Gong Shu, Tao Zhonghua, Liu Xiaoyan, Gan Lin
Research Centre for Preclinical Medicine, Luzhou Medical College, Luzhou, Sichuan 646000, P.R. China.
Biomed Rep. 2014 Jan;2(1):85-88. doi: 10.3892/br.2013.197. Epub 2013 Nov 6.
Recent studies have reported the association between the expression of oncoprotein 18 (op18) and hepatocellular carcinoma (HCC). However, any underlying mechanistic connection between op18 expression and hepatocarcinogenesis is poorly understood. In the present study, Flag-pcDNA3.1 vector and Flag-pcDNA3.1-op18 plasmid were stably transfected in SMMC7721 cells, respectively. Stable SMMC7721 control and op18 overexpression SMMC7721 cell lines were constructed and identified by western blot analysis. Using a cell counting kit-8 (CCK8), it was shown that cell proliferation was significantly increased in the op18 overexpression SMMC7721 cell group (0.60±0.05), compared with the control group (0.29±0.03) at an absorbance of 450 nm (P<0.01). Flow cytometry was used to analyze cell apoptosis by FITC-Annexin V and propidium iodide (PI) apoptosis assay kit. The results demonstrated that the percentage of apoptotic cells was inhibited to 5.80±0.33% in the op18 overexpression group, compared with 11.79±1.09% in the control group. Using FACS, single cell analysis data showed that op18 overexpression induced cell cycle arrest by inhibiting progression from G2 to M phase. The results suggest that op18 expression is closely associated with SMMC7721 cell proliferation and apoptosis, which appears to be a potential predictor of prognosis in HCC.
最近的研究报道了癌蛋白18(op18)的表达与肝细胞癌(HCC)之间的关联。然而,op18表达与肝癌发生之间任何潜在的机制联系却知之甚少。在本研究中,Flag-pcDNA3.1载体和Flag-pcDNA3.1-op18质粒分别稳定转染至SMMC7721细胞中。构建了稳定的SMMC7721对照细胞系和op18过表达SMMC7721细胞系,并通过蛋白质免疫印迹分析进行鉴定。使用细胞计数试剂盒-8(CCK8)检测发现,在450nm吸光度下,op18过表达的SMMC7721细胞组(0.60±0.05)的细胞增殖明显高于对照组(0.29±0.03)(P<0.01)。采用FITC-Annexin V和碘化丙啶(PI)凋亡检测试剂盒,通过流式细胞术分析细胞凋亡情况。结果显示,op18过表达组的凋亡细胞百分比被抑制至5.80±0.33%,而对照组为11.79±1.09%。利用荧光激活细胞分选技术(FACS)进行单细胞分析数据表明,op18过表达通过抑制从G2期到M期的进程诱导细胞周期停滞。结果表明,op18表达与SMMC7721细胞的增殖和凋亡密切相关,这似乎是肝癌预后的一个潜在预测指标。