Wang Xiao-Kang, Peng Yan, Tao Hao-Ran, Zhou Fen-Fang, Zhang Chi, Su Fei, Wang Shi-Pei, Liu Qing, Xu Li-Hua, Pan Xue-Kai, Xie Wei, Feng Mao-Hui
Department of Oncology, Zhongnan Hospital of Wuhan University, Wuhan, 430030, China.
Department of Geriatrics, The First Clinical Medical College of China Three Gorges University, The Central People's Hospital of Yichang, Yichang, 443200, China.
J Huazhong Univ Sci Technolog Med Sci. 2017 Jun;37(3):343-347. doi: 10.1007/s11596-017-1757-1. Epub 2017 Jun 6.
The sialyl Lewis X (SLe) antigen encoded by the FUT7 gene is the ligand of endotheliam-selectin (E-selectin). The combination of SLe antigen and E-selectin represents an important way for malignant tumor metastasis. In the present study, the effect of the SLe-binding DNA aptamer on the adhesion and metastasis of hepatocellular carcinoma HepG2 cells in vitro was investigated. Reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescence staining were conducted to detect the expression of FUT7 at both transcriptional and translational levels. The SLe expression in HepG2 cells treated with different concentrations of SLe-binding DNA aptamer was detected by flow cytometry. Besides, the adhesion, migration, and invasion of HepG2 cells were measured by cell adhesion assay, and the Transwell migration and invasion assay. The results showed that the FUT7 expression was up-regulated at both mRNA and protein levels in HepG2 cells. SLe-binding DNA aptamer could significantly decrease the expression of SLe in HepG2 cells. The cell adhesion assay revealed that the SLe-binding DNA aptamer could effectively inhibit the interactions between E-selectin and SLe in the HepG2 cells. Additionally, SLe-binding DNA aptamers at 20 nmol/L were found to have the similar effect to the monoclonal antibody CSLEX-1. The Transwell migration and invasion assay revealed that the number of penetrating cells on the down-side of Transwell membrane was significantly less in cells treated with 5, 10, 20 nmol/L SLe-binding DNA aptamer than those in the negative control group (P<0.01). Our study demonstrated that the SLe-binding DNA aptamer could significantly inhibit the in vitro adhesion, migration, and invasion of HepG2 cells, suggesting that the SLe-binding DNA aptamer may be used as a potential molecular targeted drug against metastatic hepatocellular carcinoma.
由FUT7基因编码的唾液酸化路易斯X（SLe）抗原是内皮细胞选择素（E-选择素）的配体。SLe抗原与E-选择素的结合是恶性肿瘤转移的重要途径。在本研究中，研究了SLe结合DNA适配体对体外肝癌HepG2细胞黏附与转移的影响。进行逆转录聚合酶链反应（RT-PCR）和免疫荧光染色以检测转录和翻译水平上FUT7的表达。通过流式细胞术检测用不同浓度的SLe结合DNA适配体处理的HepG2细胞中SLe的表达。此外，通过细胞黏附试验、Transwell迁移试验和侵袭试验来检测HepG2细胞的黏附、迁移和侵袭能力。结果显示，HepG2细胞中FUT7在mRNA和蛋白质水平上均上调。SLe结合DNA适配体可显著降低HepG2细胞中SLe的表达。细胞黏附试验表明，SLe结合DNA适配体可有效抑制HepG2细胞中E-选择素与SLe之间的相互作用。此外，发现20 nmol/L的SLe结合DNA适配体具有与单克隆抗体CSLEX-1相似的作用。Transwell迁移和侵袭试验显示，用5、10、20 nmol/L SLe结合DNA适配体处理的细胞中，Transwell膜下侧穿透细胞的数量明显少于阴性对照组（P<0.01）。我们的研究表明，SLe结合DNA适配体可显著抑制HepG2细胞的体外黏附、迁移和侵袭，提示SLe结合DNA适配体可能作为一种潜在的抗转移性肝癌分子靶向药物。
