Human cytomegalovirus (HCMV) is associated with epithelial-mesenchymal transition (EMT) in glioma cells; however, its underlying action mechanism remain ambiguous. In this study, we investigated the effects of receptor-interacting protein 2 (RIP2) and nuclear factor (NF)-κB on EMT in HCMV-infected glioma LN-18 cells. Wound healing and invasion assays were used to evaluate the migration and invasion of cells. Western blotting and immunofluorescence microscopy were used to determine the protein expression levels. We found that HCMV induced enhanced migration and invasion of LN-18 cells, activation of the RIP2/NF-κB signaling pathway, downregulation of epithelial cell marker (E-cadherin) expression, and upregulation of mesenchymal cell marker (N-cadherin and vimentin) expression. Moreover, inhibition of RIP2 or NF-κB inhibited the induction of HCMV in LN-18 cells. Therefore, HCMV induces EMT in glioma cells by promoting the activation of NF-κB signaling pathway via the upregulation of RIP2 expression.