| Objective To evaluate the role of GPER1in the rearrangement ofactin cytoskeleton and cell migration induced by tamoxifen (Tam) inhuman breast cancer MCF-7cells.Methods MCF-7tamoxifen-resistant (Tam-R) cells were derivedfrom wild-type MCF-7(MCF-7W) cells by exposure to a highconcentration of4-hydroxytamoxifen (OHT) for a short period；meanmile，G1, the specific agonist of GPER1, was used to treat MCF-7cells (G1-R).The dynamic change of F-actin was visualized by FITC-Phalloidin staining.Immunofluorescence staining were used to evaluate the expression anddistribution of E-cadherin in MCF-7W cells, Tam-R cells and G1-R cells.Pull-down assay and western blot were utilized to analyzed the activity ofsmall GTPases Rac1. Transwell assay was used to evaluate the effects ofactin cytoskeleton rearrangement on migratory ability of Tam-R cells.Results In MCF-7W cells, the F-actin concentrated along the cellmembrane and E-cadherin formed strong intercellular adhesion junction. In contrast, abnormal lamellipodia, stress fiber and reduce ofE-cadherin-mediated cell-cell adhesion were observed in Tam-R cells andG1-R cells, which were derived from MCF-7W cells exposed to1μmol/LOHT and0.5μmol/L G1for6months. The migratory ability wassignificantly elvated in Tam-R cells and G1-R cells in comparison toMCF-7W cells（P <0.05）. The GPER1specifc agonist G1promotedrearrangement of actin cytoskeleton, activation of Rac1and cell migration（P <0.05）; the GPER1specifc inhibitor G15and PI3K inhibitorWortmannin (WM) attenuated activation of Rac1, rearrangement of actincytoskeleton and migration（P <0.05） induced by tamoxifen in Tam-Rcells. The ERK1/2inhibitor U0126had few effects on the actincytoskeleton rearrangement induced by OHT.Conclusion GPER1probably mediate OHT activation of PI3K andRac1to promote rearrangement of actin cytoskeleton and cell migration,played an important role in the development of tamoxifen resistance inbreast cancer. |
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