The Chick Chorioallantoic Membrane: A Model of Molecular, Structural, and Functional Adaptation to Transepithelial Ion Transport and Barrier Function during Embryonic Development

The chick chorioallantoic membrane is a very simple extraembryonic membrane which serves multiple functions during embryo development; it is the site of exchange of respiratory gases, calcium transport from the eggshell, acid-base homeostasis in the embryo, and ion and H2O reabsorption from the allantoic fluid. All these functions are accomplished by its epithelia, the chorionic and the allantoic epithelium, by differentiation of a wide range of structural and molecular peculiarities which make them highly specialized, ion transporting epithelia. Studying the different aspects of such a developmental strategy emphasizes the functional potential of the epithelium and offers an excellent model system to gain insights into questions partly still unresolved

Specific features of the intestinal mucosa of obese Zucker rats

Metabolic syndrome is a group of obesity-related metabolic abnormalities that increase an individual’s risk of developing type 2 diabetes and cardiovascular disease. The obese Zucker rats (OZR) may represent a valuable animal model for studying several aspects of this increasingly prevalent problem in worldwide. In fact, the genetically obese (fa/fa) Zucker rats, due a recessive mutation of the leptin receptor gene (lepr), exhibit hyperphagia and develop hallmark features of metabolic syndrome, including hyperlipidemia, hypertension, insulin resistance, and increased adiposity and oxidative stress. Here, we report the preliminary results from our current studies aimed to investigate different metabolic markers in the OZR intestinal mucosa, compared with their lean counterparts (LZR). Starting from the important role attributed to carbohydrates in regulating the critical equilibrium of the intestinal environment, we applied lectin histochemistry to visualize the glycosylation pattern expressed in the OZR intestinal mucosa. The investigation was mainly focused to identification and in situ characterization of sialylated and fucosylated glycomponents which were directly demonstrated with SNA, MAL II, LTA, and UEA lectin binding. In addition, in order to look for additional and complementary information about sialic acid acetylation degree and sites, PNA and DBA lectin histochemistry was combined with sialidase predigestion, potassium hydroxide deacetylation, and differential periodate oxidation. As a parallel study, the distributional patterns of carbonic anhydrase (CA), the enzyme which is differently expressed in the gastrointestinal tract with several functions, such as regulation of cellular and extracellular acid-base homeostasis, salt absorption and fluid balance, were visualized. The immunohistochemical localization of the CA isoenzymes CAIV, CA IX, CA XII, and CA XIV was performed with the relevant specific antibodies. The complex of the data obtained suggest a marked modulation of the sialoglycoconjugate expression in the OZR intestinal epithelium, when compared with the LZR, to be considered as an interesting topic for further investigations

Lectin histochemistry for in situ profiling of rat colon sialoglycoconjugates

The growing interest in glycoconjugates expressed and released by the epithelium of the intestinal mucosa is tightly related to the multiple functional roles attributed to sialic acid and its derivatives. In the present work, biotin and HRP conjugated lectins were used to detect the sialylation pattern and to identify specific structural features of sialoderivatives in the rat colon. In particular, the occurrence and distribution of sialic acids linked a2,6 to D-Gal/D-GalNAc and a2,3 to D-Gal were directly demonstrated with SNA and MAL II binding, respectively. In addition, in order to by-pass the specificity problems of SNA and MAL II as histochemical reagents, as well as to look for additional and complementary information about acetylation degree and sites, we combined sialidase digestion, potassium hydroxide deacetylation, and differential periodate oxidation with PNA and DBA binding. The data showed the distribution and structure of sialic acidß- D-Gal(1-3)-D-GalNAc and sialic acid-D-GalNac sequences, which proved to be widely distributed as cellular components or secretory products in surface goblet cells and crypt cells of the colonic epithelium. A high degree of O-acetylation, with acetyl groups mainly at 9 and 4 positions, was found, showing an increasing gradient from the proximal to distal portion of the colon. These results, which largely reproduce the sialylation pattern in other species, contribute new insights in defining the tissue specific expression of sialoderivatives in the colonic mucosa, and testify to their high heterogeneity which the wide range of sialic acid functional correlates in the intestinal tract depend on

Mosaic lectin labelling in the quail collecting ducts

-Morphological and histoenzymological differences have been observed between intercalated and principal cells of the quail Coturnix coturnix japonica collecting ducts. The present study was designed to shed light on the lectin affinity of the collecting duct cells within cortex and medulla by the use of HRP-labelled lectins combined with glycosidase degradation. Binding of PNA and RCA-1 lectins consequent to enzymatic release of sialic acid revealed abundant sialylated carbohydrate moieties within the principal cell cytoplasm. This characteristic binding pattern differed considerably from the staining observed in the intercalated cells. Interesting information also emerged about the presence of sialoglycoconjugates having the terminal disaccharide sialic acid-B-N-acetylgalactosamine originating from the increased SBA binding and the unmodified DBA labelling after removal of sialic acid. Sequential degradation by sialidase1B-galactosidase followed by incubation with DBA offered the possibility to suspect that the receptor sugar for the penultimate Bgalactose may be N-acetylgalactosarnine. Conversely, we were not able to define the acceptor sugar for penultimate B-GalNAc owing to the lack of availability of B-N-acetylgalactosaminidase enzyme. When although further studies are clearly needed to elucidate the physiological role of the cellular sialoglycoconjugates detected, the present results already provide valuable insight into the carbohydrate composition of intercalated and principal cells in the quail collecting ducts

Variety of sialic acids occurring in the bovine sublingual gland

Sialoglycoconjugates were investigated in the bovine sublingual gland by direct visualization of sialic acid with specific lectins (LPA, SNA) and by histochemical procedures combined with sialidase digestion and lectins. The most reactive histological sructures were found to be acini which contained glycoconjugates with terminal disaccharides consisting of sialic acid linked to galactose or N-acetylgalactosamine. Resistance to periodate oxidation was interpreted as demonstrating a relevant presence of C7, C8 and Cg acetylated sialic acids. KOH-Sialidase-DBA and KOHAlcian blue sequences allowed the identification of C4 acetylated sialic acids

Distribution and evaluation of complex carbohydrates in the quail collecting ducts

The cell heterogeneity within the collecting duct epithelium of quail kidney was investigated using histochemical methods for complex carbohydrates. The selective distribution of acidic glycoconjugates allowed further discrimination between metachromatic, mucin-secreting cells and dark, proton-transporting cells. Enzymatic digestion by glycosidases was performed to characterize the carboxylated and sulphated glycoconjugates of mucin-secreting cells. The staining intensity of all histochemical reactions with and without prior treatment, was quantitated by means of scanning histophotometry and differences in absorbance values were evaluated by statistical analysis. A different distribution of sulphated components was found for the mucin-secreting cells of renal cortex and medulla, suggesting a morphofunctional heterogeneity within mucin-secreting cell population