miRNA-138 调控卵巢上皮癌细胞对顺铂敏感性的机制研究

Regulative role of miRNA-138 on platinum sensitivity of ovarian epithelial cancer cells

摘要:

目的 探讨miRNA-138 和沉默信息调节因子2 相关酶1（SIRT1）的差异表达对卵巢上皮癌顺铂化疗敏感性的影响。方法 选取2014 年1 月-2017 年1 月在牡丹江医学院红旗医院妇科就诊卵巢上皮癌患者80 例。根据患者对顺铂化疗的敏感性分为顺铂敏感组和顺铂耐药组。实时荧光定量PCR（qRT-PCR）和Western blot 检测卵巢上皮癌组织中miRNA-138 和SIRT1 的表达水平；培养SKOV3 细 胞， 分别转染miRNA-138 的类似物（miRNA-138 mimic） 和抑制物（miRNA-138 inhibitor） 后，qRTPCR检测SIRT1 miRNA 的表达变化，并采用MTT 法检测细胞对顺铂的敏感性；SIRT1 miRNA 3'-UTR 野生型（wt）和突变型（mut）萤光素酶报告质粒与miRNA-138 mimic 共转染，双荧光素酶报告系统分析荧光素酶活性。结果 铂敏感组miRNA-138 表达水平高于顺铂耐药组，而SIRT1 表达水平低于顺铂耐药组，差异有统计学意义（P <0.05）；过表达miRNA-138 可降低SIRT1 的表达，提高SKOV3 细胞对顺铂的敏感性，而抑制miRNA-138 可提高SIRT1 的表达，降低SKOV3 细胞对顺铂的敏感性，差异有统计学意义（P <0.05）；miRNA-138 mimic 与SIRT1 miRNA 3'-UTR 野生型质粒共转染，荧光素酶活性降低（P <0.05），而与SIRT1 miRNA 3'-UTR 突变型质粒共转染，荧光素酶活性无变化（P >0.05）。结论 miRNA-138 可能通过SIRT1 调控卵巢上皮癌细胞对顺铂的敏感性，可作为卵巢上皮癌顺铂化疗耐药治疗的靶点。

Abstract:

Objective To investigate the regulative effect of miRNA-138 on chemo-sensitivity of ovarian epithelial cancer to platinum and potential mechanism. Methods Totally 80 cases of patients with ovarian epithelial cancer admitted into Mudanjiang Medical College Hongqi Hospital from January 2014 to January 2017 were included in this study. Patients were divided into two groups: platinum-sensitive group and platinum-resistant group.Quantitative real-time PCR (RT-qPCR) and Western Blot were performed to detect the expression of miRNA-138 and SIRT1 in ovarian epithelial carcinoma. SKOV3 cell line were transfected with miRNA-138 analogs (miRNA-138 mimic group) and inhibitor (miRNA-138 inhibitor group). Expression of SIRT1 mRNA was measured by RT-qPCR.MTT assay was utilized for determination of sensitivity to platinum. SIRT1 mRNA 3'-UTR wild type and mutant type luciferase reporter plasmids were co-transfected with miRNA-138mimic, and double luciferase reporter system was used to analyze luciferase activity. Results The concentration of miRNA-138 in platinum-sensitive group was significantly higher than that in platinum-resistant group, while levels of SIRT1 was significantly decreased compared with platinum-resistant group (P < 0.05). Overexpression of miRNA-138 decreased the expression of SIRT1 and increased the sensitivity of tumor cells to platinum (P < 0.05), which was reversed by inhibition of miRNA-138(P < 0.05). Luciferase activity was significantly reduced in group with transfection of SIRT1 mRNA 3’-UTR wildtype plasmids, while no significant changes in luciferase activity were observed in transfected SIRT1 mRNA 3’-UTR mutant plasmid (P > 0.05). Conclusion MiRNA-138 may regulate the sensitivity of ovarian epithelial cancer cells to platinum through SIRT1 activity, which can be a potential therapeutic target of ovarian epithelial cancer.