Meng Wei, Tai Yan, Zhao Hui, Fu Binsheng, Zhang Tong, Liu Wei, Li Hua, Yang Yang, Zhang Qi, Feng Yuliang, Chen Guihua
Department of Hepatic Surgery and Liver Transplantation Center of The Third Affiliated Hospital, Organ Transplantation Institute, Sun Yat-sen University; Organ Transplantation Research Center of Guangdong Province, Guangzhou, Guangdong, China (mainland).
Guangdong Key Laboratory of Liver Disease Research, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong, China (mainland).
Med Sci Monit. 2017 Mar 14;23:1295-1304. doi: 10.12659/msm.902692.
BACKGROUND Multi-drug resistance is one of the major problems limiting the efficacy of cisplatin (CDDP) in treatment of hepatocellular carcinoma (HCC), and abnormal microRNA (miRNA) expression in drug-resistant cell lines plays an important role in liver cancer chemotherapy resistance. MATERIAL AND METHODS We established stable Hep3B and 97L HCC cell strains resistant to CDDP, both in vitro and in vivo. A combination of microRNA microarray and RT-qPCR experiments were used to screen differentially expressed miRNAs in HCC cell strains. A CCK-8 assay was carried out to detect and calculate the survival rates and relative inhibitory rates. Oligonucleotide transfection was used to confirm the regulatory function of the miRNA in HCC drug resistance. RESULTS The IC50 of Hep3B/CDDP(v), 97L/CDDP(v), Hep3B/CDDP(s), and 97L/CDDP(s) were significantly higher than that of their parental cells. Moreover, the doubling time of drug-resistant cells increased compared with their parent cells. MiRCURYTM LNA Array (v 16.0) high-throughput tests of resistant cell models and their parent cells showed that there were 5 downregulated microRNAs in the 4 drug-resistant cell lines, and we chose hsa-miR-33a-5p as our target for further study. Oligonucleotide transfection showed that miR-33a-5p overexpression increased the cisplatin sensitivity of Hep3B/CDDP(v) and 97L/CDDP(v) drug-resistant cells and reduced their resistance. Additionally, inhibition of miR-33a-5p expression reduced cisplatin sensitivity in Hep3B and 97L and increased their drug resistance. CONCLUSIONS This study confirmed that the most downregulated microRNA, miR-33a-5p, can mediate the cisplatin resistance of HCC cells, providing a new and feasible direction for research into combatting liver cancer chemotherapy resistance.
背景 多药耐药是限制顺铂（CDDP）治疗肝细胞癌（HCC）疗效的主要问题之一，耐药细胞系中微小RNA（miRNA）表达异常在肝癌化疗耐药中起重要作用。材料与方法 我们在体外和体内建立了对CDDP耐药的稳定Hep3B和97L肝癌细胞株。采用miRNA微阵列和RT-qPCR实验相结合的方法筛选肝癌细胞株中差异表达的miRNA。进行CCK-8实验检测并计算生存率和相对抑制率。采用寡核苷酸转染来确认miRNA在肝癌耐药中的调控功能。结果 Hep3B/CDDP(v)、97L/CDDP(v)、Hep3B/CDDP(s)和97L/CDDP(s)的IC50显著高于其亲本细胞。此外，耐药细胞的倍增时间与其亲本细胞相比有所增加。对耐药细胞模型及其亲本细胞进行的MiRCURYTM LNA Array（v 16.0）高通量检测表明，4种耐药细胞系中有5种微小RNA表达下调，我们选择hsa-miR-33a-5p作为进一步研究的靶点。寡核苷酸转染表明，miR-33a-5p过表达增加了Hep3B/CDDP(v)和97L/CDDP(v)耐药细胞对顺铂的敏感性并降低了其耐药性。此外，抑制miR-33a-5p表达降低了Hep3B和97L对顺铂的敏感性并增加了其耐药性。结论 本研究证实，下调最明显的微小RNA miR-33a-5p可介导肝癌细胞的顺铂耐药性，为对抗肝癌化疗耐药性的研究提供了一个新的可行方向。
