A differential response elicited in macrophages on interaction with lectins

The interaction of several lectins, both native and chemically modified, with mouse peritoneal macrophages was studied. Surface distribution and interiorization of the lectins was assessed quantitatively using their radioactively-labeled derivatives, and qualitatively by employing fluorescein-labeled lectins. On the basis of their effect on the macrophages, the lectins tested fall into two classes: lectins that induce vacuole formation in the cells (concanavalin A (ConA), wax bean agglutinin (WBA) and wheat germ agglutinin (WGA)) and lectins that in their native form do not induce vacuolation (soybean agglutinin (SBA), peanut agglutinin (PNA) and the lectin from Lotus tetragonolobus (LT)). Neuraminidase treatment of the cells did not change their response to the lectins, though in the case of SBA and PNA binding was observed only with neuraminidase-treated macrophages. Incubation of the latter cells with SBA and subsequently with ConA resulted in significantly higher vacuolation than that observed with ConA alone. Glutaraldehyde-crosslinked polymers of SBA and of PNA, which are multivalent with respect to sugar binding, induced vacuolation in neuraminidase-treated macrophages. On the other hand, succinylation of ConA, which reduces the number of sugar binding sites per mole from four to two, abolished its ability to induce vacuole formation. These data suggest that multivalency of lectins and probably also their size are important factors in inducing vacuolation, by causing extensive crosslinkage of membrane receptors which is prerequisite for triggering of vacuole formation. Quantitative binding and internalization data indicate that vacuole formation is not directly related to the number of lectin receptors on the macrophages nor to the extent of their internalization.     

Cellular heterogeneity in normal human urothelium: Quantitative studies of lectin binding

Summary A panel of 10 FITC-labelled lectins (MPA, PNA, ConA, DBA, SBA, RCA-120, WGA, UEA, GS-I, GS-II) was applied to cryosections of seven specimens of normal urothelium. Seven of the lectins (MPA, ConA, RCA, WGA, UEA, GS-I and GS-II) showed a pattern of increasing fluorescence intensity from basal to superficial cells of the urothelium whereas PNA, DBA and SBA showed more uniform binding throughout the urothelium. Urothelial cell suspensions labelled with FITC-lectins were studied by flow cytometry to quantify the variation in binding to different cells types. Three cellular subpopulations were identified in normal urothelium on the basis of their optical properties. Fluorescence intensity due to specific lectin binding was then measured separately for each subpopulation. Although there was some variation among individual cases, a general pattern emerged in this small series. WGA, RCA, and GS-II bind in large quantities to all urothelial cells while PNA, SBA, ConA and DBA show little binding. MPA, RCA, UEA and GS-I showed the most marked increase in fluorescence intensity from basal to superficial cells as observed microscopically and quantified by flow cytometry.     

Glycoconjugates in normal human kidney

Summary The avidin-biotin-peroxidase complex technique was used with 13 lectins to study the glycoconjugates of normal human renal tissue. The evaluated lectins included Triticum vulgaris (WGA), Concanavalin ensiformis (ConA), Phaseolus vulgaris leukoagglutinin and erythroagglutinin (PHA-L and PHA-E), Lens culinaris (LCA), Pisum sativum (PSA), Dolichos biflorus (DBA), Glycine max (SBA), Bandeiraea simplicifolia I (BSL-I), Ulex europaeus I (UEA-I) and Ricinus communis I (RCA-I). Characteristic and reproducible staining patterns were observed. WGA and ConA stained all tubules; PHA-L, PHA-E, LCA, PSA stained predominantly proximal tubules; DBA, SBA, PNA, SJA and BSL-I stained predominantly distal portions of nephrons. In glomeruli, WGA and PHA-L stained predominantly visceral epithelial cells; ConA stained predominantly basement membranes and UEA-I stained exclusively endothelial cells. UEA-I also stained endothelial cells of other blood vessels and medullary collecting ducts. Sialidase treatment before staining caused marked changes of the binding patterns of several lectins including a focal loss of glomerular and tubular staining by WGA; an acquired staining of endothelium by PNA and SBA; and of glomeruli by PNA, SBA, PHA-E, LCA, PSA and RCA-I. The known saccharide specificities and binding patterns of the lectins employed in this study allowed some conclusions about the nature and the distribution of the sugar residues in the oligosaccharide chains of renal glycoconjugates. The technique used in this report may be applicable to other studies such as evaluation of normal renal maturation, classification of renal cysts and pathogenesis of nephrotic syndrome. The observations herein reported may serve as a reference for these studies.     

