Water-pipe smoking promotes epithelial-mesenchymal transition and invasion of human breast cancer cells via ERK1/ERK2 pathways

Abstract

Background: With the increasing popularity of water-pipe smoking (WPS), it is critical to comprehend how WPS may affect women's health. The main goal of this study is to identify the potential outcome of WPS on human breast cancer progression.

Methods: Two breast cancer cell lines, MCF7 and BT20, were used in this investigation. We explored the outcome of WPS on cell morphology and cell invasion using inverted microscope and Biocoat Matrigel invasion chambers. On the other hand, Western blot was employed to study the expression patterns of key control genes of cell adhesion and invasion.

Results: Our data reveal that WPS induces epithelial-mesenchymal transition (EMT) of MCF7 and BT20 breast cancer cell lines; thus, WPS enhances cell invasion ability of both cell lines in comparison with their matched controls. More significantly, WPS provokes a down- and up-regulation of E-cadherin and focal adhesion kinase (FAK), respectively, which are important key regulators of cancer progression genes. Finally, our data point out that WPS incites the activation of Erk1/Erk2, which could be behind the stimulation of EMT and invasion as well as the deregulation of E-cadherin and FAK expression.

Conclusion: Our data show, for the first time, that WPS initiates EMT and stimulates cell invasion of breast cancer cells, which could incite metastatic development in breast cancer patients. Thus, we believe that further studies, both in vitro and in vivo, are required to elucidate the pathogenic outcome of WPS on cancer progression of several human carcinomas including breast.

Keywords: Breast cancer; Cell adhesion; Cell invasion; EMT; Erk1/Erk2 pathways; Water-pipe smoking.

Fig. 1

Effects of water-pipe smoking (WPS) on colony formation in human breast cancer cell lines. WPS slightly enhances colony formation of MCF7 and BT20 cell lines in comparison with their control cells. Clonogenic cell assay and WPS exposure were performed as described in “Materials and methods” section

Fig. 2

WPS stimulates epithelial–mesenchymal transition (EMT) of breast cancer cell lines, MCF7 and BT20. We note that treatment for 3 days with 200 μg/ml of WPS solution induces morphological changes from “epithelial-like” (control) cells into “fibroblast-like” (mesenchymal) phenotype, which is known as EMT

Fig. 3

Effect of WPS on cell invasion of human breast cancer cells. The results from matrigel invasion assay indicate that WPS enhances cell invasion ability of MCF7 and BT20 cell lines by approximately 35% in comparison with their control cells. The histograms show mean ± SD (P < 0.002 and 0.001, respectively; t-test was used and is considered significant with P < 0.05). The cancer cells were treated with 200 μg of WPS solution for 2 days as described in “Materials and methods” section

Fig. 4

Western blot analysis of E-cadherin, FAK and p-Erk1/Erk2 in MCF7 and BT20 cell lines under the effect of WPS. We notice that WPS decreases/increases the expression patterns of E-cadherin and FAK, respectively, in both cell lines in comparison with control cells; meanwhile, WPS stimulates Erk1/Erk2 phosphorylation in these two cell lines. GAPDH was used as a control. Cells were treated with WPS solution as explained in “Materials and methods” section as well as “Results” section

Fig. 5

Quantification of the western blot analysis of E-cadherin, FAK and Erk1/Erk2 phosphorylation as well as GAPDH in MCF7 and BT20 exposed to WPS and unexposed (control) cells. This analysis confirms the downregulation of E-cadherin in both cell lines; in parallel, WPS enhances FAK expression and Erk1/Erk2 phosphorylation in the two cell lines in comparison with their control cells. The quantification of both data by ImageJ 64-bit version 1.50b program