Expression pattern of glycoconjugates in rat retina as analysed by lectin histochemistry

The present study sought to characterize the expression and distribution of complex glycoconjugates in the rat retina by lectin histochemistry, using a panel of 21 different lectins with different carbohydrate specificities. Paraffin sections of Carnoy-fixed Sprague-Dawley rat eyes were stained with various biotinylated lectins, followed by the streptavidin-peroxidase and glucose oxidase-diaminobenzidine-nickel staining procedures. The results showed that the retinal pigment epithelium was stained intensely with LCA, Jacalin, WFA, S-WGA, PWA, DSA, UEA-I, LTA and PHA-E, suggesting that this epithelium contained glycoconjugates with alpha-Man, alpha-Glc, alpha-Gal/GalNAc, beta-GalNAc, alpha-Fuc, NeuAc and other oligosaccharide residues. The outer and inner segments of the photoreceptor layer showed different lectin binding affinities. The outer segments reacted with S-WGA and GS-II, whereas the inner segments reacted with UEA-II, UEA-I, LTA and MAA, suggesting that the inner segments contained glycoconjugates rich in alpha-Fuc and NeuAc(alpha2,3)Gal residues. PNA labelled specifically the cones and could be used as a specific marker for these photoreceptors. RCA-I, WFA, S-WGA, DSA, MAA and PHA-E reacted with both the outer and inner plexiform layers. On the other hand, UEA-I and LTA specifically labelled the outer plexiform layer, while PNA labelled the inner plexiform layer. The retinal microglial cells were labelled specifically by GS-I-B4 and SNA. Interestingly, we also observed that WFA bound specifically to Müller cells and could be used as a novel marker for this retinal glial cell. The capillaries and larger vessels in the retina and choriocapillaris reacted intensely with GS-I-B4, RCA-I, S-WGA, PWA, DSA and PHA-E. No significant differences in lectin binding were observed in the microvessels at these two sites. In summary, the present study demonstrated the expression patterns of glycoconjugates in the rat retina and that certain lectins could be used as histochemical markers for specific structural and cellular components of the rat retina.     

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.     

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.     

Glycoconjugates in normal human kidney. A histochemical study using 13 biotinylated lectins

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), Arachis hypogaea (PNA), Sophora japonica (SJA), 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.     

Cytochemical analysis of oligosaccharide processing in frog photoreceptors

Summary Lectin cytochemistry, together with exoglycosidase enzyme digestion, has been used to characterize partially glycoconjugates of several intracellular compartments in frog photoreceptors. In order to obtain uniform access of reagents to all intracellular compartments, the experiments were performed directly on semi-thin sections ofXenopus laevis retinal tissue embedded in a hydrophilic plastic resin. In the rod, the major photoreceptor intracellular binding sites for wheat germ agglutinin (WGA) are the outer segment, the Golgi complex, and other inner segment organelles which are probably involved in the transport of glycoconjugates from the Golgi complex to the outer segment. In addition, shed outer segment tips (phagosomes) are uniformly labelled with WGA. The WGA-binding sites of the outer segment and of the presumed transport organelles are resistant to neuraminidase digestion. This is consistent with the possibility that glycoconjugates (primarily opsin) are transported from the Golgi complex to the outer segment without further oligosaccharide processing. Specific staining of rod outer segments and of phagosomes is also obtained with theN-acetylglucosamine-specific lectin, succinyl-WGA (S-WGA). Outer segments and phagosomes stain the same with WGA, S-WGA and a variety of other lectins tested suggesting that no major post-Golgi oligosaccharide processing accompanies the shedding-phagocytosis event. Concanavalin A (Con A) staining of intracellular sites in rod inner segments reveals a striking difference compared to WGA staining in that the Con A binding sites are concentrated in the photoreceptor axon and presynaptic terminal. These results, and results from previous studies, indicate that the photoreceptor may utilize different mechanisms of oligosaccharide processing from the level of a single Golgi complex to the opposite ends of this cell. Furthermore, those glycoconjugates destined for the presynaptic terminal may undergo post-Golgi processing at or near their sites of insertion into the presynaptic plasma membrane.     

Distribution of lectin receptors sites in the zona pellucida of follicular and ovulated rat oocytes.

