Guo Weisheng, Zhao Lin, Wei Guangya, Liu Peng, Zhang Yu, Fu Liran
Department of Hepatobiliary Surgery, Henan Province Hospital of Traditional Chinese Medicine, The Second Affiliated Hospital of Henan University of Traditional Chinese Medicine, Zhengzhou, Henan 450002, People's Republic of China.
Department of Traditional Chinese Medicine, People's Hospital of Zhengzhou, Zhengzhou 450000, Henan, People's Republic of China.
Cancer Manag Res. 2020 Aug 14;12:7291-7303. doi: 10.2147/CMAR.S255808. eCollection 2020.
Circular RNAs have been emerging as biomarkers in diagnosis and prognosis of pancreatic ductal adenocarcinoma (PDAC). The hsa_circ_0013912 (circ_0013912) has been retrieved to be upregulated in PDAC. Here, we further investigated its role in PDAC cells, as well as its mechanism via serving as competing endogenous RNA (ceRNA) for miRNA (miR)-7-5p, which is abundant in pancreas and suppresses the development of PDAC.
The clinical human tissues were harvested from Gene Expression Omnibus (GEO) database and PDAC patients, and expression of circ_0013912 and miR-7-5p was detected by real-time quantitative PCR. The interaction between both was confirmed by dual-luciferase reporter assay, RNA immunoprecipitation and biotin-miRNA pull-down assay. Functional experiments were performed using Cell Counting Kit-8 assay, colony formation assay, fluorescence-activated cell separation method, caspase 3 activity assay kit, Western blotting, transwell assays, and xenograft tumor model.
circ_0013912 was upregulated in PDAC tumors and cells; besides, circ_0013912 upregulation was associated with TNM stage and lymph node metastasis. Silencing circ_0013912 inhibited cell viability, colony formation ability, cell cycle entrance, migration and invasion, but facilitated apoptosis rate and caspase 3 activity in PANC-1 and AsPC-1 cells, accompanied with decreased c-myc, cyclin D1 and vimentin, and increased E-cadherin. Furthermore, miR-7-5p was a target of circ_0013912. Blocking miR-7-5p could promote cell growth, migration and invasion of PANC-1 and AsPC-1 cells with circ_0013912 silencing or not. Tumor growth was also restrained by circ_0013912 downregulation.
Circ_0013912 knockdown could suppress cell growth and metastasis of PDAC cells via sponging miR-7-5p.
环状RNA已成为胰腺导管腺癌（PDAC）诊断和预后的生物标志物。已发现hsa_circ_0013912（circ_0013912）在PDAC中上调。在此，我们进一步研究了其在PDAC细胞中的作用，以及其作为微小RNA（miR）-7-5p的竞争性内源性RNA（ceRNA）的机制，miR-7-5p在胰腺中含量丰富并抑制PDAC的发展。
从基因表达综合数据库（GEO）和PDAC患者中获取临床人体组织，通过实时定量PCR检测circ_0013912和miR-7-5p的表达。通过双荧光素酶报告基因检测、RNA免疫沉淀和生物素-miRNA下拉检测证实两者之间的相互作用。使用细胞计数试剂盒-8检测、集落形成检测、荧光激活细胞分选方法、半胱天冬酶3活性检测试剂盒、蛋白质免疫印迹法、Transwell检测和异种移植肿瘤模型进行功能实验。
circ_0013912在PDAC肿瘤和细胞中上调；此外，circ_0013912上调与TNM分期和淋巴结转移相关。沉默circ_0013912可抑制PANC-1和AsPC-1细胞的活力、集落形成能力、细胞周期进入、迁移和侵袭，但促进细胞凋亡率和半胱天冬酶3活性，同时伴有c-myc、细胞周期蛋白D1和波形蛋白减少，E-钙黏蛋白增加。此外，miR-7-5p是circ_0013912的靶标。阻断miR-7-5p可促进circ_0013912沉默或未沉默的PANC-1和AsPC-1细胞的生长、迁移和侵袭。circ_0013912下调也可抑制肿瘤生长。
Circ_0013912敲低可通过海绵吸附miR-7-5p抑制PDAC细胞的生长和转移。