Lectin receptors on the cell surface membrane and the kinetics of lectin-induced cell agglutination.

Agglutination of malignant transformed hamster cells by concanavalin A (ConA) and the lectins from wheat germ (WGA) and soybean (SBA) has been automatically quantitated, by measuring the amount of light transmitted through a cell suspension. The transformed hamster cells were agglutinated by SBA only after treatment with neuraminidase. The initial rate of agglutination and the concentration of lectin (K_c) required for the half-maximum rate (V_m) has been determined. The initial rate and V_m were lower and more temperature-sensitive, and the K_c was higher, for ConA than for WGA and SBA. There was no detectable temperature-dependent phase transition for the initial rate of agglutination. The total number of receptors was lower for ConA than for WGA and SBA and the apparent association constant between lectin molecules and cell surface receptors was higher for ConA (10⁷M^?1) than for WGA and SBA (1.6 × 10⁶M^?1). The half V_m of agglutination required 75% saturation of the cell receptors for ConA, and only 13–17% saturation of the receptors for SBA and WGA. A 30% decrease in the number of SBA receptors present in agglutinable cells completely prevented their agglutination. The results indicate that there is heterogeneity of lectin receptors on the cell surface and that only a small proportion of the total number of WGA and SBA receptors have to be occupied for agglutination by these lectins.     

Trypanosoma rhodesiense Bloodstream Trypomastigote and Culture Procyclic Cell Surface Carbohydrates1, 2, 3

Bloodstream trypomastigote and culture procyclic (insect midgut) forms of a cloned T. rhodesiense variant (WRATat 1) were tested for agglutination with the lectins concanavalin A (Con A), phytohemagglutinin P (PP), soybean agglutinin (SBA), fucose binding protein (FBP), wheat germ agglutinin (WGA), and castor bean lectin (RCA). Fluorescence-microscopic localization of lectin binding to both formalin-fixed trypomastigotes and red cells was determined with fluorescein isothiocyanate (FITC)-conjugated Con A, SBA, FBP, WGA, RCA, PNA (peanut agglutinin), DBA (Dolichos bifloris), and UEA (Ulex europaeus) lectins. Electron microscopic localization of lectin binding sites on bloodstream trypomastigotes was accomplished by the Con A-horseradish peroxidase-diaminobenzidine (HRP-DAB) technique, and by a Con A-biotin/avidin-ferritin method. Trypomastigotes, isolated by centrifugation or filtration through DEAE-cellulose or thawed after cryopreservation, were agglutinated by the lectins Con A and PP with agglutination strength scored as Con A < PP. No agglutination was observed in control preparations or with the lectins WGA, FBA or SBA. Red cells were agglutinated by all the lectins tested. Formalin-fixed bloodstream trypomastigotes bound FITC-Con A and FITC-RCA but not FITC-WGA, -SBA, -PNA, -UEA or -DBA lectins. All FITC-labeled lectins bound to red cells. Con A receptors, visualized by Con A-HRP-DAB and Con A-biotin/avidin-ferritin techniques, were distributed uniformly on T. rhodesiense bloodstream forms. No lectin receptors were visualized on control preparations. Culture procyclics lacked a cell surface coat and were agglutinated by Con A and WGA but not RCA, SBA, PP and FBP. Procyclics were not agglutinated by lectins in the presence of competing sugar at 0.25 M. The expression of lectin binding cell surface saccharides of T. rhodesiense WRATat 1 is related to the parasite stage. Sugars resembling α-D-mannose are on the surface of bloodstream trypomastigotes and culture procyclics; n-acetyl-D-galactosamine and D-galactose residues are on bloodstream forms; and n-acetyl-D-glucosamine-like sugars are on procyclic stages.     

