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IGF-1 increases invasive potential of MCF 7 breast cancer cells and induces activation of latent TGF-β1 resulting in epithelial to mesenchymal transition.

Walsh LA, Damjanovski S - Cell Commun. Signal (2011)

Bottom Line: Furthermore, most studies describing the biological effects of TGF-β have been performed using high concentrations of active, soluble TGF-β, despite the fact that TGF-β is produced and secreted as a latent complex.The effects of IGF-1 appear to be mediated through signals transduced via the PI3K and MAPK pathways.In addition, increased IGF-1, together with latent TGF-β1 and active MMPs result in EMT.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, University of Western Ontario, London, Ontario, N6A 5B7, Canada. sdamjano@uwo.ca.

ABSTRACT

Introduction: TGF-β signaling has been extensively studied in many developmental contexts, amongst which is its ability to induce epithelial to mesenchymal transitions (EMT). EMTs play crucial roles during embryonic development and have also come under intense scrutiny as a mechanism through which breast cancers progress to become metastatic. Interestingly, while the molecular hallmarks of EMT progression (loss of cell adhesion, nuclear localization of β-catenin) are straightforward, the cellular signaling cascades that result in an EMT are numerous and diverse. Furthermore, most studies describing the biological effects of TGF-β have been performed using high concentrations of active, soluble TGF-β, despite the fact that TGF-β is produced and secreted as a latent complex.

Methods: MCF-7 breast cancer cells treated with recombinant IGF-1 were assayed for metalloproteinase activity and invasiveness through a matrigel coated transwell invasion chamber. IGF-1 treatments were then followed by the addition of latent-TGF-β1 to determine if elevated levels of IGF-1 together with latent-TGF-β1 could cause EMT.

Results: Results showed that IGF-1 - a molecule known to be elevated in breast cancer is a regulator of matrix metalloproteinase activity (MMP) and the invasive potential of MCF-7 breast cancer cells. The effects of IGF-1 appear to be mediated through signals transduced via the PI3K and MAPK pathways. In addition, increased IGF-1, together with latent TGF-β1 and active MMPs result in EMT.

Conclusions: Taken together our data suggest a novel a link between IGF-1 levels, MMP activity, TGF-β signaling, and EMT in breast cancer cells.

No MeSH data available.


Related in: MedlinePlus

IGF-1 and latent TGF-β1 caused morphological changes in MCF-7 cells consistent with EMT. Vehicle treated cells showed typical rounded epithelial morphology (a). Treatment of MCF-7 cells with 100 nM recombinant IGF-1 + 10 nM latent TGF-β1 caused a mesenchymal morphological phenotype characterized by a fibroblast-like appearance (g). Treatment with IGF-1 or latent TGF-β1 alone did not result in a morphological change (c, e). Pre-treatment of cells with PI3K or MAPK inhibitors abolished IGF-1 + latent TGF-β1 induced mesenchymal morphological phenotype (d, f). BB94 or TGF-β1 inhibitor prevented the morphological changes associated with IGF-1 + TGF-β1 treatment (b, h). Cytoskeleton was stained with phalloidin (red) and the nucleus with DAPI (blue). Images are representative of 3 independent experiments with consistent results.
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Figure 2: IGF-1 and latent TGF-β1 caused morphological changes in MCF-7 cells consistent with EMT. Vehicle treated cells showed typical rounded epithelial morphology (a). Treatment of MCF-7 cells with 100 nM recombinant IGF-1 + 10 nM latent TGF-β1 caused a mesenchymal morphological phenotype characterized by a fibroblast-like appearance (g). Treatment with IGF-1 or latent TGF-β1 alone did not result in a morphological change (c, e). Pre-treatment of cells with PI3K or MAPK inhibitors abolished IGF-1 + latent TGF-β1 induced mesenchymal morphological phenotype (d, f). BB94 or TGF-β1 inhibitor prevented the morphological changes associated with IGF-1 + TGF-β1 treatment (b, h). Cytoskeleton was stained with phalloidin (red) and the nucleus with DAPI (blue). Images are representative of 3 independent experiments with consistent results.

Mentions: Recent evidence suggested that signaling after the proteolytic activation of latent TGF-β1 could induce EMT in breast cancer leading to metastasis [14]. There is also a strong correlation between high levels of IGF-1 signaling and metastasis [15]. The link between IGF-1 and TGFβ1 signaling however, has not thoroughly been explored. Here we show that treating MCF-7 cells first with IGF-1 (8 h), and then subsequently with latent TGF-β1 (48 h) resulted in a change to a cellular fibroblast-like morphology consistent with EMT (Figure 2g). MCF-7 vehicle treatment (control), or treatment with IGF-1 or latent TGF-β1 alone did not induce this change in cellular morphology (Figure 2a, c, e). Furthermore, pretreatment of cells with PI3K or MAPK inhibitors demonstrated the specificity of these pathways in IGF-1 mediated latent TGF-β1 induced morphological changes as it prevented changes in morphology (Figure 2d, f). TGF-β1 specificity is highlighted by the addition of TGF-β inhibitor that blocked changes in morphology after IGF-1+latent TGF-β1 treatment (Figure 2h). Inhibition of matrix metalloproteinase's using BB94 also abolished IGF-1 mediated, latent TGF-β1 signals, as cells retained epithelial morphology (Figure 2b). These findings indicated that IGF-1 treatment may have played a role in promoting the activation of latent TGF-β1 leading to EMT.


