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. 2021 Sep 21;12:743991. doi: 10.3389/fphar.2021.743991

FIGURE 2.

FIGURE 2

Role of GRα in BPA-induced RACK1 regulation. (A) THP-1 cells transiently transfected with the Δ1 construct were pre-treated for 30 min with 20 μM mifepristone, then with 10 μM BPA for 16 h or DMSO as vehicle control (CTRL). Cells were lysed and luciferase activity was measured as described in “Materials and Methods” section. Luciferase activities are expressed as RLU% respected to non-treated construct (considered as 100%). Results are expressed as mean ± SEM, n = 3 independent experiments performed in triplicate. Statistical analysis was performed with Tukey’s multiple comparison test with *p < 0.05, ***p < 0.001 vs. control (CTRL) and §§ p < 0.01 vs. BPA 10 μM. B-C THP-1 cells pre-treated for 30 min with 20 μM mifepristone were treated for 18 h (B) or 24 h (C) with 10 μM BPA or DMSO as vehicle control (CTRL). (B) mRNA levels evaluated by qPCR (endogenous reference, 18S). (C) The image is a representative Western blot. RACK1 protein levels evaluated by Western blotting, normalized to β-tubulin expression. Each value represents the mean ± SEM, n = 3 independent experiments. Statistical analysis was performed with Tukey’s multiple comparison test with *p < 0.05, **p < 0.01, ***p < 0.001 vs control (CTRL) and §§ p < 0.01, §§§ p < 0.001 vs BPA 10 μM. (D) THP-1 cells silenced for 48 h with GRα-directed siRNA were treated for 24 h with 10 μM BPA or DMSO as vehicle control (CTRL). The image is a representative Western blot. RACK1 and GRα protein levels evaluated by Western blotting, normalized to β-tubulin expression. Statistical analysis was performed with Tukey’s multiple comparison test with *p < 0.05 vs control (CTRL) and §§ p < 0.01 vs. BPA-treated nonsilenced cells.