Flow cytometric analyses of lectin binding to rat alveolar macrophages

The binding characteristics of ten FITC-labeled plant lectins (Con-A, MPA, BPA, PNA, WGA, SBA, UEA-I, DBA, GS-I, GS-II) to lavaged rat alveolar macrophages were assessed by flow cytometry. The alveolar macrophages (AM) were incubated with varying concentrations of each lectin in a pinocytosis-inhibiting buffer. In addition to measuring lectin-associated green fluorescence, the electronic cell volumes and axial light loss characteristics of the AM were also measured flow cytometrically. These latter parameters were found to be good indicators of cell agglutination caused by some of the lectins, and, in conjunction with green fluorescence measurements, usefully serve to determine optimal or nonagglutinating lectin concentrations for flow cytometric studies. With the exception of UEA-I, all of the lectins examined bound to AM, although a wide range of binding was observed among the lectins. At subagglutinating concentrations, Con A, MPA, BPA, PNA, WGA, SBA, and GS-I bound to the AM with unimodal patterns. Histograms of lectin-associated fluorescence intensity obtained with DBA clearly presented a pattern consistent with a more complex, bimodal distribution of labeled AM, suggesting the presence of at least two subset populations. The low-intensity distribution of AM represented congruent to 70% of the cells, while the more strongly labeled subset represented congruent to 15% of the parent AM population. The remaining balance of the AM was identified as another subpopulation by the failure to detectably bind to the DBA. While GS-II bound to all of the AM, this lectin labeled about 5% of the cells much more intensely than the bulk of the population. Thus, two subset populations of AM could be resolved according to their differing avidities for the GS-II lectin.     

Lectin binding sites as markers of neonatal porcine uterine development.

To identify lectin binding sites and to determine if lectin binding patterns change with age in developing neonatal porcine uterine tissues, gilts (n = 3/day) were hysterectomized on Day 0 (birth), 7, 14, 28, 42, or 56. Lectin binding was visualized in Bouin's-fixed uterine tissues with seven biotinylated lectins (ConA, DBA, PNA, RCA-I, SBA, UEA-I, and WGA) and avidin-peroxidase staining procedures. Lectin specificities were demonstrated by pre-incubating lectins with appropriate inhibitory sugars (0.2 M). Staining intensity was evaluated visually (absent, weak, moderate, or strong) for three endometrial tissues; luminal epithelium, glandular epithelium, and stroma. Staining intensities for DBA, PNA, SBA, and WGA were not affected by neonatal age. Staining with these lectins was greater in uterine epithelium (moderate or strong) than in stroma (weak). In contrast, binding patterns for ConA, UEA-I, and RCA-I were affected by neonatal age. Strong epithelial staining associated with ConA binding was observed on all days, whereas stromal ConA staining decreased in intensity from moderate to weak after Day 14. Epithelial staining with UEA-I increased from moderate to strong after Day 28, whereas stromal UEA-I staining decreased from moderate to weak after day 28. Staining with RCA-I was homogeneous for luminal epithelium and stroma but variegated for glandular epithelium on and after Day 7. These observations indicate that a variety of lectin binding sites are present in developing neonatal porcine endometrial tissues and that developmentally related alterations in the distribution and/or orientation of glycoconjugates containing alpha-D-mannose, beta-D-galactose, beta-D-acetyl-N-galactosamine, and alpha-L-fucose residues occur between birth and Day 56 as these tissues mature.     

Lectin staining of cultured CNS microglia.

