Histochemical analysis of glycoproteins in the secretory cells in the epidermis of the head skin of Indian Major Carp,Labeo rohita

A series of histochemical procedures were employed to localise and characterise glycoprotein (GP) classes produced by the epithelial cells, the type A and the type B mucous goblet cells (MGCs) and the club cells in the epidermis of Labeo rohita. The epithelial cells secreted GPs with oxidizable vicinal diols and GPs with sialic acid residues without O-acyl substitution in low concentrations. The type A MGCs and the type B MGCs, in contrast, produced these GPs in high concentrations. Further, these MGCs produced GPs with O-sulphate esters as well. GPs with O-sulphate esters were produced in high concentration by the type A MGCs and in low concentration by the type B MGCs. The club cells produced GPs with oxidizable vicinal diols in trace amounts. Production of more than one type of GPs suggested a basis for functional discrimination in their role in the mucous secretions at the skin surface. This is considered an adaptation to environment inhabited by the fish and is discussed in relation to their role in lubrication, protection and inhibition of the invasion and proliferation of pathogenic micro-organisms.     

Histochemical analysis of glycoproteins in the secretory cells in the gill epithelium of a catfish, Rita rita (Siluriformes, Bagridae)

Glycoproteins (GPs) were visualised histochemically in the secretory cells – the mucous goblet cells (the type A and the type B), the serous goblet cells, the club cells and the epithelial cells in the gill epithelium of Rita rita. The type A mucous goblet cells, the type B mucous goblet cells and the epithelial cells elaborate GPs with oxidizable vicinal diols and GPs with sialic acid residue without O-acyl substitution. In addition, GPs with O-sulphate esters are elaborated by the type A and GPs with O-acyl sugars by the type B mucous goblet cells. GPs are absent in the serous goblet cells and are with oxidizable vicinal diols in low moieties in the club cells. The analysis of the results elucidates interesting differences in the composition and concentration of GPs in the mucus elaborated by the epithelium of the gill arches and the gill rakers; and the gill filaments and the secondary lamellae indicating the potential importance of the glycoproteins at these locations. GPs elaborated on the surfaces of the gill arches and the gill rakers could be associated to assist in feeding activities and on the surfaces of the gill filaments and the secondary lamellae in the respiratory activity.     

Glycoproteins histochemistry of the gills of Odontesthes bonariensis (Teleostei, Atherinopsidae)

The histochemistry of glycoproteins (GP) in the mucous cells of the gills of the silverside Odontesthes bonariensis was identified with: (1) oxidizable vicinal diols; (2) sialic acid and some of their chain variants, carbon 7 ((7) C), carbon 8 ((8) C) or carbon 9 ((9) C); (3) sialic acid residues without O-acyl substitution and with O-acyl substitution at (7) C, (8) C or (9) C; (4) carboxyl groups and (5) sulphate groups. A battery of seven biotinylated lectins allowed GPs sugar residues to be distinguished. Mucous cells showed the presence of neutral, sulphated and sialylated GPs. Dolichos biflorus agglutinin (DBA) and Glycine max agglutinin (SBA) showed strong positive staining; Arachis hypogaea agglutinin (PNA), Ricinus communis agglutinin-I (RCA-I) and Triticum vulgaris agglutinin (WGA) showed moderate staining, while Ulex europaeus agglutinin-I (UEA-I) was completely negative.     

