We conducted three-dimensional (3D) reconstruction of oral tongue squamous cell carcinoma (OTSCC) using serial histological sections to visualize the architecture of invasive tumors. Fourteen OTSCC cases were collected from archival paraffin-embedded specimens. Based on a pathodiagnostic survey of whole cancer lesions, a core tissue specimen (3 mm in diameter) was dissected out from the deep invasion front using a paraffin tissue microarray. Serial sections (4 μm thick) were double immunostained with pan-cytokeratin and Ki67 antibodies and digitized images were acquired using virtual microscopy. For 3D reconstruction, image registration and RGB color segmentation were automated using ImageJ software to avoid operator-dependent subjective errors. Based on the 3D tumor architecture, we classified the mode of invasion into four types: pushing and bulky architecture; trabecular architecture; diffuse spreading; and special forms. Direct visualization and quantitative assessment of the parenchymal-stromal border provide a new dimension in our understanding of OTSCC architecture. These 3D morphometric analyses also ascertained that cell invasion (individually and collectively) occurs at the deep invasive front of the OTSCC. These results demonstrate the advantages of histology-based 3D reconstruction for evaluating tumor architecture and its potential for a wide range of applications.
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