Background & Objective: Multiphoton Autofluorescence Microscopy (MPAM) and Second Harmonic Generation Microscopy (SHGM) are a promising imaging combination for revealing indicators of neoplasia and have the ability to represent the microstructural organization of the mucosa in a manner which may be compared to histology. However they provide limited field of view, which is not practical for large area surveillance. On the other hand wide-field fluorescence could provide large area surveillance of areas that may be suspicious for neoplastic features, such as through metabolic indication, but cannot provide subsurface detail. The objective of this study was to investigate the pairing of the microscopic and widefield approaches as a potential combination for identifying high risk precancers and cancer using an animal model for OSCC.
Methods: A chemically induced Golden Syrian Hamster model of precancer and oral cancer was used for these studies. A fluorescently labelled deoxy-glucose analog, 2-?deoxy-?2-?[(7-?nitro-?2,?1,?3-?benzoxadiazol-?4-?yl)amino]-?D-?glucose (2-NBDG) was used to highlight areas of high glucose uptake hypothesized to correspond to (pre)neoplastic sites due to the Warburg effect as well as additional metabolically active areas. Follow-up MPAM-SHGM was conducted on regions of interests (ROIs) to assess whether microscopy would reveal microscopic features associated with neoplasia to confirm or exclude 2-NBDG based findings. Biopsies were obtained of imaged sites and pathological grading assigned.
Results: 2-NBDG fluorescence was shown to significantly increase with conditions of dysplasia and OSCC compared to normal, however an increase was also observed for the case of inflammation. MPAM-SHGM was useful for revealing cytologic and layer based architectural abnormalities.
Conclusions: Our results show the feasibility of this combinatorial approach of large area assessment and subsurface microscopic evaluation of neoplastic oral mucosa. This finding will allow us to develop novel diagnostic approaches for oral precancers by involving a combined analysis of tissue metabolism and microstructure.