Primary hepatocellular carcinoma (HCC) often invades into vessels and has a distal metastasis at an early stage, resulting in poor prognosis and therapeutic outcome. The metastasis has been attributable to the dissemination of tumor cells into circulation as circulating tumor cells (CTCs). Moreover, cancer stem cells (CSCs) within CTCs, which are termed as circulating tumor stem cells (CTSCs), are critical for formation of distal metastatic tumors. Although CD133 and CD90 have been used to characterize and isolate CTCs or CSCs in HCC, no good marker (cocktail) has been identified so far for CTSCs in HCC. Here, we show evidence that CD90+CXCR4+ HCC cells may be CTSCs in HCC. CD90+CXCR4+ HCC cells formed tumor spheres in culture and developed tumors after serial adoptive transplantations into NOD/SCID mice, while the CD90-CXCR4-, CD90-CXCR4+ or CD90+CXCR4- cells did not. Moreover, tumor cells were significantly more frequently detected in the circulation when CD90+CXCR4+ HCC cells were subcutaneously transplanted. Further, subcutaneous transplantation of CD90+CXCR4+ HCC cells, but not transplantation of CD90-CXCR4-, CD90-CXCR4+, or CD90+CXCR4- cells significantly developed distal metastatic tumors. Together, these data suggest that CD90+CXCR4+ HCC cells may be CTSCs and selective elimination of these cells may substantially improve the current HCC therapy by reducing cancer metastasis.