Zou Yuan-Hang, Li Xue-Dong, Zhang Qi-Hao, Liu De-Zhong
Department of Hepatobiliary Surgery, The First Affiliated Hospital of Shantou University Medical College, Shantou, 515041, People's Republic of China.
Department of Orthopedics, The First Affiliated Hospital of Shantou University Medical College, Shantou, 515041, People's Republic of China.
Pathol Oncol Res. 2018 Jan;24(1):101-107. doi: 10.1007/s12253-017-0214-6. Epub 2017 Apr 10.
This study aimed to explore the effects of RACK1 gene silencing on the apoptosis and proliferation of hepatocellular carcinoma (HCC) MHCC97-H cells. After transfecting MHCC97-H cells with siRNA, RACK1 gene silencing model was established. The cells were divided into blank group, siRNA group and empty plasmid group, respectively. The mRNA and protein expressions of RACK1, cyclin D1 and BAX were determined by qRT-PCR and Western blotting. CCK-8 assay, flow cytometry and FITC-Annexin V/PI staining were used to determine cell viability, cell cycle and cell apoptosis, respectively. The results of qRT-PCR and Western blotting suggested that when compared with the blank group and the empty plasmid group, the mRNA and protein expressions of RACK1 and Cyclin D1 decreased significantly while the mRNA and protein BAX expressions increased substantially in the siRNA group (all P < 0.05). The results of CCK-8 assay revealed that the siRNA group exhibited significantly lower cell viability when compared with the blank group and the empty plasmid group (both P < 0.05); and the cell viability in the siRNA group decreased gradually with the increase of time. The results of flow cytometry and FITC-Annexin V/PI staining indicated that when compared with the blank group and the empty plasmid group, the proportion of cells in S phase was markedly lower and the apoptosis rate was significantly higher in the siRNA group (both P < 0.05). Our study suggests that inhibition of RACK1 could suppress cell proliferation and induce apoptosis in HCC MHCC97-H cells.
本研究旨在探讨RACK1基因沉默对肝癌MHCC97-H细胞凋亡和增殖的影响。用小干扰RNA（siRNA）转染MHCC97-H细胞后，建立RACK1基因沉默模型。细胞分别分为空白组、siRNA组和空质粒组。采用实时荧光定量聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测RACK1、细胞周期蛋白D1（Cyclin D1）和凋亡相关蛋白（BAX）的mRNA和蛋白表达。分别采用细胞计数试剂盒-8（CCK-8）法、流式细胞术和异硫氰酸荧光素标记的膜联蛋白V/碘化丙啶（FITC-Annexin V/PI）染色法检测细胞活力、细胞周期和细胞凋亡。qRT-PCR和蛋白质免疫印迹结果显示，与空白组和空质粒组相比，siRNA组RACK1和Cyclin D1的mRNA和蛋白表达显著降低，而BAX的mRNA和蛋白表达显著升高（均P<0.05）。CCK-8法检测结果显示，与空白组和空质粒组相比，siRNA组细胞活力显著降低（均P<0.05）；且siRNA组细胞活力随时间延长逐渐降低。流式细胞术和FITC-Annexin V/PI染色结果表明，与空白组和空质粒组相比，siRNA组S期细胞比例明显降低，凋亡率显著升高（均P<0.05）。本研究提示，抑制RACK1可抑制肝癌MHCC97-H细胞的增殖并诱导其凋亡。
