【摘要】 目的:探索LINC AC004463.6通过PI3K/AKT/BCL-2通路对肝细胞癌(hepatocellular carcinoma, HCC)细胞生物学行为的影响。方法:运用细胞转染技术构建细胞系，实时荧光定量PCR验证，运用CCK8法、Transwell实验、EdU实验和流式细胞仪检测凋亡技术观察细胞功能学变化，运用Western blotting和ELISA技术探索其可能作用机制。结果:LINC AC004463.6在HCC中异常上调，在HCC细胞系和L02细胞中均检测到LINC AC004463.6的水平。选取SMCC7721和QGY7701作为LINC AC004463.6高表达细胞。CCK8和EdU实验结果表明，LINC AC004463.6表达降低显著抑制了SMCC7721和QGY7701细胞的增殖。而LINC AC004463.6基因敲除细胞中过表达LINC AC004463.6可使细胞增殖恢复和增强。Transwell实验检测表明shRNA处理的SMCC7721和QGY7701细胞侵袭性显著降低；经Lv-Rescue质粒处理后，细胞侵袭恢复并增强。LINC AC004463.6表达降低的细胞凋亡水平较高；LINC AC004463.6基因在LINC AC004463.6基因敲低的细胞中过表达后，细胞的凋亡水平有所恢复。Western blotting和ELISA检测提示shRNA组细胞BCL-2蛋白表达水平明显下调，同时PI3K和AKT蛋白表达水平明显下降；与此相反BAX蛋白表达水平升高。结论:LINC AC004463.6可能通过PI3K/AKT/BCL-2通路改变肿瘤细胞的生物学行为。LINC AC004463.6可能是诊断HCC的一个前瞻性生物标志物。更多还原

【Abstract】 Objective:To explore the effect of LINC AC004463.6 on biological behaviour of hepatocellular carcinoma(HCC) cells via PI3 K/AKT/BCL-2 pathway.Methods: The cell lines were constructed by cell transfection technology and verified by quantitative real-time PCR. Cell functional changes were observed by CCK8 assay,Transwell assay,EdU assay and flow cytometry assay. Western blotting and ELISA were used to explore the possible mechanism of action.Results:The level of LINC AC004463. 6 was abnormally up-regulated in HCC cell lines and detected in both HCC cell lines and L02 cells. SMCC7721 and QGY7701 were selected as the highly expressed cells of LINC AC004463. 6. The results of CCK8 and EdU experiments showed that the decreased expression of LINC AC004463. 6 significantly inhibited the proliferation of SMCC7721 and QGY7701 cells. Overexpression of LINC AC004463. 6 in knockout cells can restore and enhance cell proliferation. Transwell assay showed that the invasiveness of SMCC7721 and QGY7701 cells treated with shRNA was significantly reduced. After treatment with Lv-Rescue plasmid,the cell invasion was restored and enhanced. The apoptosis level of cells with decreased expression of LINC AC004463. 6 was higher. After the overexpression of the LINC AC004463. 6 gene in knockdown cells,the apoptosis level of cells was restored to some extent. Western blotting and ELISA showed that the protein expression levels of BCL-2,PI3K and AKT were significantly decreased in the shRNA group. On the contrary,the expression level of BAX protein increased.Conclusion:LINC AC004463. 6 may alter the biological behavior of tumor cells through the PI3K/AKT/BCL-2 pathway. LINC AC004463. 6 may be a prospective biomarker for the diagnosis of HCC.更多还原