Carbohydrate binding proteins, known as lectins, bind to specific sugar groups on most membranes. We used fluorescent and light microscopy to study the interaction of various lectins with the membranes of microglia cultured from neonatal rat or fetal mouse cerebral cortices. Microglia stained intensely with GS-1, RCA, WGA, and ConA and slightly with DBA, UEA, BPA, and SBA. No staining was seen with GS-2, MPA, or PNA. Staining was specific for microglia in the mixed glial cultures and was dose dependent. In addition, microglial lectin binding could be reduced or blocked by competitive inhibition using specific sugars. Treatment of the microglia with agents such as dimethylsulfoxide (DMSO), interleukin-1 (IL-1), interferon (IFN), or lipopolysaccharide (LPS) did not eliminate lectin staining, although the degree of staining was altered. Positive staining of the microglia was also associated with a functional change for at least one lectin, i.e., ConA. Superoxide anion production by microglia was increased in the presence of ConA. Overall, binding of the lectins GS-1, RCA, WGA, and ConA can be used as an identifying tool for microglia in glial cultures, but intensity of staining varies depending on their functional state.     

FITC-labeled lectins and calcofluor white ST as probes for the investigation of the molecular architecture of cell surfaces. Studies on conjugatophycean species

Summary In a screening program with 7 FITC-labeled lectins as probes, ConA receptors were identified in all of the 28 members of theConjugatophyceae, being under investigation. In nearly all of them RCA₁₂₀ receptors, too, are expressed. In 3 species only, PNA receptors, and in 2 species UEA receptors have been detected. No binding of DBA, SBA, and WGA was observed. The receptors for ConA, RCA₁₂₀, and UEA were shown to be associated with different molecules. Each lectin exhibits a unique and specific binding pattern, both chemically, as well as with regard to the topographic distribution on cell surfaces. While ConA receptors predominantly are associated with constituents of the cell wall, RCA₁₂₀ receptors mostly form part of the surrounding mucilage; the same holds for UEA receptors. Besides a variability of topographic distribution and species-to-species variation, a cell-to-cell variation exists in many species, suggesting that the expression of a lectin receptor is due to the developmental state of the cell and/or depends on external stimuli. In conclusion, we may point out, that FITC-labeled lectins turned out to be extremely useful probes for the investigation of the molecular architecture of cell walls. Calcofluor white ST binding to fibrillar polysaccharides (most probably cellulose) was shown to be inhibited by external incrustations of the cell wall. One species does not show any reaction with calcofluor white ST at all.     

Binding of fluorescent lectins to the surface of germ cells from fetal and early postnatal mouse gonads

Fluorescent lectins were used to study the chemical nature of carbohydrate moieties present on the surface of female and male germ cells isolated from mouse gonads during fetal and early posnatal development. Concanavalin A (ConA), lens culinaris agglutinin (LCA), ricinus communis agglutinin (RCA_I) and wheat germ agglutinin (WGA) bound intensely to the germ cell plasma membrane at all stages studied. Other lectins such as ulex europaeus agglutinin (UEA_I) and agglutinin (SBA) did not bind or bound moderately (SBA to female germ cells only). Distinct developmental-related changes were observed when female germ cells were labeled with fluorescein-conjugated peanut agglutinin (PNA) or dolichos biflorus agglutinin (DBA). DBA and PNA binding was absent or weak in fetal female and male germ cells, but became intensely positive in oocytes in the immediate postnatal period. The percentage of oocytes stained with DBA increased during the first three days after birth, and from day 3–4 onwards all oocytes were strongly labeled. I suggest that these changes in lectin binding reflect changes in biochemical structure of the oocyte surface related to differentiative events occurring in the mouse ovary immediately after birth.     

