Salivary pellicles and oral health

The maintenance of oral health is dependent upon saliva that forms a thin mobile film on soft and hard tissues of the mouth. The salivary film contains an array of proteins that have been demonstrated to perform important functions including lubrication, maintenance of tooth mineralization, protection against soft tissue dehydration and modification of microbial colonization (Nieu Amerongen, AV & Veerman ECI, 2002, Oral. Dis. 8, 12-22). In recent studies by the applicants a method for the objective measurement of mucosal wetness has been validated for use in a clinical setting (Osailan et al. submitted). The protein composition of samples collected in this way has been analysed and the results indicate that mucins are retained on drier surfaces and that the statherin, a surface active salivary protein (Proctor GB, et al, 2005, Biochemical Journal, 389, 111-116) is adsorbed to oral epithelial cells in normal subjects (Pramanik et al., submitted). Statherin was previously shown to act as a boundary lubricant on the enamel surface and is a prominent component of the acquired enamel pellicle (Douglas et al., 1991, Biochem. Biophys. Res. Comm. 180, 91-97). These observations have important implications for the function of saliva as a lubricant and barrier and the alterations that occur in chronic oral dryness. Lubrication is dependent upon the rheological properties of saliva which are closely linked with the content of mucins, however, the details of the mechanism remain relatively poorly understood and the importance of an adsorbed layer of statherin and other proteins, known to have a lubricative effect on solid model surfaces in vitro (Berg et al., 2004, Biofouling 20, 65-70; Lindh L, 2002, Swed Dent J Suppl. 152, 1-57), remains unclear in vivo. An understanding of the latter is crucial to the formulation of oral health products that can effectively alleviate oral dryness. The project will: (1) examine the composition of proteins in saliva and from oral mucosal and tooth surfaces in different subjects with xerostomia, salivary hypofunction or normal controls; (2) develop in vitro models of saliva/ mucosal cell interactions and (3) in vitro measurement of lubrication. Salivas (parotid and whole mouth collected under standardized conditions) will be analysed for protein components (by SDS-PAGE, Western Blotting, ELISAs etc.), ionic components (particularly calcium) by ion-selective electrodes, rheological properties (pendant drop rheometer, Neva meter) and buffering capacity (by acid titration in open and closed systems to account for carbon dioxide absorption/ loss). Salivary proteins on mucosal surfaces (anterior tongue, buccal and hard palate) will be examined by sampling with sterile filter paper strips before and after removal of the fluid film of saliva; the latter will indicate which salivary proteins are adsorbed to surfaces. Salivary protein adsorption to harvested buccal epithelial cells (or to monolayers or full thickness multicell layers of oral epithelial cell lines, e.g. TR146) will also be assessed as a model for salivary protein adsorption to the oral epithelium. Differences between subjects in test groups and matched control groups will be determined. Important proteins identified in the above studies (likely to include statherin and mucins) will be further studied by isolating the proteins from saliva and examining more closely the interaction with oral epithelial cells in vitro. Functional assays using dye-penetration techniques or imaging of mucoadhesive agents such as chitosan will assess how well the models mimic the human pellicles. The dynamics of surface binding to model surfaces in vitro will also be assessed along with the lubricity of adsorbed proteins using in vitro models.