Mechanism Of 17β-Estradiol Induced Proliferation Of Anaplastic Thyroid Carcinoma FRO Cells Through ERA And GPER

Objective To investigate the effects of estrogen receptorα(ERα) and G protein-coupled estrogen receptor (GPER) on 17β-Estradiol (E2) induced proliferation of anaplastic thyroid carcinoma FRO cells.Methods FRO cells were treated with E2(17β-Estradiol, E2), PD98059(inhibitor of ERK1/2), LY294002(inhibitor of Akt), ICI182780(inhibitor of estrogen receptor), GPER-siRNA(GPER-small interfering RNA), and then the cell growth rates were evaluated by MTT assay, changes of cell cycle were analyzed by flow cytometry, the levels of phosphorylated ERK1/2, phosphorylated Akt, GPER, ERα, ERβ, Bcl-2 and Bax were analyzed by Western blot analysis.Results When FRO cells were treated with different concentrations (0, 10^-10 mol/L, 10^-9 mol/L, 10^-8 mol/L) of E2 for 24h, the cell growth rates were 0, (6.5±1.1)%, (10.7±1.9)% and (16.5±2.1)%. When FRO cells were treated with 10-8 mol/L of E2 for different times(0 h, 12 h, 24 h), the cell growth rates were 0, (8.2±1.9)% and (16.5±2.1)% (p<0.05) and the percentages of S/G2/M phase were (21.51±4.32)%, (37.22±4.16)% and (48.56±2.31)% (p<0.05). The levels of ERαand Bcl-2 were increased after FRO cells were treated with E2 for different times (0 h, 12 h, 24 h), while the level of Bax was decreased. ICI182780 could reduce the level of Bcl-2, but had no effects on Bax expression. When 10-8 mol/L of E2 treated FRO cells for different time points(0 min, 5 min, 10min, 15min, 30min), the level of phosphorylated ERK1/2 and phosphorylated Akt increased and the accumulation of phosphorylated ERK1/2 and phosphorylated Akt reached peak at 15 min and 10 min respectively. The expression of GPER reduced significantly when FRO cells was transfected with GPER-siRNA for 48 h. Pretreatment with PD98059 (30μmol/L), LY294002 (50μmol/L) and GPER-siRNA obliterated the inductive effects of E2 on ERK1/2 and Akt phosphorylation at 15 min and 10 min respectively in FRO cells. Meanwhile compared with E2-treated cells, pretreatment with PD98059, LY294002, GPER-siRNA and ICI182780 obliterated the inductive effects on cell growth by E2 for 24h and the cell growth rates reduced from (16.5±2.1)% to (11.2±1.3)%, (9.6±1.5)%, (7.2±1.3)% and (6.9±1.6)%.Conclusion E2 can promote proliferation of FRO cells, not only via increasing the rate of Bcl-2/Bax through ERαbut also via ERK1/2 and PI3K-Akt pathway in a GPER dependent manner.