Expression Of Endogenous Lectins Research Articles

Colonic carcinogenesis along different genetic routes: glycophenotyping of tumor cases separated by microsatellite instability/stability

Different genetic routes account for colonic carcinogenesis. However, when analyzing colon cancer specimens, separation into different groups based on genetic alterations is commonly not performed. Thus, we here initiate the comparative phenotyping considering microsatellite instability/stability for clinical specimens. The focus is given to glycan epitopes, expression of which is known to be modulated by signal-transducing proteins that act as key regulators of normal colon epithelial growth and differentiation. In addition to six plant lectins used as sensors, the presence of two adhesion/growth-regulatory galectins is studied. Overall, a considerable level of intra- and interindividual heterogeneity is revealed. Alterations in the proportion of stained cells between tumor-adjacent and malignant epithelia concerned plant lectins, which bind substituted N-glycan cores, α2,6-sialylated branch ends, core 1 O-glycans and N-acetylgalactosamine. A tendency for changes was noted between microsatellite-unstable and microsatellite-stable cases for core substitution (bisected N-glycan, presence of β1,6-branching) and status of α2,6-sialylation. Statistical significance was reached for presence of galectin-3, found to be elevated in microsatellite-stable compared to microsatellite-unstable tumors. These results emphasize the potential of distinct signaling pathways to regulate certain aspects of the glycophenotype in vivo and thus delineate a perspective to discern functionally relevant deviations in expression of endogenous lectins and their counter-receptors.

Combined analysis of tumor growth pattern and expression of endogenous lectins as a prognostic tool in primary testicular cancer and its lung metastases.

The aim of this study was to glyco- and immunohistochemically analyze expression of distinct growth/adhesion-related markers of primary testicular carcinomas and their lung metastases in relation to the risk of developing lung metastases and survival of patients, and to correlate immunohistochemical staining profile and syntactic structure analysis in order to delineate new prognostic parameters for this tumor type. Clinical features of 50 patients with primary testicular carcinomas and their corresponding lung metastases were evaluated and compared to those of a control cohort of 25 cases. The set of eight probes including labeled galectins-1 and -3, specific non-cross-reactive antibodies against galectins-1, -3, and -8 as well as anti-Ki-67, anti-bcl-2, and anti-p53 was applied to formalin-fixed, paraffin-embedded tumor sections of both primary and metastatic lesions. Syntactic structure analysis computed staining intensities and structural features of the tumor cells. These parameters were set into relation separately and in combination to clinical data including tumor stages, smoking habits, applied cytostatic therapy, disease-free interval, and survival. The risk of testis cancer patients to develop lung metastases depends in descending order on the tumor cell type (non-seminoma versus seminoma), tumor cell heterogeneity (mixed versus monomorphous cell type), age of patients, and pT stage. The extent of differential expression of galectin-related features between primary and secondary lesions was pronounced. Prognostic correlations for distinct galectin-related features were delineated in combination with data from syntactic structure analysis, for example cluster radius of galectin-3-positive tumor cells and post-surgical and total survival. Lengths of disease-free interval and total survival of patients were also correlated to characteristics obtained by syntactic structure analysis and their combination with galectin data in the first place, then to smoking habits, percentage of proliferating cells in the primary and secondary tumors, and finally to expression of certain galectins and of p53. Patients with non-seminoma testicular cancer should be thoroughly controlled for lung metastases. Regarding marker selection, our study underscores that further investigation of the growth-regulatory network of galectins is clearly warranted.

Are differences in the expression of carbohydrate-binding sites (endogenous lectins) responsible for recurrences of benign tumours of the parotid gland?

The expression of endogenous lectins in pleomorphic adenomas and papillary cystadenoma lymphomatosum (Warthin tumour) as well as in the tissue of the normal parotid gland was investigated with the aim to determine the capacity of the different cells to specifically bind carbohydrate ligand structures. The loss of sugar-binding sites of pleomorphic adenomas - lactose (diazonium derivative; diaz.), beta-N-acetylgalactosamine (diaz.), sialic acid [(2,3-epoxypropane)-4-oxybutyric acid linked; epichl.], heparin, mannose-6-phosphate (diaz.), maltose (diaz.), xylose (diaz.), melibiose (diaz.), N-acetylgalactosamine (epichl.) - was detected in the investigated tumours, leading to the hypothesis that altered carbohydrate-protein interactions have been involved in the adhesion of lost pleomorphic adenoma cells after damaging the tumour capsule. The papillary cystadenoma lymphomatosum in its cytoplasmic areas expressed significantly fewer sugar-binding sites for alpha-N-acetylgalactosamine (diaz.) and melibiose (diaz.).

