Lectin histochemistry of kidney tumours and its pathomorphological relevance

Thirty kidney tumours of various histological type were histochemically investigated under the light microscope by means of the ABC-method. We used four biotinylated lectins which are known to bind also to normal renal tubular epithelial cells of different nephron segments. The nuclear grade, the histological growth pattern, the cell type and the histogenesis of the tumours were studied. The lectin of Lotus tetragonolobus bound to nine out of ten renal cell carcinomas with low nuclear grade, but in contrast no binding was seen in eight poorly differentiated ones. Four carcinomas mimicking collecting duct epithelium were also negative after treatment with this lectin, but showed strong staining after treatment with peanut agglutinin, Dolichos biflorus agglutinin and soybean agglutinin. Five oncocytomas showed a high affinity only for Dolichos biflorus agglutinin and only one case was positive with Lotus tetragonolobus agglutinin; three tubulo-papillary adenomas of the renal cortex without any oncocytes were negative with all of the lectins used. The value of lectin histochemistry in tumour pathology and its significance in routine pathological examination of kidney tumours are discussed.     

Lectin histochemical detection of sulfoglycans in the zona pellucida of mammalian antral oocytes

Sulphated esters are important to increase effectiveness of specific biological activities of carbohydrates. Biochemical studies revealed the presence of distinct sulphated glycoproteins in mammal zona pellucida (ZP) that bind proacrosin and thus participate in the sperm-egg fusion processes. In the present study, 6 lectin-horseradish peroxidase conjugates (SBA, PNA, RCA-I, GSA-IB4, GSA-II and DBA) were used in combination with desulphation and sialidase digestion to identify sulphocarbohydrates in the terminal and/or subterminal position of oligosaccharide side chains of glycoproteins in the ZP of bovine, ovine, caprine and porcine antral oocytes. In particular, we identified the following terminal sulphoglycans located in the outer layer of the ZP only: SO4-GalNAc in bovine ZP; SO4-Galbeta1,3GalNAc in bovine and ovine ZP; SO4-Galbeta1,4GlcNAc in bovine, ovine and caprine ZP; SO4-alpha-Gal in bovine, caprine and porcine ZP. Subterminal sulphoglycans linked to sialic acid residues were evenly distributed throughout the entire thickness of the ZP: Neu5Ac-SO4-Galbeta1,3GalNAc in bovine and porcine ZP; Neu5Ac-SO4-Galbeta1,4GlcNAc in caprine ZP; Neu5Ac-SO4-alpha-Gal in porcine ZP; Neu5AcSO4-GlcNAc in bovine ZP. The results demonstrate that the chemical composition of the ZP differs among species determining the species-specificity of gamete interactions.     

Lectin binding characteristics of mouse placental cells

The lectin binding characteristics of mouse placental cells were examined. Wax embedded tissue sections of placentae from d 14 pregnant mice were stained with 26 lectins, with a wide range of sugar specificities. Cell cultures prepared from d 14 mouse placentae and cultured for 24 h were stained with 7 of the lectins to determine if they could be used as markers for the different trophoblast cells in culture. In tissue sections all placental cell populations bound lectin but no lectin bound specifically to any single trophoblast population. All the lectins which bound to layer 1 cytotrophoblast lining the maternal blood spaces of the labyrinthine placenta also bound to the fetal endothelium of the labyrinthine placenta. Binding of lectin appeared strongest on the adluminal membrane of these cell populations suggesting a role for the carbohydrate moieties in nutrient transfer. Few lectins bound to junctional zone trophoblast. Overall, the binding of lectin to cultured cells did not correlate exactly with lectin binding to the cell populations in tissue sections. The value of lectins as markers for placental cells in culture was therefore found to be limited. Our findings indicate that carbohydrate expression by at least some placental cells may vary in culture from that expressed by the cells in vivo with obvious concerns for the validity of functional in vitro studies.     

