Immune-mediated responses were the main causes of liver damage during viral hepatitis, and recently viral RNA mimetic Poly I:C was used to induce a NK cell-dominated acute hepatitis. Interleukin-17A (IL-17A), the cytokine tightly associated with various autoimmune diseases, was known to play protective or pathological roles in LPS and ConA-induced hepatitis. However, its role in NK cell-mediated acute hepatitis remains unknown. Here we demonstrated that Poly I:C treatment triggered IL-17A production from hepatic γδT cells. Neutralizing IL-17A by monoclonal antibodies reduced Poly I:C-induced intrahepatic inflammatory responses and the liver injury through decreased accumulation, activation and cytolytic activity of NK cells in the liver. Furthermore, Poly I:C didn't trigger IL-17A secretion from γδT cells directly, and Kuppfer cells were demonstrated to be the accessory cell that can secrete IL-23. Finally, our findings demonstrated a pathological role of IL-17A and γδT cells in Poly I:C-induced acute hepatitis, which provides novel insights into viral infection-induced hepatitis and may serve as potential target in clinic immunotherapy against these disease.