Jing Xu, Chen Yingjie, Chen Ye, Shi Guangyan, Lv Shuanghao, Cheng Nana, Feng Chaolin, Xin Zhen, Zhang Liping, Wu Jing
Department of Clinical Laboratory, The Second Hospital of Shandong University, Jinan, People's Republic of China.
Department of Physiology and Pathophysiology, School of Basic Medicine, Shandong University, Jinan, People's Republic of China.
Cancer Manag Res. 2020 Mar 24;12:2185-2194. doi: 10.2147/CMAR.S234586. eCollection 2020.
Cholangiocarcinoma is the second most common primary hepatobiliary malignancy with high incidence and recurrence rate. Ubiquitin-specific protease 8 (USP8) is recently reported to be involved in tumor progression. Herein, we aimed to investigate the effects of USP8 on the growth and metastasis abilities of cholangiocarcinoma cells.
The siRNA interference was used to knock down USP8 in cholangiocarcinoma cell lines QBC939 and RBE; Hucct-1 cells were transfected with pcDNA3.1-USP8 to up-regulate its expression. The effects of USP8 on cholangiocarcinoma were detected by cell function assays. We analyzed the expressions of USP8, Bcl2, Bax, cleaved caspase-3, cleaved caspase-9, Akt, p-Akt, Cyclin D1 and P70S6K by Western blot analysis.
We demonstrated that knockdown of USP8 significantly inhibited the proliferation, migration and invasion of QBC939 and RBE cells in vitro, while USP8 overexpression showed significant promoting effects on Hucct-1 cells. Moreover, silencing of USP8 also promoted apoptosis in cholangiocarcinoma cells by regulating the Bcl-2/Bax axis and Caspase cascade; up-regulation of USP8 decreased apoptosis in Hucct-1 cells. Importantly, knockdown of USP8 inhibited activation of the Akt signaling pathway by decreasing the phosphorylation level of Akt and up-regulated p53 expression, while USP8 overexpression increased activation of the Akt signaling pathway in Hucct-1 cells. Further, IGF-1 could reverse the inhibitory effects of USP8 knockdown on the Akt signaling pathway and the proliferation of QBC939 and RBE cells.
Taken together, our findings suggest that USP8 exerts an oncogenic role in the progression of cholangiocarcinoma and may be a potential therapeutic target for cholangiocarcinoma treatment.
胆管癌是第二常见的原发性肝胆恶性肿瘤，发病率和复发率都很高。泛素特异性蛋白酶8（USP8）最近被报道参与肿瘤进展。在此，我们旨在研究USP8对胆管癌细胞生长和转移能力的影响。
采用小干扰RNA（siRNA）干扰技术敲低胆管癌细胞系QBC939和RBE中的USP8；用pcDNA3.1-USP8转染Hucct-1细胞以上调其表达。通过细胞功能实验检测USP8对胆管癌的影响。采用蛋白质免疫印迹法分析USP8、Bcl2、Bax、裂解的caspase-3、裂解的caspase-9、Akt、p-Akt、细胞周期蛋白D1和P70S6K的表达。
我们证明，敲低USP8可显著抑制QBC939和RBE细胞在体外的增殖、迁移和侵袭，而USP8过表达对Hucct-1细胞有显著的促进作用。此外，沉默USP8还通过调节Bcl-2/Bax轴和半胱天冬酶级联反应促进胆管癌细胞凋亡；USP8上调可降低Hucct-1细胞的凋亡。重要的是，敲低USP8通过降低Akt的磷酸化水平抑制Akt信号通路的激活并上调p53表达，而USP8过表达增加Hucct-1细胞中Akt信号通路的激活。此外，胰岛素样生长因子-1（IGF-1）可逆转敲低USP8对Akt信号通路以及QBC939和RBE细胞增殖的抑制作用。
综上所述，我们的研究结果表明USP8在胆管癌进展中发挥致癌作用，可能是胆管癌治疗的潜在靶点。
