[1]张金刚a,齐普良a,吴 刚b,等.miR-505-3p 对胃癌细胞增殖、克隆形成及迁移、侵袭的影响及机制研究[J].现代检验医学杂志,2022,37(04):69-74+153.[doi:10.3969/j.issn.1671-7414.2022.04.014]
　ZHANG Jin-ganga,QI Pu-lianga,WU Gangb,et al.Effect and Mechanism of miR-505-3p on Proliferation, Clone Formation, Migration and Invasion of Gastric Cancer Cells[J].Journal of Modern Laboratory Medicine,2022,37(04):69-74+153.[doi:10.3969/j.issn.1671-7414.2022.04.014]
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miR-505-3p 对胃癌细胞增殖、克隆形成及迁移、侵袭的影响及机制研究()
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《现代检验医学杂志》[ISSN:/CN:]

卷:
第37卷
期数:
2022年04期
页码:
69-74+153
栏目:
论著
出版日期:
2022-07-15

文章信息/Info

Title:
Effect and Mechanism of miR-505-3p on Proliferation, Clone Formation, Migration and Invasion of Gastric Cancer Cells
文章编号:
1671-7414（2022）04-069-07
作者:
张金刚a; 齐普良a; 吴 刚b; 郭厚乐c; 王 磊d
青海省人民医院a. 急诊外科；b. 普外科；c. 肿瘤外科；d. 病理科，西宁 810000
Author(s):
ZHANG Jin-ganga;  QI Pu-lianga;  WU Gangb;  GUO Hou-lec;  WANG Leid
a.Department of Emergency Surgery；b.Department of General Surgery；c.Department of Surgical Oncology； d.Department of Pathology,Qinghai Provincial People’s Hospital, Xining 810000，China
关键词:
胃癌; 微小核糖核酸-505-3p; c-MYC; 增殖; 克隆形成; 迁移; 侵袭
分类号:
R735.2；R730.4
DOI:
10.3969/j.issn.1671-7414.2022.04.014
文献标志码:
A
摘要:
目的 检测胃癌组织及细胞中miR-505-3p 表达，并探究其对胃癌细胞增殖、克隆形成、迁移和侵袭的影响及潜在分子机制。方法 利用实时荧光定量PCR 实验（qRT-PCR）检测胃癌组织、癌旁正常组织及胃癌细胞和人正常胃黏膜细胞中miR-505-3p 相对表达；构建miR-505-3p 过表达、c-MYC 过表达和敲低表达细胞系，通过CCK-8 法，克隆斑点形成实验、Transwell 侵袭/ 迁移实验检测miR-505-3p 和c-MYC 对胃癌细胞增殖、克隆形成及迁移、侵袭的影响；裸鼠皮下成瘤实验验证miR-505-3p 对裸鼠肿瘤生长的影响；双荧光素报告实验验证miR-505-3p 和c-MYC 的靶向关系，并探究其两者间的表达调控作用；Western blot 检测miR-505-3p 和c-MYC 对Wnt/β-catenin 通路关键蛋白表达的影响。结果 临床胃癌组织中miR-505-3p 表达水平较正常癌旁组织显著下调（0.92±0.37 vs 1.74±0.59），差异有统计学意义（t=3.723，P ＜ 0.01）。胃癌细胞系MGC803（1.12±0.35），AZ521（2.31±0.24）和HGC-27（2.28±0.43）中miR-505-3p 相对表达较人正常胃黏膜细胞系GES1（4.62±0.79）明显降低，差异有统计学意义（F=26.109，P ＜ 0.001）。miR-505-3p 过表达组细胞增殖（0.92±0.27）、克隆形成（51.67±21.75）、迁移（43.25±15.47 ）、侵袭（38.53±14.76）能力较NC组（1.85±0.34，128.36±36.42，100.08±2.12，100.12±1.94）明显降低，差异均有统计学意义（t=3.131 ～ 7.166，均P ＜ 0.05）；miR-505-3p 过表达抑制了裸鼠生长。c-MYC 是miR-505-3p 的靶基因，miR-505-3p 靶向负调控c-MYC。c-MYC 过表达组细胞增殖（2.72±0.68）、迁移（147.15±20.36）、侵袭（145.46±22.73）能力较NC 组（1.85±0.34，100.08±2.12，100.12±1.94）明显增高，差异均有统计学意义（t=2.455 ～ 4.456，均P ＜ 0.05）；c-MYC 过表达可逆转miR-505-3p 对胃癌细胞增殖、迁移和侵袭的抑制作用。miR-505-3p 过表达组Wnt（0.46±0.07 ），β-catenin（0.50±0.05）蛋白相对表达水平明显低于NC 组（1.01±0.02，1.02±0.02），差异均有统计学意义（t=8.139，7.342，均P ＜ 0.001）；过表达c-MYC 能够逆转miR-505-3p 对Wnt 和β-catenin 蛋白表达的抑制作用。结论 胃癌中miR-505-3p 显著低表达，其可通过靶向调控c-MYC 表达介导Wnt/β-catenin 信号通路激活进而发挥作用参与胃癌的发生发展过程。
Abstract:
