Jin Yong-Dong, Ren Yi, Wu Ming-Wei, Chen Ping, Lu Jin
Department of Medical Oncology, Sichuan Cancer Hospital , Chengdu , China and.
Pharm Biol. 2015 Jul;53(7):1016-21. doi: 10.3109/13880209.2014.952836. Epub 2014 Dec 4.
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.
This study investigated the MDR reversal effect of shikonin induced by the overexpression of SIRT1.
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 24 h after shikonin treatment.
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.
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.
SIRT1的过表达被认为可增强HepG2细胞对辐射的抗性。紫草素是一种天然存在的萘醌化合物，具有抗癌作用并能克服癌症耐药性。
本研究调查了SIRT1过表达诱导的紫草素的多药耐药逆转作用。
通过慢病毒感染在HepG2细胞中建立SIRT1的过表达。转导5天后，使用实时定量聚合酶链反应和蛋白质印迹法检测SIRT1和MDR1/P - gp的表达。罗丹明123染色后通过流式细胞术评估耐药性。在第5天，用紫草素（10(-7)、10(-6)和10(-5) μmol/L）处理多药耐药细胞一次。通过MTT法检测细胞活力，紫草素处理24小时后通过Hoechst 33342染色和caspase - 3活性评估细胞凋亡。
SIRT1的过表达降低了罗丹明123染色，并成功构建了R - HepG2细胞系。与HepG2相比，R - HepG2中MDR1/P - gp mRNA（3.45 ± 0.35）和蛋白质（1.40 ± 0.05）的表达均上调。紫草素抑制细胞活力（从93.9 ± 2.1降至66.7 ± 1.5%），诱导R - HepG2细胞凋亡（凋亡率分别从3.5 ± 0.8升至47.5 ± 2.7%，caspase - 3活性从103.5 ± 1.9升至329.2 ± 14.9%），下调SIRT1和MDR1/P - gp的mRNA和蛋白质表达，并减少罗丹明123外排。
在本研究中，我们证明紫草素能够克服肝癌细胞中的耐药性，其机制与SIRT1 - MDR1/P - gp信号通路有关。
