Binding of lectins to "young" and "old" human erythrocytes

"Old" human erythrocytes showed a 21.2% decrease in cell surface area and a 2% decrease in the number of WGA receptor sites, but a 27% increase in the distribution density of the WGA (lectin) receptor site, when compared with "young" human erythrocytes. For a list of lectin abbreviations, see Materials and methods). Both "young" and "old" erythrocytes exhibited very weak binding activity for 125I-labeled PNA, but there was no difference in binding activity for PNA between "young" erythrocytes and "old" ones. Compared with "young" erythrocytes, decreases in the number and distribution density of receptor sites for five lectins including LPA, Con A, RCA-II, SBA and BPA on the cell surface were observed in aged erythrocytes. "Old" erythrocytes also showed a decrease in the number of PHA-E receptor sites, while the distribution density of the same receptor site remained unchanged. In view of these and other observations, it is thought that human erythrocyte aging is accompanied by elimination of some glycoconjugates which have affinity for six lectins, LPA, Con A, RCA-II, PHA-E, SBA and BPA, whereas no WGA receptor-containing glycoconjugates are released from erythrocyte membranes. Elimination of the glycoconjugates results in shrinkage of erythrocytes to reduce their cell surface areas.     

Effect of thyroid hormones on the histotopography of lectin receptors in the rat salivary gland

Using lectin-peroxidase technique, the influence of hypo- and hyperthyroidism on histotopography of glycoconjugates has been investigated in rat submandibular gland. The following lectins were used: peanut agglutinin (PNA), wheat germ agglutinin (WGA), Laburnum anagyroides lectin (LAL) and concanavalin A (con A). It has been demonstrated that hyperthyroidism is accompanied by the loss of con A, WGA and LAL receptor sites. Hypothyrodism enhanced con A binding to granular duct cells with a parallel reduction in WGA and LAL binding to these or other duct cells. Hypothyroidism as well as hyperthyroidism markedly enhanced PNA binding to duct epitheliocytes with redistribution of these lectin binding sites from the luminal surface of salivary ducts into the cytoplasm of duct cells. Possible interpretations of the observed phenomena are discussed.     

Lectin-stimulated cellular iron uptake and toxin generation in the freshwater cyanobacterium Microcystis aeruginosa

The lectin family is composed of mono- and oligosaccharide binding proteins that could activate specific cellular activities, such as cell-cell attachment and toxin production. In the present study, the effect of the external addition of lectins to culture media containing the freshwater cyanobacterium Microcystis aeruginosa on its metabolic activities, such as iron uptake and toxin production was investigated. Among the three lectins examined in this study (concanavalin A [Con A], wheat germ agglutinin [WGA] and peanut agglutinin [PNA]), PNA substantially increased the accumulated intracellular and extracellular iron content. The binding of PNA and Con A to M. aeruginosa cells was visualized via fluorescence microscopy using a lectin adjunct with fluorescein isothiocyanate, and resulted in carbohydrate and protein accumulation in the cellular capsule. Given that the highest carbohydrate accumulation was seen in the Con A system (where iron accumulation was relatively lower), carbohydrate quality is likely important factor that influences cellular iron accumulation. Since PNA specifically binds to sugars such as galactose and N-acetylgalactosamine, these saccharide species could be important candidates for intracellular and extracellular iron accumulation and transport. Microcystin biosynthesis was stimulated in the presence of PNA and WGA, whereas cellular iron uptake increased only in the presence of PNA. Thus, the iron uptake was not necessarily congruent with the upregulation of microcystin synthesis, which suggested that the positive effect of lectin on iron uptake is probably attributable to the PNA-assisted iron accumulation around the cell surface. Overall, the present study provides insights into the interactions of lectin that influence cellular metabolic activities such as iron uptake, extracellular polymeric substance accumulation, and toxin production.     

Distribution of lectin binding glycoprotein in osteoclasts

Arachis hypogaea (PNA) lectin, specific for Gal-B-1,3-GalNac disaccharide and Wheat germ (WGA) lectin, specific for (GlcNac) and terminal neuraminic acid were used to identify histiocytic giant cells, osteoclast like giant cells and osteoclasts. PNA lectin, without neuraminidase predigestion was not bound by the giant cells, while they showed a strong reaction with WGA lectin. Neuraminidase pretreatment decreased WGA lectin binding, which supports that neuraminic acid plays a role in the binding of WGA. On the other hand, neuraminidase digestion liberated large amounts of PNA binding sites in every type of giant cells examined, showing a strong, intracytoplasmic granular staining. This observation is indicative of presence of PNA binding sites masked by neuraminic acid. Instead of the intracytoplasmic PNA binding in some osteoclasts a well defined part of the cytomembrane was heavily stained. We suppose that this PNA binding part of cytomembrane equals to the zone of resorption, characterized by the ruffled border of osteoclasts. Our findings indicate that a neuraminic acid substituted PNA binding glycoprotein is synthetized both in osteoclasts and histiocytic giant cells which may indicate a common origin of these cell types.     

