Effects of caveolin-1 on the expression of epidermal growth factor in fibroblasts co-cultured with breast cancer cells

doi:10.3969/j.issn.2095-4344.2017.20.015

Abstract

Abstract:

BACKGROUND: Caveolin-1, as the most important functional protein in caveolae, is involoved in a variety of cell biological processes, and is closely related to the occurrence and development of breast cancer.

OBJECTIVE: To explore the effect of caveolin-1 on epidermal growth factor (EGF) in fibroblasts after co-cultured with breast cancer cells.

METHODS: Caveolin-1 expression in fibroblast lines ESF-1 was interfered with siRNA, and the optimal effect was determined through QRT-PCR and western blot assay. (1) The optimal silencing model of ESF-1-Caveolin-1 SiRNA-N.2 was obtained, which was co-cultured with breast cancer cells BT474 as experimental group, single-cultured ESF-1 and ESF-1-Caveolin-1 SiRNA-N.2 as controls. The expression level of EGF in ESF-1 was detected by QRT-PCR at 24 and 48 hours of culture; the expression level of EGF in the culture medium was detected by ELISA at 48 and 72 hours of culture. (2) ESF-1-Caveolin-1 SiRNA-N.2 (experimental group) and ESF-1 (control group) were respectively co-cultured with BT474, and single-cultured BT474 as blank control group. The proliferation of BT474 was detected by cell counting-kit 8 assay after 24- and 48-hour culture.

RESULTS AND CONCLUSION: The mRNA expression level of EGF in the experimental group was significantly higher than that in the other two groups (P < 0.05), and the EGF expression level in the ESF-1-Caveolin-1 SiRNA-N.2 group was significantly higher than that in the ESF-1 group (P < 0.05). The protein expression level of EGF was ranked as follows: experimental group > ESF-1-Caveolin-1 SiRNA-N.2 group > ESF-1 group (P < 0.05). The proliferation of BT474 cells was significantly increased after co-cultured with BT474 cells and ESF-1 siRNA Caveolin-1 cells, especially with BT474 cells (P < 0.05). Our findings suggest that Caveolin-1 siRNA can promote the expression of EGF in fibroblasts, especially co-cultured with breast cancer cells. Furthermore, caveolin-1 siRNA accelerates the proliferation of breast cancer cells after co-cultured with fibroblasts.

Key words: Breast Neoplasms, Fibroblasts, Epidermal Growth Factor, Tissue Engineering

CLC Number:

R318

Xiao Liang, Wang Ying, Yang Xiao-jun, Yang Jun-ping. Effects of caveolin-1 on the expression of epidermal growth factor in fibroblasts co-cultured with breast cancer cells[J]. Chinese Journal of Tissue Engineering Research, 2017, 21(20): 3202-3207.

Figures/Tables 2

References

[1] Criscitiello C,Esposito A,Curiqliano G.Tumor-stroma crosstalk:targeting stroma in breast cancer.Curr Opin Oncol. 2014;26:551-555.

[2] Williams TM,Sotgia F,Lee H,et al.Stromal and epithelial Caveolin-1 both confer a protective effect against mammary hyperplasia and tumorigenesis: Caveolin-1 antagonizes cyclin D1 function in mammary epithelial cells.Am J Pathol. 2006;169(5):1784-1801.

[3] Savage K,Lambros MB,Robertson D,et al.Caveolin-1 is overexpressed and amplified in a subset of basal-like and metaplastic breast carcinomas:a morphologic, ultrastructural, immunohistochemical, and in situ hybridization analysis.Clin Cancer Res.2007;13:90-101.

[4] Liedtkeg C,Kersting C,Burger H,et al.Caveolin-1 expression in bengin and malignant lesions of the breast.World J Surg Oncol.2007;5:110.

[5] Anwar SL,Wahyono A,Aryandono T,et al.Caveolin-1 in Breast cancer:single molecule regulation of multiple key signaling pathways.Asian Pac J Cancer Prev.2015;16:6803-6812.

[6] Drab M,Verkade P,Elger M,Kasper M,et al.Loss of caveolae,vascular dysfunction, and pulmonary defects in Caveolin-1 gene-disrupted mice.Science.2001;293: 2449-2452.

[7] Razani B,Woodman SE,Lisanti MP.Caveolae: from cell biology to animal physiology.Pharmacol Rev.2002;54: 431-467.

[8] Scherer PE,Lewis RY,Volonte D,et al.Cell-type and tissue-specific expression of caveolin-2. Caveolins 1 and 2 co-localize and form a stable hetero-oligomeric complex in vivo.J Biol Chem.1997;272(46):29337-29346.

[9] Mercier I,Jasmin JF,Pavlides S,et al.Clinical and translational implications of the caveolin gene family: lessons from mouse models and human genetic disorders.Lab Invest.2009;89: 614-623.

[10] Quest AF,Lobos-González L,Nuñez S,et al.The caveolin-1 connection to cell death and survival.Curr Mol Med.2013; 13(2):266-281.

[11] Boscher C,Nabi IR.Galectin-3-and phospho-caveolin-1 dependent outside-in integrin signaling mediates the EGF motogenic response in mammary cancer cells.Mol Biol Cell. 2013；24:2134-2145.

