Distribution Of Lectin Receptors Research Articles

Changes in the distribution of lectin receptors during capacitation and acrosome reaction in boar spermatozoa

Sperm glycocalyx modifications are known to occur during capacitation and the acrosome reaction (AR). These changes are very important for gamete recognition and fertilization in mammals but are not fully understood. The purpose of this study was to determine the distribution of surface carbohydrates in boar spermatozoa during capacitation and the AR. These processes may be associated with specific changes in the content and distribution of surface carbohydrates. Thirty-nine ejaculates from fertile boars of various breeds were analyzed. N-Acetylglucosamine and sialic acid, mannose and fucose residues were detected by fluorescence microscopy and flow cytometry using FITC-conjugated lectins. Triticum vulgaris agglutinin (WGA) bound on the head and tail of fresh sperm, and fluorescence intensity (FI) decreased in capacitated sperm (6751 to 5621 fluorescence units (FU), P<0.05), and decreased further in acrosome-reacted sperm (5240 FU, P<0.05). Concanavalia ensiformis agglutinin (Con-A) bound homogeneously on the head and the midpiece of fresh sperm with a FI of 5335 FU, and increased in capacitated sperm (5957 FU, P<0.05) mainly on the acrosomal region. In acrosome-reacted sperm, fluorescence was concentrated on the border of the acrosomal region (5608 FU, P<0.05). It was not possible to detect Ulex europaeus agglutinin (UEA) by fluorescence microscopy. However, flow cytometry revealed UEA receptors (187 FU), with a nonsignificant decreased number in capacitated (142 FU) and AR sperm (142 FU). Labeling patterns were similar in all breeds. Sperm glycocalyx modifications observed in this study provide insights to the molecular modifications accompanying capacitation and the AR. This kind of study could improve the diagnosis of reproductive problems of subfertile boars and males of other species.

Maturation-related changes in distribution of lectin receptors in baboon (Papio anubis) spermatozoa during epididymal maturation.

Changes in distribution of surface glycoproteins in baboon sperm were studied by lectin blotting techniques. In baboon, several changes in sperm surface occur during epididymal passage. These changes include increased staining of band that was observed with the WGA binding glycoproteins of 140, 80, 52, and 46 kDa; Con A bands of 66, 37, and 26 kDa; and the PNA binding glycoprotein of 114 kDa. A second change was the loss of preexisting band that was observed with RCA binding glycoproteins of 120, 80, 58, 53, 46, and 37 kDa; and the Con A band of 80 kDa. A final change was noted with Con A binding in which six bands of 8, 9, 12, 14, 18, and 21 kDa were added as the sperm matured through the cauda epididymis. These findings present new information on the changes in distribution of surface glycoproteins in baboon sperm during epididymal passage. There was some reorganization of the molecular structure of the sperm during epididymal maturation.

Distribution of lectin receptors on the plasma membranes of brain glioma cells and autologous peripheral blood mononuclear cells as a function of the degree of anaplasia

The content of thed-mannose-specific LCL-receptor that bindsd-mannose-containing lymphokines (interleukin-1 and interleukin-2) is proportional to the degree of glioma anaplasia. This may reflect the mechanism whereby brain glioma utilizes lymphokines to proliferate and to escape immunologic surveillance.

Distribution of lectin receptors in the human hyperplastic tonsil: histochemical and flow cytometric analyses.

The distribution of lectin receptors in the human tonsil was studied using 16 biotinylated lectins. The avidin-biotin-peroxidase complex (ABC) method was used on frozen and paraffin-embedded tissue sections. Cell suspensions were also analysed by dual flow cytometry using respective fluorescein isothiocyanate-conjugated lectins and phycoerythrin-labeled anti-CD3 and anti-human immunoglobulin. Frozen sections fixed with acetone and paraffin-embedded materials fixed in three solutions were compared for lectin affinity; ethanol-fixed sections gave best results followed by frozen and buffered formalin-fixed ones, then nonbuffered formalin. Con-A, RCA-1, LcH, WGA, MPA, PHA, PSA, PNA, SJA and GSA-1 reacted with all tissue components of the tonsil in immunohistochemical studies, but binding intensity was fixative dependent. Binding of Lotus and BPA to lymphocytes was limited to germinal center lymphocytes. Other tissue components were also reactive but staining intensity was weaker in Lotus compared with BPA. SBA and DBA did not react with lymphocytes, but reacted with macrophages/histiocytes, vascular endothelia, and epithelial cells. LBA and LPA were constantly negative with all tissue components irrespective of fixatives. Flow cytometric analyses showed that all but three (DBA, LBA and LPA) partially or totally stained lymphocyte surfaces. Lotus receptors were expressed exclusively on B-lymphocytes.

