Glycocalyx characterisation and glycoprotein expression of Sus domesticus epididymal sperm surface samples

2012 ◽  
Vol 24 (4) ◽  
pp. 619 ◽  
Author(s):  
Anna Fàbrega ◽  
Marta Puigmulé ◽  
Jean-Louis Dacheux ◽  
Sergi Bonet ◽  
Elisabeth Pinart
Keyword(s):  
Lectin Binding ◽  
Helix Pomatia ◽  
Surface Proteins ◽  
Surface Glycoprotein ◽  
Ulex Europaeus ◽  
Sperm Surface ◽  
Epididymal Maturation ◽  
Helix Pomatia Agglutinin ◽  
Ulex Europaeus Agglutinin I ◽  
Principal Piece

The sperm surface is covered with a dense coating of carbohydrate-rich molecules. Many of these molecules are involved in the acquisition of fertilising ability. In the present study, eight lectins (i.e. Arachis hypogae (peanut) agglutinin (PNA), Lens culimaris (lentil) agglutinin-A (LCA), Pisum sativum (pea) agglutin (PSA), Triticum vulgari (wheat) germ agglutinin (WGA), Helix pomatia agglutinin (HPA), Phaseolus vulgaris (red kidney bean) leucoagglutinin (PHA-L), Glycine max (soybean) agglutinin (SBA) and Ulex europaeus agglutinin I (UEA-I)) were investigated to identify changes in the nature and localisation of glycoproteins in boar spermatozoa migrating along the epididymal duct. Complementary procedures included measurement of global lectin binding over the surface of the viable sperm population by flow cytometry, analysis of lectin localisation on the membrane of individual spermatozoa using fluorescence microscopy and the electrophoretic characterisation of the major sperm surface glycoprotein receptors involved in lectin binding. A significant increase was found in sperm galactose, glucose/mannose and N-acetyl-d-glucosamine residues distally in the epididymis. Moreover, the sperm head, cytoplasmic droplet and midpiece were recognised by most of the lectins tested, whereas only HPA and WGA bound to the principal piece and end piece of the sperm tail. Fourteen sperm surface proteins were observed with different patterns of lectin expression between epididymal regions. The sperm glycocalyx modifications observed in the present study provide an insight into the molecular modifications associated with epididymal maturation, which may be correlated with the degree of maturation of ejaculated spermatozoa.

scholarly journals Lectin-binding properties of human breast cancer cell lines and human milk with particular reference to Helix pomatia agglutinin.

1995 ◽  
Vol 43 (3) ◽  
pp. 275-281 ◽  
Author(s):  
U Schumacher ◽  
E Adam ◽  
S A Brooks ◽  
A J Leathem
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Binding Sites ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Breast Cancer Cell Lines ◽  
Human Breast ◽  
Membrane Glycoproteins ◽  
Tissue Sections ◽  
Helix Pomatia Agglutinin

Several studies have shown binding of a variety of lectins to breast cancer cells in tissue sections. In particular, binding of the lectin from the Roman snail, Helix pomatia agglutinin (HPA), to breast cancer cells is linked with a poor prognosis. The molecular basis for lectin binding to metastatic breast cancers is not known. To elucidate this in a model system, lectin-binding patterns of seven human breast cancer cell lines were investigated, their cell membranes were isolated, and HPA binding was assessed. In addition, the influence of fixation and processing on lectin-binding sites was also investigated. Binding of lectins to the tumor cells was very heterogeneous between and within the different cell lines and was influenced by fixation and processing. However, some cell lines showed HPA-binding sites both in vivo and in tissue sections. Analysis of the isolated cell membrane glycoproteins from these cell lines on Western blots revealed that HPA can bind to several membrane glycoproteins. In contrast, human milk shows only one major milk glycoprotein that is HPA-positive. Therefore, a switch in glycosylation appears to be taking place during the transformation to a metastatic phenotype.

scholarly journals Lectinhistochemical staining of granuloma induced by bacillus Calmette-Guerin in Piaractus mesopotamicus

