Lv Jun, Ma Ling, Chen Xi-Lin, Huang Xiao-Hui, Wang Qian
Department of Hepatobiliary Surgery, the First Affiliated Hospital of Sun Yat-sen University, Guangzhou, 510080, China.
Department of Hepatobiliary Surgery, the Affiliated Hospital of Guilin Medical College, Guilin, 541001, China.
J Huazhong Univ Sci Technolog Med Sci. 2014 Jun;34(3):363-369. doi: 10.1007/s11596-014-1284-2. Epub 2014 Jun 18.
LncRNAH19 has been implicated as having both oncogenic and tumor suppression properties in cancer. LncRNAH19 transcripts also serve as a precursor for miR-675. However, it is unknown whether LncRNAH19 and miR-675 are involved in the migration and invasion of hepatocellular carcinoma (HCC) cells. The purpose of this study was to investigate the effect and mechanism of LncRNAH19 and miR-675 on migration and invasion of HCC cells. The migration and invasion of HCC cells were measured by Transwell migration and invasion assays after transfection of HCC cells with miR-675 inhibitors and LncRNAH19siRNA. The levels of LncRNAH19 and miR-675 were detected by quantitative reverse transcriptase real-time polymerase chain reaction (qRT-PCR), and the protein expression of AKT, GSK-3β and Cdc25A by Western blotting analysis. The expression levels of LncRNAH19 and miR-675 were higher in MHCC-97H cells than in L02, Huh-7 and HepG2 cells. Transwell migration assay revealed that the miR-675 inhibitor and LncRNAH19siRNA could significantly increase the migration of HCC cells (P<0.01) as compared with the control group. Transwell invasion assay demonstrated that the miR-675 inhibitor and LncRNAH19siRNA could significantly increase the invasion of HCC cells (P<0.01) as compared with the control group. Western blotting analysis showed that the expression levels of AKT and Cdc25A were significantly increased (P<0.05), and the expression level of GSK-3β was significantly decreased (P<0.05) after treatment with miR-675 inhibitors and LncRNAH19siRNA as compared with the control group. These findings suggested that inhibition of LncRNAH19 and miR-675 expression can promote migration and invasion of HCC cells via AKT/GSK-3β/Cdc25A signaling pathway.
长链非编码RNA H19在癌症中被认为具有致癌和肿瘤抑制特性。长链非编码RNA H19转录本也是miR - 675的前体。然而,长链非编码RNA H19和miR - 675是否参与肝细胞癌(HCC)细胞的迁移和侵袭尚不清楚。本研究的目的是探讨长链非编码RNA H19和miR - 675对HCC细胞迁移和侵袭的影响及机制。在用miR - 675抑制剂和长链非编码RNA H19小干扰RNA(siRNA)转染HCC细胞后,通过Transwell迁移和侵袭实验检测HCC细胞的迁移和侵袭能力。通过定量逆转录实时聚合酶链反应(qRT - PCR)检测长链非编码RNA H19和miR - 675的水平,通过蛋白质印迹分析检测AKT、糖原合成酶激酶3β(GSK - 3β)和细胞周期蛋白磷酸酶25A(Cdc25A)的蛋白表达。长链非编码RNA H19和miR - 675在MHCC - 97H细胞中的表达水平高于L02、Huh - 7和HepG2细胞。Transwell迁移实验显示,与对照组相比,miR - 675抑制剂和长链非编码RNA H19 siRNA可显著增加HCC细胞的迁移(P<0.01)。Transwell侵袭实验表明,与对照组相比,miR - 675抑制剂和长链非编码RNA H19 siRNA可显著增加HCC细胞的侵袭(P<0.01)。蛋白质印迹分析表明,与对照组相比,用miR - 675抑制剂和长链非编码RNA H19 siRNA处理后,AKT和Cdc25A的表达水平显著升高(P<0.05),GSK - 3β的表达水平显著降低(P<0.05)。这些发现提示,抑制长链非编码RNA H19和miR - 675的表达可通过AKT/GSK - 3β/Cdc25A信号通路促进HCC细胞的迁移和侵袭。