Carbohydrates of the zona pellucida (ZP) in mammals are believed to have a role in sperm-egg interaction. We have characterized the biochemical nature and distribution of the carbohydrate residues of rat ZP at the light (LM) and electron microscope (EM) levels, using lectins as probes. Immature female rats were induced to superovulate and cumulus-oocyte complexes were isolated from the oviduct, fixed with glutaraldehyde, and embedded in araldite for LM and LR-Gold for EM histochemistry. For examination of follicular oocytes, rat ovaries were fixed with glutaraldehyde and embedded in paraffin. The araldite or paraffin sections were deresined or deparaffinized, respectively, labeled with biotin-tagged lectins as probes, and avidin-biotin-peroxidase complex as visualant. For EM examination, thin LR-Gold sections were labeled with RCA-I colloidal gold complex (RCA/G) and stained with uranyl acetate. LM analyses indicate that in ovulated oocytes the ZP intensely binds peanut agglutinin (PNA); succinylated wheat germ agglutinin, (S-WGA), Griffonia simplisifolia agglutinin-I (GS-I) and soybean agglutinin (SBA), and to a lesser extent, lectins from Ricinus communis (RCA-I), Concanavaia ensiformis (Con A), Ulex europoeus (UEA-I), and wheat germ agglutinin (WGA). The neighboring cumulus cells are considerably less reactive and exhibit membrane staining only with Con A, WGA, and PNA. EM analysis of RCA/G binding revealed intensive binding to the inner layer region of the ZP and moderate binding to cytoplasmic vesicles of the cumulus cells. The ZP of follicular oocytes exhibits a different lectin binding pattern, expressed in staining strongly with PNA and S-WGA, and in a tendency of the lectin receptors to occur in the outer portion of the ZP.(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

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.     

Lectin histochemistry of brains from a murine mutant carrying a storage disorder

Summary A strain of Balb/c mice with neurovisceral storage disorder exhibits metabolic and phenotypic manifestations similar to those found in Niemann-Pick type C and D patients. The storage material in the brain reacted positively with periodate-Schiff reagent. To identify the chemical nature of the storage material we applied lectin histochemistry on paraffin-embedded and frozen sections, using biotinylated lectins and avidin-biotin-peroxidase complex. Major abnormalities were noted in the neurons and glia cells. Swollen neurons were stained heavily by Con A and S-WGA, whereas glia cells, mainly astrocytes, which were abundant both in the cerebrum and cerebellum, were positive to RCA-I, GS-I, PNA, S-WGA and WGA. The myelin tracts reacted with PNA, SBA and RCA-I but to a lesser extent in affected animals when compared to normals.Frozen brain sections stained positively only after extraction with chloroform methanol prior to the lectin treatment and revealed a lectin binding pattern similar to that of the paraffin-embedded preparations. The data presented here show that the stored glucoconjugates in the neurons are of a different chemical composition than those found in glia cells. Since only paraffin embedded sections or lipid extracted frozen sections reacted with the lectins, we suggest that the stored glucoconjugates are glycoproteins or oligosaccharides rather than glycolipids.     

Lectin histochemistry of brains from a murine mutant carrying a storage disorder

A strain of Balb/c mice with neurovisceral storage disorder exhibits metabolic and phenotypic manifestations similar to those found in Niemann-Pick type C and D patients. The storage material in the brain reacted positively with periodate-Schiff reagent. To identify the chemical nature of the storage material we applied lectin histochemistry on paraffin-embedded and frozen sections, using biotinylated lectins and avidin-biotin-peroxidase complex. Major abnormalities were noted in the neurons and glia cells. Swollen neurons were stained heavily by Con A and S-WGA, whereas glia cells, mainly astrocytes, which were abundant both in the cerebrum and cerebellum, were positive to RCA-I, GS-I, PNA, S-WGA and WGA. The myelin tracts reacted with PNA, SBA and RCA-I but to a lesser extent in affected animals when compared to normals. Frozen brain sections stained positively only after extraction with chloroform methanol prior to the lectin treatment and revealed a lectin binding pattern similar to that of the paraffin-embedded preparations. The data presented here show that the stored glucoconjugates in the neurons are of a different chemical composition than those found in glia cells. Since only paraffin embedded sections or lipid extracted frozen sections reacted with the lectins, we suggest that the stored glucoconjugates are glycoproteins or oligosaccharides rather than glycolipids.     

