Figure 5.

lnc-Hser inhibits HCs apoptosis via C5AR1. (A, B) Primary HCs were infected with LV-lnc-Hser for 72 h and further treated with 10 ng/ml TGFβ for additional 24 h. The protein level of C5AR1 was detected by western blot. GAPDH was used as an internal control (A). The mRNA level of C5, C5ar1 and Cntrl was detected by qRT-PCR (B). (C) The protein level of C5AR1 in primary HCs infected with lenti-lnc-Hser-shRNA or lenti-NC was detected by western blot. GAPDH was used as an internal control. (D) The mRNA level of C5, C5ar1 and Cntrl was detected in primary HCs infected with lenti-lnc-Hser-shRNA or lenti-NC by qRT-PCR. (E) Mice were treated with oil in combination with injection of lenti-NC (Negative Control, n = 10), or CCl₄ in combination with injection of lenti-NC (NC + CCl₄, n = 10), or oil in combination with injection of lenti-lnc-Hser-shRNA (lnc-Hser-shRNA, n = 10), or CCl₄ in combination with injection of lenti-lnc-Hser-shRNA (lnc-Hser-shRNA + CCl₄, n = 10). qRT-PCR analysis of C5, C5ar1 and Cntrl in livers of mice in each group. (F-H) lnc-Hser was stably knocked down by the CRISPR/Cas9 system with guide RNA pairs in AML12 cells. PMX205, a specific inhibitor of C5AR1, was used to treat lnc-Hser-silenced AML12 cells for 24 h. The expression of lnc-Hser, pro-fibrogenic, apoptosis-related and EMT-related genes was detected by qRT-PCR (F, G) and western blot. GAPDH was used as an internal control (H). The data are expressed as the mean ± SD for at least triplicate experiments. *p<0.05 stands for vs LV-Control, NC or Control gRNA. #p<0.05 stands for vs LV-Control + TGFβ, NC+ CCl₄ or lnc-Hser-gRNAs.