Pulmonary collectins enhance phagocytosis of Mycobacterium avium through increased activity of mannose receptor

Collectins, including surfactant proteins A (SP-A) and D (SP-D) and mannose binding lectin (MBL), are the important constituents of the innate immune system. Mycobacterium avium, a facultative intracellular pathogen, has developed numerous mechanisms for entering mononuclear phagocytes. In this study, we investigated the interactions of collectins with M. avium and the effects of these lectins on phagocytosis of M. avium by macrophages. SP-A, SP-D, and MBL exhibited a concentration-dependent binding to M. avium. The binding of SP-A to M. avium was Ca(2+)-dependent but that of SP-D and MBL was Ca(2+)-independent. SP-A and SP-D but not MBL enhanced the phagocytosis of FITC-labeled M. avium by rat alveolar macrophages and human monocyte-derived macrophages. Excess mannan, zymosan, and lipoarabinomannan derived from the M. avium-intracellular complex, significantly decreased the collectin-stimulated phagocytosis of M. avium. Enhanced phagocytosis was not affected by the presence of cycloheximide or chelation of Ca(2+). The mutated collectin, SP-A(E195Q, R197D) exhibited decreased binding to M. avium but stimulated phagocytosis to a level comparable to wild-type SP-A. Enhanced phagocytosis by cells persisted even after preincubation and removal of SP-A or SP-D. Rat alveolar macrophages that had been incubated with SP-A or SP-D also exhibited enhanced uptake of (125)I-mannosylated BSA. Analysis by confocal microscopy and flow cytometry revealed that the lung collectins up-regulated the cell surface expression of mannose receptor on monocyte-derived macrophages. These results provide compelling evidence that SP-A and SP-D enhance mannose receptor-mediated phagocytosis of M. avium by macrophages.     

Differences in lectin binding patterns of normal human endometrium between proliferative and secretory phases

Lectin binding patterns in normal human endometrium were examined by light and electron microscopy using seven different lectins (ConA, WGA, RCA, PNA, UEA-1, DBA, and SBA). For light microscopic observations, criteria based on the incidence and intensity of cells positive for the lectin staining were adopted to evaluate the different staining patterns of the proliferative and secretory endometria obtained by the avidin-biotin-peroxidase complex (ABC) technique. At the light microscopic level, ConA, WGA, and RCA stained endometrial glandular cells in both phases. The number of PNA-positive cells with the binding sites entirely limited to the apical surface tended to be reduced slightly in the secretory phase. UEA-1 weakly stained the apical surface of glandular cells in the proliferative phase but not in the secretory phase. Among the lectins used in this study, DBA and SBA displayed remarkable changes between the phases. That is, in the proliferative phase they produced only a faint or slight positive stain at the apical surface, but the incidence and intensity of DBA- and the SBA-positive glandular cells increased in the secretory phase. By electron microscopy, the reaction product of ConA was observed in the plasma membrane, endoplasmic reticulum, nuclear envelope, and the Golgi apparatus, and the binding sites of RCA and DBA were observed in the plasma and Golgi membranes. Between both phases, the reactivity of ConA and RCA showed almost no change. However, the secretory endometrial cells containing the DBA-positive Golgi apparatus were markedly increased in number compared with the proliferative ones bearing the lectin-positive organelles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Combined biochemical and cytological analysis of membrane trafficking using lectins

We have tested the application of high-mannose-binding lectins as analytical reagents to identify N-glycans in the early secretory pathway of HeLa cells during subcellular fractionation and cytochemistry. Post-endoplasmic reticulum (ER) pre-Golgi intermediates were separated from the ER on Nycodenz–sucrose gradients, and the glycan composition of each gradient fraction was profiled using lectin blotting. The fractions containing the post-ER pre-Golgi intermediates are found to contain a subset of N-linked α-mannose glycans that bind the lectins Galanthus nivalis agglutinin (GNA), Pisum sativum agglutinin (PSA), and Lens culinaris agglutinin (LCA) but not lectins binding Golgi-modified glycans. Cytochemical analysis demonstrates that high-mannose-containing glycoproteins are predominantly localized to the ER and the early secretory pathway. Indirect immunofluorescence microscopy revealed that GNA colocalizes with the ER marker protein disulfide isomerase (PDI) and the COPI coat protein β-COP. In situ competition with concanavalin A (ConA), another high-mannose specific lectin, and subsequent GNA lectin histochemistry refined the localization of N-glyans containing nonreducing mannosyl groups, accentuating the GNA vesicular staining. Using GNA and treatments that perturb ER–Golgi transport, we demonstrate that lectins can be used to detect changes in membrane trafficking pathways histochemically. Overall, we find that conjugated plant lectins are effective tools for combinatory biochemical and cytological analysis of membrane trafficking of glycoproteins.     

