Fig. 2.

Inactivation of BFAR using siRNAs in HuH7 cells. (A) HuH7 cells stably expressing PNPLA3-HiBiT were treated with pools of four siRNAs (60 pmol) each directed against PNPLA3, BFAR or with scrambled sequences (Scr), or with MG132 (10 µM). Immunoblot analysis was performed using cell lysate, as described in Materials and Methods. Signals were quantified using a LI-COR imaging system, normalized to levels of CANX, and expressed as arbitrary units (AUs). The mRNA levels of indicated genes were measured and normalized as described in Materials and Methods. All mRNA levels were compared to levels seen in cells treated with Scr siRNA using the Student’s t test. (B) Inactivation of endogenous BFAR in HuH7 cells, which express PNPLA3(148M). HuH7 cells were treated with buffer alone, or with Scr-, PNPLA3-, or BFAR-siRNAs (60 pmol). Immunoblot analysis of cell lysates and signal quantification were performed as described in panel (A). Values represent means ± SEM. Means were compared to cells treated with Scr siRNA using the Student’s t test. *P < 0.05, **P < 0.01, ***P < 0.001, †P < 0.0001. The experiments were repeated three times, and the results were similar.