Lectin histochemistry and ultrastructure of microgranular cells in Cinachyra tarentina (Porifera, Demospongiae)

A histochemical study is described that characterizes microgranular cells of the demosponge Cinachyra tarentina (C. tarentina) with the use of routine staining methods for mucosubstances, lectin histochemistry and electron microscopy. Microgranular cells are rare or absent in other species of sponges, but abundant in this species. Microgranular cells are present in both ectosome and mesohyl, particularly along the canal of the aquiferous system and around spicule holes. Inclusions of microgranular cells and the extracellular matrix were particularly positive for acidic glycoproteins with abundant sulfated ester groups and glycosidic residues containing GalNAc and Galbeta1,3GalNAc. Terminal L-fucose bound to the penultimate GalNAc residues and/or difucosylated oligosaccharides were present as well. Our results suggest that soybean lectin (SBA), peanut lectin (PNA), and winged pea lectin (WPA) are valuable markers for identifying microgranular cells of C. tarentina. Electron microscopy revealed some of the microgranular cells to contain small smooth cytoplasmic vesicles originating from the Golgi complex and few electron-dense granules, others were characterized by numerous secretory granules and vacuoles formed by vesicle fusion and connected with the plasma membrane. Our results suggest that microgranular cells in C. tarentina contribute to the synthesis of glycoprotein components of the extracellular matrix.     

Lectin histochemistry of the canine anal glands

The distribution and selectivity of complex carbohydrates in the canine anal glands were studied by means of lectin histochemistry, using PO-labeled lectins. The secretory epithelium of the anal glands and the excretory duct system exhibited large amounts of mainly neutral glycoproteins with various terminal sugars (alpha-D-mannose, beta-N-acetyl-D-glucosamine, alpha-N-acetyl-D-galactosamine, alpha-D-galactose, alpha-L-fucose, N-acetyl-neuraminic acid). Distinctly prominent in the secretion were alpha-L-fucose residues. This relatively hydrophobic sugar may in particular modify or control the viscoelastic properties of the anal gland mucus, so that a stable mucous coat of the rather dry faeces can be formed. In addition, it was obvious that the major part of the excretory duct system is also involved in secretion production, and that the essential function of the saccular dilatations of the excretory ducts is to ensure secretion maturation.     

Lectin histochemistry of glycoconjugates in the feline hair follicle and hair

The distribution of glycoconjugate in the feline hair follicle and hair was studied by light and electron microscopic histochemical methods. The hair apparatus was found to contain considerable amounts of complex carbohydrates with different saccharide residues (alpha-D-mannose, beta-D-glucose, alpha-L-fucose, beta-N-acetyl-D-glucosamine). Variations of those were detected in the plasma membrane of the hair follicle cells during the course of their differentiation and keratinization, namely, alph-D-glucose, alpha-L-fucose and beta-N-acetyl-D-glucosamine in the suprabulbar and bulbar regions. The reaction level of sialic acid residues in the plasma membrane decreased in some cell layers during the course of differentiation. The results obtained from the present study indicated that interaction between saccharide residues of neutral carbohydrates and sialyl groups during the anagen phase might contribute to cell keratinization in hair follicles and hairs. It is discussed whether the existence of glycogen in outer root sheath cells might enable these cells to provide other hair apparatus cells with energy when necessary. Moreover, it became obvious from variations in sialyl residue distribution that cell differentiation processes terminate first of all in Huxley's and Henle's layers within the suprabulbar region of the hair follicle, as followed by the hair cortex.     