Histochemical studies of intestinal epithelial goblet cell glycoproteins during the development of the human foetus

Summary Histochemical studies performed on specimens of intestine from 12 to 37-week human foetuses showed that the epithelial glycoproteins of the goblet cells of the small intestine are non-sulphated sialoglycoproteins containing neutral sugar (hexose, 6-deoxy hexose or N-acetyl hexosamine residues with Periodic acid-Schiff (PAS) reactive vicinal diols), sialic acids without O-acyl substituents, smaller and variable quantities of sialic acids with O-acyl substituents at positions C8 or C9 (or with two or three side chain substituents) and O-acyl sugars (neutral sugars with an ester substituent blocking PAS reactivity). In the lower small intestine glycoproteins containing 8 (or 9)-O-acyl sialic acids are first observed in goblet cells at the tips of the villi. As the foetus matures their quantity increases and they are found in goblet cells located along the length of the villi. Smaller quantities of O-acyl sialic acids and traces of O-acyl sugars occur in the goblet cells of the upper small intestine. The colonic goblet cells contain sulphosialoglycoproteins of two types. The first type, found in the majority of specimens, contains O-sulphate ester, neutral sugar, O-acyl sugars and 8 (or 9)-O-acyl sialic acids. The second type contains O-sulphate ester, neutral sugars, and sialic acids which are either without side chain O-acyl substituents or are a mixture of such acids and 8 (or 9)-O-acyl sialic acids; O-acyl sugars are reduced or absent. The degree of sulphation of the foetal colonic goblet cell epithelial glycoproteins differs with the region of the colon, the level of the crypt and the gestational age of the foetus in a manner consistent with that described by Lev & Orlic (1974). The detection of O-acyl sugars in foetal intestinal glycoproteins adds to the known examples of such sugars and strengthens the suggestion that they are a normal constituent of colonic epithelial glycoproteins.Part of this work was presented at the 32nd meeting of the Canadian Federation of Biological Sciences, Calgary, Alberta, June 1989 (abstract # 336).     

Histochemical procedures for the simultaneous visualization of neutral sugars and either sialic acid and its O-acyl variants or O-sulphate ester. II. Methods based upon the periodic acid-phenylhydrazine-Schiff reaction

Summary Four methods based upon the periodic acid—phenylhydrazine—Schiff reaction have been developed for the simultaneous visualization of neutral sugars with periodate oxidizablevicinal diols (hexose, 6-deoxyhexose,N-acetylhexosamine) and either sialic acids or side chainO-acyl sialic acids. In the first of these procedures, the saponification—periodic acid oxidation—2,4-dinitrophenylhydrazine—Azure A—Schiff—saponification (KOH—PA—DNPH—Az—KOH) method, all sialic acids stain Azure blue, neutral sugars with oxidizablevicinal diols stain yellow and mixtures of such components stain in various shades of green. In the second technique, periodic acid oxidation—2,4-dinitrophenylhydrazine—Azure A Schiff—saponification (PA—DNPH—Az—KOH), Azure Blue staining is confined to sialic acids without side chain substituents or which have anO-acyl substituent at position C7, while in the third method, the selective periodate oxidation—borohydride reduction—saponification—periodic acid oxidation—2,4-dinitrophenyl hydrazine—Azure A—Schiff—saponification (PA^*—Bh—KOH—PA—DNPH—Az—KOH) technique, only sialic acids withO-acyl substituents at positions C7, C8 or C9 (or which have two or threeO-acyl side chain substituents) stain Azure blue. Finally in the fourth procedure, periodic acid oxidation—2,4-dinitrophenylhydrazine—Azure A—Schiff—saponification—borohydride reduction—periodic acid oxidation—Schiff (PA—DNPH—Az—KOH—Bh—PAS), sialic acids without side chain substituents or which haveO-acyl substituents at C7 stain Azure blue, sialic acids substituted at position C8 or C9 (or which are di- or tri-substituted) stain magenta and neutral sugars stain yellow. Where mixtures of these components are present, a wide range of colours is obtained.     