2013 ◽  
pp. 3753-3758
Author(s):  
Wilson G. Manrique ◽  
Gustavo S. Claudiano ◽  
Mayra AP. Figueiredo ◽  
Thalita R. Petrillo ◽  
Paulo F. Marcusso ◽  
...  
Keyword(s):  
Wheat Germ ◽  
Close Contact ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Control Group ◽  
Bacillus Calmette Guerin ◽  
Epithelioid Cells ◽  
Dolichos Biflorus ◽  
Piaractus Mesopotamicus ◽  
Helix Pomatia Agglutinin

ABSTRACTObjetive. This study was conducted to evaluate, by means of lectinhistochemistry (LHC), the expression of carbohydrates in granulomas induced by the bacillus Calmette-Guerin (BCG) in muscle tissue of Piaractus mesopotamicus after 33 days. Material and methods. Histological sections with 3 μm thick were incubated with the following lectins :WGA (Wheat germ agglutinin), DBA (Dolichos biflorus agglutinin) and HPA (Helix pomatia agglutinin), and the results were evaluated by light microscopy. Results. Acid fast bacilli were stained by Ziehl Neelsen (ZN) and strong labeled by WGA in the cytoplasm of macrophages. Labeling with DBA was intense in fibroblasts and weak in macrophages. On the other hand, HPA binding was stronger in macrophages, especially in those that were in close contact with epithelioid cells, without evidence of binding to fibroblasts. The epithelioid cells were not labeled by the used lectins, but they were identified by Hematoxilin-Eosin (HE). The lectins labeled specific type saccharides in glycoproteins, as N-acetylglucosamine present in bacilli and macrophages, as well as N-acetyl-galactosamine in macrophages. The control group showed no inflammation or lectin binding. Conclusions. This technique may be useful in identifying receptors for WGA, DBA and the HPA lectins in epithelioid granuloma induced by BCG in P. mesopotamicus

scholarly journals Helix pomatia agglutinin lectin-binding oligosaccharides of aggressive breast cancer

2001 ◽  
Vol 95 (2) ◽  
pp. 79-85 ◽  
Author(s):  
Miriam V. Dwek ◽  
Heidi A. Ross ◽  
Andrew J. Streets ◽  
Susan A. Brooks ◽  
Elizabeth Adam ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Helix Pomatia Agglutinin ◽  
Aggressive Breast Cancer

Purification and identification of sperm surface proteins and changes during epididymal maturation

PROTEOMICS ◽  
2011 ◽  
Vol 11 (10) ◽  
pp. 1952-1964 ◽  
Author(s):  
Clémence Belleannee ◽  
Maya Belghazi ◽  
Valérie Labas ◽  
Ana-Paula Teixeira-Gomes ◽  
Jean Luc Gatti ◽  
...  
Keyword(s):  
Surface Proteins ◽  
Sperm Surface ◽  
Epididymal Maturation ◽  
Sperm Surface Proteins

scholarly journals Distinct cytoskeletal domains revealed in sperm cells.

1984 ◽  
Vol 99 (3) ◽  
pp. 1083-1091 ◽  
Author(s):  
I Virtanen ◽  
R A Badley ◽  
R Paasivuo ◽  
V P Lehto
Keyword(s):  
Ricinus Communis ◽  
Surface Membrane ◽  
Helix Pomatia ◽  
Cytoskeletal Proteins ◽  
Sperm Cells ◽  
Positive Staining ◽  
Specific Staining ◽  
Cytoskeletal Organization ◽  
Helix Pomatia Agglutinin ◽  
Principal Piece