Distribution of glycoconjugates in ion transport cells of gerbil inner ear.

Ion transport cells in gerbil inner ear were differentiated histochemically by staining glycoconjugates (GCs) with a battery of horseradish peroxidase-conjugated lectins. Strong staining with PSA and LCA showed a high content of N-linked oligosaccharides in transport cell GCs. Reactivity with PHA-L and PHA-E identified GC with triantennary and with bisected biantennary N-linked oligosaccharides, respectively, in these cells. High affinity for DSA and PWM demonstrated abundant N-acetyl lactosamine in N-linked side chains. Ion transporting epithelial cells reacting with lectins specific for N-linked oligosaccharides included strial marginal cells and outer sulcus cells of the cochlea and dark cells, transitional cells, and planum semilunatum cells of the vestibular system. In general, all of the inner ear transport epithelial cells revealed a similar lectin binding profile, with the one exception that SBA reacted strongly with ion transporting cells in the vestibular system but only weakly with those in the cochlea. Fibrocytes specialized for ion transport located in distinct areas in the suprastrial and inferior regions of the spiral ligament also stained with lectins that demonstrate N-glycosylation. However, transport fibrocytes differed from transport epithelial cells in two ways. First, they reacted e with HPA, DBA, VVA, and SJA specific for O-linkages and second, they failed to react with UEA I. The staining pattern for N-glycosylated GC resembled that for Na+, K(+)-ATPase in inner ear, suggesting a relationship between these constituents.     

Lectin histochemistry study in the human vas deferens

The oligosaccharide sequences of glycoconjugates in the normal human vas deferens and the nature of the saccharide linkage were studied by lectin histochemistry. The cytoplasm of all epithelial cell types (principal cells, basal cells, and mitochondria-rich cells) and luminal contents reacted positively with WGA, MAA, PNA, DSA, LTA, UEA-I, AAA, and ConA. The reaction was more intense in the stereocilia of principal cells. Cytoplasmic staining was diffuse except for PNA and DSA labeling which was limited to the apical cytoplasm and stereocilia of columnar cells. The cytoplasm of all cell types also reacted diffusely with HPA, although staining was weak and was not observed in the stereocilia. Positive reaction with SBA only was encountered in the stereocilia of principal cells. SNA, LTA, and DBA were unreactive. GNA-labeling showed a granular distribution in the supranuclear cytoplasm of columnar epithelial cells. Reactions with MAA, PNA, DSA, AAA, HPA and SBA disappeared after the -elimination reaction. Reactions with WGA and UEA-I decreased after -elimination or Endo-F digestion. Reactions with ConA and GNA were suppressed by Endo-F digestion. Reactions with PNA, HPA, and SBA increased after desialylation. Of all the lectins that label the luminal contents of the vas deferens, only UEA-I was not found in the luminal contents of seminiferous tubules and epididymis and, thus, this lectin would probably bind to glycoproteins secreted by the vas deferens. The chemical treatments used suggest that this secretion contains fucose residues located in both N- and O-linked oligosaccharides. The other lectins may label secreted proteins, but also structural proteins or proteins reabsorbed from the luminal fluid. The lectin- binding pattern of mitochondria-rich cells in the vas deferens differed from that found in the epididymis.     

Lectin histochemistry of human placenta

Abstract. The human placenta was studied histochemically using 23 fluorescein-isothiocyanate-labeled lectins Distinct patterns of staining, as well as some differences between first-trimester and term placenta, were discerned. Eleven lectins (HPA, VVA, BPA, HAA, SBA, PNA, GSA-I, MPA, RCA-I, RCA-II, and UEA-I) did not react with the trophoblast. Two lectins (LCA and PEA) reacted with the trophoblast of first-trimester placenta but not with the trophoblast of third-trimester placenta. The remaining ten lectins (ConA, Suc.ConA, WGA, GSA-II, LAA, STA, DBA, LBA, PHA-E, and PHA-L) reacted with the trophoblast of both first- and third-trimester placenta, and two of these lectins (ConA and Suc.ConA) reacted preferentially with the syncytiotrophoblast. Five lectins (LAA, STA, DBA, GSA-II, and LBA) reacted with nuclei of the cytotrophoblast. The nuclei of some stromal and syncytiotrophoblastic cells were also reactive. Eighteen lectins reacted with the trophoblastic basement membrane, and all reacted with Hofbauer cells and the stroma of the villi. Latin binding was influenced by the mode of fixation and tissue processing. These data show that some lectins can be used to identify components of the placental villi (e.g., basement, membrane) to characterize differences between the first- and third-trimester trophoblast, and to distinguish the cytotrophoblast from the syncytiotrophoblast.     

