The main goal of this proposal is to establish an in vitro model to study the combined effect of human papilloma viruses (HPVs) and tobacco-related carcinogens (i.e. nitroso compounds) in development of oral cancer. This study will be performed in vitro on cultured normal human oral epithelial cells. HPVs have been identified in benign, premalignant and malignant oral epithelial lesions. Numerous epidemiologic studies have implicated tobacco-related carcinogens in human oral cancer. Therefore, our hypothesis is that the combination of HPVs and tobacco-related carcinogens induce oral cancer. We will first examine the effect of each (HPV and tobacco carcinogen) on the cultured epithelial cells and then examine the effect of the combination of the two on these cells. Normal cultured epithelial cells will act as a control. The HPV and tobacco-carcinogen treated and control cells will be grown on plastic and collagen rafts. The latter will create a 3-dimensional, differentiated model which will simulate the architecture of the in vivo stratified squamous epithelium. The cells will be examined morphologically and biochemically. Morphologically, cells will be studied by phase contrast and light microscopy looking for evidence of epithelial dysplasia and/or malignancy. Biochemically, the cells will be tested for the expression of involucrin and filaggrin using monoclonal antibodies. Both of these proteins measure epithelial cell differentiation, a feature lacking in premalignant and malignant epithelial cells. The information obtained from this project will lay the groundwork for studying the process of oral epithelial cell transformation, which will be submitted as an R29 NIH grant in the near future.
