胃癌组织中高表达的miR-504通过TP53INP1调控胃癌细胞BGC-823的生物学行为  

Highly expressed miR-504 in gastric cancer tissues regulates the biological behaviors of gastric cancer cell BGC-823 through TP53INP1

作　　者：刘真义 翁国武[1] 关李稳 周真真 王力雅 冯红军 LIU Zhenyi;WENG Guowu;GUAN Liwen;ZHOU Zhenzhen;WANG Liya;FENG Hongjun(Department of Gastroenterology,Sanya Central Hospital&The Third People's Hospital of Hainan Province,Sanya 572000,Hainan,China)

机构地区：[1]海南省第三人民医院暨三亚中心医院消化内科,海南三亚572000

出　　处：《中国肿瘤生物治疗杂志》2021年第8期824-832,共9页Chinese Journal of Cancer Biotherapy

基　　金：海南省卫生计生行业科研项目资助(No.16A200142);三亚市医疗卫生科技创新项目资助(No.2017YW08)。

摘　　要：目的:探究微小RNA-504(miRNA-504)在胃癌(GC)组织中的表达水平及其对GC细胞生物学行为的调控机制。方法:收集2020年6月至2020年12月期间三亚中心医院外科收治的48例胃癌患者的肿瘤组织及癌旁组织标本,qPCR检测组织中miR-504、肿瘤蛋白53诱导型核蛋白1(tumor protein 53-induced nuclear protein 1,TP53INP1)mRNA的水平,WB法检测TP53INP1水平。体外培养人胃癌细胞BGC-823,分为对照组(正常培养的BGC-823细胞)、miR-504 mimic组、mimic-NC组、miR-504 inhibitor组、inhibitor-NC组、miR-504 inhibitor+si-NC组、miR-504 inhibitor+si-TP53INP1组,qPCR检测细胞中miR-504和TP53INP1 mRNA的表达,MTT法、流式细胞术、划痕实验和Transwell侵袭实验分别检测各组细胞的增殖、凋亡、迁移和侵袭能力,WB法检测各组细胞中增殖、迁移和侵袭相关蛋白(Cyclin D1、E-cadherin、MMP-2、MMP-9)以及TP53INP1的表达。双荧光素酶报告基因实验进一步验证miR-504与TP53INP1 mRNA的靶向关系。结果:与癌旁组织相比,胃癌组织中miR-504的表达显著升高(P<0.05),而TP53INP1 mRNA和蛋白表达水平显著降低(P<0.05或P<0.01),miR-504和TP53INP mRNA两者的表达呈负相关(P<0.01)。与对照组相比,miR-504 mimic组BGC-823细胞中miR-504的表达显著升高(P<0.05)、TP53INP1 mRNA和蛋白的表达显著降低(均P<0.05),且细胞增殖率、划痕愈合率、侵袭入Transwell小室下层的细胞数量,Cyclin D1、MMP-2、MMP-9蛋白表达均显著增加,细胞凋亡率和E-cadherin蛋白表达均显著降低(均P<0.05)。转染miR-504 inhibitor能显著下调BGC-823中miR-504的表达、上调TP53INP1 mRNA和蛋白的表达,抑制细胞的增殖、迁移与侵袭能力而促进细胞凋亡(均P<0.05);而下调TP53INP1的表达可明显减弱miR-504下调对BGC-823细胞增殖、迁移与侵袭的抑制作用(P<0.01)。miR-504高表达能明显抑制野生型TP53INP1质粒的荧光素酶活性(P<0.05)。结论:miR-504在胃癌组织中呈高表达,下调miR-504可抑制�Objective:To explore the expression level of microRNA-504(miRNA-504)in gastric cancer(GC)tissues and its regulatory mechanism in the biological behaviors of GC cells.Methods:From June 2020 to December 2020,tumor tissues and corresponding para-cancerous tissues resected from 48 patients with gastric cancer were collected in the Department of Surgical,Sanya Central Hospital.qPCR was used to detect the mRNA levels of miR-504 and tumor protein 53-induced nuclear protein 1(TP53INP1)in the tissues,and WB assay was used to detect the protein level of TP53INP1.Human gastric cancer BGC-823 cells were cultured in vitro and divided into control group(normally cultured BGC-823 cells),miR-504 mimic group,mimic-NC group,miR-504 inhibitor group,inhibitor-NC group,miR-504 inhibitor+si-NC group,and miR-504 inhibitor+si-TP53INP1 group.qRT-PCR was used to detect mRNA expression of miR-504 and TP53INP1 in each group of cells.MTT method,Flow cytometry,Scratch test,and Transwell invasion assay were used to detect the proliferation,apoptosis,migration,and invasion of the cells in each group.WB assay was used to detect the protein expression of cell proliferation, migration, and invasion-related proteins (Cyclin D1, E-cadherin, MMP-2, MMP-9) andTP53INP1 in each group. Dual-luciferase reporter gene experiment was used to further verify the targeting relationship between miR-504 and TP53INP1. Results: Compared with the para-cancerous tissues, the expression of miR-504 in gastric cancer tissues wassignificantly increased (P<0.05), and the mRNA and protein expression levels of TP53INP1 were significantly decreased (P<0.05 or P<0.01). The expression of miR-504 was negatively correlated with TP53INP1 mRNA (P<0.01). Compared with the control group, theexpression of miR-504 in the BGC-823 cells of the miR-504 mimic group was significantly increased, while the mRNA and proteinexpression of TP53INP1 was significantly decreased (all P<0.05);moreover, the cell survival rate, scratch healing rate, the number ofcells entering the lower layer of the T

关 键 词：胃癌 BGC-823细胞 miRNA-504 增殖 凋亡 迁移 侵袭 肿瘤蛋白53诱导型核蛋白1 

分 类 号：R735.2[医药卫生—肿瘤] R730.2[医药卫生—临床医学]