Objective       To investigate whether hepatic stellate cells (HSCs) can differentiate to hepatocyte-like cells in vitro

and the expression and changes of some stem cell markers during the induction.       Methods       Primary rat HSCs were isolated by the standard collagenase perfusion and purified by density gradient centrifugation

and then identified with fluorescent immunocytochemistry for Desmin and GFAP. The HSCs were treated with the cytokines as bFGF (20 μg/L)

FGF4 (20 μg/L)

HGF (20 μg/L) and IL-6 (1 μg/L) for 7 d served as experimental group

while those cultured without any cytokine for 24 h served as control group. The morphological changes of HSCs were examined by inverted phase contrast microscopy

and the hepatocyte-like phenotypes were determined by detecting the mRNA and protein expression of AFP

ALB

and other stem cell markers expression by real time-PCR and immunocytochemical assay.       Results       Our primary culture method obtained a large number of primary rat HSCs

with a positive rate to Desmin and GFAP of over 95%. After a combined induction of bFGF

FGF4

HGF and IL-6 for 7 d

HSCs showed a dramatically increased expression of AFP and ALB (P0.05) at mRNA and protein levels

with the shape changing from star-like to rotund or polygonal. But no such change was found in control cells. The combined induction also resulted in a significant decrease in mRNA expression of p75NTR (P0.05) and CD90

c-kit

sox-2 and oct-4 (P0.01). The expression of these markers at protein level was also significantly decreased or even undetectable.       Conclusion       HSCs might be of the potential as stem cells to differentiate into hepatocyte-like cells in vitro. During the induction

the expression of some stem cell markers is significantly downregulated.