摘要
目的:研究长链非编码RNA(long noncoding RNA,lncRNA)PAPPA-AS1在人肺腺癌细胞铂类耐药中的作用,并初步探讨其可能的调控机制。方法:采用RNA测序(RNA-seq)技术检测人肺腺癌细胞A549及其铂类耐药细胞中lncRNA和mRNA的差异表达谱;qRT-PCR检测lncRNA PAPPA-AS1在A549/DDP和A549/NDP细胞中的表达水平;利用siRNA技术下调铂类耐药人肺癌细胞中PAPPA-AS1的表达,CCK-8实验检测肺腺癌细胞对铂类药物的敏感性;克隆形成实验检测细胞的增殖能力;流式细胞术检测细胞凋亡分布;蛋白质印迹法检测Cleaved Caspase-3、Cleaved Caspase-9、p-PI3K、p-Akt、p-mTOR蛋白在A549/DDP和A549/NDP细胞中的表达水平。结果:与亲代敏感细胞A549相比,lncRNA PAPPA-AS1在A549/DDP和A549/NDP细胞中呈高表达;与对照组比较,si-PAPPA-AS1组的肺腺癌细胞对铂类药物的敏感性增强,细胞增殖活力下降,细胞凋亡率增加(P均<0.05);与对照组比较,si-PAPPA-AS1组的细胞中Cleaved Caspase-3和Cleaved Caspase-9蛋白表达明显升高,磷酸化PI3K、Akt和mTOR蛋白表达明显降低。结论:下调PAPPA-AS1可以通过抑制PI3K/Akt信号通路逆转肺腺癌细胞对铂类药物的抵抗性。
Objective:This study aims to investigate the role of lncRNA PAPPA-AS1 in platinum-resistant cells of lung adenocarcinoma(LAD)and to explore its possible regulatory mechanism.Methods:The differential expression profiling of lncRNA and mRNA in human lung adenocarcinoma(LAD)parental A549 cells and platinum-resistant cells were detected by RNA-seq;the expression of lncRNA PAPPA-AS1 in A549/DDP and A549/NDP cells were detected by fluorescence quantitative PCR;After using siRNA to down-regulate the expression of PAPPA-AS1 in platinum-resistant lung adenocarcinoma cells,the Chemosensitivity of LAD cells to platinumdrugs was detected by CCK-8 assay;the proliferation ability of platinum-resistant cells was detected by cloning formation assay;flow cytometry was used to detect the apoptosis of platinum-resistant cells;Western blotting was used to detect the expression of Cleaved Caspase-3,Cleaved Caspase-9,p-PI3K,p-Akt and p-mTOR protein levels in A549/DDP and A549/NDP cells.Results:Compared with human LAD parental A549cells,lncRNA PAPPA-AS1 was highly expressed in A549/DDP and A549/NDP.Compared with the control group,LAD platinum-resistant cells in the si-PAPPA-AS1 group showed increased sensitivity to drugs,decreased cell proliferation,and increased apoptosis(all P<0.05).Compared with the control group,the expression of Cleaved Caspase-3 and Cleaved Caspase-9 protein levels in si-PAPPA-AS1cells were significantly up-regulated,and the expression of phosphorylated PI3K,Akt and mTOR protein levels weredown-regulated.Conclusion:Down-regulation of PAPPA-AS1 can reverse platinumresistance of LAD via inhibiting the PI3K/Akt signaling pathway.
作者
施辰
曹海霞
徐陈欣
张辉
娄芮
吴建中
马蓉
冯继锋
SHI Chen;CAO Hai-xia;XU Chen-xin;ZHANG Hui;LOU Rui;WU Jian-zhong;MA Rong;FENG Ji-feng(Department of Oncology,The Affiliated Cancer Hospital of Nanjing Medical University,Nanjing Jiangsu 210009;Research Center for Clinical Oncology Jiangsu Cancer Hospital,Nanjing Jiangsu 210009,China)
出处
《江苏大学学报（医学版）》
CAS
2020年第3期215-220,228,共7页
Journal of Jiangsu University：Medicine Edition
基金
江苏省自然科学基金资助项目(BK20141016)
江苏省医学重点学科项目(ZDXKA2016009)
江苏省卫计委科研项目(Z2018047)。