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.     

A histochemical study of the distribution of lectin binding sites in the developing oocytes of the lancelet Branchiostoma belcheri

The distribution of carbohydrate moieties in lancelet (Branchiostoma belcheri) oocytes has been studied at different stages of development, using a peroxidase-labeled lectin incubation technique, the PAS-reaction and Alcian Blue staining. Binding sites of 5 lectins, indicating the presence of different sugar moieties (Wheat germ agglutinin (WGA) for N-acetylglucosamine, Concanavalin A (Con A) for glucose/mannose, Helix pomatia agglutinin (HPA) for N-acetyl-D-galactosamine, Ricinus communis agglutinin (RCA-I) for galactose and Ulex europaeus agglutinin (UEA-I) for fucose), were identified and were shown to undergo considerable variation during oocyte development. In the previtellogenic stage, HPA, RCA-I and UEA-I were not identified on the oocyte surface, but WGA and Con A gave strongly positive reactions at this site. In the cytoplasm, 4 lectins (Con A, HPA, RCA-I and UEA-I) gave a weak or moderate reaction, and Con A was also observed in the perinuclear region. In vitellogenic oocytes, these 4 lectins were found to also bind to the nuclear envelope, karyoplasm and nucleolus, and, with the exception of Con A, could also be found in the nuclei of more mature stages. The cytoplasmic yolk granules and Golgi vesicles of the vitellogenic oocyte, were moderately positive for Con A, HPA, RCA-I and UEA-I, but HPA, RCA-I and UEA-I were only weakly bound at the oocyte surface. In mature oocytes, all 5 lectins bound moderately or strongly to yolk granules and cell surface. HPA, RCA-I and UEA-I bound moderately or strongly to various nuclear compartments. Thus, carbohydrate content varied with the development and maturation of the oocytes, and the PAS results were in agreement with the lectin-binding results. Charged carbohydrate residues were observed in the egg envelope and Golgi bodies.These results suggest that the appearence of Con A-, HPA-, RCA-I- and UEA-I-binding glycoconjugates in the nuclei of developing oocytes show a varying pattern indicating different phases of nuclear activity which correlate with different carbohydrate synthetic activities of the oocyte.     

Pregnancy-related changes in the mouse oviduct and uterus revealed by differential binding of fluoresceinated lectins

The binding of 20 fluorescein isothiocyanate (FITC)-labeled lectins to various portions of the pregnant and non-pregnant murine oviduct and uterus was studied by fluorescence microscopy. Five lectins (from Ricinus communis (RCA-I), Maclura pomifera (MPA), Triticum vulgare (wheat germ-WGA), Bauhinia purpurea (BPA), and Ulex europeus (UEA-I] reacted differentially with the epithelium of pregnant as compared with the non-pregnant uterus. The binding of RCA-I, MPA and WGA delineated pregnancy-related changes in the distal oviduct and colliculus tubaris. WGA recognized also pregnancy related changes in the proximal oviduct. The reactivity of the remaining 15 lectins did not distinguish the pregnant and non-pregnant oviduct and uterus, although some of them served to identify specific components of the mouse genital tract. Thus, Soybean lectin (SBA) reacted almost exclusively with the colliculus tubaris. UEA-I alone reacted exclusively with the epithelium of the non-pregnant uterus. RCA-II reacted preferentially with the epithelium of the oviduct and uterus as compared with its weak reactivity with the stroma. Two lectins (from Pisum sativum and Lens culinaris) reacted selectively with stromal cells of the uterus and oviduct. Present data indicate that the differential binding properties of these FITC-labeled lectins can be exploited to identify certain components of the mouse oviduct and uterus and to indicate changes in the cell surface and/or cytoplasm in these structures during pregnancy.     

