肿瘤相关成纤维细胞中雌激素/GPER/PKA/FGF2信号轴促进乳腺癌细胞侵袭

余腾骅; 王智亮; 彭美茜; 邱宇安

doi:10.16016/j.1000-5404.201904110

您当前的位置：

首页 >

文章列表页 >

肿瘤相关成纤维细胞中雌激素/GPER/PKA/FGF2信号轴促进乳腺癌细胞侵袭

基础医学 | 更新时间：2026-01-09

- 肿瘤相关成纤维细胞中雌激素/GPER/PKA/FGF2信号轴促进乳腺癌细胞侵袭
- Estrogen/GPER/PKA/FGF2 signal axis in cancer-associated fibroblasts enhances the invasion of breast cancer cells
- 第三军医大学学报 2019年41卷第18期页码：1730-1737
- 作者机构：
  
  1.330029 南昌，江西省肿瘤医院:乳腺肿瘤外科
  2.400010 重庆，重庆医科大学附属第二医院妇产科
  3.400016重庆，重庆医科大学临床检验诊断学教育部重点实验室
  4.330029 南昌，江西省肿瘤医院:重症医学科
- 作者简介：
  
  [ 邱宇安, E-mail:qya96245@163.com QIU Yuan, E-mail:qya96245@163.com
- 基金信息：
  
  国家自然科学基金青年科学基金(81702641);江西省自然科学基金杰出青年基金(2018ACB21042);江西省自然科学基金青年基金(20171BAB215046)
- DOI：10.16016/j.1000-5404.201904110
  中图分类号： R73-37;R730.23;R737.9
- 收稿：2019-04-14，
  
  修回：2019-05-28，
  
  纸质出版：2019-09-30
- 稿件说明：
移动端阅览
余腾骅, 王智亮, 彭美茜, 等. 肿瘤相关成纤维细胞中雌激素/GPER/PKA/FGF2信号轴促进乳腺癌细胞侵袭[J]. 第三军医大学学报, 2019,41(18):1730-1737.

Tenghua YU, Zhiliang WANG, Meiqian PENG, et al. Estrogen/GPER/PKA/FGF2 signal axis in cancer-associated fibroblasts enhances the invasion of breast cancer cells[J]. Journal of Third Military Medical University, 2019, 41(18): 1730-1737.
余腾骅, 王智亮, 彭美茜, 等. 肿瘤相关成纤维细胞中雌激素/GPER/PKA/FGF2信号轴促进乳腺癌细胞侵袭[J]. 第三军医大学学报, 2019,41(18):1730-1737. DOI： 10.16016/j.1000-5404.201904110.

Tenghua YU, Zhiliang WANG, Meiqian PENG, et al. Estrogen/GPER/PKA/FGF2 signal axis in cancer-associated fibroblasts enhances the invasion of breast cancer cells[J]. Journal of Third Military Medical University, 2019, 41(18): 1730-1737. DOI： 10.16016/j.1000-5404.201904110.

摘要

目的

探讨共培养下乳腺癌肿瘤相关成纤维细胞(cancer-associated fibroblasts，CAFs)中细胞质G蛋白偶联雌激素受体(G protein-coupled estrogen receptor，GPER)活化对成纤维细胞生长因子2(fibroblast growth factors 2，FGF2)的表达调控机制及其对肿瘤细胞侵袭能力的影响。

方法

细胞免疫荧光法检测单独及与雌激素受体阳性(estrogen receptor positive

)乳腺癌MCF-7或ER阴性(ER

)乳腺癌MDA-MB-468细胞共培养下CAFs中GPER的细胞内定位；荧光定量PCR法与ELISA法分别检测不同处理后CAFs细胞内FGF2的mRNA表达及细胞上清液中的分泌水平；Western blot法检测CAFs细胞中GPER下游ERK、AKT及PKA信号通路活化情况；Transwell小室实验检测不同药物处理下乳腺癌细胞侵袭能力的改变。

结果

两种亚型的乳腺癌细胞均可诱导CAFs中GPER由细胞核转位入细胞质。共培养条件下，17-β雌二醇(E2)可显著促进CAFs中FGF2的mRNA表达及细胞上清液中的分泌量，突变GPER氨基酸序列中的核输出信号(nucleus export signal

