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Fig. 4 | Breast Cancer Research

Fig. 4

From: TNFAIP3 is required for FGFR1 activation-promoted proliferation and tumorigenesis of premalignant DCIS.COM human mammary epithelial cells

Fig. 4

AP20187-induced TNFAIP3 expression is dependent on ERK1/2 activation in DCIS-iFGFR1 cells. a TNFAIP3 mRNA expression. DCIS-Ctrl (Control) and DCIS-iFGFR1 cells were treated with vehicle, FGFR1/2/3 inhibitor LY2874455 (LY, 500 nM) or FGFR1/2/3/4 inhibitor AZD4547 (AZD, 500 nM) as indicated for 1 h. Then, AP20187 (AP, 100 nM) was added to treat cells for another 6 h. TNFAIP3 mRNA was analyzed by real-time RT-qPCR and normalized to β-actin mRNA. Data were mean ± standard deviation (SD) of three independent experiments. * and ****p < 0.05 and 0.0001 vs. vehicle-treated group by one-way ANOVA. b Immunoblotting analysis. DCIS-Ctrl and DCIS-iFGFR1 cells were treated as described above for Panel A, except that cells were treated for another 24 h after adding AP20187. The ratios of TNFAIP3 band intensity to GAPDH band intensity were calculated from three repeating experiments. *p < 0.05 between vehicle- and Ap20187-treated groups by one-way ANOVA. c and d DCIS-iFGFR1 cells were treated with vehicle, MEK inhibitor PD0325901 (100 nM) or ERK1/2 inhibitor GDC0994 (1 μM) for 1 h, then AP20187 (100 nM) was added to treat the indicated cells for another 24 h. TNFAIP3 mRNA was analyzed by real-time RT-qPCR and normalized to β-actin mRNA. ** and ***p < 0.01 and p < 0.001 vs. vehicle-treated group by one-way ANOVA. (Panel C). Immunoblotting was performed by using antibodies against TNFAIP3, p-RSK, total RSK, p-ERK1/2, and total ERK1/2. The ratios of TNFAIP3 band intensity to β-actin band intensity were calculated from three repeating experiments. *p < 0.05 between vehicle- and AP20187-treated groups by one-way ANOVA; no significant differences between vehicle-treated group and all other inhibitor-treated groups (Panel D)

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