| Objective To explore the way that 17β-estradiol induce endometriosis by ERαandβ-catenin.Methods Endometriosis stroma cells(ESC) from normal people were aparated and culture in vitro. Part 1, The ESC were treated by 17β-estradiol (17β-E2 )with various concentrations for 48h, expression ofβ-catenin mRNA and protein of these ESC were detected by RT-PCR and Western blot; Meanwhile, treated ESC with 17β-E2+ICI182, 780(10-6MOL/L), then detected theβ-catenin expression by the same method; Part 2, ESC were treated with 17β-estradiol with 10-8 mol/L, another group was treated together with ER inhibitor ICI182.780, compared there invasion ability with blank group by invasive assay; simultaneous, after transfected withβ-catenin siRNA cells were treated with 17β-E2 together with negative transfected group and blank group, then observed the invasion capability. Part 3 Used dual innumofluorescence study to confirm whether ERαand Lef-1/Tcf-3 ,β-catenin and Lef-1/Tcf-3 have colocalization. Immunoprecipitation to detected protein-protein interactions and the expression of ERα/Lef-1/Tcf-3 complex andβ-catenin/Lef-1/Tcf-3 complex after treated with 17β-estradiol indifferent time.Results 1. RT-PCR and Western blot showed that 17β-E2 stimulated the expression ofβ-catenin mRNA and protein in ESC in a dose-dependent manner, and the effect was maximum at 10-8 mol/L, while ICI182.780 could inhibit this upregulation; 2. The invasive assay show that the invasive capability of normal endometrium stroma cells were enhances by17β-estradiol but ICI182.780 andβ-catenin siRNA could reverse this appearance; 3. Dual innumofluorescence and immunoprecipitation show that ERαand Lef-1/Tcf-3,β-catenin and Lef-1/Tcf-3 colocated and interacted in ESC. With the treatment of 17β-estradiol, expression ofβ-catenin and ERαwas upregulated and expression of Lef-1 and Tcf-3 were higher than control group.Conclusion Estrogen may active Wnt/β-catenin signaling pathway via ERαand mediate endometriosis by this way. |
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