heparitin-sulfate and Laryngeal-Neoplasms

A sensitive and accurate quantitative assay for the measurement of minor amounts of chondroitin/dermatan sulfate and heparan sulfate that does not require specific apparatus or reagents is described. The assay involves labeling of chondroitin sulfate A following reaction of carboxyl groups with biotin hydrazide in the presence of carbodiimide. ELISA plate wells were coated with glutaraldehyde and then spermine was coupled to it via a Schiff's base bond. In such activated wells, the biotinylated molecules were readily bound and detected after the interaction with avidin-peroxidase conjugates and the subsequent enzymic assay. Chondroitin/dermatan sulfate and heparan sulfate competed this interaction in a linear manner. Disaccharides derived from chondroitin sulfate A did not act as competitors, while heparan sulfate disaccharides showed significant competition. From the competition, before and after digestion with either chondroitinase ABC or heparitinases, the amounts of chondroitin sulfate and heparan sulfate in a sample could be calculated. The assay was applied for the determination of sulfated glycosaminoglycans in normal and cancerous human laryngeal cartilage samples. By using this procedure, the accurate determination, especially, of heparan sulfate in a mixture of glycosaminoglycans was achieved, which otherwise would require the use of very expensive technology.

Topics: Binding, Competitive; Cartilage; Enzyme-Linked Immunosorbent Assay; Glycosaminoglycans; Heparitin Sulfate; Humans; Laryngeal Neoplasms

Epithelial structures are separated from the stroma by a basement membrane (BM) which serves as a barrier for the epithelial cells. Invasive tumour growth as in laryngeal carcinoma, involves the degradation of this barrier. In our present immunohistochemical analysis, we evaluated both the quantitative and the qualitative changes in the BM composition of 50 laryngeal carcinomas. This analysis comprised the localisation of the BM components collagen IV, laminin, and fibronectin. Furthermore, we applied antibodies against BM components that have not, or only very rarely, been analysed in malignant squamous cell neoplasms--particularly collagen VII and heparan sulfate proteoglycan (HSPG). In this study we provide considerable evidence that varying amounts of preserved BM material can be found in laryngeal carcinoma of different grades of tumour differentiation. Especially by comparison of the different staining patterns for collagen IV and collagen VII, we recognised far more gaps in the staining of the BM by collagen VII than by collagen IV. This fact is underlined by the observation that even in G1 carcinomas collagen VII showed in almost 40% of the cases a complete loss of BM staining. In general, we found a correlation between the amount of preserved BM deposition and the grade of tumour differentiation. This fact may underline the significance of the immunohistochemically detectable amount of BM components as a prognostically relevant parameter.

Topics: Adult; Aged; Basement Membrane; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Differentiation; Cell Division; Cell Transformation, Neoplastic; Collagen; Female; Fibronectins; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Immunoenzyme Techniques; Laminin; Laryngeal Neoplasms; Male; Middle Aged; Neoplasm Invasiveness; Proteoglycans

The basement lamina of the epithelium of the true vocal cords of 34 inpatients suffering from chronic laryngitis, Reinke's oedema and squamous cell carcinoma have been investigated with the electron and immunofluorescence microscope. In chronic laryngitis, the lamina fibroreticularis is thickened (due to collagen type VII), corresponding to the clinical finding. In this layer mobile cells of the connective tissue can be found. In cases of Reinke's oedema it is the lamina densa which might be thickened. In this disease, also lamina densa-like material can protrude into the lamina fibroreticularis, and the number of anchoring filaments is increased. In squamous cell carcinoma we found the basement lamina irregularly arranged and folded. The lamina densa was always interrupted by numerous small gaps and in some areas the basement membrane could not be identified over a long distance. Lamina densa-like material was also found between the tumour cells within the epithelium. With the immunofluorescence microscope this material was proven as laminin, collagen type IV and heparan sulfate proteoglycan. Our investigation shows that malignant as well as benign lesions of the true vocal cords are characterised by distinct fine structural findings even concerning the basement lamina.

Topics: Adult; Aged; Basement Membrane; Biomarkers, Tumor; Carcinoma, Squamous Cell; Collagen; Epithelium; Female; Fibronectins; Fluorescent Antibody Technique; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Hoarseness; Humans; Laryngeal Edema; Laryngeal Neoplasms; Laryngitis; Larynx; Male; Microscopy, Electron; Middle Aged; Neoplasm Invasiveness; Proteoglycans; Vocal Cords