Accumulating evidence implies that cell fusion is one of the driving forces of cancer invasion and metastasis. However, considerably less is still known about the triggering factors and underlying mechanisms associated with cancer-host cell fusion, particularly in inflammatory tumor microenvironment. In this study, we confirmed that inflammatory factor TNF-α could enhance fusion between squamous cell carcinoma cells 9 (SCC-9) and human umbilical vein endothelial cells (HUVEC). Further study revealed that TNF-α could promote up-regulation of syncytin-1 in SCC-9 and its receptor neutral amino acid transporter type 2 (ASCT-2) in HUVEC. Syncytin-1 acted as an important downstream effector in TNF-α-enhanced cancer-endothelial cell fusion. TNF-α treatment also led to the activation of Wnt/β-catenin signal pathway in SCC-9. The activation of Wnt/β-catenin signal pathway was closely associated with the up-regulation of syncytin-1 in SCC-9 and increased fusion between SCC-9 and HUVEC while blocking of Wnt/β-catenin signal pathway resulted in the corresponding down-regulation of syncytin-1 accompanied by sharp decrease of cancer-endothelial cell fusion. Taking together, our results suggest that Wnt/β-catenin signal pathway activation-dependent up-regulation of syncytin-1 contributes to the pro-inflammatory factor TNF-α-enhanced fusion between oral squamous cell carcinoma cells and endothelial cells.