摘要
目的:探讨环状RNA MRPS35(circMRPS35)对胃癌(GC)细胞增殖、凋亡、迁移和侵袭的调控机制。方法:体外培养人GC细胞系(HGC-27、MGC-803、MKN45和AGS)和正常胃上皮GES-1细胞,实时荧光定量PCR (RT-qPCR)检测circMRPS35、mi R-130a-3p和锌环指蛋白3 (ZNRF3)m RNA表达。另取MGC-803细胞,分为对照组、pc-NC组、pc-circMRPS35组、pc-circMRPS35+miR-NC组、pc-circMRPS35+miR-130a-3p组,采用Lipofectamine 3000进行质粒转染。RT-qPCR检测circMRPS35、mi R-130a-3p和ZNRF3 m RNA表达,Western blot检测ZNRF3蛋白表达,CCK-8法、流式细胞术检测细胞增殖与凋亡,划痕实验和Transwell小室实验检测细胞迁移与侵袭能力,裸鼠移植瘤实验探究circMRPS35对GC细胞体内生长的影响。双荧光素酶报告基因检测mi R-130a-3p与circMRPS35或ZNRF3的靶标关系。结果:GC细胞系中circMRPS35和ZNRF3 m RNA呈低表达,mi R-130a-3p呈高表达(均P<0.05)。过表达circMRPS35可降低mi R-130a-3p,上调ZNRF3 m RNA和蛋白水平,抑制细胞增殖、迁移和侵袭,并促进细胞凋亡(均P<0.05);circMRPS35过表达对GC细胞恶性行为和裸鼠移植瘤生长的抑制作用可被mi R-130a-3p mimic逆转(P<0.05)。双荧光素酶实验结果显示,过表达mi R-130a-3p可降低circMRPS35-WT和ZNRF3-WT的荧光素酶活性(P<0.05)。结论:circMRPS35可能通过mi R-130a-3p/ZNRF3轴抑制GC细胞的增殖、迁移和侵袭,并促进细胞凋亡。
Objective: To investigate the regulatory mechanism of circular RNA MRPS35(circMRPS35) on the proliferation,apoptosis, migration and invasion of gastric cancer(GC) cells. Methods: Human GC cell lines(HGC-27, MGC-803, MKN45 and AGS)and normal gastric epithelial GES-1 cells were cultured in vitro, real-time quantitative PCR(RT-qPCR) was used to detect the m RNA expressions of circMRPS35, mi R-130a-3p and zinc and ring finger 3(ZNRF3). MGC-803 cells were also taken and grouped into control group, pc-NC group, pc-circMRPS35 group, pc-circMRPS35 combined with mi R-NC group, and pc-circMRPS35 combined with mi R-130a-3p group. Plasmid transfection using Lipofectamine 3000. RT-qPCR was used to detect the m RNA expression of circMRPS35,mi R-130a-3p and ZNRF3. Western blot was used to detect ZNRF3 protein expression, CCK-8 method and flow cytometry were used to detect cell proliferation and apoptosis, scratch assay and Transwell chamber assay were used to detect cell migration and invasion abilities, nude mouse xenograft experiments were performed to investigate the effect of circMRPS35 on the growth of GC cells in vivo.Dual-luciferase reporter genes were implemented to detect the target relationship of mi R-130a-3p with circMRPS35 or ZNRF3. Results:In GC cell lines, the m RNA expression of circMRPS35 and ZNRF3 was low, and the expression of mi R-130a-3p was high(all P<0.05).Overexpression of circMRPS35 was able to reduce the expression of mi R-130a-3p, up-regulate the m RNA and protein levels of ZNRF3,inhibit cell proliferation, migration and invasion, and promote cell apoptosis(all P<0.05). The inhibitory effect of circMRPS35 overexpression on malignant behavior of GC cells and tumor growth in nude mice can be reversed by mi R-130a-3p mimic(P<0.05). The results of dual luciferase experiment showed that overexpression of mi R-130a-3p was able to reduce the luciferase activities of circMRPS35-WT and ZNRF3-WT(P<0.05). Conclusion: CircMRPS35 may inhibit the proliferation, migration and invasion of GC cells and promote apoptosis through the mi R-130a-3p/ZNRF3 axis.
作者
刘文飞
秦洪真
董翼
郭玉洋
胡文军
方勇
LIU Wen-fei;QIN Hong-zhen;DONG Yi;GUO Yu-yang;HU Wen-jun;FANG Yong(General Surgery,The 305 Hospital of PLA,Beijing,100017,China)
出处
《现代生物医学进展》
CAS
2022年第21期4020-4026,共7页
Progress in Modern Biomedicine
基金
北京市自然科学基金项目(71620715)
解放军第305医院院级课题(17YQ013)。
