AEG1促进肝癌细胞HepG2分泌细胞因子IL-6、IL-1β的实验研究

AEG1 promotes the secretion of IL-6 and IL-1β in liver cancer HepG2 cells

目的研究AEG1能否促进肝癌细胞HepG2分泌细胞因子IL-6、IL-1β,探讨AEG1是否参与改变肿瘤微环境。方法用脂质体稳定转染法分别转染pcDNA3.1(-)-AEG1(AEG1全长质粒)、psilencer 2.0-shAEG1(AEG1干扰质粒)至HepG2细胞,相对应以转染空质粒pcDNA3.1(-)、psilencer 2.0的HepG2细胞为对照组;采用实时荧光定量RT-PCR(qRT-PCR)、Western blotting检测AEG1 mRNA和蛋白表达变化;采用实时荧光定量RT-PCR和酶联免疫吸附实验(ELISA)检测细胞因子IL-6、IL-1β的表达和分泌。结果与转染相应空载体的阴性对照组和未转染的空白对照组相比,分别转染AEG1全长质粒/AEG1干扰质粒后,HepG2细胞中AEG1的蛋白表达明显增高/降低。转染AEG1全长质粒的HepG2细胞中IL-6、IL-1βmRNA均较对照组和空白组明显增强(P均<0.05),细胞培养介质中IL-6、IL-1β蛋白浓度也显著高于对照组(P均<0.05);转染AEG1 shRNA的HepG2细胞中IL-6、IL-1βmRNA均较对照组和空白组降低(P均<0.05),细胞培养介质中IL-6、IL-1β蛋白浓度也低于对照组(P均<0.05)。结论成功构建稳定过表达和沉默AEG1的HepG2细胞,AEG1能调控IL-6、IL-1β的表达和分泌。

Objective To investigate whether AEG1 could promote the secretion of inflammatory cytokine IL-6 and IL-1β in liver cancer HepG2 ceils. Methods The pcDNA3.1 （ - ）-AEG1 and psileneer 2.0-shAEG1 plasmids were transfected into HepG2 cells to get stably AEGl-overexpression and knockdown cells. Expression of AEG1 mRNA and protein were detected by qRT-PCR and Western blotting. The expression of IL-6 and IL-1β mRNA were assayed by qRT-PCR and the concentrations of IL-6 and IL-1β in supernatants were detected by ELISA. Results Compared with empty vector control and blank control, AEG1 protein was increased in HepG2 cells transfected with pcDNA 3.1 （ - ）- AEG1 and decreased in cells transfected with psilencer 2.0-shAEG1. The expression of IL-6 and IL-1β mRNA were in- creased in AEGl-overexpression HepG2 cells and decreased in AEGl-knockdown cells. The secretion of IL-6 and IL-1β were increased in supernatant of AEGl-overexpression cells and decreased in AEGl-knockdown cells. Conclusion The AEGl-overexpression HepG2 cells and AEGl-knockdown HepG2 ceils were established and detected by Western blot- ting. AEG1 could promote the secretion of inflammatory cytokine IL-6 and IL-1β in liver cancer HepG2 ceils.