Objective The expression of miR-505-3p in gastric cancer tissues and cells was detected, and its effect on proliferation, clonogenesis, migration and invasion of gastric cancer cells and its potential molecular mechanism were explored. Methods QRT-PCR was used to detect the relative expression of miR-505-3p in gastric cancer tissues, normal adjacent tissues, gastric cancer cells and human normal gastric mucosa cells. MiR-505-3p overexpression, c-MYC overexpression and knockdown expression cell lines were constructed, and the effects of miR-505-3p and c-MYC on proliferation, clonal formation, migration and invasion of gastric cancer cells were detected by CCK-8 assay, clone spot formation assay and Transwell invasion/migration assay. The effect of miR-505-3p on tumor growth in nude mice was detected by subcutaneous tumorigenesis assay. Dual fluorescein reporting assay was used to verify the targeting relationship between miR-505-3p and c-MYC, and explored the expression regulation between the two. Western blot was used to detect the effects of miR-505-3p and c-MYC on the expression of key proteins in the Wnt/β-catenin pathway. Results The expression level of miR-505-3p in gastric cancer tissues was significantly down-regulated compared with normal adjacent tissues (0.92±0.37 vs 1.74±0.59）, the difference was statistically significant（t=3.723, P ＜ 0.01）. The relative expression of miR-505-3p in gastric cancer cell line MGC803 (1.12±0.35), AZ521 (2.31±0.24), and HGC-27 (2.28±0.43) was significantly lower than that in human normal gastric mucosa cell line GES1 (4.62±0.79)，the differences was statistically significant (F=26.109, P ＜ 0.001). MiR-505-3p overexpression group showed proliferation (0.92±0.27), clonal formation (51.67±21.75)，migration (43.25±15.47) and invasion ability (38.53±14.76 ) were significantly decreased compared with NC group（1.85±0.34，128.36±36.42，100.08±2.12，100.12±1.94），the differences were statistically significant (t=3.131 ~ 7.166, all P ＜ 0.05). MiR-505-3p overexpression inhibited the growth of nude mice. C-MYC was the target gene of miR-505-3p, and miR-505-3p negatively regulates c-MYC. C-MYC overexpression group showed proliferation (2.72±0.68), migration (147.15±20.36) and invasion (145.46±22.73 ) compared with NC group （1.85±0.34，100.08±2.12，100.12±1.94），the differences were statistically significant (t=2.455~ 4.456, all P ＜ 0.05). c-MYC overexpression can reverse the inhibitory effect of miR-505-3p on proliferation, migration and invasion of gastric cancer cells. The relative expression levels of Wnt (0.46±0.07) and β-catenin (0.50±0.05) in miR-505-3p overexpression group were significantly lower than those in NC group（1.01±0.02，1.02±0.02）, the differences were statistically significant (t=8.139，7.342, all P ＜ 0.001).Overexpression of c-MYC could reverse the inhibitory effect of miR-505-3p on the expression of Wnt and β-catenin proteins. Conclusion MiR-505-3p was significantly underexpressed in gastric cancer, which plays a role in the occurrence and development of gastric cancer through the targeted regulation of c-MYC expression mediating the activation of Wnt/β-catenin signaling pathway.

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备注/Memo

备注/Memo:
基金项目：青海省2018 年医药卫生科技项目课题（指导性项目）（No.Y2018-21）。
作者简介：张金刚（1980-），男，本科，主治医师，急诊外科，研究方向: 胰腺肿瘤，E-mail: zjgyou@126.com。
通讯作者：齐普良（1987-），男，硕士，主治医师，急诊外科，研究方向: 胰腺肿瘤，E-mail: qipuliang@163.com。
更新日期/Last Update: 1900-01-01