Figure 4.

CXCL6 Activates Hepatic Stellate Cell (HSCs) by Stimulating TGF‐β and Downstream Smad3‐BRD4. A and B, Kupffer cells (KCs) were treated with CXCL6 (100 ng/mL) or phosphate‐buffered saline (PBS) with the addition of CXCR1/2 antagonist SCH527123 (10 μmol/L), EGFR antagonist Afatinib (10 μmol/L), or DMSO as a control for 18 h. KC conditioned medium was then transferred onto 3‐d HSCs and cultured for 8 h. α‐SMA and TGF‐β concentrations in HSC‐T6 cells were detected by ELISA. C and D, α‐SMA, SMAD3, BRD4, C‐MYC and EZH2 mRNA and protein levels in HSC‐T6 cells cultured with KC conditioned medium were detected by RT‐PCR and western blotting. E, KCs were treated with CXCL6 (100 ng/mL), TGF‐β (10 ng/mL) or PBS with the addition of TGF‐β receptor antagonist SB431542 (10 μmol/L), TGF‐β‐neutralizing antibody 1D11 (4 μg/mL) or DMSO as a control for 18 h. KC conditioned medium was then transferred onto 3‐d HSC‐T6 cells and cultured for 8 h. α‐SMA concentration in HSCs was detected by ELISA. F and G, Relative mRNA and protein levels in HSC‐T6 cells cultured with KC conditioned medium were detected by RT‐PCR and western blotting (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001 vs control group, ^## P < 0.01, ^### P < 0.001 vs CXCL6 + DMSO group