137 Gamete Recognition and Sexual Isolation in a Red Alga,Aglaothamnion Byssoides (Rhdophyta)

The binding of fluorescein isothiocyanate (FITC) conjugated lectins to gametes of Aglaothamnion byssoides Itono during the fertilization was studied by the use of confocal microscope. The physiological effects of lectins and carbohydrates on gamete binding were also examined. Three lectins, concanavalin A (ConA), Soybean agglutinin (SBA) and wheat germ agglutinin (WGA) bound to the surface of spermatia, but each lectin labeled different region of the spermatium. SBA bound only to the spermatial appendages but ConA bound to the whole spermatial surface except spermatial appendages. WGA labeled narrow region which connects spermatial body and appendages. During fertilization, ConA and WGA specific substances on the spermatial surface moved towards the area contacting with trichogyne and accumulated on the surface of fertilization canal. Spermatial binding to trichogynes was inhibited by pre‐incubation of spermatia with SBA, while trichogyne receptors were blocked by the complementary carbohydrate, N‐acetyl‐D‐galactosamine. WGA and its complementary carbohydrate had little effect on gamete binding. For searching the step of sexual isolation, crossing experiment was performed between Aglaothamnion byssoides and twelve other red algal species. Results showed that the gamete recognition was genus‐specific: the gametes bound freely with their partners of the same genus. When two species from same genus were crossed, sexual isolation occurs gradually during the fertilization process. Therefore, sexual isolation in red algae appears to be determined by multi‐step process and gamete binding is the initial step.     

Lectin-binding pattern of neuroepithelial and respiratory epithelial cells in the mouse nasal cavity

Summary Sections from the nasal cavity of 12-day-old Swiss albino mice (NMRI strain) were subjected to lectin histochemistry. A panel of biotinylated lectins (Con A, WGA, s-WGA, PNA, SBA, DBA and UEA I) and a horseradish peroxidase-conjugated lectin (GSA II) showed marked differences in binding to the respiratory and the neuroepithelial cells. SBA (affinity for galactose andN-acetylgalactosamine), PNA (galactose) and WGA (sialic acids andN-acetylglucosamine) labelled the receptor neurons in the olfactory and vomeronasal epithelium. DBA (N-acetylgalactosamine) labelled a subgroup of about 5% of the olfactory receptor neurons, but most neurons in the vomeronasal organ. UEA I (fucose) and s-WGA (N-acetylglucosamine) intensely labelled the entire nerve cell population in the vomeronasal organ, but in the olfactory epithelium the labelling with these lectins was stratified. In the respiratory epithelium the ciliated cells were labelled with WGA and s-WGA, while the secretory cells bound most of the lectins. Thus different sugars are exposed on the surface of the different types of epithelia in the nasal cavity, providing a basis for selectivity in microbial attacks on these areas.     

Lectin activity of the major surfactant protein (SP-A) may participate in, but is not required for, binding to rat type II cells.

Pulmonary surfactant is a complex mixture of lipids and proteins, of which surfactant protein A (SP-A) is the most abundant glycoprotein. The SP-A molecule has several distinct structural features that include a lectin-like domain, sharing structural features with other mammalian lectins. We have tested the hypothesis that lectin activity of the SP-A molecule is required for the binding to its receptor on the surface of alveolar Type II cells. By using colloidal gold immunocytochemistry in conjunction with electron microscopy, we evaluated the ability of mannosylated proteins to inhibit canine SP-A binding to rat Type II cells in vitro. After preincubation of SP-A with the mannosylated protein horse-radish peroxidase (HRP), SP-A was incubated with isolated filter-grown Type II cells. HRP did not alter the binding of SP-A to the Type II cell surface. Evidence that SP-A did bind to HRP was shown by coincident observation of gold-labeled SP-A and HRP precipitates. These results provide visual evidence that the lectin activity associated with SP-A is not required for its binding to receptor on rat alveolar Type II epithelial cells.     

