Sheng Long, Xiong Maoming, Li Cong, Meng Xiangling
Department of General Surgery, The First Affiliated Hospital of Anhui Medical University, Hefei, 230022, China,
Pathol Oncol Res. 2014 Jul;20(3):541-8. doi: 10.1007/s12253-013-9726-x. Epub 2013 Dec 11.
Multidrug resistance (MDR) in hepatocellular carcinoma (HC) significantly impedes the effect of chemotherapy and is considered as a primary reason leading to its recurrences and metastasis. The aim of present study was to explore new molecular targets for the reversal of MDR in HC by screening the adriamycin (ADM)-induced, human MDR-resistant HC cell subline Bel-7402/ADM. Small interfering RNAs (siRNAs) of four (MDR1si326, MDR1si1513, MDR1si2631 and MDR1si3071) targeting MDR1 were designed and transfected into Bel-7402/ADM cell strains. The experiments involved the following: mRNA expression of MDR1 gene by RT-PCR, P-glycoprotein (P-gp) expression by Western blot, intracellular ADM accumulation flow cytometry, and IC50 of ADM by a cytotoxic MTT assay. Four siRNAs reversed MDR in HC mediated by MDR1 to varying degrees. The expression level of MDR1 mRNA in cells of MDR1si326 or MDR1si2631 group (0.190 ± 0.038 or 0.171 ± 0.011) was more decreased. The expression level of P-gp in cells of MDR1si326 group was the lowest. The accumulation of ADM in cells of MDR1si326 or MDR1si2631 group (77.0 ± 3.5 or 75.4 ± 2.9) was more increased. The IC50 of cells to ADM was lowest in MDR1si326 group (11.32 ± 0.69 mg/L). Compared with other three siRNAs, MDR1si326 performed the optimal reversal effect of drug resistance in human HC Bel-7402/ADM.
肝细胞癌（HC）中的多药耐药（MDR）显著阻碍化疗效果，被认为是导致其复发和转移的主要原因。本研究旨在通过筛选阿霉素（ADM）诱导的人MDR耐药HC细胞亚系Bel-7402/ADM，探索逆转HC中MDR的新分子靶点。设计了针对MDR1的四种小干扰RNA（siRNA，即MDR1si326、MDR1si1513、MDR1si2631和MDR1si3071）并转染到Bel-7402/ADM细胞株中。实验包括：通过RT-PCR检测MDR1基因的mRNA表达，通过蛋白质印迹法检测P-糖蛋白（P-gp）表达，通过流式细胞术检测细胞内ADM蓄积，以及通过细胞毒性MTT试验检测ADM的半数抑制浓度（IC50）。四种siRNA不同程度地逆转了由MDR1介导的HC中的MDR。MDR1si326或MDR1si2631组细胞中MDR1 mRNA的表达水平降低得更多（分别为0.190±0.038或0.171±0.011）。MDR1si326组细胞中P-gp的表达水平最低。MDR1si326或MDR1si2631组细胞中ADM的蓄积增加得更多（分别为77.0±3.5或75.4±2.9）。MDR1si326组细胞对ADM的IC50最低（11.32±0.69 mg/L）。与其他三种siRNA相比，MDR1si326在人HC Bel-7402/ADM中表现出最佳的耐药逆转效果。
