甲基硝基亚硝基胍对人正常胃黏膜GES-1细胞恶变的影响及其机制研究

Effect of methylnitronitrosoguanidine(MNNG) on the malignant transformation of human gastric mucosa GES-1 cells and its mechanism

目的观察甲基硝基亚硝基胍(MNNG)对人正常胃黏膜GES-1细胞恶变的影响,并探讨其可能的机制。方法采用不同浓度梯度MNNG作用于人正常胃黏膜GES-1细胞,培养24h后采用CCK-8法检测各组细胞增殖情况,Transwell小室检测细胞侵袭及迁移能力;48h后采用Annexin V-FITC/PI双染法检测细胞凋亡情况,实时荧光定量PCR及Western blotting检测细胞NF-κB p65 m RNA及蛋白表达情况。结果选择诱导胃黏膜GES-1细胞恶变的MNNG浓度为2×10–4mol/L。MNNG能明显促进GES-1细胞的增殖和侵袭转移,并抑制其凋亡(P<0.01);MNNG诱导胃黏膜GES-1细胞后,细胞内NF-κB p65 m RNA及蛋白表达水平均上调(P<0.05)。结论 MNNG可促进人正常胃黏膜GES-1细胞的增殖及侵袭转移,抑制细胞凋亡,诱导细胞恶变,其机制可能与激活NF-κB有关。

Objective To investigate the effects of methylnitronitrosoguanidine （MNNG） on the malignant transformation of human gastric mucosa GES-1 cells and explore the possible molecular mechanism. Methods The GES-1 cells was treated by 9 different concentrations of MNNG for 24h, then removed MNNG and culture cell normally. The cell proliferation was detected by CCK-8 assay after 24h. The effects of MNNG on invasion and migration ability were detected by Transwell invasion and migration tests after 24h. The apoptosis was detected by Annexin V-FITC/PI FCM after 48h, and the expression levels of NF-KB p65 mRNA and protein were measured by real-time fluorescent quantitative PCR and Western blotting respectively. Results 2 x 10-4mol/L MNNG was chose for the induction of malignant transformation of GES-1 cells by the statistic analysis, 2 x 104mol/L MNNG could increase the proliferation, promote the invasion and migration, suppress the apoptosis of GES-1 cells （P〈0.01）. The expression levels of NF-KB p65 mRNA and protein were upregulated in GES-1 cells after treatment with MNNG （P〈0.05）. Conclusions MNNG can accelerate the proliferation, invasion and migration of GES-1 cells and inhibit apoptosis. This effect may be related to the activation of NF-KB.