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目的探讨乙醛脱氢酶1A1(ALDH1A1)基因表达对胃癌细胞的生物学行为影响及机制。方法通过脂质体将shRNA-ALDH1A1载体转染到胃癌MKN-45细胞,48 h后,采用RT-PCR及蛋白印迹(WB)法检测转染后MKN-45细胞中ALDH1A1蛋白及mRNA表达;四唑盐(MTT)法检测细胞活力;Transwell法检测胃癌细胞迁移及侵袭能力;WB检测细胞中基质金属蛋白酶2(MMP2)、MMP9及Wnt/β-catenin信号通路表达情况。结果shRNA-ALDH1A1转染组M KN-45细胞中ALDH1A1蛋白及mRNA表达量[分别为(0.09±0.01)、(0.08±0.01)]明显低于对照组[分别为(0.89±0.09)、(0.48±0.05)];shRNA-ALDH1A1组M KN-45细胞活力(0.40±0.04)低于对照组(0.73±0.07),细胞迁移、侵袭数目[分别为(39.78±3.24)、(42.53±4.25)个/视野]少于对照组[分别为(98.86±9.03)、(90.52±9.14)个/视野];shRNA-ALDH1A1组MKN-45细胞中MMP2、MMP9、Wnt1、Wnt5a及β-catenin表达水平[分别为(0.10±0.01)、(0.10±0.01)、(0.28±0.03)、(0.12±0.01)、(0.19±0.02)]明显低于对照组[分别为(0.49±0.04)、(0.95±0.09)、(1.29±0.12)、(0.52±0.04)、(0.56±0.05)],差异均具有统计学意义(P<0.01)。结论 ALDH1A1基因沉默可明显降低胃癌细胞MKN-45活力、抑制细胞迁移、侵袭能力,其机制可能与抑制Wnt/β-catenin信号通路有关。
Objective To investigate the effect of ALDH1A1 gene expression on the biological behavior of gastric cancer cells and its mechanism. METHODS: The shRNA-ALDH1A1 vector was transfected into gastric cancer MKN-45 cells by lipofectamine 2000. After 48 h, the expression of ALDH1A1 protein and mRNA in MKN-45 cells was detected by RT-PCR and Western blotting. The cell viability was assayed by MTT assay. The migration and invasion ability of gastric cancer cells was detected by Transwell assay. The expressions of MMP2, MMP9 and Wnt / β-catenin were detected by WB. Results The expression of ALDH1A1 protein and mRNA in shRNA-ALDH1A1 transfection group [(0.09 ± 0.01) and (0.08 ± 0.01)] were significantly lower than those in the control group [(0.89 ± 0.09) and ± 0.05)]. The viability of M KN-45 cells in shRNA-ALDH1A1 group was significantly lower than that in control group (0.40 ± 0.04 vs 0.73 ± 0.07, 39.78 ± 3.24 and 42.53 ± 4.25, respectively / Visual field] was less than that of the control group [(98.86 ± 9.03), (90.52 ± 9.14) / visual field, respectively]; the expression levels of MMP2, MMP9, Wnt1, Wnt5a and β-catenin in MKN-45 cells of shRNA-ALDH1A1 group (0.10 ± 0.01), (0.28 ± 0.03), (0.12 ± 0.01), (0.19 ± 0.02)] were significantly lower than those in the control group [(0.49 ± 0.04) and (0.95 ± 0.09, , (1.29 ± 0.12), (0.52 ± 0.04) and (0.56 ± 0.05) respectively. There were significant differences between the two groups (P <0.01). Conclusion Silencing ALDH1A1 can significantly decrease the activity of MKN-45 in gastric cancer cells, and inhibit the migration and invasion of gastric cancer cells. The mechanism may be related to the inhibition of Wnt / β-catenin signaling pathway.