Characteristics of the distribution of lectin receptors in intrahepatic cholangiocellular carcinoma

The receptors of peanut agglutinin (PNA), Dolichos biflorus agglutinin (DBA) and Ulex europaeus agglutinin I (UEA-I) were localized in intrahepatic cholangiocellular carcinoma, hepatocellular carcinoma, intrahepatic bile ducts and normal, cirrhotic and pericarcinomatous liver using the avidin-biotin-peroxidase complex method. It was found that epithelial cells of normal bile ducts had many UEA-I receptors, fewer DBA receptors and no PNA receptors. The positive rates of PNA, UEA-I and DBA receptors in 18 cases of intrahepatic cholangiocellular carcinoma were 88.9%, 61.1% and 33.3% respectively, which were significantly higher than those in hepatocellular carcinoma (16.0%, 4.0% and 4.0% respectively). Hepatocytes in normal, cirrhotic and pericarcinomatous liver had no receptors for these three lectins. It is suggested that lectin receptor distribution in intrahepatic cholangiocellular carcinoma is obviously different from that in normal bile duct cells and in hepatocellular carcinoma, and might be used as an auxiliary index in its clinical diagnosis.

Expression of glycoconjugates in intrahepatic cholangiocellular carcinoma

Twenty cases of intrahepatic cholangiocellular carcinoma (IHCCC) were studied by lectin histochemistry for their glycoconjugate expression. Combined alcian blue-peroxidic acid Schiff (ABPS) staining was also made for mucins in these tissues. Results showed that epithelial cells of intrahepatic bile ducts contained varied quantity of DBA, WGA, LCA, Con A, PHA, RCAI, BLAI, SJA, SBA, PSA and UEAI receptors but no PNA receptors. Lectin receptors were present at varying rates; 100% (Con A), 95% (PSA), 90% (LCA), 95% (WGA), 30% (BSAI), 35% (DBA), 65% (PHA), 74% (PNA), 85% (RCAI), 35% (SBA), 25% (SJA) and 55% (UEAI), respectively. The positive rates in well-differentiated IHCCC were significantly higher than those in either moderately or poorly-differentiated IHCCC. The distribution of lectin receptors in IHCCC was obviously different from that in peri-carcinomatous, cirrhotic and normal liver, and also differed from that in epithelial cells of normal intrahepatic bile ducts. All larger ducts were stained by ABPS and were mainly blue colour, while only 80% of IHCCC were positive for ABPS staining. Most of them were blue, others were red or purple. The results suggest that the expression of glycoconjugates had changed after the neoplastic transformation of bile duct cells.

Distribution of lectin receptors in postimplantation mouse embryos at 6–8 days gestation

We have examined the pattern of binding of eleven lectins--BSL-II, WGA, LPA, Con A, DBA, SBA, LTA, UEA-I, MPA, PNA, and RCA-I, with specificity for a range of saccharides, to postimplantation mouse embryos from 6 to 8 days of gestation. The lectins were used to stain sections of ethanol-fixed paraffin-embedded and formaldehyde-fixed gelatin-embedded embryonic material. Our observations reveal a complex pattern of lectin binding to both cell surfaces and cytoplasm. Many of the lectins bind particularly to the outer surface of visceral endoderm (e.g., DBA, WGA, SBA, and RCA-I) and to the surface of the proamniotic cavity (e.g., RCA-I, PNA, and WGA). In the newly formed mesenchyme of primitive-streak-stage embryos, galactose and N-Ac-neuraminic acid are present but lectins with specificity for other sugars either did not bind to the cells or bound only in small amounts.

Ultrastructural localization of lectin receptors on the luminal and abluminal aspects of brain micro-blood vessels.

