Cheng Rongjie, Li Nan, Yang Shuyan, Liu Lei, Han Shiyu
Department of Obstetrics and Gynaecology, The Fourth Affiliated Hospital of Harbin Medical University, Harbin City, Heilongjiang Province, People's Republic of China,
Department of Pathology, The Fourth Affiliated Hospital of Harbin Medical University, Harbin City, Heilongjiang Province, People's Republic of China.
Onco Targets Ther. 2018 Oct 18;11:7245-7253. doi: 10.2147/OTT.S179937. eCollection 2018.
Long non-coding RNAs (lncRNAs) play important roles in cancer initiation and development. The purpose of the present study was to determine the functions and mechanisms of lncRNA ZEB1-AS1 in human cervical cancer (CC).
A total of 106 pairs of CC tissues and adjacent normal epithelial tissues were collected from CC patients who underwent resection. Three human CC cell lines (HeLa, C33A and SiHa) and a normal cervical cell line Crl-2614 and were transfected with human ZEB1-AS1 cDNA, or empty vector as the control. Then, cells were transfected with ZEB1-AS1-specific small interfering RNA (si-ZEB1-AS1), ZEB1-specific siRNA (si-ZEB1) or negative siRNA control (si-NC). The transfection efficiency was confirmed by RT-qPCR analysis. qPCR was applied to determine the qualification of RNA. Cell proliferation was investigated by MTT assay. The apoptosis rate of cells was detected by flow cytometer. Cell invasion was detected by transwell assay. Western blot was applied to determine the expression of proteins. CC xenografts in 12 male BALB/c athymic nude mice were established. And the tumor volumes were measured by vernier caliper.
We found that ZEB1-AS1 expression was remarkably increased in human CC tissue samples and cell lines, and its expression levels were closely associated with poor prognosis of CC patients. Moreover, we found that knockdown of ZEB1-AS1 inhibited the proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) of CC cells in vitro and suppressed CC xenograft tumor growth in vivo. Mechanistically, we found that knockdown of ZEB1-AS1 significantly inhibited ZEB1 expression, and knockdown of ZEB1 could rescue the effects of ZEB1-AS1 overexpression in CC cells.
In conclusion, our findings indicated that ZEB1-AS1 serves an oncogenic role in CC, which might become a potential prognostic indicator and therapeutic target in CC.
长链非编码RNA（lncRNA）在癌症的发生和发展中发挥重要作用。本研究旨在确定lncRNA ZEB1-AS1在人宫颈癌（CC）中的功能及机制。
从接受切除术的CC患者中收集106对CC组织及相邻正常上皮组织。将人ZEB1-AS1 cDNA或空载体（作为对照）转染至三种人CC细胞系（HeLa、C33A和SiHa）以及一种正常宫颈细胞系Crl-2614。然后，用ZEB1-AS1特异性小干扰RNA（si-ZEB1-AS1）、ZEB1特异性siRNA（si-ZEB1）或阴性siRNA对照（si-NC）转染细胞。通过RT-qPCR分析确认转染效率。应用qPCR确定RNA质量。通过MTT法研究细胞增殖。用流式细胞仪检测细胞凋亡率。通过Transwell法检测细胞侵袭。应用蛋白质印迹法确定蛋白质表达。在12只雄性BALB/c无胸腺裸鼠中建立CC异种移植瘤。用游标卡尺测量肿瘤体积。
我们发现ZEB1-AS1在人CC组织样本和细胞系中的表达显著增加，其表达水平与CC患者的不良预后密切相关。此外，我们发现敲低ZEB1-AS1可在体外抑制CC细胞的增殖、迁移、侵袭和上皮-间质转化（EMT），并在体内抑制CC异种移植瘤生长。机制上，我们发现敲低ZEB1-AS1可显著抑制ZEB1表达，敲低ZEB1可挽救ZEB1-AS1过表达对CC细胞的影响。
总之，我们的研究结果表明ZEB1-AS1在CC中起致癌作用，可能成为CC潜在的预后指标和治疗靶点。
