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. 2022 Aug 13;23(16):9086. doi: 10.3390/ijms23169086

Figure 3.

Figure 3

Involvement of EGFR in TNF-α-induced CCL20 expression. (A) Cells were pretreated with 10 μM AG1478 for 1 h and then incubated with 5 ng/mL TNF-α for 12 h. The conditioned media were utilized to determine the CCL20 level via an ELISA kit. (B) Cells were pretreated with 10 μM AG1478 for 1 h and then incubated with 5 ng/mL TNF-α for the indicated time. The mRNA levels and promoter activity of CCL20 were determined by real-time PCR (2 h) and promoter assay (4 h), respectively. (C) Cells were transfected with scrambled or EGFR siRNA and then incubated with TNF-α for 2 h. The mRNA levels of CCL20 were determined by real-time PCR. The protein levels of EGFR were determined by Western blot with GAPDH as a loading control. (D) Cells were pretreated without or with 1 μg/mL TNFR1 Ab or 10 μM AG1478 for 1 h and then treated with TNF-α for the indicated time (0, 5, 10, 15, 30, and 60 min). The phosphorylation of EGFR was determined by Western blot with GAPDH as a loading control. Data are expressed as mean ± S.E.M. of three independent experiments (n = 3). # p < 0.05, as compared with the cells exposed to vehicle alone; or significantly different as indicated.