miR-186-5p targeting Sal inhibits cisplatin resistance in non-small-cell lung cancer cells (NSCLCs)
miR-186-5p targeting Sal inhibits cisplatin resistance in non-small-cell lung cancer cells (NSCLCs)
复制标题
靶向 SIX1 的 miR-186-5p 抑制非小细胞肺癌细胞 (NSCLC) 的顺铂耐药性
DOI：
10.4149/neo_2019_190511n420
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发表时间：
2020-01-01
期刊：
影响因子：
3
通讯作者：
Luo, H.
中科院分区：
文献类型：
--
作者：
Liu, X.;Zhou, X.;Luo, H.
据报道，miR-186-5p与非小细胞肺癌细胞(NSCLC)的肿瘤发生和紫杉醇耐药有关。然而，miR-186-5p是否参与了肺癌(包括非小细胞肺癌)对二氨基二氯铂(DDP)的耐药尚不清楚。RT-qPCR和Western印迹检测miR-186-5p和SIX1基因的表达。体外用WTT法测定DDP的半数抑制浓度(IC50)和细胞增殖。用流式细胞仪和Transwell法检测细胞的凋亡率和迁移侵袭能力。在Diana Tools软件上预测miR-186-5p与SIX1的靶向结合，并通过双荧光素酶报告实验和RNA免疫沉淀证实。体内实验采用异种移植监测肿瘤生长情况。在耐DDP的NSCLC组织和细胞(A549/DDP和H1299/DDP)中miR-186-5p的表达下调。在功能上，miR-186-5p过表达可抑制A549/DDP和H1299/DDP细胞的增殖、迁移和侵袭，促进细胞凋亡率。从机制上讲，SIX1被认为是miR-186-5p的下游靶点，并在A549/DDP和H1299/DDP细胞中高表达。同样，四喜基因敲除也可抑制耐DDP的NSCLC细胞的增殖、迁移和侵袭，促进细胞凋亡率，而miR-186-5p下调可逆转这一作用。此外，A549/DDP细胞诱导的移植瘤对顺铂耐药，在DDP治疗下，miR-186-5p过表达可抑制肿瘤生长。总之，在体外和体内，miR-186-5p的上调可能通过靶向SIX1来抑制顺铂耐药的NSCLC细胞的耐药性。
miR-186-5p has been reported to be implicated in tumorigenesis and paclitaxel resistance in non-small-cell lung cancer cells (NSCLCs). However, it remains undisclosed whether miR-186-5p takes a part in chemoresistance against diaminodichloroplatinum (cisplatin, DDP) in lung cancers, including NSCLC. Expression of miR-186-5p and sine oculis homeobox 1 (SIX1) was detected using RT-qPCR and western blot. In vitro, 50% inhibitory concentration (IC50) of DDP and cell proliferation were measured by WTT assay. The rate of apoptosis and abilities of migration and invasion were evaluated with flow cytometry and Transwell assay. The target binding between miR-186-5p and SIX1 was predicted on Diana tools software and confirmed by dual-luciferase reporter assay and RNA immunoprecipitation. In vivo experiments, xenogeneic transplantation was conducted to monitor the tumor growth. Expression of miR-186-5p was downregulated in DDP resistant NSCLC tissues and cells (A549/DDP and H1299/DDP). Functionally, miR-186-5p overexpression could inhibit cell proliferation, migration and invasion, and promoted apoptosis rate in A549/DDP and H1299/DDP cells. Mechanically, SIX1 was identified as a downstream target for miR-186-5p and was highly expressed in A549/DDP and H1299/DDP cells. Similarly, SIXI knockdown could also suppress DDP resistant NSCLC cell proliferation, migration and invasion, and promote apoptosis rate, which was reversed by miR-186-5p downregulation. Moreover, xenograft tumors induced by A549/DDP cells exerted cisplatin resistance, and miR-186-5p overexpression could inhibit tumor growth under DDP treatment. In conclusion, upregulation of miR-186-5p suppresses cisplatin resistance in DDP resistant NSCLC cells both in vitro and in vivo presumably by targeting SIX1.