Oncogenes and expression of endogenous lectins and glycoconjugates

It is now well established that malignant transformation of eucaryotic cells is concomitant with typical alterations of glycosylation and the expression pattern of endogenous lectins. In parallel, oncogene transfection studies revealed a correlation between the expression of some of these genes, the transformed state and perhaps metastasis. These observations lead to the idea that oncogenes may control the expression of enzymes involved in the biosynthetic pathway of cell membrane glycoconjugates and the expression of endogenous lectins. Indeed, several contributions have shown that cells upon transfection with activated oncogenes of the ras family become invasive and/or metastatic and have their membrane glycoproteins modified. Information on the molecular mechanism of this postulated oncogene regulation is still lacking. Because of the diversity of the functions of oncogene-encoded proteins, further experiments dealing with other activated oncogenes may help in deciphering the regulation of expression of glycoconjugates and endogenous lectins together with their functions.

Oncogenes and expression of endogenous lectins and glycoconjugates

Oncogenes and expression of endogenous lectins and glycoconjugates

Quantitative microscopy of mouse colon 26 cells growing in different metastatic sites

Quantitative microdensitometry and computerised interactive image analysis were used to compare the expression of endogenous lectins by cells of mouse colon 26 carcinomas, growing either as primary tumours or metastases, in five different anatomic sites (caecum, liver, lung, spleen, s.c.). Endogenous lectins were visualised in tissue sections using the ABC peroxidase technique with a panel of 17 biotinylated neoglycoproteins representing a variety of carbohydrates found in glycoproteins, glycolipids and proteoglycans. Clear-cut site-associated differences in endogenous lectin expression were detected in cancer cells growing in all five sites. The patterns of these changes were complex and shifts in expression of different lectins were independently variable in both direction and amount. In addition to site-associated variations, differences in lectin expression were also detected in the liver and lungs, between cells in spontaneous metastases and cells in colonies generated by direct injection of cancer cells into the bloodstream. The results demonstrate quantitative, as distinct from qualitative, differences developing in cancer cell populations after delivery of cells to different target organs. The differences between liver and lung metastases are in accord with analogous site-associated differences in metastatic patterns produced by colon carcinoma cells in mice and in humans.

Quantitation of endogenous lectin expression in 3LL tumors, growing subcutaneously and in the kidneys of mice

By means of microdensitometry and image analysis, quantitative measurements were compared of endogenous lectin expression in tissue sections from 3LL carcinomas growing in the subcutis and kidneys of mice following the direct injection of cancer cells. The lectins were visualized by means of a modification of the ABC-peroxidase technique. In comparison with the subjective, visual inspection of similar materials reported previously, these quantitative techniques not only confirmed, but also provided additional information on site-associated changes in 3LL cell populations. Previously, 2 classes of cancer cells were recognized within the tumors on the basis of staining intensity, compared with 3 classes in the present study and, in addition, the relative areas occupied by these 3 densitometric classes were determined by image-analysis. Both "low" and "high" integrated intensities were significantly greater in the kidney tumors in 11 of 24 cases, and in 6 of 24 cases of s.c. tumors. The areas occupied by "high" and "low" intensity cells were significantly higher in 14 of 24 kidney tumors, and in 2 of 24 s.c. tumors. The results indicate often marked differences in endogenous lectin expression, between 3LL cancer cells from a common source, after direct delivery and growth in 2 different anatomic sites. These results indicate the necessity of taking into account analogous changes, when comparing these and other properties of cancer cell populations in metastases with those in primary tumors, with respect to both the generation of site-associated differences and their possible site-associated abrogation.

Mouse zygotes express endogenous lectins

Cell interactions during mouse development have been shown to involve carbohydrate-containing macromolecules (glycoconjugates). We have therefore used a series of fluorescein-labelled synthetic glycoproteins to determine if mouse oocytes and zygotes also express sugar binding molecules (endogenous lectins) which might participate in such interactions. Unfertilized secondary oocytes did not express endogenous lectins at 4 degrees C but a low level of expression of fucose, mannose, and galactose-binding activity could be detected at 37 degrees C. In contrast, the zygote clearly expressed three classes of endogenous lectins, with preferential binding for i) fucose or mannose, ii) glucose or galactose, and iii) lactose. The expression of these lectins was much reduced at 4 degrees C and maximal binding at 37 degrees C was achieved only after 2 h incubation. We therefore conclude that a low level of endogenous lectin expression in the mouse oocyte is greatly enhanced after fertilisation and that, at both stages, expression, or the detection of expression, is markedly temperature dependent.

