Fig. 2.

3Cl-AHPC inhibits CYP7A1 expression in a SHP-dependent manner resulting in decreased bile acid levels. A and B, HepG2 cells were transfected with plasmids as indicated (for plasmid amounts, see Materials and Methods) and treated with 3Cl-AHPC (3-Cl) or vehicle for 10 or 20 h, and reporter assays were performed. The values for firefly luciferase activities were normalized by dividing by values for β-galactosidase activities. The mean and sem are plotted (n = 3). C, HepG2 cells were treated with 3Cl-AHPC as indicated, and relative (rel) mRNA levels of CYP7A1 were measured by q-RT-PCR and normalized to those of 36B4, sem (n = 3). D, HepG2 cells were transfected with small hairpin (sh) pSUPER vector as indicated and 60 h later, cells were treated with 3Cl-AHPC overnight, and mRNA levels of SHP and CYP7A1 were measured by q-RT-PCR and normalized to those of 36B4, sem (n = 3). Consistent results were observed in two independent (each in triplicate) assays. E, Experimental outlines. HepG2 cells were treated with vehicle, 3Cl-AHPC, or GW4064, as a control, for 24 h, and cells were collected for measuring CYP7A1 mRNA levels and bile acid levels (F). A–F, Statistical significance was measured using the Student's t test. *, P < 0.05; **, P < 0.01. NS, Nonsignificant; DMSO, dimethylsulfoxide.