摘要
目的探讨RNAi沉默中性粒细胞相关载脂蛋白(neutrophil gelatinase-associated lipocalin,NGAL)基因表达对胃癌细胞增殖凋亡的影响及机制。方法提取胃癌组织及相应癌旁组织中的总RNA及总蛋白,RT-PCR及Western blotting检测NGAL基因的mRNA及蛋白表达;将NC-siRNA组、NGAL-siRNA转染生长至对数期的人胃癌BGC823细胞,未转染任何siRNA的作为对照组,48 h后收集细胞,Western blotting检测NGAL、Ki67、PCNA、Cleaved caspase3、β-catenin、Cyclin D1蛋白表达;CCK8实验检测细胞增殖;流式细胞术检测细胞凋亡。结果胃癌组织中NGAL基因的mRNA及蛋白表达均显著高于癌旁组织(P<0.01);NC-siRNA组NGAL的蛋白表达与对照组比较,差异无统计学意义(P>0.05),NGAL-siRNA组NGAL的蛋白表达显著低于对照组(P<0.01);与对照组及NC-siRNA组比较,NGAL-siRNA组细胞存活率及Ki67、PCNA、β-catenin、Cyclin D1蛋白表达显著降低,细胞凋亡率及Cleaved caspase3蛋白表达显著升高(P<0.01)。结论 NGAL基因在胃癌中高表达,RNA干扰其表达后可降低人胃癌BGC823细胞增殖及促进细胞凋亡,其机制与抑制Wnt/β-catenin信号通路有关。
Objective To investigate the effect and its mechanism of RNAi silencing NGAL gene expression on the proliferation and apoptosis of gastric cancer cells. Methods Total RNA and total protein were extracted from gastric cancer and paracancerous tissues, and expressions of NGAL gene mRNA and protein were detected by RT-PCR and Western blotting ; NC-siRNA, NGAL-siRNA were transfected into the logarithmic phase of growth of human gastric cancer BGC823 cells, and not transfeeted with any siRNA as the control group, cells were collected transfected for 48 hours, the expressions of NGAL, Ki67, PCNA, Cleaved caspase3, β-catenin, Cyclin D1 protein were detected by Western blotting; cell proliferation was detected by CCK8 assay; cell apoptosis was analyzed by flow cytometry. Results The expressions of NGAL gene mRNA and protein in gastric cancer tissues were significantly higher than those in adjacent tissues （P 〈 0.01 ） ; the expression of NGAL protein in NC-siRNA group and control group had no significant difference （P 〉 0.05） , the expression of NGAL protein in NGAL-siRNA group was significantly lower than that in the control group （P 〈 0.01 ） ; compared with the control group and NC-siRNA group, the cell survival rate and expressions of Ki67, PCNA, β-catenin, Cyclin D1 protein in NGAL-siRNA group were significantly decreased, cell apoptosis rate and Cleaved caspase3 protein were significantly increased （P 〈0.01）. Conclusion NGAL gene is highly expressed in gastric cancer, and RNA interference NGAL expression can decrease the proliferation and promote apoptosis of human gastric cancer cell line BGC823, which may be related to the inhibition of Wnt/β-eatenin signaling pathway.
出处
《胃肠病学和肝病学杂志》
CAS
2017年第12期1360-1364,共5页
Chinese Journal of Gastroenterology and Hepatology
