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Transforming growth factor-β1 induces EMT by the transactivation of epidermal growth factor signaling through HA/CD44 in lung and breast cancer cells.

Li L, Qi L, Liang Z, Song W, Liu Y, Wang Y, Sun B, Zhang B, Cao W - Int. J. Mol. Med. (2015)

Bottom Line: Hyaluronan (HA) has been shown to induce EMT through either TGF-β1 or EGF signaling and to be a regulator of the crosstalk between these two pathways in fibroblasts.In this study, in order to clarify whether HA has the same effect in tumor cells, we utilized the lung cancer cell line, A549, and the breast cancer cell line, MCF-7, and found that the effects of stimulation with TGF-β1 were more potent than those of EGF in regulating the expression of EMT-associated proteins and in enhancing cell migration and invasion.In conclusion, our data demonstrate that TGF-β1 induces EMT by the transactivation of EGF signaling through HA/CD44 in lung and breast cancer cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Tianjin Medical University Cancer Institute and Hospital, Tianjin 300060, P.R. China.

ABSTRACT
Epithelial-mesenchymal transition (EMT), a process closely related to tumor development, is regulated by a variety of signaling pathways and growth factors, such as transforming growth factor-β1 (TGF-β1) and epidermal growth factor (EGF). Hyaluronan (HA) has been shown to induce EMT through either TGF-β1 or EGF signaling and to be a regulator of the crosstalk between these two pathways in fibroblasts. In this study, in order to clarify whether HA has the same effect in tumor cells, we utilized the lung cancer cell line, A549, and the breast cancer cell line, MCF-7, and found that the effects of stimulation with TGF-β1 were more potent than those of EGF in regulating the expression of EMT-associated proteins and in enhancing cell migration and invasion. In addition, we observed that TGF-β1 activated EGF receptor (EGFR) and its downstream AKT and extracellular signal-regulated kinase (ERK) pathways. Furthermore, we found that TGF-β1 upregulated the expression of hyaluronan synthases (HAS1, HAS2 and HAS3) and promoted the expression of CD44, a cell surface receptor for HA, which interacts with EGFR, resulting in the activation of the downstream AKT and ERK pathways. Conversely, treatment with 4-methylumbelliferone (4-MU; an inhibitor of HAS) prior to stimulation with TGF-β1, inhibited the expression of CD44 and EGFR, abolished the interaction between CD44 and EGFR. Furthermore, the use of shRNA targeting CD44 impaired the expression of EGFR, deactivated the AKT and ERK pathways, reversed EMT and decreased the migration and invasion ability of cells. In conclusion, our data demonstrate that TGF-β1 induces EMT by the transactivation of EGF signaling through HA/CD44 in lung and breast cancer cells.

No MeSH data available.


Related in: MedlinePlus

Transforming growth factor-β1 (TGF-β1) is an important inducer of epithelial-mesenchymal transition (EMT) compared with epidermal growth factor (EGF). (A) Western blot analysis of EMT-related proteins in A549 and MCF-7 cells treated with TGF-β1 (5 ng/ml) or EGF (10 ng/ml). (B) Immunofluorescence staining was conducted to detect E-cadherin (green) and N-cadherin (red) expression in the A549 and MCF-7 cells, which were treated with EGF or TGF-β1 for 24 h. The nuclei were visualized with 4′,6-diamidino-2-phenylindole staining (DAPI, blue). Images were taken at ×200 magnification. (C) RT-PCR was used to measure the mRNA levels of EGFR and TGF-β1 receptor (TβR) following stimulation with TGF-β1. Western blot analysis detected the activation of phosphorylated (p-) Smad, downstream of TGF-β1, and the activation of p-AKT and p-ERK, downstream of EGF/EGFR in the A549 and MCF-7 cells following stimulation with TGF-β1 or EGF.
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f1-ijmm-36-01-0113: Transforming growth factor-β1 (TGF-β1) is an important inducer of epithelial-mesenchymal transition (EMT) compared with epidermal growth factor (EGF). (A) Western blot analysis of EMT-related proteins in A549 and MCF-7 cells treated with TGF-β1 (5 ng/ml) or EGF (10 ng/ml). (B) Immunofluorescence staining was conducted to detect E-cadherin (green) and N-cadherin (red) expression in the A549 and MCF-7 cells, which were treated with EGF or TGF-β1 for 24 h. The nuclei were visualized with 4′,6-diamidino-2-phenylindole staining (DAPI, blue). Images were taken at ×200 magnification. (C) RT-PCR was used to measure the mRNA levels of EGFR and TGF-β1 receptor (TβR) following stimulation with TGF-β1. Western blot analysis detected the activation of phosphorylated (p-) Smad, downstream of TGF-β1, and the activation of p-AKT and p-ERK, downstream of EGF/EGFR in the A549 and MCF-7 cells following stimulation with TGF-β1 or EGF.

Mentions: Significant alterations in the expression of EMT-associated proteins (E-cadherin, N-cadherin, Snail, Twist and ZEB1) were observed following stimulation of the cells with TGF-β1 and EGF, although it was clearly evident that TGF-β1 induced EMT to a greater extent, as its effects on the epxression of these proteins were more potent (Fig. 1A and B).


