Anna Maria Bondi

Glycoconjugate composition of mammalian parotid glands elucidated in situ by lectins and glycosidases.

Sugar specific lectins (PNA, RCA I, LPA, SBA, DBA, GSA IB4, GSA II, WGA, LTA, UEA I, Con A, LCA) with and without prior selective glycosidase digestion (sialidase, alpha-fucosidase, alpha-mannosidase, beta-N-acetylglucosaminidase, alpha- and beta-galactosidase, beta-glucosidase) were used in order to investigate the distribution of native accessible carbohydrates and obtain information dealing with the composition of terminal disaccharides within glycoconjugates present in acinar compartments and ductal segments of mammalian (mouse, rat, hare, and rabbit) parotid glands. Glycoconjugates containing variable amounts of mannose, glucose, N-acetylgalactosamine and N-acetylglucosamine were present in the parotid glands of all species. However, these carbohydrate chains exhibited a different composition of terminal sequences within each type of gland. For example, sialylated components having the terminal dimers sialic acid-galactose and sialic acid-N-acetylgalactosamine were found in all acinar cells, whereas fucoglycoconjugates with terminal disaccharide fucose-galactose were localized in the rat striated ducts and hare acinar cells. The terminal sequence alpha-galactose-beta-galactose was demonstrated in the mouse acinar cells. Finally, glycoconjugates characterized by the terminal dimer beta-galactose-N-acetylgalactosamine were demonstrated in the mouse acinar and ductal cells and the rat ductal ones. Thus, present findings outlined and further confirmed the possibility to elucidate the oligosaccharide structure in situ using lectin histochemistry combined with enzymatic degradation.

Ultrastructural Histochemical Study on Glycoconjugates of the Submandibular Gland of Rabbits

An ultrastructural histochemical study was carried out on submandibular glands of rabbits. Stainings were performed with dialysed iron (DI), high iron diamine (HID), tannic acid uranyl acetate (TA-U), tannic acid-ferric chloride (TA-F) sequence, and periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method. It was demonstrated that neutral glycoproteins are present in the cells with dark granules of the preterminal tracts, and that neutral and acid glycoproteins are present in the cells with light granules of the terminal tracts. Result are discussed and compared to other previously obtained histochemical and biochemical data.

Variability and ultrastructural histochemical localization of sulphates in the salivary glands of rodents and Lagomorpha.

Stainings were effected on an ultrastructural histochemical level to localize sulphomucins in the submandibular and sublingual glands of growing mice and rabbits. Sulphates behave in an entirely different way in the salivary glands of Lagomorpha and in those of Rodents. In growing rabbits sulphates can be demonstrated in both glands; at maturity they can be demonstrated only in the sublingual gland and no longer in the submandibular gland. In the salivary glands of Rodents, sulphates cannot be demonstrated histochemically in new born subjects or in adults. The histochemical results are compared to the biochemical ones, and discrepancies, where present, are discussed.

Fine localization of sulphated and non-sulphated glycoconjugates in the rabbit oviduct during the estrous cycle.

An ultrastructural histochemical investigation was carried out to ascertain the localization of the glucidie components and to verify the histochemical modifications during the physiological estrogenic cycle of rabbit oviduct. Stainings were performed with high iron diamine (HID), low iron diamine (LID), dialized iron (DI), tannic acid-uranyl acetate (TA-U), and periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) methods. The findings indicated that the secretory activity is more intense in the isthmus than in the ampulla. Neutral glycoconjugates are localized both in the secreting elements of the ampulla and in those of the isthmus, while carboxylated and sulphated glycosaminoglycans are localized only in the secretory granules of the secreting elements of the isthmus.

Visualization of carbohydrate chains in rabbit salivary glands by means of enzymatic degradation and plant lectins

We investigated the structure of glycoconjugates contained within the secretory end-pieces and ductal segments in the rabbit submandibular and sublingual glands. Glycosidic sequences were examined by means of enzymatic degradation with specific glycosidases (sialidase, alpha-fucosidase, beta-galactosidase, alpha-mannosidase) followed by lectin binding with PNL-HRP, WPL-HRP, WGL-HRP, SBL-HRP, Con A-HRP. It was found that this procedure represents a valid tool for studying carbohydrates, in so far as their characterization and localization were based only on colour reactions. In particular, this research showed that sialic acid was present in the terminal dimers sialic acid-beta-galactose and sialic acid-N-acetyl-D-galactosamine within the submandibular gland, whereas in the sublingual gland it was only present as the sequence sialic acid-beta-galactose. Conversely, fucose had as the subterminal sugar N-acetyl-D-glucosamine in both glands. Also, elucidations about structural sequences concerning other non-terminal sugars were obtained.

