Figure 5.

C/EBPβ is critical for HSC activation in response to TGFβ or IRE1α signaling. A, LX-2 cells were infected with a lentivirus expressing an shRNA against C/EBPβ (sh-C/EBPβ) or an NT control. Two clonal cell lines were engineered and were assessed by immunoblotting for expression of C/EBPβ isoforms (LAP1/2 and LIP). HSC70 served as a loading control. Quantification is shown below. B and C, sh-C/EBPβ cells (clones 1 and 2) or NT cells were treated with TGFβ (5 ng/ml) for 24 h. Cell lysates were harvested and assessed by immunoblotting for collagen I, FN, C/EBPβ, and HSC70 (loading control). Quantification is shown below. D, sh-C/EBPβ or NT cells were treated with TGFβ (5 ng/ml), after which mRNA was harvested and qPCR was performed to assess fibronectin, procollagen 1α1 and 1α2, or αSMA expression. E, DoxIRE1α cells were infected with lentivirus expressing shRNA targeting C/EBPβ (sh-C/EBPβ) or an NT control, and clonal cell populations were selected. Cells were treated with doxycycline for 24 h, after which cell lysates were harvested and assessed by immunoblotting for IRE1α, fibronectin, collagen I, αSMA, C/EBPβ, and HSC70 (loading control). Quantification is shown adjacent. Statistics were performed using ANOVA followed by Tukey post hoc analysis (*, p < 0.05; **, p < 0.01; ***, p < 0.001). n ≥ 3 biological replicates for each experiment. Error bars, S.E.