Ginsenosides Rhl and Rh2 induced differentiation in cultured F9 teratocarcinoma stem cells. To elucidate the mechanism of differentiation-inducing effects of Rhl and Rh2, we examined the possible involvement of glucocorticoid receptor (GR) in the differentiation effects of these agents because of their structural similarity with glucocorticoids. Treatment of F9 cells with Rhl or Rh2 induced nuclear translocation of GR as did dexamethasone, a synthetic glucocorticoid. Furthermore, using gel shift assay, we determined a binding protein in F9 cells induced by Rhl or Rh2 using glucocorticoia response element (GRE). Protein complex was detected in the nuclear extracts of F9 cells and increased in Rhl or Rh2 treated F9 cells. To confirm whether this protein complex is a GR, we performed the competition assay with unlabled GRE as a specific competitor. In addition, supershift assay using a GR antibody showed supershift bands in Rhl or Rh2-treated F9 cells. Moreover, overexpression of GR by cotransfection of GR expression vector and GRE-luciferase vector enhanced the transactivation activity of GRE promoter in the presence of Rhl or Rh2 and further confirmed by examining the inhibitory effect of RU486, a glucocorticoid antagonist with a high affinity for the GR. These results suggest that the differentiation-inducing action of ginsenosides Rhl and Rh2 in F9 cells is due to the interaction with a GR.