摘要
目的探讨黑色素瘤缺乏因子2(AIM2)对胰腺癌恶性细胞生物学行为的影响及其作用机制。方法通过慢病毒转染技术构建AIM2稳定过表达的胰腺癌细胞株BxPC-3和PANC-1, 并分别通过定量反转录聚合酶连锁反应(qRT-PCR)和蛋白质印迹法(Western blot)验证AIM2的mRNA及蛋白水平。通过BrdU增殖实验、划痕实验、Transwell侵袭实验、细胞凋亡以及细胞周期实验检测过表达AIM2对胰腺癌细胞株功能的影响。通过转录组测序探究AIM2在胰腺癌中的作用机制并进行验证, 使用基因表达差异分析评估多组样本之间的差异, 组间差异采用t检验分析。结果慢病毒转染可成功构建AIM2稳定过表达的胰腺癌细胞株。BrdU增殖实验结果表明, BxPC-3和PANC-1细胞LV-AIM2-OE组的增殖能力显著低于LV-NC组(吸光度值:0.502±0.006比0.543±0.005、0.956±0.032比1.235±0.091, t=5.568、2.903, P<0.05);划痕及Transwell实验结果表明, 过表达AIM2后BxPC-3、PANC-1细胞的迁移[(51.960±4.347)%比(43.070±3.546)%、(33.380±1.562)%比(37.950±2.441)%, t=1.584、1.578, P>0.05]及侵袭能力[(66.730±2.588)个比(60.800±1.506)个、(119.900±4.410)个比(112.900±2.461)个, t=1.981、1.399, P>0.05]不受影响;流式细胞术分析细胞凋亡及细胞周期显示, BxPC-3、PANC-1细胞株LV-AIM2-OE组的早期凋亡[(6.073±0.206)%比(2.825±0.111)%、(5.230±0.247)%比(4.165±0.385)%, t=13.890、2.425, P<0.05)]以及总凋亡[(16.420±0.662)%比(10.650±0.169)%、(25.920±0.829)%比(20.710±2.697)%, t=8.441、2.116, P<0.05]比例均显著高于LV-NC组, 并且AIM2过表达后可改变细胞周期分布。转录组测序(RNA-Seq)结果显示, BxPC-3和PANC-1细胞LV-AIM2-OE组中蛋白激酶B(Akt)3基因表达水平显著下调;蛋白印迹结果显示, LV-AIM2-OE组的磷酸化Akt(p-Akt)蛋白水平显著低于LV-NC组(0.647±0.027比1.451±0.030、0.408±0.019比0.978±0.026, t=19.810、17.650, P<0.001), 并且人第10号染色体同源丢失性磷酸酶张力蛋白基�
Objective To explore the effect of absent in melanoma 2(AIM2)on the biological function of pancreatic cancer cells and its potential mechanism to provide a new idea for molecular targeted therapy of pancreatic cancer.Methods The pancreatic cell strains overexpressing AIM2 were constructed by lentiviral transfection technology,and the RNA and protein levels of AIM2 were verified by quantitative real-time polymerase chain reaction(qRT-PCR)and Western blotting respectively.The BrdU proliferation experiment,Wound healing experiment,Transwell experiment,cell apoptosis,and cell cycle test were used to detect the difference in biological function in the pancreatic cancer cell lines between the AIM2 overexpression group(LV-AIM2-OE)and control group(LV-NC).Transcriptome sequencing was used to further compare and verify the gene expression differences between AIM2 overexpressing pancreatic cancer cells and the control group.The differences between groups were analyzed by t test.Results The results of cell function experiments showed that the proliferation ability of BxPC-3 and PANC-1 cells in the LV-AIM2-OE group was significantly lower than that in the LV-NC group(absorbance value:0.502±0.006 vs.0.543±0.005,0.956±0.032 vs.1.235±0.091,t=5.568,2.903,P<0.05).Wound healing and Transwell experiments showed that the migration ability of BXPC-3,PANC-1 cells after expressing AIM2[(51.960±4.347)%vs.(43.070±3.546)%,(33.380±1.562)%vs.(37.950±2.441)%,t=1.584,1.578,P>0.05]and invasion ability[66.730±2.588 vs.60.800±1.506,119.900±4.410 vs.112.900±2.461,t=1.981,1.399,P>0.05]were not affected.Flow cytometry analysis of apoptosis and cell cycle showed that the proportion of early apoptosis[(6.073±0.206)%vs.(2.825±0.111)%,(5.230±0.247)%vs.(4.165±0.385)%,t=13.890,2.425,P<0.05]and total apoptosis[(16.420±0.662)%vs.(10.650±0.169)%,(25.920±0.829)%vs.(20.710±2.697)%,t=8.441,2.116,P<0.05]of BxPC-3 and PANC-1 cell lines in the LV-AIM2-OE group was significantly higher than that in the LV-NC group,and the overexpression of AIM2
作者
郑盼盼
郑晓
蒋敬庭
Zheng Panpan;Zheng Xiao;Jiang Jingting(Department of Tumor Biological Treatment,the Third Affiliated Hospital of Soochow University Jiangsu Engineering Research Center for Tumor Immunotherapy Institute of Cell Therapy,Soochow University,Changzhou 213003,China)
出处
《中华实验外科杂志》
CAS
北大核心
2023年第8期1562-1565,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金资助项目(81972869、32270955)
江苏省中医药科技发展计划专题研究项目(ZT202115)。
关键词
黑色素瘤缺乏因子2
胰腺癌
生物功能
凋亡
Absent in melanoma 2
Pancreatic cancer
Biological function
Apoptosis