Glomerulocystic kidney disease in a Belgian Malinois dog: an ultrastructural, immunohistochemical, and lectin-binding study

Renal cysts in the cortex of a juvenile Belgian Malinois dog with acute renal failure were studied by means of light, scanning and transmission electron microscopy, immunohistochemistry for intermediate filaments, and binding for wheat germ agglutinin (WGA), peanut agglutinin (PNA), and Maclura pomifera agglutinin (MPA) lectins to determine the morphological and histochemical features of the epithelial cells of these cysts. The cysts were renal corpuscles with expanded urinary space. Glomerular tufts were small with poorly developed capillary loops and increased mesangial matrix. Continuity with the proximal tubule was evident in some cystic glomeruli. Two cell types lined Bowman's capsule. One was squamous with a central cilium and microvilli. The other had morphological and histochemical features of immature podocytes (parietal podocytes). These cells were round and protruded into the urinary space; they had thick cytoplasmic projections that resembled foot processes of podocytes, microvilli, and filtration slits. The parietal podocytes expressed vimentin and cytokeratins and had affinity for WGA as do normal immature podocytes. These features suggest that the parietal podocytes are derived by metaplasia of the parietal cells. The basement membrane of Bowman's capsule was irregularly thickened and showed multifocal glycosylation changes with lectin histochemistry (WGA, PNA, MPA) in areas adjacent to the parietal podocytes. Histologic and ultrastructural findings in this dog are consistent with glomerulocystic kidney disease. This is the second report of canine glomerulocystic kidney disease. Features are similar to those of the human counterpart, but it is unclear whether genetic defects cause the disease in the dog. The presence of parietal podocytes in all cysts suggests that abnormal differentiation may play an important role in the pathogenesis of this type of polycystic kidney disease.     

Lectin Histochemical Evalution of Neuronal Glycosaminoglycan Content Following Bone Marrow Transplantation in Dogs with MPS I

Abstract

The storage of glycosaminoglycans (GAGs) within cells of the central nervous system of dogs with mucopolysaccharidosis I (MPS I) was studied. Lectins that detect these stored GAGs were applied to formalin fixed tissues from 5 MPS I-affected dogs, 2 of which were previously treated by littermate bone marrow transplantation (BMT), and from 2 unaffected littermate controls.

In all transplant recipients there was reduced reactivity of the central nervous system (CNS) neurons with soybean agglutinin and horse gram agglutinin indicating that BMT results in reduced levels of those GAGs stored excessively in MPS I. Wheat germ agglutinin was found to react less strongly with cells from treated or untreated dogs despite its predicted binding to sugars in GAGs that are known to accumulate in MPS I. These findings, coupled with previously reported total brain GAG content, show that neuronal storage of GAG is reduced following BMT in canine MPS I. (The J Histotechnol 14:183, 1991)     

Mucoid carcinomas of the breast: a study using mucin histochemistry and peanut lectin

The mucin secreted by mucoid carcinomas of the breast has been studied in a series of 20 tumours by histochemical methods and peroxidase labelled peanut lectin. The histochemical techniques showed that the carcinomas could be divided into two groups depending on whether the mucin was composed predominantly of acid or neutral mucopolysaccharides. The specific binding of peanut lectin to β -galactosyl groups, in combination with neuraminidase treatment, enabled further divisions to be made and so demonstrated the heterogenous nature of the mucin secreted by mucoid carcinomas and also the value of using lectins as specific probes in histological methods.     

Lectin and mucin histochemistry as an aid to cancer surveillance in ulcerative colitis

In a preliminary study, we assessed 10 lectins for the identification of dysplasia in colectomy specimens from patients with ulcerative colitis. Peanut agglutinin (PNA) binding was found in all cases of dysplasia. In the main study the relationship between PNA staining, high iron-diamine/alcian blue (HID-AB) histochemistry, and dysplasia was investigated in 115 pre-operative colonoscopic biopsies and the subsequent resection specimens from patients with ulcerative colitis complicated by carcinoma (n = 6) and patients undergoing proctocolectomy for failure of medical management (n = 8). Peanut lectin was of no value in the assessment of pre-malignant changes or cancer risk. However, the HID-AB stain appears to clarify the interpretation of less severe pre-malignant changes and may be usefully applied to the interpretation of colonoscopic biopsies for cancer surveillance in ulcerative colitis.     