Antibodies against different cytoskeletal proteins were used to study the cytoskeletal organization of human spermatozoa. A positive staining with actin antibodies was seen in both the acrosomal cap region and the principal piece region of the tail. However, no staining was obtained with nitrobenzoxadiazol-phallacidin, suggesting that most of the actin was in the nonpolymerized form. Most of the myosin immunoreactivity was confirmed to a narrow band in the neck region of spermatozoa. Tubulin was located to the entire tail, whereas vimentin was only seen in a discrete band-like structure encircling the sperm head, apparently coinciding with the equatorial segment region. Surface staining of the spermatozoa with fluorochrome-coupled Helix pomatia agglutinin revealed a similar band-like structure that co-distributed with the vimentin-specific staining. Instead, other lectin conjugates used labeled either the acrosomal cap region (peanut and soybean agglutinins), both the acrosomal cap and the postacrosomal region of the head (concanavalin A), or the whole sperm cell surface membrane (wheat germ and lens culinaris agglutinins and ricinus communis agglutinin l). In lectin blotting experiments, the Helix pomatia agglutinin-binding was assigned to a 80,000-mol-wt polypeptide which, together with vimentin, also resisted treatment with Triton X-100. Only the acrosomal cap and the principal piece of the tail were decorated with rabbit and hydridoma antibodies against an immunoanalogue of erythrocyte alpha-spectrin (p230). p230 appeared to be the major calmodulin-binding polypeptide in spermatozoa, as shown by a direct overlay assay of electrophoretic blots of spermatozoa with 125I-calmodulin. The results indicate that spermatozoa have a highly specialized cytoskeletal organization and that the distribution of actin, spectrin, and vimentin can be correlated with distinct surface specializations of the sperm cells. This suggest that cytoskeleton may regulate the maintenance of these surface assemblies and, hence, affect the spermatozoan function.

Lectin binding of F9 embryonal carcinoma cells: evidence for population heterogeneity and developmentally regulated high-Mr cell surface proteins

1989 ◽  
Vol 92 (4) ◽  
pp. 561-568
Author(s):  
J. Tienari ◽  
I. Virtanen ◽  
E. Lehtonen
Keyword(s):  
Cell Surface ◽  
Embryonal Carcinoma ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Population Level ◽  
Electrophoretic Analysis ◽  
Surface Proteins ◽  
Population Heterogeneity ◽  
Cell Surface Proteins ◽  
F9 Cells

Undifferentiated F9 embryonal carcinoma (EC) cells bound fluorochrome-coupled Helix pomatia agglutinins (HPA) and peanut agglutinins (PNA) homogeneously, but were distinctly heterogeneous in their binding of Dolichos biflorus agglutinin (DBA) conjugates. Upon chemically induced differentiation the proportion of cells binding the DBA conjugates increased, but a distinct heterogeneity in the intensity of binding remained among the parietal endoderm (PE)-like F9 derivatives. These cells were heterogeneous in their binding of HPA conjugates as well, and many of them failed to bind PNA conjugates, apparently due to sialylation of the PNA-binding sites. Electrophoretic analysis of lectin-binding glycoproteins in the detergent-soluble fraction of the cells revealed the appearance of a doublet of polypeptides of Mr 300,000-400,000 upon differentiation induced by retinoic acid (RA). In addition, an Mr 220,000 polypeptide appeared upon differentiation induced by RA and dibutyryl cyclic AMP (dbcAMP). These polypeptides were obtained from both metabolically labelled and surface-labelled cells. A major secreted glycoprotein, which comigrated with laminin, bound to DBA. This suggests that laminin secreted by the differentiated F9 derivatives contains O-glycosidic saccharides. The results show that even though differentiation of F9 cells leads to changes in their binding of fluorochrome-coupled lectins, these lectin conjugates reveal distinct population heterogeneity among undifferentiated and differentiated F9 cells and are hence likely to be of limited value in the characterization of individual cells. At the whole cell population level, on the other hand, affinity binding to lectins reveals the appearance of high-Mr cell surface proteins in differentiating F9 cells.

scholarly journals Buffalo sperm surface proteome profiling reveals an intricate relationship between innate immunity and reproduction

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Vipul Batra ◽  
Vanya Bhushan ◽  
Syed Azmal Ali ◽  
Parul Sarwalia ◽  
Ankit Pal ◽  
...  
Keyword(s):  
Male Fertility ◽  
Female Reproductive Tract ◽  
Surface Proteins ◽  
Glycosylation Pattern ◽  
Sperm Surface ◽  
Beta Defensins ◽  
Surface Proteome ◽  
Related Proteins ◽  
Sperm Surface Proteins

