Glut-1 shRNA及诱导自噬对HEp-2细胞的增殖、凋亡和迁移的影响

Glut-1 shRNA and induced autophagy inhibits cell proliferation,apoptosis and migration of laryngeal carcino⁃ma HEp-2 cells

目的葡萄糖转运蛋白-1(Glut-1)短发夹RNA(shRNA)及诱导自噬是否可以影响喉癌HEp-2细胞生长、迁移。方法构建Glut-1 shRNA稳转株(HEp-2细胞),分为Control组、阴性对照(NC)组、Glut-1 shRNA组、NC+雷帕霉素(RAP)组、Glut-1 shRNA+RAP组、NC+过表达(Lenti)-Beclin1组、Glut-1 shRNA+Lenti-Beclin1组等七组;采用CCK-8、流式细胞仪、Transwell实验分别检测Glut-1 shRNA、自噬诱导剂RAP、Beclin-1过表达载体对喉癌HEp-2细胞增殖、凋亡、迁移的影响。结果CCK-8结果显示:72 h后,与NC组比较,Glut-1 shRNA组、NC+RAP组和NC+Lenti-Beclin1组的细胞均明显减少(t分别=26.00、24.74、27.57,P均<0.05);与Glut-1 shRNA组比较,Glut-1 shRNA+RAP组、Glut-1 shRNA+Lenti-Beclin1组的细胞均明显减少(t分别=12.60、14.99,P均<0.05);与NC+RAP组比较,Glut-1 shRNA+RAP组的细胞明显减少(t=9.85,P<0.05);与NC+Lenti-Beclin1组比较,Glut-1 shRNA+Lenti-Beclin1组的细胞明显减少(t=12.80,P<0.05)。细胞流式检测显示:与NC组比较,Glut-1 shRNA组、NC+RAP组和NC+Lenti-Beclin1组细胞凋亡率明显增多(t分别=10.12、8.89、8.51,P均<0.05)。与Glut-1 shRNA组比较,Glut-1 shRNA+RAP组、Glut-1 shRNA+Lenti-Beclin1组细胞凋亡率明显增多(t分别=21.01、28.13,P均<0.05)。与NC+RAP组比较,Glut-1 shRNA+RAP组的细胞凋亡率明显增多(t=15.67,P<0.05)。与NC+Lenti-Beclin1组比较,Glut-1 shRNA+Lenti-Beclin1组的细胞凋亡率明显增多(t=18.78,P<0.05)。Transwell实验结果显示:与Glut-1 shRNA组比较,Glut-1 shRNA+RAP组、Glut-1 shRNA+Lenti-Beclin1组细胞数明显减少(t分别=10.71、9.53,P均<0.05);与NC+RAP组比较,Glut-1 shRNA+RAP组的细胞数减少(t=11.31,P<0.05);与NC+Lenti-Beclin1组比较,Glut-1 shRNA+Lenti-Beclin1组的细胞数明显减少(t=11.73,P<0.05)。结论Glut-1 shRNA及诱导自噬可以抑制HEp-2细胞的增殖、促进凋亡、降低迁移。

Objective To investigate whether Glut-1 shRNA and induced autophagy can affect the growth and migration of laryngeal carcinoma HEp-2 cells.Methods Stable HEp-2 cells transferred with shRNA Glut-1 were constructed.The experimental groups were divided into seven groups:control group,negative control groups(NC),Glut-1 shRNA group,NC+Rapamycin(RAP)group,Glut-1shRNA+RAP group,NC+Lenti-Beclin1 group,Glut-1 shRNA+RAP group,and Glut-1 shRNA+Lenti-Beclin1 group.CCK-8,flow cytometry and Transwell methods were used to detect the effect of Glut-1 shRNA,RAP and overexpression of Beclin1 on the growth,apoptosis and migration of laryngeal carcinoma HEp-2 cells,respectively.Results The results of CCK-8 showed that cell proliferation of Glut-1 shRNA or NC+RAP group or and NC+Lenti-Beclin1 group was inhibited compared to NC group at 72h(t=26.00,24.74,27.57,P<0.05).Compared to Glut-1 shRNA,the cell proliferation of HEp-2 cells of Glut-1 shRNA+RAP group and Glut-1 shRNA+Lenti-Beclin1 group was significantly inhibited(t=12.60,14.99,P<0.05).Compared to NC+RAP group,the cell proliferation of HEp-2 cells of Glut-1 shRNA+RAP group was significantly inhibited(t=9.85,P<0.05).Compared to NC+Lenti-Beclin1 group,the cell proliferation of HEp-2 cells of Glut-1 shRNA+Lenti-Beclin1 group was significantly inhibited(t=12.80,P<0.05).The results of flow cytometry showed that apoptosis of HEp-2 cells of Glut-1 shRNA group,NC+RAP group and NC+Lenti-Beclin1 group was significantly increased compared to NC group,respectively(t=10.12,8.89,8.51,P<0.05).Compared to Glut-1 shRNA group,the apoptosis rate of Glut-1 shRNA+RAP group and Glut-1 shRNA+Lenti-Beclin1 group was significantly increased(t=21.01,28.13,P<0.05).Compared to NC+RAP group,the apoptosis rate of Glut-1 shRNA+RAP group was significantly increased(t=15.67,P<0.05).Compared to NC+Lenti-Beclin1 group,the apoptosis rate of Glut-1 shRNA+Lenti-Beclin1 group increased significantly(t=18.78,P<0.05).The results of Transwell showed that the number of cells migration of Glut-1 shRNA+RAP group and Glut-1 shRNA+Lenti-Beclin1 group was significantly decreased compared to Glut-1 shRNA group(t=10.71,9.53,P<0.05).Compared with NC+RAP group,the number of cells migration of Glut-1 shRNA+RAP group was decreased(t=11.31,P<0.05).Compared with NC+Lenti-Beclin1 group,the number of cells migration of Glut-1 shRNA+Lenti-Beclin1 group was significantly decreased(t=11.73,P<0.05).Conclusion Glut-1 shRNA and induced autophagy can inhibit the proliferation of HEp-2 cells,promote apoptosis and reduce migration.