Carbohydrate determinants of organs of the reproductive tract of the mouse according to the results of using lectins with different specificities

Histotopography of lectin receptor sites in adult mice ovary, oviduct, uterus, testis and epididymis has been investigated on light-optic level by means of lectin-peroxidase technique. Paraffin sections are treated with peanut agglutinin (PNA), soybean agglutinin (SBA), wheat germ agglutinin (WGA) and Laburnum anagyroides lectin (LAL), conjugated with horseradish peroxidase. Concanavalin A (Con A) receptor sites are visualized by indirect method. The usefulness of lectins for selective histochemic evaluation of definite organ structures is demonstrated. Zona pellucida, luteocytes, oviductal and uterine epitheliocytes are rich in receptor sites for all lectin used in the investigation. The most intense binding to zonae pellucidae glycoconjugates possess WGA and LAL, to luteocytes--PNA, SBA and LAL, to oviductal and uterine epitheliocytes--Con A and LAL. The preferential SBA binding to the acrosomal system and plasma membranes of spermatogenic cells is demonstrated. Changes in lectin-binding patterns during the maturation of intraovarian oocytes and spermatogenic cells are also studied. The perspectives of practical application of the results obtained, as well as trends in further lectin histochemistry investigations of the reproductive system are discussed.     

Distribution of lectin binding glycoprotein in osteoclasts

Summary Arachis hypogaea (PNA) lectin, specific for Gal-B-1, 3-GalNac disaccharide and Wheat germ (WGA) lectin, specific for (GlcNac) and terminal neuraminic acid were used to identify histiocytic giant cells, osteoclast like giant cells and osteoclasts. PNA lectin, without neuraminidase predigestion was not bound by the giant cells, while they showed a strong reaction with WGA lectin. Neuraminidase pretreatment decreased WGA lectin binding, which supports that neuraminic acid plays a role in the binding of WGA. On the other hand, neuraminidase digestion liberated large amounts of PNA binding sites in every type of giant cells examined, showing a strong, intracytoplasmic granular staining. This observation is indicative of presence of PNA binding sites masked by neuraminic acid. Instead of the intracytoplasmic PNA binding in some osteoclasts a well defined part of the cytomembrane was havily stained. We suppose that this PNA binding part of cytomembrane equals to the zone of resorption, characterized by the ruffled border of osteoclasts. Our findings indicate that a neuraminic acid substituted PNA binding glycoprotein is synthetized both in osteoclasts and histiocytic giant cells which may indicate a common origin of these cell types.     

Lectin binding pattern in normal human labial mucosa

Summary The pattern of lectin binding in normal human labial mucosa was examined by light and electron microscopy using eight different lectins (ConA, LCA, WGA, UEA-1, RCA-1, SBA, DBA and PNA) and compared with the patterns in normal human skin and oesophageal mucosa. As seen by light microscopy, ConA, LCA, and WGA stained cell membranes in all layers of the mucosae. RCA-1 stained the plasma membrane of cells in the basal and middle layers, whereas cells in the superficial layers showed little positive staining. UEA-1, SBA, and PNA stained the cells in the middle layers weakly in some cases. No positive staining for DBA was seen. By electron microscopy, reaction product indicating ConA-binding sites was observed in the plasma membrane, cisternae of the endoplasmic reticulum, nuclear envelope and the Golgi apparatus. Binding of LCA, WGA, and RCA-1 was observed in the plasma membrane. These results show that the binding pattern of PNA, SBA, and RCA-1 in labial mucosa is different from that in the normal skin or oesophageal mucosa, although the labial mucosal epithelium, epidermis, and oesophageal epithelium are all stratified squamous epithelia. These differences in the cell-surface sugar residues are likely to be related to the possible functional differences in these tissues.     