Lectin histochemistry for the study of glycoconjugate residues on colonic adenocarcinoma

TheMaackia amurensis leukoagglutinin (MAL),Sambucus nigra L.I (SNA I), andAmaranthus caudatus (amaranthin; ACL) lectins were used for the study of glycoconjugate residues on normal colonic mucosa and adenocarcinoma by the lectin gold technique. MAL labeled normal colonic mucosa but SNA I did not. ACL stained normal mucosa with regional differences. In contrast, all three lectins labeled most adenocarcinomas. The labeling pattern of MAL was variable but that of SNA I and ACL bound the apical surface of carcinoma cells. Some cells were labeled by SNA I but not by MAL, suggesting thatde novo expression of an α2, 6 sialytrans-ferase was accompanied by malignant transformation. Pretreatment of the sections with galactose oxidase-Schiff sequence abolished the ACL labeling, indicating colonic mucosa and carcinoma cells exhibiting a cryptic form of the T antigen.     

Lectin histochemistry of the rat lymph node: visualisation of stroma, blood vessels, sinuses, and macrophages. A contribution to the concept of an immune accessory role of sinus-lining endothelia

The lectin Chelidonium majus agglutinin (CMA) was previously shown to visualise endothelia of all blood vessels and those lining sinuses of red pulp, stromal reticular meshwok (RM) and dendritic cells of lymphatic follicles in white pulp of the spleen in rats. The aim of the present study was the analysis of CMA and some other lectins in labelling RM, vascular structures and macrophages in lymph nodes of rats. It appeared that CMA stained the entire RM, dendritic cells, lining cells of sinuses and all types of blood vessels. Sinus-lining cells of lymph nodes were labelled with CMA and mannose-, GalNac-, and sialic acid-specific lectins. Moreover, lymph node macrophages were labelled above all by mannose specific lectins. The broad lectin-binding pattern of sinuses--not observed in rat spleen- and CMA-reactivity of both sinus-lining and dendritic cells corroborates the hypothesis that lymph node sinus-lining endothelia are precursors or a special type of immune accessory cells.     

Immunohistochemistry and lectin histochemistry in sarcomatoid renal cell carcinoma: a comparison with classical renal cell carcinoma

We investigated the expression of various cell markers in renal cell carcinoma, concentrating particularly on the sarcomatoid variety, using lectin and immunohistochemical techniques. The sarcomatoid variant showed stronger staining in a higher proportion of cases for vimentin and reduced positivity for epithelial membrane antigen, in comparison with classical renal cell carcinoma. All sarcomatoid tumours reacted with at least one cytokeratin, enabling them to be distinguished from true renal sarcomas; this is of diagnostic value when a panel of markers is used. Overall a similar pattern of markers is seen in sarcomatoid and classical renal cell carcinoma using lectin and immunohistochemistry, suggesting that the sarcomatoid variant arises as a metaplastic change rather than having a different histogenesis.     

Lectin histochemistry of gastrointestinal glycoconjugates in the greater horseshoe bat, Rhinolophus ferrumequinum (Schreber, 1774)

Mucins in the gastrointestinal tract of Rhinolophus ferrumequinum were investigated by histochemistry and lectin histochemistry to evaluate morphofunctional variations of different regions and their possible physiological and evolutionary implications. Histochemical methods included periodic acid-Schiff (PAS), Alcian blue (AB) at pH 2.5 and 1.0 and high-iron-diamine AB pH 2.5. Binding of lectins Con A, DBA, WGA, LTA, LFA, PNA and SBA; LFA, PNA and SBA with prior sialidase treatment; and paradoxical Con A were evaluated. The oesophagus lacked glands. The stomach was divided into a short cardias, a wide fundus and a brief pylorus. The surface muciparous cells secreted sulpho- and sialomucins with N-acetylgalactosamine (GalNAc) residues, N-acetyllactosamine and (beta1,4 N-acetylglucosamine)(n) chains. Towards the pylorus, N-acetylgalactosamine residues disappeared and acidity decreased. Cardiac glands, neck cells in the fundic glands, pyloric and duodenal Brunner's glands all shared neutral, stable class-III mucins, mainly with N-acetylgalactosamine sequences. The intestine was divided into a duodenum, a jejuno-ileum and a short rectum. The goblet cells produced sulpho- and sialomucins with sialylated N-acetylgalactosamine sequences, (beta1,4 N-acetylglucosamine)(n) and N-acetyllactosamine, whose sialylation increased towards the rectum. The main features of the mucins are probably associated with the requirements of fast absorption and food passage and in protection against mechanical and pathogenic injuries.     

