Estrogen/progestogen Receptor

Kaempferol and ER /PR inhibitor suppress the migration and Rho activity of MCF-7 cells. Notes: (A) MCF-7 ER +/PR+ breast cancer cells were allowed to migrate in response to 20 μmol/L kaempferol and/or 0.1 nmol/L AZD9496 (ER inhibitor) and 200 μmol/L megestrol acetate (PR inhibitor) for 6 h. Results are presented as mean ± SD of 5 independent experiments. (B, C) MCF-7 cells were incubated with 20 μmol/L kaempferol and/or 0.1 nmol/L AZD9496 and 200 μmol/L MA for 1 h, and then subjected to the RhoA (B) and Rac1 (C) activity assays. The relative levels of Rho activity were normalized to the average value of controls. Results are presented as mean ± SD of 3 independent experiments. Abbreviations: AZD, AZD9496; Ctrl, control; ER, estrogen receptor; MA, megestrol acetate; ns, no significance; PR, progesterone receptor.

PTPH1 confers breast cancer cell sensitivity to fulvestrant. E and F, PTPH1 overexpression increases the growth inhibition by fulvestrant. PTPH1 was overexpressed by a Tet-on system or a stable transfection, and resultant cells were incubated with fulvestrant as indicated for about 2 weeks. Colony formed was stained and counted. Results shown are normalized to its own solvent control of Vector and PTPH1-overexpressed cells, respectively (means ± SD; n = 3–5) with insets showing PTPH1 overexpression. *, versus vector or no Tet cells for E and F.

Effects of DPN, LY500307, Raloxifene and Tamoxifen on cell viability in BSO-treated FRDA fibroblasts. BSO concentration was 1 mM and all steroid concentrations were 100 nM. Depicted are mean ± SD for n= 8 per group. * indicated p<0.05 versus BSO alone-treated cells.

a, Quantification of sterol levels in OPCs (oligodendrocyte progenitor cells) treated with the indicated molecules at 2 μM.

Myogenic differentiation assay to determine the GR specificity of DEX by using RU-486 (10 uM). Immunofluorescence detection of MyHC (red) and DAPI counterstaining of nuclei (blue) was used to detect myotubes. The scale bar is 50 um.

E, Analysis of BrCSC viability upon treatment with tamoxifen (tam) and fulvestrant (fulv) at the indicated doses. The experiments were performed in triplicates. ** P<0.01.

The analgesic effect of koumine was reversed in sciatic nerve chronic constriction injury rats by intrathecal treatment with medroxyprogesterone acetate. Medroxyprogesterone acetate (MPA) or dimethyl sulfoxide (DMSO, vehicle) was administered via an intrathecal catheter 10 days after sciatic nerve chronic constriction injury (CCI) surgery. After 30 min, koumine (7 mg kg−1 bw) or normal saline (NS) was administered by subcutaneous (s.c.) injection. The mechanical withdrawal threshold (MWT) of the hind paws was measured 1 h after completion of the drug or vehicle administration. The data are presented as the mean ± SEM (n = 6 per group) and were analyzed by one-way ANOVA followed by Bonferroni post hoc test at each time point. *P < 0.05 vs. the DMSO + NS group; # P < 0.05 vs. the DMSO + koumine group.

Metformin alleviated EP-induced decidualization of endometrial stromal cells. (A–D) Endometrial stromal cells were treated with or without estradiol (10 nM) and progesterone (1 μM) in the absence or presence of metformin (1 mM) for 10 days or 14 days. Then, mRNA levels of IGFBP-1 and prolactin in endometrial stromal cells were examined by RT-qPCR assay (A and B). Secreted protein levels of IGFBP-1 and prolactin into media were tested by matching ELISA kits (C and D). #* P < 0.05.

Representative images of OPCs treated for 72 h with the indicated small molecules. All molecules were used at 300 nM. Scale bar, 100 μm.

(a) Inhibitory activities of IDH1R132H enzyme lead to decreased production of D-2HG in IDH1R132H mutant HT1080 cells after treatment with clomifene for 48 h, and D-2HG was measured by LC/MS method. (b) HT1080 cells were treated for 24 h in the different concentrations of clomifene and then processed for FACS by using Annexin V/propidium iodide staining. (c) Treatment of clomifene significantly reduced methylation levels of histone lysine residues in HT1080 cells. All data are the average of results from triplicate experiments.

CDK4/6 inhibitor resistance promotes diminished ER expression and activity. (a and b) Proliferation of MCF-7 and MR cells in the presence of different concentrations of ICI (a) or 4-hydroxytamoxifen (4-OHT) (b) was measured by the alamarBlue assay and plotted as % inhibition of proliferation against log concentration of inhibitor. Each data point represents the average value±s.d. of six replicates obtained in three independent experiments.

Total cellular protein was harvested after 8 days and the protein levels of adipogenic factors were measured using western blot analysis with specific antibodies (B).