NES)可显著抑制以上变化(

0.05)。CAFs单独培养下则无以上现象。同时，

E2处理共培养下CAFs细胞后显著活化细胞内GPER/AKT与GPER/PKA信号通路，促进下游p-AKT与p-PKA的蛋白表达(

0.05)。突变GPER中NES序列后可显著抑制下游PKA信号的活化，反而促进p-ERK的表达(

0.05)。用PKA特异性抑制剂(H-89)可明显阻断CAFs中E2诱导的FGF2表达(

0.05)，AKT特异性抑制剂(WM)则不能。CAFs中E2刺激下的FGF2旁分泌明显提高乳腺癌细胞侵袭能力，该效应被FGF2中和抗体所阻断(

0.05)。

结论

雌激素活化CAFs中细胞质GPER依赖性的下游PKA信号旁分泌FGF2进而促进乳腺癌细胞侵袭，上述通路或在肿瘤进展中发挥重要作用。

Abstract

Objective

To investigate the effect of activated cytoplasmic G protein-coupled estrogen receptor (GPER) from co-cultured cancer-associated fibroblasts (CAFs) in the regulation of fibroblast growth factor 2 (FGF2) and the invasion of breast cancer cells.

Methods

The intracellular GPER localization in CAFs cultured alone or co-cultured with ER

positive (MCF-7) and ER

negative (MDA-MB-468) breast cancer cells was examined using immunofluorescence assays. Real-time quantitative PCR and enzyme-linked immunosorbent assay (ELISA) were utilized to detect the expression of FGF2 mRNA in the CAFs and FGF2 secretion in the supernatant

respectively. The activation of GPER downstream ERK

AKT and PKA signal pathways was determined by Western blotting. The changes in the invasiveness of breast cancer cells were examined by Transwell assays.

Results

Co-culture with the 2 breast cancer cells both induced translocation of GPER from the cell nucleus to the cytoplasm in the CAFs. In the cell co-cultures

17-β estradiol (E2) significantly increased FGF2 levels in the CAFs and the supernatant

and this effect was obviously blocked by mutation of the nucleus export signal (NES) in GPER amino acid sequence (

0.05); these results could not be observed in the CAFs cultured alone. E2 treatment of the co-cultured CAFs resulted in obvious activation of GPER/AKT and GPER/PKA signal pathways (

0.05) and increased the protein expression of p-AKT and p-PKA; Mutation of the NES in GPER sequence significantly inhibited PKA signaling and up-regulated the expression of p-ERK (

0.05). The E2-induced increase of FGF2 expression in CAFs was significantly blocked by the application of the PKA inhibitor H-89 but not by the AKT inhibitor WM (

0.05). More importantly

the secreted FGF2 from E2-treated CAFs significantly promoted the invasion of the breast cancer cells

which was abolished by the neutralizing antibody of FGF2.

Conclusion

Estrogen enhances the activation of the downstream PKA signaling in a cytoplasmic GPER-dependent manner in the CAFs

and the enhanced FGF2 paracrine promotes the invasion of breast cancer cells. These signal pathways may play important roles in the progression of breast cancer.

关键词

Keywords

references

LAPPANO R, MAGGIOLINI M. GPER is involved in the functional liaison between breast tumor cells and cancer-associated fibroblasts (CAFs)[J]. J Steroid Biochem Mol Biol, 2018, 176: 49-56. DOI:10.1016/j.jsbmb.2017.02.019.

SUN K X, TANG S F, HOU Y X, et al . Oxidized ATM-mediated glycolysis enhancement in breast cancer-associated fibroblasts contributes to tumor invasion through lactate as metabolic coupling[J ] . EBio Medicine, 2019, 41: 370-383. DOI:10.1016/j.ebiom.2019.02.025.

YU T H, LIU M R, LUO H J, et al . GPER mediates enhanced cell viability and motility via non-genomic signaling induced by 17β-estradiol in triple-negative breast cancer cells[J ] . J Steroid Biochem Mol Biol, 2014, 143: 392-403. DOI:10.1016/j.jsbmb.2014.05.003..

YIN H, ZHU Q, LIU M R, et al . GPER promotes Tamo-xifen-resistance in ER + breast cancer cells by reduced Bim proteins through MAPK/Erk-TRIM2 signaling axis[J ] . Int J Oncol, 2017, 51(4): 1191-1198. DOI:10.3892/ijo.2017.4117..

HSU L H, CHU N M, LIN Y F, et al . G-protein coupled estrogen receptor in breast cancer[J ] . IJMS, 2019, 20(2): 306. DOI: 10.3390/ijms20020306.

