SAPCD2对肺腺癌A549细胞生物学功能的影响及机制研究  

Effect and mechanism of SAPCD2 on the biological function of lung adenocarcinoma A549 cells

作　　者：赵哲[1,2] 穆培娟 张冬[1] ZHAO Zhe;MU Peijuan;ZHANG Dong(Department of Respiratory and Critical care Medicine,The First Affiliated Hospital of Baotou Medical College of Inner Mongolia University of Science and Technology,Baotou,Inner Mongolia 014010,P.R.China;Baotou Medical College of Inner Mongolia University of Science and Technology,Baotou,Inner Mongolia 014010,P.R.China)

机构地区：[1]内蒙古科技大学包头医学院第一附属医院呼吸与危重症医学科,内蒙古包头014010 [2]内蒙古科技大学包头医学院,内蒙古包头014040

出　　处：《中国呼吸与危重监护杂志》2023年第7期506-514,共9页Chinese Journal of Respiratory and Critical Care Medicine

基　　金：包头医学院青年科技人才发展计划(BYJJ-DXX 2022025)。

摘　　要：目的探讨SAPCD2在肺腺癌细胞中的表达情况,研究SAPCD2调控Hippo信号通路影响肺腺癌细胞增殖、侵袭、迁移、凋亡的影响及其机制研究。方法采用实时荧光定量聚合酶链反应(quantitative real-time polymerase chain reaction,qRT-PCR)、免疫印迹法(Western blot)分别检测4种肺癌细胞(HCC827、H1650、SK-MES-1、A549)及人正常肺上皮细胞(BESA-2B)中SAPCD2 mRNA和蛋白质表达水平,然后从中选取SAPCD2表达量相对较高的肺癌细胞来进行后续实验。实验分为正常对照组(NC组)、si-SAPCD2组和通路抑制剂组(si-SAPCD2+XMU-MP-1组)。通过干扰小RNA(small interfering RNA,siRNA)技术沉默SAPCD2 mRNA,采用qRT-PCR法检测转染后肺癌细胞SAPCD2的表达,采用克隆平板实验检测沉默后肺癌细胞的增殖情况,采用流式细胞术检测沉默后的肺癌细胞的凋亡情况,再通过细胞周期实验观察肺癌细胞在不同时期细胞数量,采用Transwell实验分析沉默SAPCD2对肺癌细胞迁移及侵袭的影响,采用Western blot检测ki-67、Bcl-2、Caspase-3、NF2、P-MST1、P-LATS1、P-YAP、YAP、TAZ蛋白表达。结果SAPCD2在肺腺癌A549细胞中表达水平最高(P<0.01)。沉默SAPCD2后A549细胞的增殖能力明显下降(P<0.01),抑制了A549细胞的迁移(P<0.05)及侵袭(P<0.01),促进A549细胞凋亡(P<0.01);超过半数的细胞停留在G0/G1期,与NC组相比,A549细胞在G0/G1期的细胞明显增多(P<0.01),G2/M期及S期的细胞明显减少(P<0.01),早期凋亡的细胞比例明显增加(P<0.01)。Western blot结果显示沉默SAPCD2后与NC组相比会下调ki-67、Bcl-2、YAP、TAZ蛋白的表达(P<0.01),上调Caspase-3、NF2、P-MST1、P-LATS1、P-YAP蛋白的表达(P<0.01)。结论SAPCD2在肺腺癌A549细胞的表达较正常肺上皮细胞BESA-2B明显升高,促进A549细胞增殖、迁移及侵袭,抑制细胞凋亡,其机制可能与Hippo信号通路被抑制有关。Objective To investigate the expression of SAPCD2 in the lung adenocarcinoma cells,and to study the effect of SAPCD2 regulating Hippo signaling pathway on the proliferation,invasion,migration and apoptosis of the lung adenocarcinoma cells and its mechanism.Methods Quantitative real-time PCR(qRT-PCR)and Western blot were used to detect the expression levels of SAPCD2 mRNA and protein in four types of lung cancer cells(HCC827,H1650,SK-MES-1,A549)and human normal lung epithelial cells(BESA-2B),respectively.Then,lung cancer cells with relatively high levels of SAPCD2 expression were selected for subsequent experiments.The experiment cells were divided into a normal control group(NC group),a si-SAPCD2 group,and a pathway inhibitor group(si-SAPCD2+XMU-MP-1 group).Firstly,SAPCD2 mRNA was silenced using small interfering RNA(siRNA)technology,and then qRT-PCR was used to detect the expression of SAPCD2 in transfected lung cancer cells;using clone plate assay to detect the proliferation of lung cancer cells after silencing;using flow cytometry to detect the apoptosis of lung cancer cells after silencing;observe the number of lung cancer cells at different stages through cell cycle experiments;then Transwell experiment was used to analyze the effect of silencing SAPCD2 on the migration and invasion of lung cancer cell migration.Finally,Western blot was used to detect the expression of ki-67,Bcl-2,Caspase-3,NF2,P-MST1,P-LATS1,P-YAP,YAP,and TAZ proteins.Results SAPCD2 had the highest expression level in lung adenocarcinoma A549 cells(P<0.01).Silencing SAPCD2 significantly decreased the proliferation ability of A549 cells(P<0.01),inhibited their migration(P<0.05)and invasion(P<0.01),and promoted A549 cell apoptosis(P<0.01);more than half of the cells remained in the G0/G1 phase.Compared with the NC group,A549 cells showed a significant increase in G0/G1 phase cells(P<0.01),a significant decrease in G2/M and S phase cells(P<0.01),and a significant increase in the proportion of early apoptotic cells(P<0.01).Western blot results

关 键 词：SAPCD2 肺腺癌 增殖 迁移 侵袭 Hippo信号通路 

分 类 号：R734.2[医药卫生—肿瘤]