Kaempferol and ER /PR inhibitor suppress the migration and Rho activity of MCF-7 cells. Notes: (A) MCF-7 ER +/PR+ breast cancer cells were allowed to migrate in response to 20 μmol/L kaempferol and/or 0.1 nmol/L AZD9496 (ER inhibitor) and 200 μmol/L megestrol acetate (PR inhibitor) for 6 h. Results are presented as mean ± SD of 5 independent experiments. (B, C) MCF-7 cells were incubated with 20 μmol/L kaempferol and/or 0.1 nmol/L AZD9496 and 200 μmol/L MA for 1 h, and then subjected to the RhoA (B) and Rac1 (C) activity assays. The relative levels of Rho activity were normalized to the average value of controls. Results are presented as mean ± SD of 3 independent experiments. Abbreviations: AZD, AZD9496; Ctrl, control; ER, estrogen receptor; MA, megestrol acetate; ns, no significance; PR, progesterone receptor.

PTPH1 confers breast cancer cell sensitivity to fulvestrant. E and F, PTPH1 overexpression increases the growth inhibition by fulvestrant. PTPH1 was overexpressed by a Tet-on system or a stable transfection, and resultant cells were incubated with fulvestrant as indicated for about 2 weeks. Colony formed was stained and counted. Results shown are normalized to its own solvent control of Vector and PTPH1-overexpressed cells, respectively (means ± SD; n = 3–5) with insets showing PTPH1 overexpression. *, versus vector or no Tet cells for E and F.

Effects of DPN, LY500307, Raloxifene and Tamoxifen on cell viability in BSO-treated FRDA fibroblasts. BSO concentration was 1 mM and all steroid concentrations were 100 nM. Depicted are mean ± SD for n= 8 per group. * indicated p<0.05 versus BSO alone-treated cells.

a, Quantification of sterol levels in OPCs (oligodendrocyte progenitor cells) treated with the indicated molecules at 2 μM.

Myogenic differentiation assay to determine the GR specificity of DEX by using RU-486 (10 uM). Immunofluorescence detection of MyHC (red) and DAPI counterstaining of nuclei (blue) was used to detect myotubes. The scale bar is 50 um.

E, Analysis of BrCSC viability upon treatment with tamoxifen (tam) and fulvestrant (fulv) at the indicated doses. The experiments were performed in triplicates. ** P<0.01.

The analgesic effect of koumine was reversed in sciatic nerve chronic constriction injury rats by intrathecal treatment with medroxyprogesterone acetate. Medroxyprogesterone acetate (MPA) or dimethyl sulfoxide (DMSO, vehicle) was administered via an intrathecal catheter 10 days after sciatic nerve chronic constriction injury (CCI) surgery. After 30 min, koumine (7 mg kg−1 bw) or normal saline (NS) was administered by subcutaneous (s.c.) injection. The mechanical withdrawal threshold (MWT) of the hind paws was measured 1 h after completion of the drug or vehicle administration. The data are presented as the mean ± SEM (n = 6 per group) and were analyzed by one-way ANOVA followed by Bonferroni post hoc test at each time point. *P < 0.05 vs. the DMSO + NS group; # P < 0.05 vs. the DMSO + koumine group.

Metformin alleviated EP-induced decidualization of endometrial stromal cells. (A–D) Endometrial stromal cells were treated with or without estradiol (10 nM) and progesterone (1 μM) in the absence or presence of metformin (1 mM) for 10 days or 14 days. Then, mRNA levels of IGFBP-1 and prolactin in endometrial stromal cells were examined by RT-qPCR assay (A and B). Secreted protein levels of IGFBP-1 and prolactin into media were tested by matching ELISA kits (C and D). #* P < 0.05.

Representative images of OPCs treated for 72 h with the indicated small molecules. All molecules were used at 300 nM. Scale bar, 100 μm.

(a) Inhibitory activities of IDH1R132H enzyme lead to decreased production of D-2HG in IDH1R132H mutant HT1080 cells after treatment with clomifene for 48 h, and D-2HG was measured by LC/MS method. (b) HT1080 cells were treated for 24 h in the different concentrations of clomifene and then processed for FACS by using Annexin V/propidium iodide staining. (c) Treatment of clomifene significantly reduced methylation levels of histone lysine residues in HT1080 cells. All data are the average of results from triplicate experiments.

CDK4/6 inhibitor resistance promotes diminished ER expression and activity. (a and b) Proliferation of MCF-7 and MR cells in the presence of different concentrations of ICI (a) or 4-hydroxytamoxifen (4-OHT) (b) was measured by the alamarBlue assay and plotted as % inhibition of proliferation against log concentration of inhibitor. Each data point represents the average value±s.d. of six replicates obtained in three independent experiments.

Total cellular protein was harvested after 8 days and the protein levels of adipogenic factors were measured using western blot analysis with specific antibodies (B).