异牡荆素调节miR-107/CCND1轴对胰腺癌细胞增殖、凋亡、迁移和侵袭的影响  

Influences of isovitexin on the proliferation,apoptosis,migration and invasion of pancreatic cancer cells by regulating the miR-107/CCND1 axis

作　　者：玄鸿雁 阚璇 朱振胜 XUAN Hongyan;KAN Xuan;ZHU Zhensheng(Department of Oncology,Jinan Hospital of Integrated Chinese and Western Medicine,Shandong Jinan 271100,China)

机构地区：[1]济南市中西医结合医院肿瘤科,山东济南271100

出　　处：《现代肿瘤医学》2023年第15期2756-2764,共9页Journal of Modern Oncology

基　　金：山东省中医药科技项目(编号:Z-2022003)。

摘　　要：目的:探讨异牡荆素(IVX)对胰腺癌(PCa)细胞增殖、凋亡、迁移和侵袭的影响及分子机制。方法:体外培养人PCa细胞系(PANC-1、AsPC-1、BxPC-3、Capan-1和SW 1990)和人正常胰腺导管上皮细胞(HPDE),实时荧光定量PCR(qRT-PCR)和蛋白质印迹(Western blot)检测细胞中微小RNA-107(miR-107)和细胞周期蛋白D1(CCND1)的表达。采用MTT法检测IVX对PANC-1细胞的毒性作用;将PANC-1细胞分为正常对照(NC)组、5μmol/L IVX组、10μmol/L IVX组、20μmol/L IVX组、20μmol/L IVX+anti-NC组、20μmol/L IVX+anti-miR-107组。细胞克隆形成实验检测细胞增殖活性;流式细胞术检测细胞凋亡和细胞周期;划痕愈合实验和Transwell小室实验检测细胞迁移、侵袭;双荧光素酶报告基因检测和RNA结合蛋白免疫沉淀(RIP)实验验证PANC-1细胞中miR-107和CCND1的靶向关系;裸鼠成瘤实验、免疫组织化学(IHC)染色和qRT-PCR探究IVX对PCa细胞体内肿瘤生长和Ki-67、CCND1、miR-107表达的影响。结果:在PCa细胞系中miR-107呈低表达,CCND1呈高表达。IVX以浓度依赖性方式抑制PANC-1细胞增殖;5、10、20μmol/L IVX以浓度依赖性方式抑制细胞增殖、迁移和侵袭,诱导细胞周期停滞和细胞凋亡,上调miR-107表达,降低CCND1 mRNA和蛋白水平(均P<0.05);且下调miR-107的表达可显著减弱IVX对PCa细胞恶性生物学行为的抑制作用;双荧光素酶报告基因检测和RIP实验证实CCND1是miR-107的靶标。裸鼠移植瘤实验结果显示,IVX可显著抑制体内移植瘤生长和Ki-67、CCND1表达,并上调miR-107表达(均P<0.05)。结论:IVX可能通过上调miR-107,抑制CCND1表达,进而抑制PCa细胞的增殖、迁移和侵袭,并诱导细胞凋亡。Objective:To investigate the influences and molecular mechanism of isovitexin(IVX)on the proliferation,apoptosis,migration and invasion of pancreatic cancer(PCa)cells.Methods:Human PCa cell lines(PANC-1,AsPC-1,BxPC-3,Capan-1 and SW 1990)and human normal pancreatic ductal epithelial cells(HPDE)were cultured in vitro,the expression of microRNA-107(miR-107)and cyclin D1(CCND1)in cells was detected by quantitative real-time PCR(qRT-PCR)and Western blot.MTT assay was used to detect the toxicity of IVX on PANC-1 cells.PANC-1 cells were separated into normal control(NC)group,5μmol/L IVX group,10μmol/L IVX group,20μmol/L IVX group,20μmol/L IVX+anti-NC group and 20μmol/L IVX+anti-miR-107 group.Cell clone formation assay was performed to detect cell proliferation activity.Flow cytometry was performed to detect apoptosis and cell cycle.Scratch healing assay and Transwell chamber assay were performed to detect cell migration and invasion.The targeting relationship between miR-107 and CCND1 in PANC-1 cells was verified by dual-luciferase reporter gene detection and RNA-binding protein immunoprecipitation(RIP)experiments.Nude mouse tumorigenesis experiments,immunohistochemistry(IHC)staining and qRT-PCR were performed to explore the influences of IVX on tumor growth and expression of Ki-67,CCND1 and miR-107 in PCa cells in vivo.Results:The expression of miR-107 was low and the expression of CCND1 was high in PCa cell line.IVX inhibited the proliferation of PANC-1 cells in a concentration-dependent manner.5,10 and 20μmol/L IVX inhibited cell proliferation,migration and invasion in a concentration-dependent manner,induced cell cycle arrest and apoptosis,up-regulated miR-107 expression,and decreased CCND1 mRNA and protein levels(all P<0.05).Down-regulating the expression of miR-107 could obviously reduce the inhibitory effect of IVX on the malignant biological behaviors of PCa cells.Dual-luciferase reporter assays and RIP experiments confirmed that CCND1 was a target of miR-107.The results of nude mice xenograft experiments

关 键 词：胰腺癌 异牡荆素 微小RNA-107 细胞周期蛋白D1 增殖 凋亡 迁移 侵袭 

分 类 号：R735.9[医药卫生—肿瘤]