Lectin- or glycoprotein-colloidal gold complexes were used for detection of specific monosaccharide residues in mouse brain micro-blood vessels (MBVs). The lectins tested recognize the following residues: beta-D-galactosyl (Ricinus communis agglutinin-120, RCA-1), alpha-N-acetylgalactosaminyl (Helix pomatia agglutinin, HPA), alpha-D-mannosyl and alpha-D-glucosyl (Concanavalin A, Con A), sialoglycoconjugates (Limax flavus agglutinin, LFA), N-acetylglucosaminyl and sialyl (wheat germ agglutinin, WGA), and alpha-L-fucosyl (Ulex europeus agglutinin, UEA-1). Use of these lectin-gold complexes and ultrathin sections of Lowicryl K4M-embedded tissue makes it possible to gain insights into localization of lectin receptors in the entire cross-section of MBV walls. Receptors for all lectins, except UEA-1, were found on both luminal and abluminal fronts of the endothelial cells (ECs). Differential labeling of luminal and abluminal fronts of ECs with some lectins (Con A, HPL) is considered to reflect the polarity of the endothelium. Some differences noted in the distribution of lectin receptors in the wall of representatives of three types of MBVs (capillaries, arterioles, and venules) are thought to be associated with different functions performed by the above-mentioned segments of the microvasculature in maintenance of the blood-brain barrier.

Differentiated microdomains on the luminal surface of capillary endothelium: distribution of lectin receptors.

Lectins conjugated with either peroxidase or ferritin were used to detect specific monosaccharide residues on the luminal front of he fenestrated endothelium in the capillaries of murine pancreas and intestinal mucosa. The lectins tested recognize, if accessible, the following residues: alpha-N-acetylgalactosaminyl (soybean lectin), beta-D-galactosyl (peanut agglutinin [PA] and Ricinus communis agglutinin-120 [RCA]), beta-N-acetylglucosaminyl and sialyl residues (wheat germ agglutinin [WGA]), alpha-L-fucosyl (lotus tetragonolobus lectin), and alpha-D-glucosyl and beta-D-mannosyl (concanavalin A [ConA]). Thi labeled lectins were introduced by perfusion in situ after thoroughly flushing with phosphate-buffered saline the microvascular beds under investigation. Specimens were fixed by perfusion, and subsequently processed for peroxidase detection and electron microscopy. Control experiments included perfusion with: (a) unlabeled lectin before lectin conjugate; (b) labeled lectin together with the cognate hapten sugar, and (c) horseradish peroxidase or ferritin alone. Binding sites were found to be relatively homogeneously distributed on the plasmalemma proper, except for Lotus tetragonolobus lectin and Con A, which frequently bound in patches. Plasmalemmal vesicles, transendothelial channels, and their associated diaphragms were particularly rich in residues recognized by RCA and PA (beta-D-galactosyl residues) and by WGA (beta-N-acetylglucosaminyl residues). Receptors for all lectins tested appeared to be absent or considerably less concentrated on fenestral diaphragms. The results reported here extend and complement previous findings on the existence of microdomains generated by the preferential distribution of chemically different anionic sites (Simionescu et al., 1981, J. Cell Biol., 9:605-613 and 614-621).

The distribution of lectin receptors on the plasma membrane of the fertilized sea urchin egg during first and second cleavage

The Con A receptors in the plasma membrane of the fertilized sea urchin egg are uniformly distributed. With the onset of first division, the fluorescent lectin-receptor complexes become localized in the cleavage furrow. A similar asymmetric pattern is seen if the cleaving cells are fixed before the addition of fluorescent Con A. After cleavage is complete, fluorescent Con A is lost from the furrow and accumulates at the poles furthest from the plane of cleavage. This asymmetry is retained through second cleavage; thus the individual blastomeres have a surface memory of their original orientation with respect to each other.

Lectin receptors on cells isolated from the turtle retina.