Safety in the histochemistry laboratory

Histochemistry and histopathology laboratories are extremely dangerous workplaces owing to the abundance of chemical hazards that constitute major safety and health concerns. A brief review is presented of some of the hazards to which laboratory personnel are exposed. These include fixatives, solvents, embedding media, dyes and stains, and histochemical substrates. Laboratory staff should be fully informed of the dangers and potential long-term health hazards and take the necessary precautionary measures. Despite the abundance of chemical hazards in the histochemistry laboratory, the overall number of work accidents and injuries remains low.     

Use of enzyme histochemistry in the diagnosis of pancreatoblastoma

The macroscopic, light microscopic and ultrastructural findings in a case of pancreatoblastoma in a neonate are presented. No evidence for exocrine or endocrine secretion could be found by histochemistry, immunocytochemistry or electron microscopy. However, enzyme histochemical techniques to localize activity of 'glucose-6-phosphatase', acid phosphatase, esterase and esteroprotease supported both a pancreatic origin for the tumour and a biphasic pattern of differentiation.     

Lectin histochemistry reveals SNA as a prognostic carbohydrate-dependent probe for invasive ductal carcinoma of the breast: a clinicopathological and immunohistochemical auxiliary tool

Increased sialylation and β1,6-branched oligosaccharides has been associated with a variety of structural changes in cell surface carbohydrates, most notably in tumorigenesis. Lectins are defined as proteins that preferentially recognize and bind carbohydrate complexes protruding from glycolipids and glycoproteins. This interaction with carbohydrates can be as specific as the interaction between antigen and antibody. Due to this type of interaction lectins have been used as experimental auxiliary tools in histopathological diagnosis of cancer. This study was designed to evaluate the differential expression of sialic acids and β1,6-N-acetylglucosaminyltransferase V (MGAT5) in invasive (IDC) and in situ (DCIS) ductal carcinoma of the breast and its possible application as prognostic biomarkers. A possible transition between pre-malign and malign lesions was evaluated using DCIS samples. Biopsies were analyzed regarding the expression of MUC1, p53, Ki-67, estrogen receptor, progesterone receptor, HER-2 and MGAT5. α2,6-linked sialic acids residues recognized by SNA lectin was overexpressed in 33.3% of IDC samples and it was related with Ki-67 (p=0.042), PR (p=0.029), lymphnodes status (p=0.017) and death (p=0.011). Regarding survival analysis SNA was the only lectin able to correlate with specific-disease survival and disease-free survival (p=0.024 and p=0.041, respectively), besides, it presents itself as an independent variable by Cox Regression analysis (p= 0.004). Comparing IDC and DCIS cases, only SNA showed different staining pattern (p=0.034). The presence of sialic acids on tumor cell surface can be an indicative of poor prognosis and our study provides further evidence that SNA lectin can be used as a prognostic probe in IDC and DCIS patients.     

