Bufalin, as a Chinese traditional anti-tumor agent, has been studied about inhibiting proliferation and promoting apoptosis of liver cancer, however, there are few reports on immune modulating function. We used the human liver cancer cell lines along with 91 pathologically-verified postoperative hepatocellular carcinoma (HCC) specimens to assess immune modulating function of bufalin. We found that bufalin directly balances stimulatory and inhibitory receptors on the surface of NK cells and indirectly activates natural killer (NK) cells by inhibiting MICA shedding, which prevented immune escape and indirectly enhanced NKG2D-dependent immune surveillance. This study showed that bufalin can directly or indirectly regulate the immune response, which provides a new theoretical basis for the clinical application of "Huachansu injection".Keywords: ADAM9; Bufalin; Hepatocellular carcinoma; MICA; Natural killer cells.