Changes of glycoconjugates in human hepatocellular carcinoma

Receptors of 12 lectins in 25 cases of human hepatocellular carcinomas (HCC) were histochemically investigated by avidin-biotin-peroxidase complex (ABC) method. Liver tissues of five cirrhotic patients and five normal subjects were used as controls. SJA receptor was absent both in HCC and controls, while LCA and PSA receptors were present in all tissues studied here. Receptors of DBA, PHA, PNA, UEAI and SBA which did not bind to normal, cirrhotic and pericarcinomatous liver tissues had the positive rates of 4%, 44%, 16%, 4% and 12% in HCC, respectively. Four lectins which strongly bound to the non-cancer liver tissues had their receptors in 96% (ConA, WGA, RCAI) and 36% (BSAI) of HCC. The pretreatment of tissue sections with neuraminidase abolished most of WGA receptors and exposed some PNA binding sites. There were many differences in lectin distribution between HCC and noncancer liver tissues. The changes of glycoconjugates in HCC were discussed.     

Contribution of fungi to chronic dermatitis and lymphangitis in patients with filariasis]

Adenolymphangitis is a common occurrence in filarial lymphedema. Damage to the lymphatics by F. bancrofti is followed by obliteration of lymph vessel and lymph stasis. Obstruction of lymphatics prevents the microbs skin penetration. Presented studies were performed to evaluate the role of fungi colonization of the skin among patients with filarial lymphedema. The fungal colonization of skin in patients with filarial lymphedema may be an important reason for chronic inflammatory disorders.     

Lectin binding by structures of the submandibular gland of the rat during postnatal ontogenesis in the presence of thyroid pathology

Redistribution of lectin receptor sites in rat submandibular gland under hypo- and hyperthyroidism has been investigated using lectin-peroxidase technique. Lectin preparation include con A, peanut agglutinin (PNA), wheat germ agglutinin (WGA) and fucose-specific lectin from Laburnum anagyroides bark (LAB). Submandibular gland is excised on the 1st, 10th, 20th and 40th days of postnatal life and in adult rats. Hyperthyroidism, as well as hypothyroidism cause a considerable decrease of con A, WGA and LAB binding with simultaneous enhancement of PNA binding in all investigated groups of the animals. The only exception from this regularity is noted for con A: hypothyroidism in adult rats and on the 40th day of postnatal development causes enhancement of this lectin binding to granular duct epitheliocytes. Selective staining of mast cells with PNA and of separate neurocytes with con A is also demonstrated; these cells staining is affected neither by hyperthyroidism, nor by hypothyroidism. Possible interpretation of the observed phenomena is discussed.     

Changes of glycoconjugates in human hepatocellular carcinoma

Summary Receptors of 12 lectins in 25 cases of human hepatocellular carcinomas (HCC) were histochemically investigated by avidin-biotin-peroxidase complex (ABC) methol. Liver tissues of five cirrhotic patients and five normal subjects were used as controls. SJA receptor was absent both in HCC and controls, while LCA and PSA receptors were present in all tissues studied here. Receptors of DBA, PHA, PNA, UEA_I and SBA which did not bind to normal, cirrhotic and pericarcinomatous liver tissues had the positive rates of 4%, 44%, 16%, 4% and 12% in HCC, respectively. Four lectins which strongly bound to the non-cancer liver tissues had their receptors in 96% (ConA, WGA, RCA_I) and 36% (BSA_I) of HCC. The pretreatment of tissue sections with neuraminidase abolished most of WGA receptors and exposed some PNA binding sites. There were many differences in lectin distribution between HCC and noncancer liver tissues. The changes of glycoconjugates in HCC were discussed.     

Lectin binding in skeletal muscle. Evaluation of alkaline phosphatase conjugated avidin staining procedures

Summary Cryostat sections from rat gracilis muscles were incubated with different biotinylated lectins: Con A (Concanavilin A), WGA (Wheat germ agglutinin), SBA (soybean agglutinin), GS I and GS II (Griffonia simplicifolia agglutinin), LCA (Lens culinaris agglutinin), PNA (peanut agglutinin) and PSA (Pisum sativum agglutinin). The sections were subsequently treated with alkaline phosphatase conjugated avidin. The lectin binding sites were visualized after incubation in substrate media containing: (1) 5-bromo-4-chloro indoxyl phosphate and Nitro Blue tetrazolium or copper sulphate; (2) naphthol AS-MX phosphate or naphthol AS-BI phosphate and various types of diazonium salts; (3) -naphthylphosphate and Fast Blue BB; (4) -glycerophosphate according to the method of Gomori. The results obtained with the alkaline phosphatase methods were compared with those seen with a streptavidin-horseradish peroxidase procedure. Several chromogen protocols for visualizing alkaline phosphatase activity showed differences in the ability to detect lectin binding sites. A sarcoplasmic reaction was evident for Con A, GS II, WGA, LCA, and PSA after incubation in the indoxyl phosphate medium. Sarcoplasmic reaction for GS II was also noticed after incubation with naphthol AS-MX Fast Blue BB and -glycerophosphate. The latter substrate also gave rise to a sarcoplasmic Con A reaction. With the indoxylphosphate tetrazolium salt method some muscle fibres showed a very strong intracellular reaction after incubation with Con A and GS II while the staining intensity was weak in other fibres. The same muscle fibres were stained with PAS. No sarcoplasmic reactions were observed with either naphthol phosphate media or with the diaminobenzidine peroxidase methods. Further, the staining of the muscle fibre periphery, connective tissue, and capillaries was intensified using the indoxyl method. The indoxylphosphate-tetrazolium salt method seems to be suitable for future investigations of lectin binding sites in muscle sections.     