A new method for the histochemical demonstration of O-acyl sugars in human colonic epithelial glycoproteins

Summary A new general method has been developed for the specific histochemical identification ofO-acyl sugars in any epithelial glycoprotein. These sugars include hexose, 6-deoxyhexose andN-acetylhexosamine with an ester substituenent(s) located on a potentialvicinal diol(s). In the procedure reported [the periodic acid-borohydride reduction-saponification-selective periodate oxidation-borohydride reduction-periodic acid-Schiff (PA-Bh-KOH-PA-Bh*-Bh-PAS) method] the initial PA-Bh treatment rendersvicinal diols located on either sialic acid or neutral sugars PAS unreactive. In the subsequent steps ester substituents are removed from bothO-acyl sugars andO-acyl sialic acids by saponification (KOH), sialic acidvicinal diols are selectively removed by the PA^*-Bh sequence andO-acyl sugars are stained with the PAS technique. This method has the advantage that the results are obtained with a single section and the results are either positive or negative. Consequently, it is superior to the three indirect methods investigated because it does not require an observer to compare the intensity or the shade of the staining obtained with serial sections.Using the PA-Bh-KOH-PA^*-Bh-PAS method we have demonstrated, for the first time, thatO-acyl sugars occur in the epithelial goblet cell glycoproteins of adult human colon. The effect of the presence ofO-acyl sugars on the interpretation of a number of other methods for the histochemical investigation of glycoproteins is discussed. It is recommended that the results obtained with the PA-Bh-KOH-PA^*-Bh-PAS method be evaluated before histochemical procedures for the investigation of neutral sugars andO-acyl sialic acids are selected.     

A new histochemical method for the selective periodate oxidation of total tissue sialic acids

Summary Evaluation of the intensity of the periodic acid—Schiff (PAS) staining produced following oxidation for 1 h at 4°C with 0.4mm periodic acid in approximately 1m hydrochloric acid indicated that this reagent completely oxidized all available sialic acid residues of either the sialo- or sialosulphoglycoproteins of human and rat colon or the sialoglycoproteins of rat sublingual gland. These conditions produced no visible Schiff staining of either neutral macromolecules orvicinal diols located on hexose, 6-deoxyhexose orN-acetylhexosamine residues (neutral sugars) of sialo- and sialosulphoglycoproteins. Furthermore, there was no extraction of epithelial glycoproteins or de-O-acylation of side chain substituted sialic acid residues. These data demonstrate that 0.4mm periodic acid in approximately 1m hydrochloric acid can be used as a specific reagent for the selective visualization of sialic acids in the PAS procedure.Studies of the mechanism of the oxidation of neutral sugars with 0.4mm periodic acid in approximately 1m hydrochloric acid indicated that their lack of PAS reactivity was not due to the production of Schiff unreactive hemiacetals or hemialdals. It is suggested that the selectivity of 0.4mm periodic acid in approximately 1m hydrochloric acid is a result of an increase in the rate of the oxidation of the sialic acid residues together with a decrease in the rate of oxidation of neutral sugars.     

Histochemical procedures for the simultaneous visualization of neutral sugars and either sialic acid and its side chain O-acyl variants or O-sulphate ester. I. Methods based upon the selective periodate oxidation of sialic acids

Summary Five new methods, based upon the selective oxidation of sialic acid residues with 0.4mm periodic acid in approximately 1m hydrochloric acid at 4°C for 1 h (PA^*), have been devised for the simultaneous visualization of neutral sugars and either sialic acid and its side chainO-acyl variants orO-sulphate ester. In the first of these, the selective periodate oxidation—borohydride reduction—saponification—selective periodate oxidation—Thionin Schiff—saponification—borohydride reduction—periodic acid—Schiff (PA^*—Bh—KOH—PA^*—T—KOH—Bh—PAS) technique, sialic acids withO-acyl substituents at C7, C8 or C9 (or which have two of three side chainO-acyl substituents) stain blue while neutral sugars with periodate-sensitivevicinal diols (hexose, 6-deoxyhexose, andN-acetylhexosamine) stain magenta. The second method, the saponification—selective periodate oxidation—Thionin Schiff—saponification—borohydride reduction—periodic acid—Schiff (KOH—PA^*—T—KOH—Bh—PAS), stains all sialic acids blue and neutral sugars magenta. In the third procedure, the selective periodate oxidation—Thionin Schiff—borohydride reduction—periodic acid—Schiff—saponification (PA^*—T—Bh—PAS—KOH) method, sialic acids without side chain substituents (or which have anO-acyl substituent at C7) stain blue and neutral sugars stain magenta. In the fourth method, the saponification-selective periodate oxidation—borohydride reduction—Alcian Blue pH 1.0—periodic acid—Schiff (KOH—PA^*—Bh—AB1.0—PAS) technique,O-sulphate esters stain aquamarine blue and neutral sugars stain magenta. In all of these techniques mixtures of the components stain in various shades of purple. Performance of the KOH—PA^*—Bh—AB1.0—PAS technique without the Alcian Blue pH 1.0 step provides a method for the selective identification of neutral sugars in macromolecules that also contain sialic acids.     