Lectin Histochemistry on Glycoconjugates of the Epidermis and Dermal Glands of Xenopus laevis (Daudin, 1802)

We performed an investigation at the light microscopical level of the differential distribution of lectin-binding sites among cells of the epidermis and glandular domains of the African clawed frog Xenopus laevis. Using a panel of biotinylated lectins (Con-A. PSA, LCA, UEA-I, DBA, SBA, SJA, RCA-I, BSL-I, WGA, s-WGA, PHA-E and PHA-L) and an avidin–biotin–peroxidase complex (ABC), we have identified specific binding patterns. The results show that expression of saccharide moieties in Xenopus epidermal keratinocytes is related to the stage of cellular differentiation, different cell layers expressing different sugar residues. Moreover, oliogosaccharides with “identical” biochemically defined sugar compositions can be distinguished. The method allowed further characterization of complex glycoconjugates of dermal glands. In view of these results, the ABC technique and the biotinylated lectins employed in the present study are believed to be a reliable method for the precise localization of saccharide residues of glycoconjugates present in ectothermic vertebrates.     

Lectin histochemistry of glycolipid storage diseases on frozen and paraffin-embedded tissue sections

Lectin histochemical studies were performed on frozen and paraffin-embedded brain tissue sections from six cases of galactosylceramide lipidosis (i.e., globoid cell leukodystrophy, or Krabbe's disease) in Twitcher mice and one case of canine infantile GM1-gangliosidosis. The globoid cells in Krabbe's disease stained with Ricinus communis agglutinin-I (RCA-I), peanut agglutinin (PNA), and Bandeirea simplicifolia agglutinin-I (BS-I) in frozen sections. However, paraffin sections and frozen sections pretreated with chloroform-methanol or xylene, from the same animals, stained with Concanavlia ensiformis agglutinin (ConA), wheat germ agglutinin (WGA), and succinylated-WGA (S-WGA), in addition to staining with RCA-I, PNA, and BS-I. The affected neurons of canine infantile GM1-gangliosidosis stained only with RCA-I in frozen sections. In paraffin sections, however, these cells were negative with RCA-I but positive with BS-I, ConA, Dolichos biflorus agglutinin (DBA), soybean agglutinin (SBA) and Ulex europaeus agglutinin (UEA-I) in paraffin sections. These results indicate that in paraffin processing of glycolipid storage disease tissue, some lectin receptors are lost and others are unmasked. The retained receptors can be stained with specific lectins and could serve as markers to characterize and differentiate among the various glycolipid storage diseases.     

Carbohydrate-binding peptides from several anti-H(O) lectins.

Peptide fragments have been obtained from L-fucose-binding anti-H(O) lectins [Lotus tetragonolobus lectin (LTA) and Ulex europeus lectin I (UEA-I)] and di-N-acetylchitobiose-binding anti-H(O) lectins [Ulex europeus lectin II (UEA-II) and Laburnum alpinum lectin I (LAA-I)] by treatment with endoproteinase Asp-N or Lys-C. The peptide fragments were fractionated by affinity chromatography on a column of Fuc-Gel for LTA and UEA-I, and on a column of N-acetyl-D-glucosamine oligomer-Sepharose for UEA-II and LAA-I. The peptides with affinity for these columns were identified by peptide sequencing. All of these retarded peptides were found to be parts of the metal-binding regions of these lectins. It is strongly suggested that these peptides represent the carbohydrate-binding and metal ion-binding sites of legume lectins, respectively.     

Comparative lectin-binding patterns in the epidermis and dermal glands of Bufo bufo (L.) and Xenopus laevis (Daudin)

Bouin-fixed and paraffin-embedded sections from the dorsal skin of Bufo bufo and Xenopus Laevis were subjected to lectin histochemistry. A panel of biotinylated lectins (Con-A, PSA, LCA, UEA-I, DBA, SBA, SJA, RCA-I, BSL-I, WGA, s-WGA, PHA-E and PHA-L) and an avidin-biotin-peroxidase complex (ABC) showed a species-specific compartmentalization of saccharides to certain parts of the epidermis and glandular domains. Some marked histochemical differences between the two examined species adapted to fully aquatic (X. laevis) or semiterrestrial (B. bufo) environments may be relevant of a relationship existing between habitat selection and the glycosaminoglycans content of the skin. In addition the technique used in this paper may be applicable for further studies of the carbohydrate composition in various tissues of lower vertebrates.