沉默APN对胶质瘤细胞U251增殖、侵袭和凋亡的影响  

Effects of silencing APN gene on proliferation,invasion and apoptosis of human glioma U251 cells

作　　者：董云朋 程远[1] 陈晋[1] DONG Yunpeng;CHENG Yuan;CHEN Jin(Department of Neurosurgery,the Second Affiliated Hospital of Chongqing Medical University,Chongqing,400010,China)

机构地区：[1]重庆医科大学附属第二医院神经外科,重庆400010

出　　处：《第三军医大学学报》2020年第9期908-914,共7页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81401505)。

摘　　要：目的探究aminopeptidase N(APN)对胶质瘤细胞U251增殖能力、迁移侵袭能力和凋亡等生物学行为的影响及可能的作用机制。方法通过RNA干扰技术沉默胶质瘤细胞U251中APN的表达后,运用RT-qPCR的方法检测APN mRNA表达,采用Western blot检测APN蛋白表达水平,CCK-8检测各组U251细胞的增殖活性;采用流式细胞仪检测细胞凋亡和细胞周期;采用细胞划痕实验检测U251细胞的迁移能力,Transwell实验检测细胞侵袭能力;采用Western blot检测STAT3通路蛋白t-STAT3、p-STAT3、Bcl-2和MMP-2的蛋白表达水平。结果与空白对照组和阴性对照组相比,沉默U251中APN的表达后APN在基因和蛋白表达水平下降(P<0.01),沉默APN后U251的增殖活性明显下降(P<0.05),细胞周期停滞在G0/G1期,并且凋亡率明显增加(P<0.01);另外沉默APN的表达后U251细胞的迁移能力和侵袭能力明显下降(P<0.01);沉默APN后STAT3磷酸化水平下降,引起下游蛋白Bcl-2和MMP-2等蛋白表达下降(P<0.05)。结论沉默APN可通过STAT3信号途径抑制胶质瘤细胞U251增殖、迁移和侵袭,诱导凋亡。Objective To determine the effect of aminopeptidase N(APN)on the proliferation,invasion,and apoptosis in human glioma U251 cells and to investigate the possible mechanism.Methods After APN gene expression was silenced in glioma cells U251 by RNAi,its expression at mRNA and protein levels was detected by RT-qPCR and Western blotting,respectively.Then,CCK-8 assay,flow cytometry,wound healing test and transwell assay were used to detect the proliferation ability,cell cycle and apoptosis,migration and invasion activity in the U251 cells.The protein expression of STAT3 signal pathway proteins t-STAT3,p-STAT3,MMP-2 and Bcl-2 was measured by Western blotting.Results Compared with blank control and NC-siRNA cells,the expression of APN was decreased at mRNA and protein levels in APN-siRNA cells(P<0.01).Silencing of APN gene also resulted in inhibited proliferation ability(P<0.05),cell cycle arrested at G0/G1 phase,enhanced apoptotic rate(P<0.01),and decreased migration and invasion abilities(P<0.01)in U251 cells.What’s more,the inhibited expression of p-STAT3 caused the decreased expression of Bcl-2 and MMP-2(all P<0.05).Conclusion Silencing of APN could inhibit the proliferation,migration and invasion,and induce apoptosis in glioma U251 cells via STAT3 signaling pathway.

关 键 词：胶质瘤 AMINOPEPTIDASE N RNA干扰 STAT3 细胞凋亡 侵袭 

分 类 号：R394.3[医药卫生—医学遗传学] R730.23[医药卫生—基础医学]