Histochemical identification of 9-O-acyl sialic acids: studies of bovine submandibular and rat sublingual gland and human colon

Summary Formalin-fixed tissue specimens containing glycoproteins with side chain O-acylated sialic acids were used to re-examine, compare and evaluate the usefulness of three methods based on the periodic acid-borohydride reduction-saponification-periodic acid-Schiff sequence (PA-Bh-KOH-PAS) for the histochemical identification of 9-O-acyl sialic acids (9-O-AcSA). Method I, modified from Vehet al. (1979), involved a comparison of the staining intensely obtained when both oxidation steps of the PA-Bh-KOH-PAS sequence were carried out with the selective oxidation technique of Volzet al. (1987) with that obtained when the initial oxidation step was carried out with 0.5m periodic acid for 4h at room temperature. Methods II and III, modified from Reidet al. (1978), involved an initial PA-Bh step under oxidation conditions that cleaved all the vicinal diols associated with neutral sugars and side chain unsubstituted and 7-O-acyl sialic acids. The Schiff staining obtained following subsequent re-oxidation with either 0.5m (method II) or 1% periodic acid (method III) for 4h at room temperature (PA-Bh-PAS procedure) identifies 9-O-AcSa.The results of this study indicate that (a) bovine submandibular gland acinar cell glycoproteins contain 9-O-AcSA as well as sialic acids which have ester substituents at C7 or C8, or which are di-(C7C8, C7C9, C8C9) or tri-(C7C8C9) substituted, (b) the side chain O-acyl sialic acids of the glycoproteins of Sprague Dawley rat sublingual gland acinar cells are entirely or almost entirely 9-O-AcSA and (c) it is likely that the majority of the human adult and foetal glycoproteins studied contain small quantities of 9-O-AcSA mixed with sialic acids which are substituted at C7 or C8 or which have two or three side chain O-acyl substituents. However, the interpretation of the results are complicated by observations that indicate that (a) treatment with 0.5m periodic acid either extracts or removes sialic acids from bovine submandibular gland glycoproteins, (b) some human colonic epithelial glycoproteins apparently contain a component other than 9-O-AcSA that oxidises slowly with periodic acid and (c) 1% periodic acid for 2h at room temperature oxidises a small but significant quantity of 9-O-AcSA, thus reducing the intensity of staining in methods II and III. It is concluded that when adequately controlled, methods I, II and III are capable of detecting 9-O-AcSA in glycoproteins containing large quantities of the sialic acid. However, these methods may not detect small quantities of 9-O-AcSA in the presence of large quantities of sialic acids which have O-acyl substitutents at positions C7 or C8 or which have two (C7C8, C7C9, C8C9) or three (C7C8C9) side chain O-acyl substituents. Thus, caution should be used when interpreting data that indicates the absence of 9-O-AcSA.     

