从GATA2、GATA6探究环状RNA调控胃癌自噬和凋亡的机制

Mechanism of autophagy and apoptosis regulated by cyclic RNA in gastric cancer from GATA2GATA6

目的通过GATA2、GATA6探究环状RNA调控胃癌自噬和凋亡的机制。方法以人胃癌SGC-7901细胞(中国科学院上海细胞生物学研究所)为研究对象,建立干扰RNA转染模型,通过流式细胞术检测细胞周期改变、膜联蛋白V-异硫氰酸荧光素(Annexin V-FITC)/碘化丙锭(PI)双染检测细胞凋亡、倒置相差显微镜、透射电镜观察细胞形态,并应用MDC染色对细胞自噬进行测定。结果反转录-聚合酶链反应(RT-PCR)以及蛋白质印迹法(Western blot)实验结果表明,胃癌细胞株GATA2与GATA6的mRNA与蛋白质的表达水平(0.84±0.18)、(0.87±0.13)、(0.85±0.19)、(0.87±0.11)均高于胃黏膜细胞的(0.21±0.03)、(0.23±0.05)、(0.19±0.01)、(0.18±0.06)。与人胃黏膜细胞GES1比较,胃癌细胞株中GATA2与GATA6表达水平均显著上调,表明GATA2与GATA6在胃癌中具有较高表达水平。对自噬相关蛋白的检测发现,GATA2、GATA6干扰后胃癌细胞自噬水平显著降低,分别为(0.87±0.03)、(0.84±0.05)(P<0.05),而透射电镜发现,GATA2、GATA6水平降低后,胃癌细胞中自噬小体随之减少。结论干扰GATA2/GATA6表达可以降低胃癌细胞自噬水平,促进细胞凋亡,从而影响胃癌细胞的增殖能力。

Objective To explore the mechanism of cyclic RNA regulating autophagy and apoptosis in gastric cancer by GATA2 and GATA6.Methods Human gastric cancer SGC-7901 cells were transfected with interfering RNA.Cell apoptosis,inverted phase contrast microscopy and transmission electron microscopy were detected by flow cytometry,Annexin V-fluoresceine isothiocyanate(FITC)/propidium iodide(PI)double staining,and autophagy was detected by MDC staining.Results The results of reverse transcriptase-polymerase chain reaction(RT-PCR)and Western blotting showed that the expression levels of mRNA and protein in gastric cancer cell lines GATA2 and GATA6 were(0.84±0.18),(0.87±0.13),(0.85±0.19),(0.87±0.11),which was higher than those in gastric mucosa cells of(0.21±0.03),(0.23±0.05),(0.19±0.01),(0.18±0.06).Compared with human gastric mucosal cell GES1,the expression levels of GATA2 and GATA6 in gastric cancer cell lines were significantly up-regulated,indicating that GATA2 and GATA6 had higher expression levels in gastric cancer.Autophagy-related proteins were detected in gastric cancer cells after GATA2 and GATA6 interference,which was(0.87±0.03),(0.84±0.05,P<0.05).Transmission electron microscopy(TEM)showed that the levels of GATA2 and GATA6 decreased,and autophagy bodies in gastric cancer cells decreased accordingly.Conclusion Interference with GATA2/GATA6 expression can reduce the level of autophagy and promote apoptosis of gastric cancer cells,thereby affecting the proliferation of gastric cancer cells.