Lectin binding sites in duodeno-jejunal mucosae from coeliac children

Summary The distribution pattern of some labelled lectins (WGA, SBA, PNA, DBA, ConA, LCA) has been investigated in small intestinal mucosa of coeliac children (119 cases) and normal (short stature) and pathological (postenteritis syndrome) controls. The oligosaccharide side chains of glycoproteins present in the cytoplasm and in the brush border of enterocytes, goblet cells and luminal crypt surface have been revealed. The most important differences in lectin binding reactivity between coeliac and controls mucosae are discussed.     

Lectin binding sites in duodeno-jejunal mucosae from coeliac children

The distribution pattern of some labelled lectins (WGA, SBA, PNA, DBA, ConA, LCA) has been investigated in small intestinal mucosa of coeliac children (119 cases) and normal (short stature) and pathological (postenteritis syndrome) controls. The oligosaccharide side chains of glycoproteins present in the cytoplasm and in the brush border of enterocytes, goblet cells and luminal crypt surface have been revealed. The most important differences in lectin binding reactivity between coeliac and controls mucosae are discussed.     

Studies on fertilization in the ascidians : II. Lectin binding to the gametes of ciona intestinalis

In the present work we have compared the binding of fluorescein-conjugated lectins (concanavalin A (ConA), wheat germ agglutinin (WGA), fucose binding protein (FBP) and soybean agglutinin (SBA)) to the sperm surface and to the egg and its envelopes of Ciona intestinalis. Only WGA is bound to the follicle cells: yet this lectin has no binding sites on the sperm surface. Both ConA and FBP are bound by the chorion, the oolemma and the sperm surface. However, while ConA reacts only with the sperm head, FBP is bound both to the head and to the flagellum. Experiments on the effect of ConA and FBP on the fertilization reaction have been carried out. The role of the lectin-binding sites that are shared by the surfaces of both gametes is discussed in connection with the nature of the sperm-binding sites.     

Immuno- and lectin histochemistry of epithelial subtypes and their changes in a radiation-induced lung fibrosis model of the mini pig

Cell types of lung epithelia of mini pigs have been studied using a panel of monoclonal and polyclonal antibodies against cytokeratins (CKs) and vimentin and three lectins before and after radiation-induced fibrosis. In normal tissues, CK18 specific antibodies reacted above all with type II alveolar epithelial cells, while CK7 and pan CK-specific antibodies stained the whole alveolar epithelium. In bronchial epithelial cells, CKs 7, 8, 18 and focally CKs 4 and 13 as well as vimentin were found. Cell specificity of the CK pattern was confirmed by double label immunofluorescence using type II cell-specific Maclura pomifera (MPA) lectin, type I cell specific Lycopersicon esculentum (LEA) lectin and capillary endothelium-binding Dolichos biflorus (DBA) lectin. In experimental pulmonary fibrosis, enhanced coexpression of CK and vimentin was observed in bronchial epithelium. Subtypes of alveolar epithelial cells were no longer easily distinguishable. CK18 was found to be expressed in the entire alveolar epithelium. The gradual loss of the normal alveolar epithelial marker, as seen by the binding of MPA to type I-like cells, of LEA to type II-like cells and the partial loss of MPA-binding to type II cells, was paralleled by the appearance of CK4, typical for squamous epithelia, and the occurrence of DBA-binding in epithelial cells. Implications of these results for general concepts of intermediate filament protein expression and lectin binding in the fibrotic process are discussed.