Carbohydrates in the epidermal mucous cells of a fresh-water fish Mastacembelus pancalus (mastacembelidae,Pisces) as studied by electron-microscopic cytochemical methods

Carbohydrates in the mucous cells of the epidermis of the fish Mastacembelus pancalus were studied by means of electron-microscopic cytochemical methods using physical development procedures. Three types of mucous cells (types A-C) were differentiated on the basis of the reactivities of the secretory products elaborated by them. The carbohydrate contents of mucous globules predominantly comprised sulfate esters and traces of oxidizable vicinal diols in type-A cells, oxidizable vicinal diols in type-B cells, and moderate amounts of both sulfate esters and oxidizable vicinal diols in type-C cells. Glycogen particles were also found to occur in the cytoplasm of these cells, and glycoproteins containing oxidizable vicinal diols were visualized in Golgi cisternae, rough endoplasmic reticulum, nuclear envelopes, and plasma membranes. In the type-A and type-B cells situated in the superficial layers of the epidermis, extensive cisternae of the Golgi apparatus and copious rough endoplasmic reticulum suggested the active syntheses of secretory contents, in contrast to the type-C mucous cells, which displayed poor development of these organelles, in the deeper layers.     

Histochemical studies of the colonic epithelial glycoproteins of the normal rabbit

Summary Two general classes of glycoproteins have been identified in the colonic epithelial cells of New Zealand white rabbits. Each is associated with an ultrastructurally distinct secretory cell. The first of these classes is found in cells, termed vesiculated columnar cells, characterized by electron-translucent vesicles, a small rough endoplasmic reticulum-Golgi complex and prominent microvilli. The glycoproteins of the vesiculated cells contain abundantO-sulphate ester, sialic acids with ester substituents at positions C-8 or C-9 (or with two or three side chain substituents) and neutral sugars withvicinal diols whose periodate oxidation is prevented by anO-acyl ester substituent(s). The second class of glycoproteins occurs in goblet cells characterized by electron-dense vesicles, an abundant rough endoplasmic reticulum, a well-developed Golgi apparatus and few, if any, microvilli. Goblet cells along the entire length of the crypts contain neutral sugars with periodate-oxidisablevicinal diols and a ferriferricyanide-reactive component. Cells in the upper halves of the crypts also contain components that are sulphated, Schiff-reactive and acid-fast. In the lower halves of the crypts, the goblet cells contain smaller quantities of the above components plus sialic acids, some of which possibly have anO-acyl substituent located at position C-8 or C-9 (or which have two or three side chainO-acyl substituents). It is suggested that the function of the glycoproteins from the vesiculated columnar cells is protective and that from the goblet cells is lubricative.     

Histochemical identification of side chain substituted O-acylated sialic acids: the PAT-KOH-Bh-PAS and the PAPT-KOH-Bh-PAS procedures

Summary Two new methods, based on the original periodic acid-Thionin Schiff-saponification-periodic acid-Basic Fuchsin Schiff (PAT-KOH-PAS) technique of Cullinget al. (1976), have been devised for the histochemical identificantion of side-chainO-acylated sialic acids. In the first of these, the periodic acid-Thionin Schiff-saponification-borohydride reduction-periodic acid-Basic Fuchsin Schiff (PAT-KOH-Bh-PAS) procedure, the specificity of the original PAT-KOH-PAS technique was improved by: (a) extending, when necessary, the initial period of periodate oxidation, (b) increasing the period of exposure to Thionin Schiff reagent from 30 min to 4 h, (c) using a Thionin Schiff reagent prepared by a different method, (d) interposing a borohydride reduction step between the saponification and PAS steps and, (e) extending the period of oxidation in the final PAS step from 10 to 30 min. In the second procedure, the periodic acid-phenylhydrazine-Thionin Schiffborohydride reduction-periodic acid-Basic Fuchsin Schiff (PAPT-KOH-Bh-PAS), based on the periodic acid-phenylhydrazine-Schiff (PAPS) technique of Spicer (1961), blue Thionin Schiff staining was confined to sialic acid residues with oxidizable side chainvicinal diols by interposing a treatment with 0.5% (w/v) aqueous phenylhydrazine hydrochloride for 2 h at room temperature between the initial periodic acid oxidation and the Thionin Schiff steps of the PAT-KOH-Bh-PAS procedure. These procedures are discussed within the general framework of the methods available for the histochemical identification of side-chainO-acylated sialic acids.     

