The present invention relates to the field of polypeptide medicine, and relates to polypeptides against hepatitis B virus X Protein and uses thereof. Specifically, the present invention relates to polypeptides comprising D-amino acids, which have a function of inhibiting hepatitis B virus X protein, and inhibit HBx activities, inhibit replication of hepatitis B virus DNA and expression of related antigens (e.g., HBeAg) at molecular level, cellular level and animal level, and further inhibit hepatitis and hepatic cirrhosis caused by hepatitis B virus infection and liver cancer that occurs on the basis of hepatic cirrhosis. The polypeptides can be widely used in the prevention and treatment of liver diseases after hepatitis B infection, including hepatitis, hepatic cirrhosis, and liver cancer.