LUO H J, LIU M R, LUO S J, et al . Dynamic monitoring of GPER-mediated estrogenic effects in breast cancer associated fibroblasts: An alternative role of estrogen in mammary carcinoma development[J ] . Steroids, 2016, 112: 1-11. DOI: 10.1016/j.steroids.2016.03.013.

YU T, YANG G, HOU Y, et al . Cytoplasmic GPER trans-location in cancer-associated fibroblasts mediates cAMP/PKA/CREB/glycolytic axis to confer tumor cells with multidrug resistance[J ] . Oncogene, 2017, 36(15): 2131-2145. DOI: 10.1038/onc.2016.370.

AKL M R, NAGPAL P, AYOUB N M, et al . Molecular and clinical significance of fibroblast growth factor 2 (FGF 2 /bFGF) in malignancies of solid and hematological cancers for personalized therapies[J ] . Oncotarget, 2016, 7(28): 44735-44762. DOI:10.18632/oncotarget.8203..

TRAER E, JAVIDI-SHARIFI N, AGARWAL A, et al . Ponatinib o vercomes FGF 2 -mediated resistance in CML patients without kinase domain mutations[J ] . Blood, 2014, 123(10): 1516-1524. DOI: 10.1182/blood-2013-07-518381..

SEO J H, YU J H, SUH H, et al . Fibroblast growth factor-2 induced by enriched environment enhances angiogenesis and motor function in chronic hypoxic-ischemic brain injury[J ] . PLoS ONE, 2013, 8(9): e74405. DOI: 10.1371/journal.pone.0074405.

QIAO A X, GU F, GUO X J, et al . Breast cancer-associated fibroblasts: their roles in tumor initiation, progression and clinical applications[J ] . Front Med, 2016, 10(1): 33-40. DOI: 10.1007/s11684-016-0431-5.

CIRILLO F, PELLEGRINO M, MALIVINDI R, et al . GPER is involved in the regulation of the estrogen-metabolizing CYP 1 B 1 enzyme in breast cancer[J ] . Oncotarget, 2017, 8(63): 106608-106624. DOI: 10.18632/oncotarget.22541..

DE FRANCESCO E M, SIMS A H, MAGGIOLINI M, et al . GPER mediates the angiocrine actions induced by IGF 1 through the HIF-1α/VEGF pathway in the breast tumor microenvironment[J ] . Breast Cancer Res, 2017, 19(1): 129. DOI: 10.1186/s13058-017-0923-5..

FILARDO E J, QUINN J A, FRACKELTON A R Jr, et al . Estrogen action via the G protein-coupled receptor, GPR30: stimulation of adenylyl cyclase and cAMP-mediated attenuation of the epidermal growth factor receptor-to-MAPK signaling axis[J ] . Mol Endocrinol, 2002, 16(1): 70-84. DOI: 10.1210/mend.16.1.0758..

TREMBLAY P G, SIRARD M A. Transcriptomic analysis of gene cascades involved in protein kinase A and C signaling in the KGN line of human ovarian granulosa tumor cells[J]. Biol Reprod, 2017, 96(4): 855-865. DOI: 10.1093/biolre/iox024.

GIULIANELLI S, RIGGIO M, GUILLARDOY T, et al . FGF 2 induces breast cancer growth through ligand-independent activation and recruitment of ERα and PRBΔ4 isoform to MYC regulatory sequences[J ] . Int J Cancer, 2019: ijc.32252. DOI: 10.1002/ijc.32252..

MIURA K, OBA T, HAMANAKA K, et al . FGF 2 -FGFR1 pathway activation together with thymidylate synthase upregulation is induced in pemetrexed-resistant lung cancer cells[J ] . Oncotarget, 2019, 10(11): 1171-1192. DOI: 10.18632/oncotarget.26622..

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

ATP1A2通过Src/PI3K/Akt信号通路抑制人乳腺癌细胞的侵袭和迁移

基于中国人群的雌激素受体表达新分层（低/中/高）可更精准地预测HR⁺/HER2^-早期乳腺癌患者的预后

二氢杨梅素通过抑制mTOR信号通路抑制乳腺癌细胞迁移和侵袭

BIRC5基因rs9904341多态性与乳腺癌易感性的关联研究

长链非编码ATB通过抑制miR-200c的表达促进乳腺癌细胞的侵袭转移