Chemical and histochemical studies of normal and diseased human gastrointestinal tract. V. A differential diagnostic method for the histochemical classification of glycoproteins

Summary A differential diagnostic scheme is described for the division of colonic epithelial glycoproteins into eleven histochemically distinct classes. The scheme depends upon the use of seven histochemical techniques which, collectively, permit the differential staining ofO-sulphate ester, sialic acid and its side chainO-acyl variants and vicinal diols located on carbohydrate residues other than sialic acids. Elements of the scheme also provide a general approach to the classification of epithelial glycoproteins in anatomic sites other than the colon.Application of the scheme permitted the classification of the epithelial glycoproteins in the mucosa 0.5–5.0 cm from human colonic tumours and provided direct confirmation of previous observations that changes from normal in the relative proportions of either side chainO-acylated sialic acids or sialic acids andO-sulphate esters can occur independently of one another.     

VARIATIONS IN THE SUBSTITUTED 3‐LINKED MANNANS CLOSELY ASSOCIATED WITH THE SILICIFIED WALLS OF DIATOMS1

The polysaccharides from cleaned frustules of the diatoms Pinnularia viridis (Nitzsch) Ehrenberg, Craspedostauros australis Cox, Thalassiosira pseudonana Hasle et Heimdal, and Nitzschia navis‐varingica Lundholm et Moestrup were extracted with hot alkali that dissolved the silica and were characterized by constituent sugar and linkage analyses. The polysaccharides from P. viridis were investigated further by permethylation, partitioning according to solubility, desulfation, and CD₃I‐methylation. Yields of carbohydrate in the hot alkali extracts ranged from 0.9% to 1.8% w/w based on the dry weight of the silica. Mannose was the dominant sugar in the polysaccharides from all four species (54–69 mol% of constituent sugars), although 14 other monosaccharides, including neutral sugars (glucose, galactose, xylose, arabinose, rhamnose, fucose), acidic sugars (glucuronic acid, galacturonic acid, 2‐O‐methylglucuronic acid), and O‐methylated neutral sugars (2‐O‐methylrhamnose, 3‐O‐methylrhamnose, 2,3‐di‐O‐methylrhamnose, 3‐O‐methylxylose, 4‐O‐methylxylose) were also detected in varying proportions among the four samples. The polysaccharides were predominantly composed of a 3‐linked mannopyranose backbone with a prevalence of linkage and/or substitution at O‐2 of the 3‐linked mannopyranosyl residues, and they were polyanionic, bearing uronic acid residues and/or sulfate esters. There were, however, species‐specific differences in the degree and position of substitution on the mannan backbone, the type and substitution patterns of the anionic substituents, and the type and linkage patterns of sugars other than mannose. Although definitive functions for these polysaccharides in diatom biology remain uncertain, a possible role in biosilicification is discussed.     

Chemical and histochemical studies of normal and diseased human gastrointestinal tract. II. A comparison between histologically normal small intestine and Crohn's disease of the small intestine