IGF-1 increases invasive potential of MCF 7 breast cancer cells and induces activation of latent TGF-β1 resulting in epithelial to mesenchymal transition.

Walsh LA, Damjanovski S - Cell Commun. Signal (2011)

IGF-1 and latent TGF-β1 caused morphological changes in MCF-7 cells consistent with EMT. Vehicle treated cells showed typical rounded epithelial morphology (a). Treatment of MCF-7 cells with 100 nM recombinant IGF-1 + 10 nM latent TGF-β1 caused a mesenchymal morphological phenotype characterized by a fibroblast-like appearance (g). Treatment with IGF-1 or latent TGF-β1 alone did not result in a morphological change (c, e). Pre-treatment of cells with PI3K or MAPK inhibitors abolished IGF-1 + latent TGF-β1 induced mesenchymal morphological phenotype (d, f). BB94 or TGF-β1 inhibitor prevented the morphological changes associated with IGF-1 + TGF-β1 treatment (b, h). Cytoskeleton was stained with phalloidin (red) and the nucleus with DAPI (blue). Images are representative of 3 independent experiments with consistent results.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3104381&req=5

Figure 2: IGF-1 and latent TGF-β1 caused morphological changes in MCF-7 cells consistent with EMT. Vehicle treated cells showed typical rounded epithelial morphology (a). Treatment of MCF-7 cells with 100 nM recombinant IGF-1 + 10 nM latent TGF-β1 caused a mesenchymal morphological phenotype characterized by a fibroblast-like appearance (g). Treatment with IGF-1 or latent TGF-β1 alone did not result in a morphological change (c, e). Pre-treatment of cells with PI3K or MAPK inhibitors abolished IGF-1 + latent TGF-β1 induced mesenchymal morphological phenotype (d, f). BB94 or TGF-β1 inhibitor prevented the morphological changes associated with IGF-1 + TGF-β1 treatment (b, h). Cytoskeleton was stained with phalloidin (red) and the nucleus with DAPI (blue). Images are representative of 3 independent experiments with consistent results.
Mentions: Recent evidence suggested that signaling after the proteolytic activation of latent TGF-β1 could induce EMT in breast cancer leading to metastasis [14]. There is also a strong correlation between high levels of IGF-1 signaling and metastasis [15]. The link between IGF-1 and TGFβ1 signaling however, has not thoroughly been explored. Here we show that treating MCF-7 cells first with IGF-1 (8 h), and then subsequently with latent TGF-β1 (48 h) resulted in a change to a cellular fibroblast-like morphology consistent with EMT (Figure 2g). MCF-7 vehicle treatment (control), or treatment with IGF-1 or latent TGF-β1 alone did not induce this change in cellular morphology (Figure 2a, c, e). Furthermore, pretreatment of cells with PI3K or MAPK inhibitors demonstrated the specificity of these pathways in IGF-1 mediated latent TGF-β1 induced morphological changes as it prevented changes in morphology (Figure 2d, f). TGF-β1 specificity is highlighted by the addition of TGF-β inhibitor that blocked changes in morphology after IGF-1+latent TGF-β1 treatment (Figure 2h). Inhibition of matrix metalloproteinase's using BB94 also abolished IGF-1 mediated, latent TGF-β1 signals, as cells retained epithelial morphology (Figure 2b). These findings indicated that IGF-1 treatment may have played a role in promoting the activation of latent TGF-β1 leading to EMT.

Bottom Line: Furthermore, most studies describing the biological effects of TGF-β have been performed using high concentrations of active, soluble TGF-β, despite the fact that TGF-β is produced and secreted as a latent complex.The effects of IGF-1 appear to be mediated through signals transduced via the PI3K and MAPK pathways.In addition, increased IGF-1, together with latent TGF-β1 and active MMPs result in EMT.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, University of Western Ontario, London, Ontario, N6A 5B7, Canada. sdamjano@uwo.ca.

ABSTRACT

Introduction: TGF-β signaling has been extensively studied in many developmental contexts, amongst which is its ability to induce epithelial to mesenchymal transitions (EMT). EMTs play crucial roles during embryonic development and have also come under intense scrutiny as a mechanism through which breast cancers progress to become metastatic. Interestingly, while the molecular hallmarks of EMT progression (loss of cell adhesion, nuclear localization of β-catenin) are straightforward, the cellular signaling cascades that result in an EMT are numerous and diverse. Furthermore, most studies describing the biological effects of TGF-β have been performed using high concentrations of active, soluble TGF-β, despite the fact that TGF-β is produced and secreted as a latent complex.

Methods: MCF-7 breast cancer cells treated with recombinant IGF-1 were assayed for metalloproteinase activity and invasiveness through a matrigel coated transwell invasion chamber. IGF-1 treatments were then followed by the addition of latent-TGF-β1 to determine if elevated levels of IGF-1 together with latent-TGF-β1 could cause EMT.

Results: Results showed that IGF-1 - a molecule known to be elevated in breast cancer is a regulator of matrix metalloproteinase activity (MMP) and the invasive potential of MCF-7 breast cancer cells. The effects of IGF-1 appear to be mediated through signals transduced via the PI3K and MAPK pathways. In addition, increased IGF-1, together with latent TGF-β1 and active MMPs result in EMT.

Conclusions: Taken together our data suggest a novel a link between IGF-1 levels, MMP activity, TGF-β signaling, and EMT in breast cancer cells.

No MeSH data available.


Related in: MedlinePlus