[12] Thomas NB,Hutcheson IR,Campbell L,et al.Growth of hormone-dependent MCF-7 breast cancer cells is promoted by constitutive caveolin-1 whose expression is lost in an EGF-R-mediated manner during development of tamoxifen resistance.Breast Cancer Res Treat.2010;119(3):575-591.

[13] Shi XY,Xiong LX,Xiao L,et al.Downregulation of caveolin?1 upregulates the expression of growth factors and regulators in co?culture of fibroblasts with cancer cells.Mol Med Rep. 2016;13(1):744-752.

[14] 石小玉,肖亮,熊丽霞,等.Caveolin-1-siRNA对与乳腺癌细胞共培养的成纤维细胞自噬体表达影响的研究[J].中国细胞生物学学报,2015,37(3):212-220.

[15] Sloan EK,Ciocca DR,Pouliot N,et al.Stromal cell expression of Caveolin-1 predicts outcome in breast cancer.Am J Pathol. 2009;174:2035-2043.

[16] Williams TM,Medina F,Badano I,et al.Caveolin-1 gene disruption promotes mammary tumorigenesis and dramatically enhances lung metastasis invivo: role of Cav-1 in cell invasiveness and matrix met alloproteinase (MMP-2/9) secretion.J Biol Chem.2004;279: 51630-51646.

[17] Hayashi K,Matsuda S,Machida K,et al.Invasion activating Caveolin-1 mutation in human scirrhous breast cancers. Cancer Res.2001;61:2361-2364.

[18] Goetz JG,Lajoie P,Wiseman SM,et al.Caveolin-1 in tumor progression: the good,the bad and the ugly.Cancer Metastasis Rev.2008;27:715-735.

[19] Lock R,Debnath J.Extracellular matrix regulation of autophagy. Curr Opin Cell Biol.2008;20:583-588.

[20] Fiucci G,Ravid D,Reich R,et al.Caveolin-1 inhibits anchorage-independent growth, anoikis and invasiveness in MCF-7 human breast cancer cells.Oncogene.2002;21: 2365-2375.

[21] Engelman JA,Wykoff CC,Yasuhara S,et al.Recombinant expression of Caveolin-1 in oncogenically transformed cells abrogates anchorage-independent Growth.Biol Chem.1997; 272:16374-16381.

[22] Glait C,Ravid D,Lee SW,et al.Caveolin-1 controls BRCA1 gene expression and cellular localization in human breast cancer cells.FEBS Lett.2006;580:5268-5274.

[23] Lanuti M,Liu G,Goodwin JM,et al.A functional epidermal growth factor(EGF) polymorphism,EFG serum levels,and esophageal adenocarcinoma risk and outcome.Clin Cancer Res,2008,14:3216-3222.

[24] Kim J,Kong J,Chang H,et al.EGF induces epithelial- mesenchymal transition through phospho-Smad2/3-Snail signaling pathway in breast cancer cells.Oncotarget.2016.doi: 10.18632/oncotarget.13116.[Epub ahead of print]

[25] Kainulainen V,Sundwall M,Maatta JA,et al.A natural ErbB4 isform that does not activate phosphoinositide 3-kinase mediates proliferation but not survival or chemotaxis.J Biol Chem.2000;275:8641-8649.

[26] Shlessinger J.Cell signaling by receptor tyrosine kinases.Cell. 2002;103:211-225.

[27] Mouneimne G,Desmarais V,Sidani M,et al.Spatial and temporal control of cofilin activity is required for directional sensing during chemotaxis.Curr Biol.2006;16:2193-2205.

[28] Chan AY,Raft S,Bailly M,et al.EGF stimulates an increase in actin nucleation and filament number at the leading edge of the lamellipod in mammary adenocarcinoma cells.J Cell Sci. 1998;111:199-211.

[29] Wang W,Eddy R,Condeelis J.The cofilin pathway in breast cancer invasion and metastasis.Nat Rev Cancer.2007;7: 429-440.

[30] Al Moustafa AE,Achkhar A,Yasmeen A.EGF-receptor signaling and epithelial-mesenchymal transiton in hunman carcinomas.Front Biosci(Schol ED).2012;4:671-684.

[31] Boccaccio C,Luraqhi P,Comoqlio PM.MET-mediated resistance to EGFR inhibitors:an old liaison rooted in colorectal cancer stem cells.Cancer Res.2014;74:3647-3651.

[32] Luan TY,Zhu TN,Cui YJ,et al.Expression of caveolin-1 is correlated with lung adenocarcinoma proliferation, migration, and invasion.Med Oncol.2015;32(7):207.

[33] Moreno-Càceres J,Caja L,Mainez J,et al.Caveolin-1 is required for TGF-β-induced transactivation of the EGF receptor pathway in hepatocytes through the activation of the metalloprotease TACE/ADAM17.Cell Death Dis.2014;5: e1326.

[34] Yu XW,Lin S,Du HZ,et al.Synergistic combination of DT-13 and topotecan inhibits human gastric cancer via myosin IIA-induced endocytosis of EGF receptor in vitro and in vivo.Oncotarget.2016;7:32990-3003.

[35] Agelaki S,Spiliotaki M,Markomanolaki H,et al.Caveolin-1 regulates EGFR signaling in MCF-7 breast cancer cells and enhances gefitinib-induced tumor cell inhibition.Cancer Biol Ther.2009;8915:1470-1477.