Summary Comparative chemical and histochemical studies were performed on formalin-fixed, surgical specimens of human small intestine from cases of Crohn's disease and normal controls. The sialic acids of the crude glycoproteins isolated from normal ileum were significantly less neuraminidase-susceptible and more C₄ substituted (P<0.01) than those of the glycoproteins isolated either from normal upper small intestine (duodenum and jejunum) or from cases of Crohn's disease of the ileum. Fractionation yielded two major sialic acid-containing fractions, eluting from DEAE-cellulose with 0.2m or 0.3m sodium chloride. Both fractions contained fucose, galactose, glucosamine and galactosamine in addition to sialic acids both with and withoutO-acyl substituents at position C₄ and/or in the side-chain (side-chainO-acylated sialic acids were also detected by histochemical procedures). The fractions differed significantly from one another with respect to the neuraminidase susceptibility of their sialic acids (P<0.01), the percentage of C₄ (P<0.01) and side-chain substituted sialic acids (P<0.05), and the molar fucose-sialic acid ratio (P<0.05). TheO-acyl substitution patterns of the sialic acids of both the 0.2m and 0.3m fractions of the upper small intestine glycoproteins differed significantly from those of the corresponding fractions from normal ileum, while the sialic acids of the 0.2m fractions from Crohn's disease of the ileum differed significantly from normal with respect to neuraminidase susceptibility (P<0.01) and percentage C₄ substitution (P<0.01); the 0.3m fractions differed only in the percentage of sialic acids substituted at C₄. The differences between the sialic acids from the normal and Crohn's disease specimens were shown to be independent of either the anatomical origin of the specimen or the histopathological sub-group of the Crohn's disease specimens; no significant differences were noted between the sub-groups but all the sub-groups differed from normal.     

Invasion of <Emphasis Type="Italic">Trypanosoma cruzi</Emphasis> into host cells is impaired by <Emphasis Type="Italic">N</Emphasis>-propionylmannosamine and other <Emphasis Type="Italic">N</Emphasis>-acylmannosamines

The etiologic agent of Chagas’ disease, Trypanosoma cruzi, is widely distributed in South America, affecting millions of people with thousands of deaths every year. Adherence of the infectious trypomastigote to host cells is mediated by sialic acid. T. cruzi cannot synthesize sialic acids on their own but cleave them from the host cells and link them to glycans on the surface of the parasites using the trans-sialidase, a GPI-anchored enzyme. The infectivity of the protozoan parasites strongly depends on the activity of this enzyme. In this report, we investigated whether the transfer of sialic acids from the host to the parasites can be attenuated using novel sialic acid precursors. The cell line 86-HG-39 was infected with T. cruzi and treated with defined N-acylmannosamine analogues bearing an elongated N-acyl side-chain. By treatment of these cells the number of T.cruzi infected cell was reduced up to 60%. We also showed that the activity of the bacterial sialidase C was reduced with N-glycan substrates with elongated N-acyl side chains of the terminal sialic acids. The affinity of this sialidase decreased with the length of the N-acyl side-chain. The data presented suggest that N-acyl modified sialic acid precursors can change the transfer of sialic acids leading to modification of infection. Since the chemotherapy of this disease is inefficient and afflicted by side effects, the need of effective drugs is lasting. These findings propose a new path to prevent the dissemination of T. cruzi in the human hosts. These compounds or further modified analogues might be a basis for the search of new agents against Chagas’ disease.     

Histochemical detection of sialic acid residues using periodate oxidation

Synopsis The use of low concentrations of periodate for the detection of sialic acid residues in tissue sections has been investigated. Oxidation of aqueous solutions of sugar glycosides with 0.4mm periodate revealed that sialic acid was oxidized more rapidly than other sugars found in glycoproteins. Sequential treatment of tissue sections with 0.4mm periodate for 30 min followed by Schiff's reagent stained sialic acid residues but other sugar components were not stained under these conditions.     