Transforming growth factor-β1 induces EMT by the transactivation of epidermal growth factor signaling through HA/CD44 in lung and breast cancer cells.

Li L, Qi L, Liang Z, Song W, Liu Y, Wang Y, Sun B, Zhang B, Cao W - Int. J. Mol. Med. (2015)

Transforming growth factor-β1 (TGF-β1) is an important inducer of epithelial-mesenchymal transition (EMT) compared with epidermal growth factor (EGF). (A) Western blot analysis of EMT-related proteins in A549 and MCF-7 cells treated with TGF-β1 (5 ng/ml) or EGF (10 ng/ml). (B) Immunofluorescence staining was conducted to detect E-cadherin (green) and N-cadherin (red) expression in the A549 and MCF-7 cells, which were treated with EGF or TGF-β1 for 24 h. The nuclei were visualized with 4′,6-diamidino-2-phenylindole staining (DAPI, blue). Images were taken at ×200 magnification. (C) RT-PCR was used to measure the mRNA levels of EGFR and TGF-β1 receptor (TβR) following stimulation with TGF-β1. Western blot analysis detected the activation of phosphorylated (p-) Smad, downstream of TGF-β1, and the activation of p-AKT and p-ERK, downstream of EGF/EGFR in the A549 and MCF-7 cells following stimulation with TGF-β1 or EGF.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494581&req=5

f1-ijmm-36-01-0113: Transforming growth factor-β1 (TGF-β1) is an important inducer of epithelial-mesenchymal transition (EMT) compared with epidermal growth factor (EGF). (A) Western blot analysis of EMT-related proteins in A549 and MCF-7 cells treated with TGF-β1 (5 ng/ml) or EGF (10 ng/ml). (B) Immunofluorescence staining was conducted to detect E-cadherin (green) and N-cadherin (red) expression in the A549 and MCF-7 cells, which were treated with EGF or TGF-β1 for 24 h. The nuclei were visualized with 4′,6-diamidino-2-phenylindole staining (DAPI, blue). Images were taken at ×200 magnification. (C) RT-PCR was used to measure the mRNA levels of EGFR and TGF-β1 receptor (TβR) following stimulation with TGF-β1. Western blot analysis detected the activation of phosphorylated (p-) Smad, downstream of TGF-β1, and the activation of p-AKT and p-ERK, downstream of EGF/EGFR in the A549 and MCF-7 cells following stimulation with TGF-β1 or EGF.
Mentions: Significant alterations in the expression of EMT-associated proteins (E-cadherin, N-cadherin, Snail, Twist and ZEB1) were observed following stimulation of the cells with TGF-β1 and EGF, although it was clearly evident that TGF-β1 induced EMT to a greater extent, as its effects on the epxression of these proteins were more potent (Fig. 1A and B).

Bottom Line: Hyaluronan (HA) has been shown to induce EMT through either TGF-β1 or EGF signaling and to be a regulator of the crosstalk between these two pathways in fibroblasts.In this study, in order to clarify whether HA has the same effect in tumor cells, we utilized the lung cancer cell line, A549, and the breast cancer cell line, MCF-7, and found that the effects of stimulation with TGF-β1 were more potent than those of EGF in regulating the expression of EMT-associated proteins and in enhancing cell migration and invasion.In conclusion, our data demonstrate that TGF-β1 induces EMT by the transactivation of EGF signaling through HA/CD44 in lung and breast cancer cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Tianjin Medical University Cancer Institute and Hospital, Tianjin 300060, P.R. China.

ABSTRACT
Epithelial-mesenchymal transition (EMT), a process closely related to tumor development, is regulated by a variety of signaling pathways and growth factors, such as transforming growth factor-β1 (TGF-β1) and epidermal growth factor (EGF). Hyaluronan (HA) has been shown to induce EMT through either TGF-β1 or EGF signaling and to be a regulator of the crosstalk between these two pathways in fibroblasts. In this study, in order to clarify whether HA has the same effect in tumor cells, we utilized the lung cancer cell line, A549, and the breast cancer cell line, MCF-7, and found that the effects of stimulation with TGF-β1 were more potent than those of EGF in regulating the expression of EMT-associated proteins and in enhancing cell migration and invasion. In addition, we observed that TGF-β1 activated EGF receptor (EGFR) and its downstream AKT and extracellular signal-regulated kinase (ERK) pathways. Furthermore, we found that TGF-β1 upregulated the expression of hyaluronan synthases (HAS1, HAS2 and HAS3) and promoted the expression of CD44, a cell surface receptor for HA, which interacts with EGFR, resulting in the activation of the downstream AKT and ERK pathways. Conversely, treatment with 4-methylumbelliferone (4-MU; an inhibitor of HAS) prior to stimulation with TGF-β1, inhibited the expression of CD44 and EGFR, abolished the interaction between CD44 and EGFR. Furthermore, the use of shRNA targeting CD44 impaired the expression of EGFR, deactivated the AKT and ERK pathways, reversed EMT and decreased the migration and invasion ability of cells. In conclusion, our data demonstrate that TGF-β1 induces EMT by the transactivation of EGF signaling through HA/CD44 in lung and breast cancer cells.

No MeSH data available.


Related in: MedlinePlus