上调XB130表达对人肝癌HepG2细胞凋亡和增殖的影响

Effect of XB130 on human hepatocellular carcinoma HepG2 cells

摘要:

目的观察XB130 表达上调后人肝癌细胞HepG2 增殖和凋亡的变化，并探讨其作用机制。方法Western blot及实时荧光定量聚合酶链反应（qRT-PCR）检测肝癌细胞株中XB130表达；构建过表达质粒pCMVEGFP-XB130 转染HepG2 细胞，实验分为pCMV-EGFP-XB130 组（pCMV-EGFP-XB130 转染），空白对照组（EGFP 转染）。以免疫荧光、Western blot法及qRT-PCR 法检测上调效率。Western blot 法检测PI3K/Akt通路相关基因表达；采用MTT法检测转染后细胞增殖，流式细胞术检测细胞凋亡。结果XB130 蛋白及其mRNA在肝癌细胞株Hep3B、Huh7、HepG2 和MHCC97H 中均有表达，在HepG2 细胞株中表达量最低（ F=6.342 和5.424，均P=0.001）。与EGFP 组比较，pCMV-EGFP-XB130 组中p-p21、p-p27、p-FOXO3a 及Pro Capase-9蛋白表达上调（t =4.652，3.558，6.743 和3.021，均P<0.05）；XB130 表达上调后在72 及96 h HepG2 细胞增殖能力增强（ t=4.752 和8.542，均P=0.001）；XB130 表达上调后HepG2 细胞凋亡率降低（t =7.467，P =0.001）。结论XB130 表达上调通过PI3K/Akt信号通路促进肝细胞癌细胞增殖，抑制凋亡。

Abstract:

Objective To investigate the effect of XB130 on proliferation and apoptosis of human hepatocellular carcinoma HepG2 cell lines. Methods Western blot and qRT-PCR were utilized to determine expression of XB130 in hepatocellular carcinoma cells. Over expressed plasmid pCMV-EGFP-XB130 was transfected into HepG2 cells. Expression of XB130 was identified by Immunofluorescence, Western blot, and qRT-PCR. PI3K/Akt pathway related proteins were measured by Western blot; proliferation and apoptosis rate were measured by MTT assay and Flow cytometry, respectively. Results XB130 protein and mRNA were detected in different hepatocellular carcinoma cell-lines including Hep3B, Huh7, HepG2 and MHCC97H while HepG2 showed the lowest concentration of XB130 (P = 0.001). p-p21, p-p27, p-FOXO3a and pro-Caspase 9 protein in pCMV-EGFP-XB130 group was up-regulated significantly when compared with EGFP control group(t = 4.652, 3.021, 3.558 and 6.743, P< 0.05); XB130 overexpression increased cellular proliferation (P = 0.001) while decreased apoptosis at the 72nd and 96th ( P= 0.001) in HepG2. Conclusion Up-regulation of XB130 increases proliferation while inhibits apoptosis via modulating PI3K/Akt signal pathway in hepatocellular carcinoma HepG2.