Differential binding sites of peroxidase-labelled lectins in the submandibular gland of sucking and adult cats

The glycosidic residues in the submandibular gland of cat at different ages were studied using a battery of 6 lectins conjugated with horseradish peroxidase as histochemical reagents. The submandibular glands of sucking and adult cats showed diversities in the reaction intensity or in the localization of binding sites. The morphologically distinct secretory tracts appear to produce different types of oligosaccharides in sucking and adult subjects. The sugars that can be defined as growth markers in the cat submandibular gland seem to be alpha-L-fucose, N-acetyl-D-glycosamine and beta-galactose. The data originated from this research have been compared with biochemical data previously obtained by the authors on the cat submandibular gland during growth.

Ultrastructural localization of complex carbohydrates present in the sublingual gland of rabbits

Stainings with dialized iron (DI), high-iron diamine-thiocarbohydrazide-silver proteinate HID-TCH-SP), tannic acid-uranylacetate (TA-U), tannic acid-ferric chloride (TA-F), and periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) were applied to samples of sublingual glands of rabbits. Comparing the results obtained with what had been previously demonstrated biochemically, it is possible to assert that the faintly electron-opaque, granules of the cells of the preterminal tracts contain sulphated glycoconjugates because of the presence of sulphated hexosamines and hexoses, acid glycoconjugates because of the presence of sialic acid and uronic acids, and neutral glycoconjugates because of the presence of hexoses.

Attempt to visualize an endogenous lectin recognizing α-L-fucose immunohistochemically in the rabbit oviduct

An immunohistochemical study was conducted in order to visualize an endogenous lectin, having specificity for alpha-L-fucose residues, in the ampullary and isthmic tracts of the rabbit oviduct. 2 different sequences (S1 and S2) were tested using antilectin UEA I antibody. Reactive sites towards S2 sequence were localized at ciliated cell level in the ampulla; in the isthmus, instead, the staining was present both in ciliated and secretory cells. A role in the sugar immobilization and gamete and zygote protection was postulated for the endogenous lectin evidenced.

Developmental expression of glycocomponents in the chick chorioallantoic membrane

Widespread interest has focused on the research of the chorioallantoic membrane (CAM) and its functional contribution to gaseous exchange, calcium reabsorption, water and electrolyte transport during chick embryogenesis. Nevertheless, very little information is available on the glycoconjugate components of this extra-embryonic structure. In the present study, we investigated by lectin histochemistry, the glycosylation pattern expressed in the CAM epithelia during embryonic development. Occurrence of sialic acid-associated glycoproteins was detailed by either specific lectins, which discriminate α2,3 and α2,6 sialoderivatives, or sialidase digestion combined with appropriate lectins to identify the sialic acid acceptor sugars. Lectin affinities proved to depend greatly on differentiation of the CAM epithelia which showed highest expression of binding sites during the second half of incubation up to hatching. Differences emerged between the chorionic and the allantoic epithelium, regarding qualitative, quantitative and temporal expression of sugar moieties. A cell type-specific distribution of glycocomponents was found in the chorionic epithelium where lectin binding sites were specifically located in the villus cavity cells. In the allantoic epithelium, high and heterogeneous occurrence of sialoglycoconjugates as well as specific presence of fucose residues were evidenced mostly in the granule cells. We conclude from these findings that various glycoconjugates in the CAM could participate in different physiological functions characteristic of the chorionic and the allantoic epithelium.

Soluble arylsulphatases in the rabbit salivary glands. A light and electron microscopy study.

A histochemical study was effected, by light and electron microscopy, on the activity of arylsulphatases A and B in the submandibular and sublingual glands of male and female rabbits, distinguishing, among the latter, between the physiological estrus and anestrus condition. The results obtained demonstrated that the enzymatic activity is more intense in the sublingual gland and that the estrus condition causes an increase in this activity. No correlation was found between the topochemical distribution of the soluble arylsulphatases and that of sulphates.