concanavalin-a and maclurin

The mammalian zona pellucida contains several glycoproteins whose oligosaccharide moieties are known to play a key role in the interaction with spermatozoa. Since zona pellucida defects may represent one of the most likely causes of failed fertilization in human in-vitro reproduction, we have studied the carbohydrate composition and distribution over the human zona pellucida by means of lectins. Donated, not inseminated cumulus-oocyte complexes, from cohorts with high fertilization rates, and fertilization-failed oocytes from cohorts inseminated with proven fertile donor semen, were analysed using 11 fluorescein-labelled lectins, on deplasticized semi-thin epoxy sections. Results showed that wheat germ agglutinin (WGA), Maclura pomifera (MPA) and Pisum sativum (PSA) bound to the extracellular matrix bordering the zona pellucida-corona radiata interface of cumulus-oocytes complexes, while the zona pellucida was labelled by WGA, Concanavalin A (ConA) and PSA. WGA labelling and correlative electron microscopy on the cumulus-oocyte complexes demonstrated that this lectin is a useful tool to trace the cortical granule distribution in the human oocyte. Surprisingly, in the failed-fertilized oocytes the zona pellucida was also labelled by MPA and showed three different patterns: (i) labelling of the zona pellucida outer surface; (ii) uniform labelling; (iii) labelling of an outer zona pellucida layer with variable thickness. Comparative analysis of WGA and MPA labelling on single failed-fertilized oocytes demonstrated that MPA zona pellucida patterns are not related to the cortical reaction. The nature and meaning of the MPA pattern of failed-fertilized oocytes were discussed in the light of zona pellucida defects impairing sperm receptivity.

Topics: Adult; Carbohydrates; Concanavalin A; Female; Fertilization in Vitro; Fluorescein; Fluorescent Dyes; Humans; Lectins; Male; Microscopy, Electron; Microscopy, Fluorescence; Oocytes; Plant Lectins; Wheat Germ Agglutinins; Zona Pellucida

The membrane structure of bronchial ciliated epithelium and lymphoepithelium was studied in rats by lectin histochemistry. The lymphoepithelium, contrary to bronchial ciliated epithelium, did not contain terminal beta-D-galactose residues. Moreover L-fucose and beta-D-Gal (1-3)-D-GalNAc residues, being masked, could be visualized only after enzymatic digestion of terminal sialic acid. These structural differences in membranes provide a basis for the different functions of bronchial lymphoepithelium.

Topics: Animals; Bronchi; Cell Membrane; Concanavalin A; Epithelial Cells; Epithelium; Histocytochemistry; Lectins; Lymphoid Tissue; Male; Membrane Glycoproteins; Microscopy, Electron; Peanut Agglutinin; Plant Lectins; Rats; Rats, Sprague-Dawley; Wheat Germ Agglutinins

Secretion of surfactant phosphatidylcholine has been extensively studied and there is evidence that it is a regulated process that can be influenced by a variety of physiological factors and pharmacological agents. In contrast, secretion of the major surfactant protein, surfactant protein A (SP-A), has been investigated to much lesser extent. It is not known whether SP-A secretion is constitutive or regulated and, if regulated, whether its regulation is similar to that of phosphatidylcholine. To address those questions we measured SP-A secretion in primary cultures of type II pneumocytes under conditions identical to those used to study phosphatidylcholine secretion. Freshly isolated cells from adult rats were cultured overnight, washed, and then incubated in fresh medium in the presence and absence of surfactant phospholipid secretagogues. As previously reported for phosphatidylcholine, SP-A secretion was linear with time for up to 4 h. However, the rate of SP-A secretion, approximately 6% of total SP-A (cells+medium) released into the medium per hour, was more than sixfold greater than that of the lipid. Although freshly isolated cells contained 70% more SP-A than cells that were cultured overnight, the rate of SP-A secretion was not significantly different. Secretion of SP-A by freshly isolated or cultured type II cells was not increased by a combination of ATP, terbutaline, the adenosine A2 receptor agonist 5'(N-ethylcarboxyamido)adenosine, 12-O-tetradecanoylphorbol-13-acetate, and ionomycin at concentrations that optimally stimulated phosphatidylcholine secretion. We conclude that secretion of the major lipid and protein components of surfactant are independently regulated.

Topics: Adenosine; Adenosine Triphosphate; Adenosine-5'-(N-ethylcarboxamide); Animals; Blotting, Western; Cells, Cultured; Concanavalin A; Electrophoresis, Polyacrylamide Gel; Immune Sera; Ionomycin; Lectins; Lung; Molecular Weight; Phosphatidylcholines; Plant Lectins; Proteolipids; Pulmonary Surfactant-Associated Protein A; Pulmonary Surfactant-Associated Proteins; Pulmonary Surfactants; Rats; Receptors, Purinergic P2; Spectrophotometry; Terbutaline; Tetradecanoylphorbol Acetate

Secretion of surfactant phospholipids can be inhibited by the surfactant-associated protein SP-A. SP-A was reported to stimulate lipid uptake by type II cells, whereas surfactant secretagogues were reported to have the same effect in isolated perfused lungs. We examined the effect of such secretagogues on uptake of liposomes containing L-alpha-[2-palmitoyl-9,103H(N)]-dipalmitoyl phosphatidylcholine in primary cultures of type II cells. As SP-A contains a lectinlike domain and other lectins were reported to inhibit surfactant secretion, we also examined the effect of such lectins on lipid uptake. At concentrations at which they maximally stimulate phosphatidylcholine secretion in type II cells, several secretagogues had no effect on liposome uptake. Maclura pomifera agglutinin (MPA) stimulated uptake approximately 10-fold with a concentration eliciting 50% maximum stimulation (EC50) of 17 micrograms/ml. The effect of MPA on uptake was considerably greater than that of SP-A. However, although the stimulatory effect of ATP on phosphatidylcholine secretion was almost completely antagonized by SP-A, it was maximally inhibited only 75% by MPA. The concentration eliciting 50% maximum inhibition (IC50) for MPA inhibition of secretion was 0.5 micrograms/ml. Concanavalin A, another lectin, had no effect on lipid uptake but completely inhibited secretion. These data show that a lectin other than SP-A can stimulate phospholipid uptake by type II cells cultured on plastic and suggest that surfactant secretion and reuptake are independently regulated processes.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Animals; Cells, Cultured; Concanavalin A; Cytochalasins; Endocytosis; Kinetics; L-Lactate Dehydrogenase; Lectins; Liposomes; Male; Microscopy, Fluorescence; Plant Lectins; Pulmonary Alveoli; Pulmonary Surfactants; Rats; Rats, Inbred Strains