Phaseolus vulgaris leukoagglutinating lectin-binding reactivity in human diffuse large B-cell lymphoma and its relevance to the patient's clinical outcome: lectin histochemistry and lectin blot analysis

Many reports have suggested a strong correlation between certain lectin-binding patterns and biological behavior in various tumors. To clarify a relationship between lectin-binding reactivity and survival of patients with diffuse large B-cell lymphoma (B-DLCL), 57 cases with B-DLCL were analyzed by lectin histochemistry and lectin blot method with or without treatment of neuraminidase or acidic hydrolytic conditions. B-DLCL cases were grouped into three types based on the data on lectin-binding reactivity under neuraminidase-treated or untreated conditions: (i) Group A (non-reactive type); (ii) Group B (sialylated type); and (iii) Group C (non-sialylated type). Among various lectins, Phaseolus vulgaris-L (L-PHA) binding reactivity showed that the survival of patients with Group A + B or Group B was significantly shorter than that of patients with Group C. Lectin blot analysis revealed failure of L-PHA-binding to 32 kd and 29 kd glycoproteins, which may be attributable to the masking of L-PHA-binding sites by sialylation or the lack of L-PHA-binding sites, leading to the short survival of patients with B-DLCL. L-PHA-binding reactivity may be a useful marker for the evaluation of survival of patients with B-DLCL.     

Morphological and histochemical variations of mucous and oxynticopeptic cells in the stomach of the seps, Chalcides chalcides

Mucous and oxynticopeptic cells in the gastric mucosa of the seps, Chalcides chalcides (Linnaeus, 1758) were examined by standard histochemical staining methods and by lectin histochemistry. The epithelial mucous cells lining the surface of the stomach and the mucous cells of the fundic glands elaborated mainly neutral glycoproteins with β(1,4)GlcNAc oligomers, GalNAc glycosidic residues and Galβ1,3GalNAc terminal sequences. The mucous cells of the fundic glands were stained specifically with the Paradoxical Con A method. The mucosecreting cells of the pyloric glands produced neutral glycoproteins, with β(1,4)GlcNAc oligomers, GalNAc residues and Galβ1,3GalNAc terminal sequences. Terminal L-fucose bound to the penultimate GlcNAc residues, and/or difucosylated oligosaccharides were also present. The pyloric glands did not stain with the Paradoxical Con A procedure. The morphology of the oxynticopeptic cells changes from the oral to the aboral region of the fundic mucosa. In the oral fundic tract the oxynticopeptic cells showed cytoplasm filled with zymogen granules, while in the aboral fundic region these cells contained few zymogen granules and showed cytoplasm full of empty vesicles, typical of the acid secreting cells. A secretion gradient of proteolytic enzymes and hydrochloric acid along the fundic mucosa of the seps can be hypothesised.     

Lectin histochemistry of the lymphoid organs of the chicken

Cellular interactions within the immune system are in part mediated via the carbohydrate-rich coat of the cell membrane, the glycocalyx, of which the terminal carbohydrate residues are of particular functional importance. Thus, these carbohydrate residues from thymus, bursa of Fabricius, spleen and bone marrow of 2- and 30-day-old chickens were investigated by lectin histochemistry. In the thymus, mannose as well as N-acetyl-glucosamine (glcNAc)-specific lectins labelled macrophages, epithelial reticulum cells and lymphocytes within the cortex. In the bursa of Fabricius, the brush border of the lining epithelium, the macrophages and the endothelium were labelled by mannose-specific lectins. The follicle-associated epithelium was labelled by a broad spectrum of lectins. Epithelial cells that separated the cortex from the medulla and large mononuclear cells in the cortex were only being labelled by N-acetyl-galactosamine (galNAc)-specific and glcNAc-specific lectins, respectively. In the spleen, lymphocytes of the peri-ellipsoid lymphocyte sheaths and macrophages of the red pulp were labelled by lectins of nearly all sugar specificities. In general, glycotopes of these organs were more intensively labelled in the 2-day-old chicken than in the 30-day-old chicken, indicating changes in glycotope expression during post-hatching development. Thus, cells of the avian immune system are as rich and diverse in their lectin binding sites as their mammalian counterparts, indicating that similar carbohydrate lectin interactions between cells and matrices take place in birds as well.