[1]刘 冲,张 静a,李 胜,等.miR-335 通过下调Fra-1 表达抑制人乳腺癌细胞增殖机制的实验研究[J].现代检验医学杂志,2022,37(05):76-80+104.[doi:10.3969/j.issn.1671-7414.2022.05.016]
　LIU Chong,ZHANG Jinga,LI Sheng,et al.Research of miR-335 Inhibited Proliferation of Human Breast Cancer Cells by Down-Regulated Fra-1 Expression[J].Journal of Modern Laboratory Medicine,2022,37(05):76-80+104.[doi:10.3969/j.issn.1671-7414.2022.05.016]
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miR-335 通过下调Fra-1 表达抑制人乳腺癌细胞增殖机制的实验研究()
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《现代检验医学杂志》[ISSN:/CN:]

卷:
第37卷
期数:
2022年05期
页码:
76-80+104
栏目:
论著
出版日期:
2022-09-15

文章信息/Info

Title:
Research of miR-335 Inhibited Proliferation of Human Breast Cancer Cells by Down-Regulated Fra-1 Expression
文章编号:
1671-7414（2022）05-076-06
作者:
刘 冲1; 张 静2a; 李 胜1; 赵 奇2b
1. 湖北理工学院附属妇幼保健院（儿童医院）/ 鄂东医疗集团黄石市妇幼保健医院医学检验科，湖北黄石 435000；2. 黄石爱康医院a. 医学检验科；b. 医学骨科，湖北黄石 435000
Author(s):
LIU Chong1; ZHANG Jing2a; LI Sheng1; ZHAO Qi2b
1.Department of Clinical Laboratory，Hubei Institute of Technology Affiliated Maternal and Child Health Care Hospital (Children’s Hospital)/Huangshi Maternal and Child Health Care Hospital of Edong Healthcare， Hubei Huangshi 435000, China；2a.Department of Clinical Laboratory；2b.Department of Orthopedics, Huangshi Aikang Hospital，Hubei Huangshi 435000, China
关键词:
微小核糖核酸-335; Fos 相关抗原1; 乳腺癌; 基因表达
分类号:
R737.9；R730.43
DOI:
10.3969/j.issn.1671-7414.2022.05.016
文献标志码:
A
摘要:
目的　探讨微小核糖核酸（microRNA，miR）-335 对人乳腺癌细胞增殖能力的影响，以及其可能存在的机制。方法　体外培养乳腺癌细胞及正常乳腺组织细胞，检测miR-335 及Fros 相关抗原1(fros related antigent-1,Fra-1) 的表达量；体外转染miR-335 mimics 后，通过细胞活力检测试剂盒(cell counting kit-8,CCK-8) 检测细胞的增殖能力，检测Fra-1 的表达变化及下游细胞增殖相关蛋白分子基质金属蛋白酶9（matrix metalloproteinase-9，MMP-9）、血管内皮生长因子C(vascular endothelial growth factor- C,VEGF-C) 的表达量；生物信息学预测及双荧光霉素报告基因验证Fra-1 为miR-335作用靶基因。结果　乳腺癌细胞中miR-335 的表达量（0.755±0.034）低于正常乳腺细胞（1.495±0.029），Fra-1 蛋白的表达量（2..347±0.120）高于正常乳腺细胞（1.319±0.038），差异均有统计学意义（t=0.075，4.191, 均P ＜ 0.001）。转染miR-335 mimic 后48，72 h 乳腺癌细胞增殖能力（0.881±0.062，0.887±0.082）低于对照组细胞（1.326±0.051，1.493±0.038），差异均有统计学意义（t=44.096，59.307, 均P ＜ 0.001）；转染miR-335 mimic 后乳腺癌细胞中Fra-1的蛋白表达量（0.567±0.091）低于未转染细胞（2.347±0.204），差异有统计学意义（t=3.763, P<0.001）；转染miR-335 mimic 后细胞中MMP-9（0.469±0.027），VEGF-C（0.540±0.041）的蛋白表达量低于对照组（0.958±0.058，1.024±0.171），差异均有统计学意义（t=1.953，7.856, 均P ＜ 0.001）；荧光霉素报告基因实验结果显示miR-335 下调Fra-1 启动子核心转录区域（-164 ～ -52nt）的转录活性。结论　miR-335 通过抑制MMP-9 和VEGF-C 的表达，并下调Fra-1 启动子转录活性，抑制了体外乳腺癌细胞的增殖。
Abstract:
Objective　To explore the effect of microRNA (miR) -335 on the proliferation of human breast cancer cells and its possible mechanism. Methods　Breast cancer cells and normal breast tissue cells were cultured in vitro, and the expression of miR-335 and Fros related antigent-1（Fra-1）were detected. MiR-335 mimics were transfected into breast cancer cells, the proliferation ability of breast cancer cells was detected by cell counting kit-8 (CCK-8). Real-time PCR and Western blotting were used to detecte the expression of Fra-1 mRNA and protein, and detected the expression of cell proliferation-related molecules matrix metalloproteinase 9(MMP-9) and vascular endothelial growth factor-C(VEGF-C); Bioinformatics prediction and luciferase reporter gene assay were comfirmed that Fra-1 was the potential target genes of miR-335. Result　The expression level of miR- 335 in breast cancer cells （0.755±0.034）, which was lower than that in normal breast cells (1.495±0.029)，the expression level of Fra-1 protein （2.347±0.120）, which was higher than that in normal breast cells (1.319±0.038), the differences were statistically significant (t=0.075,4.191, all P ＜ 0.001). MiR-335 mimic was transfected into breast cancer cells and cultured for 48 and 72 hours, the proliferation ability of breast cancer cells were 0.881±0.062 and 0.887±0.082, which were lower than that of control cells by 1.326±0.051 and 1.493±0.038, the differences were statistically significant(t=44.096, 59.307, all P ＜ 0.001). The protein expression of Fra-1 in breast cancer cells which transfected with miR-335 mimic was 0.567±0.091, which was lower than that untransfected cells (2.347±0.204), the difference was statistically significant (t=3.763，P ＜ 0.001). MiR-335 mimic was transfected into breast cancer cells ,the protein expressions of MMP-9 and VEGF-C in the cells were 0.469±0.027 and 0.540±0.041, which were lower than the control group by 0.958±0.058 and 1.024±0.171, the differences were statistically significant(t=1.953，7.856，all P ＜ 0.001). Luciferase reporter gene assay showed that miR-335 could down-regulate the transcription activity of the core transcription region of Fra-1 promoter (-164 ～ -52nt). Conclusion　MiR-335 inhibited the proliferation of breast cancer cells in vitro by down-regulating the expression of MMP-9, VEGF-C and the transcriptional activity of Fra-1 promoter.

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备注/Memo

备注/Memo:
基金项目：湖北省卫生健康科研基金项目（WJ2019H203）。
作者简介：刘冲（1987-），男，硕士，主管技师，主要从事肿瘤分子诊断研究，E-mail:39662238@qq.com。
通讯作者：赵奇，硕士，主治医师，E-mail：649921632@qq.com。
更新日期/Last Update: 2022-09-15