关键词: LncRNA BLACAT1, miR-324-5p, MAP4K3, 肝癌, 增殖, 迁移, 侵袭

Abstract: Objective To investigate the effect of long non-coding RNA bladder cancer-associated transcript 1 (LncRNA BLACAT1) on the proliferation, migration and invasion of human hepatoma cells by regulating miR-324-5p / mitogen-activated protein kinase kinase kinase kinase 3 (MAP4K3) axis. Methods Real-time quantitative PCR (RT-qPCR) was used to detect the expression of BLACAT1, miR-324-5p and MAP4K3 mRNA in the hepatocellular carcinoma tissues, paracancerous tissues, and human normal hepatocytes and hepatoma cell lines ( Huh-7, SMMC7721, MHCC97 L). The liver cancer cells Huh-7 were divided into 5 groups: control group, si-NC group, si-BLACAT1 group, si-BLACAT1 + miR-NC group, and si-BLACAT1 + miR-324-5p inhibitor group. RT-qPCR was used to detect the mRNA expression levels of BLACAT1, miR-324-5p and MAP4K3 in cells of each group. CCK-8 assay was used to detect the cell proliferation. Wound-healing assay and transwell assay was used to detect the cell migration and cell invasion. Western blotting was used to detect the expression levels of MAP4K3, PCNA, MMP2, MMP9, c-Myc and Cyclin D1 in cells. The dual-luciferase reporter assays were used to verify the relationship between BLACAT1 and miR-324-5p, and the relationship between miR-324-5p and MAP4K3. Results The mRNA expression levels of BLACAT1 and MAP4K3 in hepatoma tissues and human hepatoma cell lines were greatly increased, and the expression level of miR-324-5p was greatly decreased (P < 0. 05). When compared with the control group and the si-NC group, the mRNA expression levels of BLACAT1, MAP4K3, the A450 value, the migration and invasion rate, and the expression levels of PCNA, MMP2, MMP9, MAP4K3, c-Myc and Cyclin D1 proteins were greatly decreased in the si-BLACAT1 group (P < 0. 05). The expression level of miR-324-5p was greatly increased ( P < 0. 05). Silencing of miR-324-5p attenuated the inhibitory effect of BASCAT1 knockdown on the proliferation, migration and invasion in Huh-7 cells. BASCAT1 targeted and negatively regulated the expression of miR-324-5p, and miR-324-5p targeted and regulated the MAP4K3. Conclusion Knockdown of BLACAT1 may down-regulate the expression of MAP4K3 by targeting miR-324-5p, thereby inhibiting the proliferation, migration and invasion of liver cancer cells. 

Key words: LncRNA BLACAT1, miR-324-5p, MAP4K3, liver cancer, proliferation, migration, invasio