Differentiated epithelial cells show substantial lineage plasticity upon severe tissue injuries. In chronically injured mouse livers, part of hepatocytes become Sry-HMG box containing 9 (Sox9) (+) epithelial cell adhesion molecule (-) hepatocyte nuclear factor 4 alpha(+)biphenotypic hepatocytes. However, it is not clear whether all Sox9(+) hepatocytes uniformly possess cellular properties as hepatocyte progenitors. Here, we examined the microarray data comparing Sox9(+) hepatocytes with mature hepatocytes and identified CD24 as a novel marker for biphenotypic hepatocytes. Immunohistochemical analyses showed that part of Sox9(+) hepatocytes near expanded ductular structures expressed CD24 in the liver injured by 3,5-diethoxycarbonyl-1,4-dihydro-collidine (DDC) diet and by bile duct ligation. Indeed, Sox9(+) hepatocytes could be separated into CD24(-) and CD24(+) cells by fluorescence activated cell sorting. The ratio of CD24(+) cells against CD24(-) ones in Sox9+ hepatocytes gradually increased while DDC-injury progressed and colony-forming capability mostly attributed to CD24(+) cells. Although hepatocyte markers were remarkably downregulated in of Sox9(+) CD24(+) hepatocytes, they re-differentiated into mature hepatocytes in vitro and in vivo. Our current results demonstrate that the emergence of biphenotypic hepatocytes is a sequential event including the transition from CD24(-) and CD24(+) status, which may be a crucial step for hepatocytes to acquire progenitor properties.