Uptake and metabolism of carbohydrates by epidermal tissue

Isolated epidermis of Commelina communis L. and Tulipa gesneriana L. assimilated ¹⁴CO₂ into malic acid and its metabolites but not into sugars or their phosphates; epidermis could not reduce CO₂ by photosynthesis and therefore must be heterotrophic (Raschke and Dittrich, 1977). If, however, isolated epidermis of Commelina communis was placed on prelabelled mesophyll (obtained by an exposure to ¹⁴CO₂ for 10 min), radioactive sugars appeared in the epidermis, most likely by transfer from the mesophyll. Of the radioactivity in the epidermis, 60% was in sucrose, glucose, fructose, 3-phosphoglyceric acid and sugar phosphates. During a 10-min exposure to ¹⁴CO₂, epidermis in situ incorporated 16 times more radioactivity than isolated epidermal strips. Isolated epidermis of Commelina communis and Tulipa gesneriana took up ¹⁴C-labelled glucose-1-phosphate (without dephosphorylation), glucose, sucrose and maltose. These substances were transformed into other sugars and, simultaneously, into malic acid. Carbons-1 through-3 of malic acid in guard cells can thus be derived from sugars. Radioactivity appeared also in the hydrolysate of the ethanol-insoluble residue and in compounds of the tricarboxylic-acid cycle, including their transamination products. The hydrolysate contained glucose as the only radioactive compound. Radioactivity in the hydrolysate was therefore considered an indication of starch. Starch formation in the epidermis began within 5 min of exposure to glucose-1-phosphate. Autoradiograms of epidermal sections were blackened above the guard cells. Formation of starch from radioactive sugars therefore occurred predominantly in these cells. Epidermis of tulip consistently incorporated more ¹⁴C into malic and aspartic acids than that of Commelina communis (e.g. after a 4-h exposure to [¹⁴C]glucose in the dark, epidermis, with open stomata, of tulip contained 31% of its radioactivity in malate and aspartate, that of Commelina communis only 2%). The results of our experiments allow a merger of the old observations on the involvement of starch metabolism in stomatal movement with the more recent recognition of ion transfer and acid metabolism as causes of stomatal opening and closing.Abbreviation G-1-P glucose-1-phosphate     

Teichulosonic acid,an anionic polymer of a new class from the cell wall of <Emphasis Type="Italic">Actinoplanes utahensis</Emphasis> VKM Ac-674<Superscript>T</Superscript>

The cell wall of Actinoplanes utahensis VKM Ac-674^T contains two anionic polymers: teichoic acid 1,3-poly(glycerol phosphate) that is widespread in cell walls of Gram-positive bacteria; and a unique teichulosonic acid belonging to a new class of bioglycans described only in microorganisms of the Actinomycetales order. The latter polymer contains residues of di-N-acyl derivative of sialic acid-like monosaccharide — 5,7-diamino-3,5,7,9-tetradeoxy-L-glycero-β-L-manno-non-2-ulosonic or pseudaminic acid (Pse) which bears the N-(3,4-dihydroxybutanoyl) group (Dhb) at C7. This polymer has irregular structure and consists of fragments of two types, which differ in substitution of the Dhb residues at O4 either with β-D-glucopyranose or with β-Pse residues. Most of the β-Pse residues (∼80%) are glycosylated at position 4 with α-D-galactopyranose residues in both types of fragments. The glucose, galactose, and Dhb residues are partly O-acetylated. The structures of the polymers were established by chemical and NMR spectroscopy methods.     

The acid and enzymic hydrolysis of O-acetylated sialic acid residues from rabbit Tamm–Horsfall glycoprotein

Rabbit Tamm-Horsfall glycoprotein and bovine submaxillary glycoprotein were both found to contain sialic acid residues which are released at a slow rate by the standard conditions of acid hydrolysis. These residues are also resistant to neuraminidases from Vibrio cholerae and Clostridium perfringens. This behaviour was attributed to the presence of O-acetylated sialic acid, since the removal of O-acetyl groups by mild alkaline treatment normalized the subsequent release of sialic acid from rabbit Tamm-Horsfall glycoprotein by acid and by enzymic hydrolysis. Determination of the O-acetyl residues in rabbit Tamm-Horsfall glycoprotein indicated that on average two hydroxyl groups of sialic acid are O-acetylated, and these were located on the polyhydroxy side-chain of sialic acid or on C-4 and C-8. These findings confirm the assumption that certain O-acetylated forms of sialic acid are not substrates for bacterial neuraminidases. Several explanations have been suggested to explain the effect of O-acetylation of the side-chain on the rate of acidcatalysed hydrolysis of sialic acid residues.