Lectin histochemistry of rabbit nephron

In order to investigate the usefulness of lectin histochemistry to detail nephronal segmentation we used 12 different biotinylated lectins (Con-A, DBA, GS-I, LCA, PNA, PWN, RCA-I, RCA-II, SWGA, SBA, UEA-I, and WGA) and Avidin-Biotin-Peroxidase (ABC) system on formalin-fixed and paraffin-embedded rabbit kidney sections. Each lectin, except UEA-I which did not stain any nephron structure, shows a different staining pattern along the nephron. Con-A, LCA, and RCA-I display a diffuse staining, while BS-I, RCA-II, SWGA, PWN, DBA, SBA and PNA are selective markers for specific nephron tracts. Furthermore, it is possible, according to the WGA binding pattern, to differentiate the convoluted part of the proximal tubule into two parts, named Segment A and Segment B. Lectin histochemistry on formalin-fixed and paraffin-embedded rabbit kidney sections displays a specific binding pattern along the rabbit nephron and shows interesting morphofunctional correlations.     

Bacterial infections of skin and soft tissues in filariasis

Adenolymphangitis is a common occurrence in filarial lymphedema. Damage to the lymphatics and lymph nodes by F. bancrofti is followed by obliteration of lymph vessels and lymph stasis. Obstruction of lymphatics prevents the bacteria penetrating skin to be evacuated with lymph stream to regional lymph nodes. Colonization of dermis, subcutis and lymphatics evokes clinical symptoms of adenolymphangitis. The question arises which strains of bacteria are responsible for the acute and chronic types of adenolymphangitis. The most probable strains responsible for this condition belong to the cocci and probably the bacillus strains.     

Mode of cell surface marker changes during retinoic acid-induced differentiation in pluripotent embryonal carcinoma cells

Pluripotent teratocarcinoma cell line, 311, was cultured in the presence of retinoic acid (RA) and studied for the processes of early marker changes associated with cell differentiation. The cell populations that have lost peanut agglutinin (PNA), Lotus tetragonolobus agglutinin (LTA) or wheat germ agglutinin (WGA) receptor increased in proportion to the period since the start of RA treatment. The kinetics of the appearance of these receptor-negative cell populations suggests that the differentiating cells lose lectin receptors in the order of PNA, LTA and WGA. However, the changes in F9 antigen(s) and LTA receptor occurred at an equal frequency in PNA+ and PNA- cells, indicating that, although the loss of lectin receptors takes place in a distinct order, the change in each receptor itself proceeds independently of the state of other lectin receptors.     

Anatomic distribution of lectin-binding sites in mouse testis and epididymis

Testis and epididymis of sexually mature mice were studied histochemically using 25 fluorescein-isothiocyanate-labeled lectins. Several lectin-specific binding patterns were recognized. Thus, HAA, HPA, GSA-I, and UEA-II reacted only with spermatozoa. PNA, GSA-II, SBA, VVA, BPA, RCA-I, and RCA-II reacted with spermatozoa and spermatocytes. WGA, PEA, LCA, and MPA reacted with spermatogonia, spermatocytes, and spermatozoa in increasing order of intensity. ConA, Suc. ConA, LAA, STA, LTA, LPA, PHA-E, PHA-L, UEA-I, and LBA reacted with all spermatogenic cells with equal intensity. In the epididymis, 12 lectins reacted uniformly with the epithelial cells lining all segments of this organ. One lectin (VVA) did not react with epididymal lining cells. The remaining 12 lectins reacted in a specific manner with portions of the head, body, or tail, thus selectively outlining different portions of the epididymis. RCA-I and RCA-II selectively accentuated the so-called halo cells of the epididymis. These findings provide a detailed map of lectin-binding sites in the mouse testis and epididymis and show that certain lectins can be used as specific markers for spermatogenic cells and segments of the epididymis.     

