Squamous cell carcinoma (SCC) is a malignant tumor that shows morphologic and phenotypic similarities to normally differentiated squamous epithelium. Thus, it may be an ideal model for seeking a marker of tumor-initiating cells (TICs) based on their morphology. Using the human SCC cell line A431, we found that, as a paradigm of cancer stem cells: (1) podoplanin(+) cells generate both podoplanin(+) and podoplanin cells; (2) podoplanin(-) cells rarely generate podoplanin(+) cells: (3) podoplanin(+) cells have higher colony formation efficiency and tumorigenicity than podoplanin- cells; (4) localization and morphology of podoplanin(+) cells in a xenografted tumor derived from podoplanin(+) cells are similar with those in human oral SCC tissue or normal epithelium. Furthermore, podoplanin(+) A431 cells share sonic hedgehog and CD44 expression with stem cells in normal squamous epithelium. Hence, we concluded that podoplanin is a novel marker to enrich TICs with stem-cell-like properties from SCC cell line A431. (C) 2008 Elsevier Inc. All rights reserved.