Differential lectin binding to cellular membranes in the epidermis of the newborn rat.

Abstract

Three fluorescein-labeled lectins have been shown to exhibit specificity for the surface of cells in different layers of the epidermis in the newborn rat. An isolectin from seeds of Bandeiraea simplicifolia with specificity for a-D-galactosyl end groups labeled the basal and lower spinous cells; a lectin from Ulex europaeus exhibiting specificity for a-L-fucosyl units outlines the surface of spinous cells, and a second lectin from B. simplicifolia, with specificity for N-acetyl-Dglucosamine, labels the cornified cells. Appropriate blocking experiments have confirmed the specific nature of the binding.The investigation of the differentiative process in mammalian epidermis would be assisted by the availability of a technique that would specifically identify different strata. Lectins, which have an affinity for specific sugars (cf. refs. 1 and 2), have been used to probe the nature of the complex carbohydrates associated with cell surfaces (cf. ref.3) and to investigate the role of the cell surface in cellular functions such as growth (4, 5), adhesion (6), pinocytosis (7), antigenicity (8), fertilization (9), and differentiation (10).It seemed possible that specific lectins could be found that would associate with the cell surface during various stages of epidermal differentiation if changes in surface carbohydrates, occurred during this process. Fluorescein-labeled concanavalin A attaches to the surfaces of cells in all layers of the human epidermis (11) and oral mucosa (12) except the stratum corneum. The pattern of binding for concanavalin A and pemphigus antibody are similar (13); however, there are differences in the binding of lectins to the cell surface of carcinomas and wounds in the human oral epithelium (14). In the normal oral mucosa (15), human epidermis (16), and gastrointestinal mucosa of the rat (17, 18), keratinocytes in different differentiative stages exhibit specificity for the binding of particular lectins to their surfaces. The loss of the nucleus from differentiating erythrocytes of the rat results in a change in the labeling of the surface by concanavalin A (10).Three lectins have now been identified that do exhibit specificity for the surface of cells in different layers of the epidermis in the newborn rat. One lectin, with specificity for a-D-galactosyl groups, binds to basal and lower spinous cells; another, specific for a-L-fucosyl units, outlines spinous cells, and the third, specific for N-acetyl-D-glucosamine, labels cornified cells. FITC-Ulex I was diluted to 125-150 g/ml. As controls, Lfucose (Sigma) or N-acetyl-D-glucosamine (Aldrich) at a concentration of 0.5 M in phosphate-buffered saline was preincubated with the lectin. Dorsal skin from newborn rats (CFN strain) was stretched over the surface of a watch glass and frozen to -30'C. Small strips (3 mm X 5 mm) were subsequently cut with a scalpel and set into Tissue-Tek II O.C.T. embedding medium (Miles Laboratories, Naperville, IL), exposing a cross section of skin. Sections were cut 4-6 Mm thick in a cryostat and placed o...

Loss of epithelial cell surface carbohydrates during experimental oral carcinogenesis in the rat

“…Lectin specificity was confirmed as described elsewhere and included pre-incubation of lectins with the appropriate competing oligosaccharide and degradation of the tissue sections with specific glycosidases prior to lectin application (Brabec et al, 1980;Prime et al, 1985b). The purification of FITC-GS-I-B4 and FITC-UEA-I was carried out prepurchase by Sigma (USA) using purified affinity gel electrophoresis.…”

“…Recently, we described the topographical binding of the lectins Griffonia simplicifolia (GS-I-B4; specific for a-Dgalactosyl groups) and Ulex europeus (UEA-I; specific for a-L-fucosyl groups) in rat oral mucosa and demonstrated specificity of the labelling to basal/parabasal and spinous epithelial cells, respectively (Prime et al, 1986b). The lectins GS-I-B4 and UEA-I, therefore, can be used as markers of epithelial differentiation (Brabec et al, 1980) and this makes them ideal probes for the study of cell surface carbohydrate changes in carcinogenesis.Rodent tumours, comparable with human oral carcinomas (Prime et al, 1986a) can be induced by 4-nitroquinoline-Noxide (4NQO) (Wallenius & Lekholm, 1973). Since the tumours develop in all animals after approximately 9 months, cell surface changes from normality through premalignancy to overt malignancy are likely to be present.…”

“…Recently, we described the topographical binding of the lectins Griffonia simplicifolia (GS-I-B4; specific for a-Dgalactosyl groups) and Ulex europeus (UEA-I; specific for a-L-fucosyl groups) in rat oral mucosa and demonstrated specificity of the labelling to basal/parabasal and spinous epithelial cells, respectively (Prime et al, 1986b). The lectins GS-I-B4 and UEA-I, therefore, can be used as markers of epithelial differentiation (Brabec et al, 1980) and this makes them ideal probes for the study of cell surface carbohydrate changes in carcinogenesis.…”

Lectin-binding sites in basal cell epitheliomas and carcinomas.

“…Lectins isolated from plant and animal sources interact specifically with corresponding sugar residues in polysaccharides of tissue sections (6,24) . Histochemical lectin-binding in keratinocytes was mainly at the cellular surface and in intercellular materials in normal stratified squamous epithelia, pemphigus, and certain pathologic conditions (1,4,5) . Preceding papers from our laboratory have noted that the distribution of lectin-binding sites in normal epithelial structures might be indicative of the expression of stratification and differentiation of keratinocytes, and this expression of lectin-binding patterns in squamous epithelia was much more conspicuous in the keratinized epithelia than in the non-keratinized ones (11) .…”

Cationic Colloidal Gold Staining of Acidic Glycoconjugates in Mouse Paneth Cells.

“…Recent interest in acidic glycoconjugates stems from the significance of their roles in cell recognition and differentiation (ROTH et al, 1971;GAHMBERG and HAKOMORI, 1973;EDELMAN, 1976;SKUTELSKY and FARQUHAR, 1976;BRABEC et al, 1980). Several postembedding staining methods, such as cationized ferritin (CF: DANON et al, 1972;THIERY and OVTRACHT, 1979), cationic colloidal gold (CCG: SKUTELSKY and ROTH,1986;KASHIO et al, 1992KASHIO et al, , 1993 and cationic colloidal iron (CCI: OHTSUKA et al, 1993) methods, have been developed to enable the detection of anionic sites in cells and tissues using electron microscopy.…”