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Metastasis is one of the main causes of poor prognosis for hepatocellular carcinoma (HCC), which has been linked to cell death resistance.Autophagy is an important survival mechanism under the condition of cell stress.Autophagy is hypothesized to play a role in HCC metastasis because of its pro-survival effect.HCC cell lines with stable autophagy inhibition were established through lentivirusmediated-siRNA targeting Beclin-1 and Atg5.A nude mouse model of pulmonary metastasis was developed using HCC cell lines with and without autophagy.Lung histopathological examination and small animal imaging indicated that autophagy inhibition significantly decreased the incidence of HCC pulmonary metastasis in vivo.Further experiments indicated that autophagy inhibition did not affect cell invasiveness or migration but attenuated the anoikis-resistance and lung colonization of HCC cells.Analysis of the changes in autophagic activity during metastasis showed that autophagy appeared to be activated in tumor colonization but not in cell invasion, migration, or detachment.