The aim of this study is to investigate the mechanism of hedgehog signaling pathway on CD166 positive oral squamous cell carcinoma (OSCC).OSCC cell lines, HN13 and HN6, were used in this study.Cell proliferation results showed HH increased CD166 level and GANT61 inhibited CD166 Level in a time dependent manner.Transiently transfections with Plasmids of CD166, GLI1, SHH, mutant GLI2 were used.The results showed CD166 expressions were increased after transfection of CD166 plasmid, however the expression levels of GLI1, GLI2 and SHH proteins were not changed.Then we used siRNA methods to knock down the genes of CD166.The results showed HH treatment prolonged the half-time of CD166 protein.We conclude that Hedgehog signaling pathway up-regulates and accumulates CD166 expression in OSCC by increasing protein stability and prolongs half-time of CD166.The study will help to explain the role of the HH/GLI axis in CD166 positive OSCC cells.