Department of Pathology, The First Affiliated Hospital, Zhengzhou University, Zhengzhou, Henan 450052, P.R. China.
Int J Oncol. 2017 Aug;51(2):657-667. doi: 10.3892/ijo.2017.4034. Epub 2017 Jun 8.
Esophageal squamous cell carcinoma (ESCC) is one of the most common malignancies in the world, marked by dysphagia and weight loss, bringing great suffering to patients. HNF1A‑AS1 (HAS1), a long non-coding RNA (lncRNA), has been identified prevalently involved in various human cancers. However, the exact effects and molecular mechanisms of HAS1 in ESCC progression are still elusive. In this study, upregulated expression of HAS1 was detected in ESCC tissues and four human ESCC cell lines (KYSE70, KYSE450, EC109 and EC970) compared with normal tissues and cell lines. Small interfering RNA (siRNA)-mediated knockdown of HAS1 largely suppressed cell proliferation and promoted cell apoptosis in KYSE70 and EC109 cells. The decreased expression of proliferation marker proteins and elevated level of apoptosis marker proteins further verified that HAS1‑siRNA suppressed cell viability in ESCC cells. Besides, the silence of HAS1 strongly reduced the wound closing rate and the number of invasive cells compared with control group. HAS1-siRNA also restrained the expression of migration marker proteins matrix metalloproteinase-9 (MMP-9) and vascular endothelial cell growth factor (VEGF). In addition, miR‑214 was predicted as a direct target of HAS1 by bioinformatics analysis. Downregulated expression of miR‑214 was elevated in KYSE70 and EC109 cells transfected with HAS1-siRNA. Subsequently, elevated expression of miR‑214 was suppressed by co-transfecting with miR‑214 inhibitor in EC109 cells pretreated with HAS1-siRNA. The result of luciferase activity assay showed that luciferase activity was strongly weakened by the combination of LncR-HAS1 WT and miR‑214 mimic. Moreover, the expression of SOX-4, a predicted target gene of miR‑214, was suppressed by HAS1-siRNA and was increased by miR‑214 inhibitor. HAS1-siRNA counteracted the effect of miR‑214 inhibitor on cell viability and mobility in EC109 cells. Finally, the in vivo experiment revealed that HAS1-siRNA abated the role of miR‑214 inhibitor in promoting tumor growth and metastasis. miR-214 also mediated the effect of HAS1 on upregulating the expression of SOX-4 in vivo. Taken together, our study indicated a HAS1-miR‑214-SOX-4 pathway in regulating the growth and metastasis of ESCC, providing a promising target for ESCC therapy.
食管鳞状细胞癌（ESCC）是世界上最常见的恶性肿瘤之一，其特征为吞咽困难和体重减轻，给患者带来极大的痛苦。HNF1A 反义 RNA1（HAS1）是一种长链非编码 RNA（lncRNA），已被确定广泛参与各种人类癌症。然而，HAS1 在 ESCC 进展中的确切作用和分子机制仍不清楚。在这项研究中，与正常组织和细胞系相比，ESCC 组织和四种人 ESCC 细胞系（KYSE70、KYSE450、EC109 和 EC970）中检测到 HAS1 的表达上调。与对照组相比，用小干扰 RNA（siRNA）介导的 HAS1 敲低在 KYSE70 和 EC109 细胞中大量抑制细胞增殖并促进细胞凋亡。增殖标记蛋白表达降低和凋亡标记蛋白水平升高进一步证实 HAS1-siRNA 抑制 ESCC 细胞的活力。此外，与对照组相比，HAS1-siRNA 强烈降低了伤口闭合率和侵袭细胞的数量。HAS1-siRNA 还抑制了迁移标记蛋白基质金属蛋白酶 9（MMP-9）和血管内皮生长因子（VEGF）的表达。此外，生物信息学分析预测 miR-214 是 HAS1 的直接靶标。与 HAS1-siRNA 转染的 KYSE70 和 EC109 细胞相比，miR-214 的表达下调。随后，在 HAS1-siRNA 预处理的 EC109 细胞中转染 miR-214 抑制剂后，miR-214 的表达升高得到抑制。荧光素酶活性测定结果表明，LncR-HAS1 WT 和 miR-214 模拟物的组合强烈减弱了荧光素酶活性。此外，miR-214 预测靶基因 SOX-4 的表达被 HAS1-siRNA 抑制，而被 miR-214 抑制剂增加。HAS1-siRNA 拮抗了 miR-214 抑制剂对 EC109 细胞活力和迁移的影响。最后，体内实验表明，HAS1-siRNA 减弱了 miR-214 抑制剂在促进肿瘤生长和转移中的作用。miR-214 还介导了 HAS1 在体内上调 SOX-4 表达的作用。总之，我们的研究表明，HAS1-miR-214-SOX-4 通路在调节 ESCC 的生长和转移中起作用，为 ESCC 的治疗提供了一个有前途的靶点。
