摘要Background and aims:Berberine is one of the most promising clinically tested natural alkaloids,and immunotherapy using natural killer(NK)cells is a potentially effective treatment for hepatocellular carcinoma(HCC).This study aims to elucidate the effect of berberine on the anti-HCC effect of NK92-MI cells.Materials and methods:Human HCC SMMC-7721 and Hep3B cells were co-incubated with NK92-MI cells,berberine(60 or 80 μmol/L),or their combination for 36 h.To evaluate the killing effect of NK92-MI cells on HCC cells,the release of lactate dehydrogenase(LDH)in HCC cells was measured.The anti-tumor effects of berberine,NK92-MI cells,and their combinations were evaluated by MTS,EdU,Tunel,and Western blot assays.A male BALB/c nude mouse subcutaneous tumor model was used to investigate the anti-HCC effect of berberine and NK92-MI cells in vivo.Results:The LDH assay showed that berberine enhanced the cytotoxicity of NK92-MI cells on HCC cells.The combination of berberine and NK92-MI cells demonstrated more obvious anti-HCC effect than did the berberine or NK92-MI single treatment in inhibiting cell proliferation and inducing apoptosis both in vitro and in vivo.Mechanistically,the expression of programmed cell death-ligand 1(PD-L1)in HCC cells was upregulated after co-culture with NK-92MI cells.PD-L1 expression was knocked down,thereby inhibiting the proliferation and promoting apoptosis of HCC cells,and inhibited by berberine that blocked the secretion of interferon gamma(IFN-γ),thereby enhancing the anti-tumor effect of NK92-MI cells.Conclusions:Current data show that the immunomodulatory role of berberine is to enhance the cyto-toxic effect of NK92-MI cells and inhibit tumor immune escape by reducing the expression of PD-L1.Our study provides a rationale for the clinical application of berberine in combination with NK cells for the treatment of HCC.