Anatomic distribution of lectin-binding sites in mouse testis and epididymis

Abstract. Testis and epididymis of sexually mature mice were studied histochemically using 25 fluorescein-isothiocyanate-labeled lectins. Several lectin-specific binding patterns were recognized. Thus, HAA, HPA, GSA-I, and UEA-I1 reacted only with spermatozoa. PNA, GSA-11, SBA, VVA, BPA, RCA-I, and RCA-I1 reacted with spermatozoa and spermatocytes. WGA, PEA, LCA, and MPA reacted with spermatogonia, spermatocytes, and spematozoa in increasing order of intensity. ConA, SUC. ConA, LAA, STA, LTA, LPA, PHA-E, PHA-L, IJEA-I, and LBA reacted with all spermatogenic cells with equal intensity. In the epididymis, 12 lectins reacted uniformly with the epithelial cells lining all segments of this organ. One lectin (VVA) did not react with epididymal lining cells. The remaining 12 lectins reacted in a specific manner with portions of the head, body, or tail, thus selectively outlining different portions of the epididymis. RCA-I and RCA-I1 selectively accentuated the so-called halo cells of the epididymis. These findings provide a detailed map of lectin-binding sites in the mouse testis and epididymis and show that certain lectins can be used as specific markers for spermatogenic cells and segments of the epididymis.     

Lectin histochemistry of epidermal glandular cells in the earthworm Lumbricus terrestris (Annelida Oligochaeta)

Carbohydrate residues were localized in the glandular cells of the epidermis of Lumbricus terrestris by lectin histochemistry. The following biotinylated lectins were used: ConA, PNA, WGA, UEA-I. Each lectin has a specific binding pattern in the epidermal glandular cells. The ConA binding is evident in the orthochromatic mucous cells; PNA in the metachromatic mucous cells; WGA in the neuroendocrine-like cells; UEA-I in the cuticle. The epidermal glandular cells possess specific sites for the different lectins in relation to their functional characteristics. Therefore, these sugar residues indicate different behaviours of the cells in epidermal functions related to ion transport, receptor-secretory processes and defence.     

Postnatal development of lectin binding sites in the rat ventral prostate

Summary Five Fluorescein-isothiocyanate (FITC)-labelled lectins were used to study the postnatal development of carbohydrate constituents in the rat ventral prostate: Concanavalin A (Con A), wheat germ agglutinin (WGA), peanut agglutinin (PNA),Dolichos biflorus agglutinin (DBA) andRicinus communis agglutinin I (RCA-I) With all the lectins, tested, except RCA-I, specific binding sites could be shown for every stage of differentiation in the glandular epithelium. Binding sites for Con A, WGA, PNA and DBA were found from day 10 to 13 post partum onwards. Each lectin showed a characteristic localization. Binding sites for the lectins used changed to different extents during the following two weeks. After the 24th day post partum no further changes in the lectin binding pattern could be found. The development of the lectin binding properties showed that the changes in carbohydrate-containing constituents of the prostate correlate with the beginning of prostatic secretion and to prostatic epithelial differentiation. In the periacinar stroma the development of the lectin binding pattern was similar to that in the glandular epithelium. The changes of stromal binding sites for Con A and WGA during epithelial differentiation may reflect the changes of epithelial-stromal interactions in the prostate.     

Characterization of carbohydrates of adult Echinococcus granulosus by lectin-binding analysis

The identification of lectin-binding structures in adult worms of Echinococcus granulosus was carried out by lectin fluorescence; the distribution of carbohydrates in parasite glycoconjugates was also studied by lectin blotting. The lectins with the most ample recognition pattern were ConA, WGA, and PNA. ConA showed widespread reactivity in tegument and parenchyma components, including the reproductive system, suggesting that mannose is a highly expressed component of the adult glycans. Although reproductive structures appeared to be rich in N-acetyl-D-glucosamine (GlcNAc)-N-acetyl neuraminic acid (NeuAc) and galactose (Gal) as demonstrated by their strong reactivity with WGA and PNA, respectively, some differences were observed in their labeling patterns. This was very clear in the case of the vagina, which only reacted with WGA. Furthermore, WGA and ConA both had reactivity with the excretory canals. RCA, the other Gal binding lectin used, only reacted with the tegument, suggesting that widespread PNA reactivity with the reproductive system is related to the presence of the D-Gal-beta-(1,3)D-GalNAc terminal structure. UEA I failed to bind to any parasite tissues as determined by lectin fluorescence, whereas DBA and SBA showed a very faint staining of the tegument. However, in transferred glycans, N-acetyl-D-galactosamine (GalNAc) and fucose (Fuc) containing glycoproteins were distinctly detected.