In spite of the fact that tracheobronchial mucus glycoproteins (TBMGP's) play a critical role in protecting the respiratory tract from inhaled particulate material and have been implicated in the pathophysiology of several fairly common diseases such as chronic bronchitis, asthma, and cystic fibrosis, our understanding of their chemical structure is quite rudimentary. Reports of disease related changes in the carbohydrate and sulfate composition of TBMGP's, particularly to cystic fibrosis, imply that carbohydrate structures are altered. What these changes are, how they affect the physical properties of TBMGP's, and the biological processes responsible for the changes are all unknown. The short range goal of this proposal is to determine exactly how much difference there really is between TBMGP's from normal compared to those from persons with cystic fibrosis. Glycoproteins will be obtained primarily by tracheal explaint tissue culture techniques. Reduced oligosaccharides will be released by Beta-elimination and divided into neutral, sialylated, and sulfated fractions. Each fraction will then be analyzed by one or more high pressure liquid chromatographic (HPLC) methods to give an oligosaccharide pattern. Differences will be established by comparing the pattern for qualitative and quantitative changes. Preliminary results with human TBMGP's and hog gastric mucin indicate that HPLC can resolve at least 30-40 different oligosaccharides. For scientific credibility the pattern analysis must be supplemented with compositional and structural analysis of specific oligosaccharides. Compositions will be determined by HPLC of benzoylated methyl glycosides as described in the Preliminary Results section. Structural analysis will use mass spectoscopy, NMR, and the classical techniques of carbohydrate chemistry, modified to allow use of benzoylation and HPLC for detecting reaction products. The primary question to be answered is whether disease related changes in TBMGP's reflect the disease state itself or some secondary effect of hypersecretion such as, for example, and altered ratio of TBMGP synthesis in goblet cells compared to submucosal glands.
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