Abstract Background Low conception rate (CR) despite insemination with morphologically normal spermatozoa is a common reproductive restraint that limits buffalo productivity. This accounts for a significant loss to the farmers and the dairy industry, especially in agriculture-based economies. The immune-related proteins on the sperm surface are known to regulate fertility by assisting the spermatozoa in their survival and performance in the female reproductive tract (FRT). Regardless of their importance, very few studies have specifically catalogued the buffalo sperm surface proteome. The study was designed to determine the identity of sperm surface proteins and to ascertain if the epididymal expressed beta-defensins (BDs), implicated in male fertility, are translated and applied onto buffalo sperm surface along with other immune-related proteins. Results The raw mass spectra data searched against an in-house generated proteome database from UniProt using Comet search engine identified more than 300 proteins on the ejaculated buffalo sperm surface which were bound either by non-covalent (ionic) interactions or by a glycosylphosphatidylinositol (GPI) anchor. The singular enrichment analysis (SEA) revealed that most of these proteins were extracellular with varied binding activities and were involved in either immune or reproductive processes. Flow cytometry using six FITC-labelled lectins confirmed the prediction of glycosylation of these proteins. Several beta-defensins (BDs), the anti-microbial peptides including the BuBD-129 and 126 were also identified amongst other buffalo sperm surface proteins. The presence of these proteins was subsequently confirmed by RT-qPCR, immunofluorescence and in vitro fertilization (IVF) experiments. Conclusions The surface of the buffalo spermatozoa is heavily glycosylated because of the epididymal secreted (glyco) proteins like BDs and the GPI-anchored proteins (GPI-APs). The glycosylation pattern of buffalo sperm-surface, however, could be perturbed in the presence of elevated salt concentration or incubation with PI-PLC. The identification of numerous BDs on the sperm surface strengthens our hypothesis that the buffalo BDs (BuBDs) assist the spermatozoa either in their survival or in performance in the FRT. Our results suggest that BuBD-129 is a sperm-surface BD that could have a role in buffalo sperm function. Further studies elucidating its exact physiological function are required to better understand its role in the regulation of male fertility.

Glycosylation of developing human glomeruli: lectin binding sites during cell induction and maturation.

1987 ◽  
Vol 35 (1) ◽  
pp. 33-37 ◽  
Author(s):  
H Holthöfer ◽  
I Virtanen
Keyword(s):  
Binding Sites ◽  
Mesangial Cells ◽  
Ricinus Communis ◽  
Lectin Binding ◽  
Helix Pomatia ◽  
Cell Types ◽  
Basement Membranes ◽  
Phenotypic Change ◽  
Neuraminidase Treatment ◽  
Nephron Development

Expression of cellular glycoconjugates during differentiation of human fetal kidney was studied using fluorochrome-labeled lectins. Each lectin revealed a characteristic binding pattern during the phenotypic change of the nephrogenic mesenchyme and during distinct stages of nephron development. The uninduced mesenchymal cells were positive for Pisum sativum (PSA), Concanavalin A (ConA), Wistaria floribunda (WGA), and Ricinus communis (RCA-I) lectins. However, these lectins failed to react with the uninduced cells of the S-shaped bodies, whereas Maclura pomifera (MPA), Triticum vulgaris (WGA) and, after neuraminidase treatment, Arachis hypogaea (PNA) agglutinins bound intensely to the presumptive podocytes. During later stages of nephrogenesis, MPA positively on the podocytes weakened and could not be observed in adult kidney glomeruli. Binding sites for Helix pomatia (HPA) agglutinin in glomeruli were also expressed only transiently during nephrogenesis. During further development PSA, ConA, WFA, and RCA-I reacted with mesangial cells in addition to the glomerular basement membranes. The segment-specific lectin binding patterns of the tubuli emerged in parallel with the appearance of brush border and Tamm-Horsfall antigens of the proximal and distal tubuli. The results show that nephron site-specific saccharides appear in a developmentally regulated manner and in parallel with morphologic maturation of the nephron. Lectins therefore appear to be useful tools for study of induction and maturation of various nephron cell types.

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