Objective: To detect the expression level of YES-associated protein 1 (YAP) in hepatocellular carcinoma (HCC) cell lines and investigate its effects on the proliferation activity and the sensitivity to sorafenib in HCC cells. Methods: Western blot was used to detect the protein expression levels of YAP in SMMC-7721, SK-Hep-1, HepG-2, Huh7 and the normal liver cell line L-O2. YAP specific small interfering RNA (si-YAP) or YAP expression plasmid were transfected in SK-Hep-1 or Huh7 cells, respectively. Cell counting kit-8 (CCK-8) test was used to detect the cell proliferation activity and the cell cycle test was conducted by flow cytometry. SK-Hep-1 and SK-Hep-1 si-YAP cells were subcutaneously injected into the nude mice which were sequentially treated by intragastric administration of sorafenib, and the tumor growth in vivo were observed and compared. Results: The expression of YAP was upregulated in HCC cell lines. Deletion of YAP expression significantly decreased the survival rate of SK-Hep-1 cells [(78.5±0.3)% vs (92.3±0.2)%, P=0.025]. Knockdown of YAP significantly increased the percentage of G(0)/G(1)-phase cells [ (65.4±3.3) % vs (55.7±3.4) %, P=0.039]. On the contrary, upregulation of the YAP expression in Huh7 cells significantly increased the cell survival rate [(81.2±1.3)% vs (62.5±1.1)%, P=0.013] and reduced the percentage of G(0)/G(1)-phase cells [(38.2±3.8)% vs (48.8±2.9)%, P=0.019]. The survival rate of SK-Hep-1 cells treated by si-YAP combined with sorafenib was (31.13±1.79)%, significantly lower than (48.87±0.58) % of SK-Hep-1 cells treated by sorafenib alone (P=0.001), while overexpression of YAP attenuated the inhibitory effect of sorafenib on the survival of Huh7 cells [(69.98±2.94) % vs (53.53±1.93)%, P=0.001]. The tumor weights of SK-Hep-1 group, sorafenib alone group, SK-Hep-1 si-YAP group and SK-Hep-1 si-YAP combined with sorafenib group were (0.96±0.08) g, (0.62±0.08) g, (0.70±0.06) g and (0.27±0.02) g, respectively. The tumor weights of sorafenib alone group and SK-Hep-1 si-YAP group were significantly lower than that of SK-Hep-1 group (P=0.012 and P=0.031, respectively). The tumor weight of SK-Hep-1 si-YAP combined with sorafenib group was significantly lower than that of SK-Hep-1 si-YAP group (P=0.001). Conclusions: The expression of YAP is upregulated in HCC cell lines, which regulates the proliferation, cell cycle, and sensitivity to sorafenib of HCC cells. YAP is a potential molecular target for HCC treatment.
目的： 检测YAP在肝癌细胞中的表达水平，并探讨其对肝癌细胞增殖活性以及对索拉非尼敏感性的影响。 方法： 采用Western blot方法检测人肝癌细胞株SMMC－7721、SK－Hep－1、HepG－2、Huh7和正常人肝细胞株L－O2中YAP的蛋白表达水平。以YAP的小干扰RNA或过表达质粒分别转染SK－Hep－1和Huh7细胞，采用细胞计数试剂盒8(CCK－8)法检测细胞的增殖能力，流式细胞术检测细胞周期。将SK－Hep－1和SK－Hep－1 si－YAP细胞接种于裸鼠皮下，并以索拉非尼灌胃，观察移植瘤与对照组的差异。 结果： YAP蛋白在肝癌细胞株中表达升高。下调SK－Hep－1细胞的YAP表达后，细胞存活率为(78.5±0.3)%，与对照组[(92.3±0.2)%]比较明显下降，差异有统计学意义(P＝0.025)；G(0)/G(1)期细胞占(65.4±3.3)%，与对照组[(55.7±3.4)%]比较明显升高，差异有统计学意义(P＝0.039)。上调Hun7细胞的YAP表达后，细胞存活率为(81.2±1.3)%，与对照组[(62.5±1.1)%]比较明显升高，差异有统计学意义(P＝0.013); G(0)/G(1)期细胞占(38.2±3.8)% ，与对照组[(48.8±2.9)%]比较明显下降，差异有统计学意义(P＝0.019)。si－YAP＋索拉非尼组SK－Hep－1细胞的存活率为(31.13±1.79)%，低于索拉非尼组[(48.87±0.58)%]，差异有统计学意义(P<0.0001)。PC3.1－YAP＋索拉非尼组的细胞存活率为(69.98±2.94)%，高于索拉非尼组[(53.53±1.93)%]，差异有统计学意义(P<0.0001)。裸鼠移植瘤模型实验显示，SK－Hep－1组、SK－Hep－1＋索拉非尼组、SK－Hep－1 si－YAP组和SK－Hep－1 si－YAP＋索拉非尼组瘤重分别为(0.96±0.08)g、(0.62±0.08)g、(0.70±0.06)g和(0.27±0.02)g。SK－Hep－1＋索拉非尼组和SK－Hep－1 si－YAP组瘤重均低于SK－Hep－1组(P值分别为0.012和0.031)，SK－Hep－1 si－YAP＋索拉非尼组瘤重低于SK－Hep－1 si－YAP组(P＝0.001)。 结论： YAP在肝癌细胞系中表达上调，其表达变化影响细胞增殖能力和细胞周期进程，并影响肝癌对索拉非尼的敏感性。YAP是肝癌治疗的一个潜在分子靶点。.
Keywords: Carcinoma, hepatocellular; Drug sensitivity; Sorafenib; YAP.