Effect of shikonin on multidrug resistance in HepG2: The role of SIRT1

Abstract

Overexpression of SIRT1 is considered to enhance the resistance of HepG2 cells to irradiation. Shikonin, a naturally occurring naphthoquinone compound, displays anticancer effects and circumvents cancer drug resistance. Objectives: This study investigated the MDR reversal effect of shikonin induced by the overexpression of SIRT1. Materials and methods: The overexpression of SIRT1 in HepG2 cells was established by lentivirus infection. Five days after transduction, real-time quantitative polymerase chain reaction and western blotting were used to detect the expression of SIRT1 and MDR1/P-gp. Drug resistance was also evaluated by flow cytometry after rhodamine-123 staining. On day 5, the multidrug resistance cells were treated by shikonin (10-7, 10-6, and 10-5 μmol/L) one time. The cell viability was detected by the MTT assay, and apoptosis was evaluated by Hoechst 33342 staining and caspase-3 activity 24h after shikonin treatment. Results: Overexpression of SIRT1 decreased rhodamine-123 staining and successfully produced the R-HepG2 cell line. Compared with HepG2, the expression of MDR1/P-gp mRNA (3.45±0.35) and protein (1.40±0.05) were both upregulated in R-HepG2. Shikonin inhibited cell viability (from 93.9±2.1 to 66.7±1.5%), induced apoptosis of R-HepG2 (apoptotic ratio from 3.5±0.8 to 47.5±2.7%, caspase-3 activity from 103.5±1.9 to 329.2±14.9%, respectively), downregulated the mRNA and protein expression of SIRT1 and MDR1/P-gp, and decreased rhodamin 123 efflux. Discussion and conclusion: In the present study, we demonstrated that shikonin is able to overcome drug resistance in hepatocellular carcinoma cells, and the mechanism is related to the SIRT1-MDR1/P-gp signaling pathway.