopatogénesis de la tuberculosis. Permiten la adhesión, penetración y persistencia de la micobacteria en el macrófago; de igual manera, participan en los mecanismos de activación de estas células y la producción de citocinas relevantes durante la respuesta inmune. En esta revisión, examinamos las características de las principales estructuras sacarídicas de la superficie de la micobacteria y su relación con la modulación de la respuesta inmune.

The glycosylated compounds of Mycobacterium tuberculosis envelope play an important role in the immunopathogenesis of tuberculosis. These molecules are involved in the binding to the host cell surface followed by their internalization and persistence in macrophages; likewise they take part in the macrophage's activation and the production of cytokines that are relevant during the immune response against mycobacteria. In this review we examine the molecular characteristics of the main mycobacterial cell surface saccharide structures and their relation with the modulation of the immune response to tuberculosis.