Spatial differences of endogenous lectin expression within the cellular organization of the human heart: A glycohistochemical, immunohistochemical, and glycobiochemical study

Protein-carbohydrate recognition may be involved in an array of molecular interactions on the cellular and subcellular levels. To gain insight into the role of proteins in this type of interaction, surgically removed specimens of human endomyocardial tissue were processed for histochemical and biochemical analysis. The inherent capacity of these sections to bind individual sugar moieties, which are constituents of the carbohydrate part of cellular glycoconjugates, was assessed using a panel of biotinylated neoglycoproteins according to a standardized procedure. Together with appropriate controls, it primarily allowed localization of endogenous lectins. Differences in lectin expression were observed between layers of endocardial tissue, myocardial cell constituents, connective-tissue elements, and vascular structures. The endocardium proved to be positive with beta-galactoside-bearing probes; with neoglycoproteins carrying beta-xylosides, alpha-fucosides, and galactose-6-phosphate moieties; and with probes containing a carboxyl group within the carbohydrate structure, namely sialic acid and glucuronic acid. In contrast, only fucose-and maltose-specific receptors were apparent in the elastic layers of the endocardium. Aside from ascertaining the specificity of the protein-carbohydrate interaction by controls, i.e., lack of binding of the probe in the presence of the unlabelled neoglycoprotein and lack of binding of the labelled sugar-free carrier protein, respective sugar receptors were isolated from heart extracts by using histochemically effective carbohydrates as immobilized affinity ligand. Moreover, affinity chromatography using immobilized lactose as affinity ligand as well as the use of polyclonal antibodies against the predominant beta-galactoside-specific lectin of heart demonstrated that the lactose-specific neoglycoprotein binding was due to this lectin. Remarkably, the labelled endogenous lectin, preferred to plant lectins for detecting ligands of the endogenous lectin, localized ligands in tissue parts where the lectin itself was detected glycohistochemically as well as immunohistologically. This demonstration of receptor-ligand presence in the same system is a further step toward functional assignment of the recorded protein-carbohydrate interaction. Overall, the observed patterns of lectin expression may serve as a guideline to elucidate the precise physiological relevance of lectins and to analyze pathological conditions comparatively.

Site-associated expression of endogenous tumor lectins.

The expression of endogenous lectins was compared in mouse Lewis lung carcinomas growing in the kidney or subcutaneously. A panel of carbohydrates coupled to a biotinylated carrier molecule (bovine serum albumin), biotinylated desialylated glycoproteins and sulfated polysaccharides were used in histochemical assays to detect the presence and distribution of carbohydrate receptors. Heterogeneous staining patterns were observed with most carbohydrates in sections of tumor tissue from both anatomic sites, and staining intensities also varied within each section. At least 2 populations of cells were identified at each site, of which one had receptors for all carbohydrates, while the other had no receptors for melibiose, sialic acid, or alpha-glucosides (maltose and glucose). There were quantitative differences in expression of endogenous lectins by tumors growing s.c. or in the kidney; 3LL cells growing in the kidney bound 6 out of 10 carbohydrates to a greater extent than 3LL cells in s.c. tumors. Conversely, 3LL cells in s.c. tumors bound heparin and asialofetuin to greater extents than cancer cells in kidney tumors. Biochemical analyses of detergent extracts of tissues subjected to affinity chromatography and subsequent SDS-PAGE revealed quantitative and also qualitative differences in lectins between tumors growing in the 2 anatomic sites.

Expression of endogenous lectins in human small-cell carcinoma and undifferentiated carcinoma of the lung

Endogenous carbohydrate-binding proteins (lectins) were detected in specimens of tumor tissue (undifferentiated carcinoma and xenografted small-cell carcinoma) from human lung. Fractionation of salt and detergent extracts on different sets of Sepharose columns covalently derivatized with lactose, asialofetuin, melibiose, mannan, and fucose, successive elution with a chelating agent and a specific sugar, and analysis of the eluates by gel electrophoresis, resulted in the characterization of the profiles of endogenous carbohydrate-binding proteins. All preparations were devoid of enzymatic activity. Comparison between the patterns of the two types of lung carcinoma showed significant qualitative differences, e.g. the presence of fucose-binding proteins of apparent molecular weights 60,000 and 80,000 in the undifferentiated carcinoma, and the presence of β-galactoside-binding proteins of apparent molecular weights 18,000 and 22,000 in the small-cell lung carcinoma. These proteins were not detectable in normal lung tissue. Such differences, documented for the first time for human lung tumors, are of potential importance as a step towards a lectin-based refinement of lung-cancer diagnosis and therapy.