The presence of specific oligosaccharides on the surface of retinal cells was examined by incubating FITC-labeled lectins with cells dissociated from papain-treated turtle retinas. The pattern and intensity of binding was found to vary among the cells examined. With Con A, there was strong surface staining of both rods and cones, with an intense ring of fluorescence above the nucleus. The bipolar and ganglion cells also showed strong surface labeling. In Müller (glial) cells there was intense fluorescence in the apical, microvillous region. In contrast, the horizontal cells and their axons showed weak staining. When RCA-60, RCA-120, and WGA were incubated with photoreceptors, bipolar cells, or horizontal cells, little fluorescence was visible. However, all three lectins bound strongly to the Müller cells. In contrast, the Lotus lectin did not bind to any of the cells examined. In all the cases, lectin binding was inhibited by the appropriate haptene sugar. Further, prior treatment cells with neuraminidase did not alter lectin binding to any cell type. These results suggest differences in the distribution of lectin receptors among specific cell types, and particularly between neurons and glial cells in the vertebrate retina.

The effect of amphotericin B on lectin-induced aggregation of cell surface receptors

The mobility of concanavalin A (ConA) and ricin receptors from NS20 neuroblastoma and C 6 glioma cells was studied using an electrophoretic technique. Cells attached to a solid support were exposed to an electrical field (12V cm −1) at room temperature. The distribution of lectin receptors on the cell surface was revealed by fluorescent conjugates of lectins and microscopic observation of the fixed cells. This technique allowed the estimation of the mobilities of lectin receptors either in free or liganded form, depending on the time at which the cells are labeled with lectins (either after or before electrophoresis). In line with previous observations [1] it is shown that in their free form ConA and ricin receptors are mobile all over the cell surface. Ligand binding induced an apparent receptor immobilization. Immobilization of ricin receptors from C 6 glioma cells could be induced either by the multivalent or the monovalent form of the lectin indicating that cross-linking of receptors by the ligand did not play a predominant role in the process of receptor immobilization. Amphotericin B but not ionophores like valinomycin or gramicidin blocked ligand-induced receptor immobilization. It is concluded from this observation that the effect of amphotericin B is not related to its ionophoretic properties but more likely to its capacity to interact with membrane cholesterol. When cells were incubated at 37 °C extensive patching of lectin receptors could be observed. This process was also inhibited by amphotericin B. A model is proposed to account for a role of cholesterol in ligand-induced receptor immobilization and patching.

Distribution and mobility of lectin receptors on synaptic membranes of identified neurons in the central nervous system.

The distribution and mobility of concanavalin A (Con A) and Ricinus communis agglutinin (RCA) receptors (binding sites) on the external surfaces of Purkinje, hippocampal pyramidal, and granule cells and their attached boutons were studied using ferritin-lectin conjugates. Dendritic fields of these cells were isolated by microdissection and gently homogenized. Cell fragments and pre- and postsynaptic membranes were labeled with the ferritin-lectin conjugates at a variety of temperatures, and the distribution of lectin receptors was determined by electron microscopy. Both classes of these lectin receptors were concentrated at nearly all open and partially open postsynaptic junctional membranes of asymmetric-type synapses on all three neuron types. Con A receptors were most concentrated at the junctional membrane region, indicating that the mature neuron has a specialized nonrandom organization of carbohydrates on its outer surface. Lectin receptors located on postsynaptic junctional membranes appeared to be restricted in their mobility compared to similar classes of receptors on extrajunctional membrane regions. Labeling with ferritin-RCA and -Con A at 37 degrees C produced clustering of lectin receptors on nonjunctional surfaces; however, Con A and RCA receptors retained their nonrandom topographic distribution on the postsynaptic junctional surface. The restricted mobility of lectin receptors was an inherent property of the postsynaptic membrane since the presynaptic membrane was absent. It is proposed that structures in the postsynaptic density may be transmembrane-linked to postsynaptic receptors and thereby determine topographic distribution and limit diffusion of specialized synaptic molecules. Speicalized receptor displays may play an important role in the formation and maintenance of specific synaptic contacts.

Intramembrane particle distribution and lectin binding of glioblastoma cells after long term subculture

Human glioblastoma cells in long-term monolayer culture showed an even distribution of intramembrane particles (IMP) on all surfaces of the plasma membrane; junctional complexes were rarely observed and rectilinear arrays were not seen. Cells treated with Con A-ferritin and Ricin II-ferritin showed an even distribution of lectin receptors and under conditions used no capping occurred. Lectin-ferritin complexes were taken up into pinocytotic vesicles. Cleaved preparations of Ricin II-ferritin treated cells showed no change in the distribution of IMP.