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Objective: Mumps virus (MuV) infection is highly testis tropism and severely impairs male fertility.Mechanisms behind the MuV-induced innate immune responses in testicular cells have yet to be intensively investigated.This study aimed to investigate the underlying mechanisms of MuV-induced the testicular innate immune responses.Methods: WT and TLR2/mice were used as models.Realtime qRT-PCR was used to determine gene expression at mRNA level.ELISA, Western blotting and immunohistochemistry were used to analyze gene products at protein level.Results: This study showed that MuV induced innate immune responses in Sertoli and Leydig cells by triggering TLR2 and retinoic acid-inducible gene I (RIG-I) signaling.By contrast, MuV did not induce the innate immune responses in male germ cells.Notably, Sertoli cells produced relatively high levels of proinflammatory cytokines and chemokine, including TNF-, IL-6, MCP-1, and IP-10, compared with Leydig cells in response to MuV infection.Whereas, Leydig cells produced relatively high levels of IFN-and IFN-β compared with Sertoli cells.The MuV-induced innate immune responses were significantly reduced in TLR2/or RIG-Ⅰ knockdown cells.Furthermore, the local injection of MuV into the testis triggered the innate immune responses in vivo.Conclusion: The results suggest that MuV mainly induces inflammatory responses in Sertoli cells, and predominantly induces innate antiviral responses in Leydig cells.Data provide novel insights into the mechanisms by which MuV induces the testicular innate immune responses in mice.