Journal of Immunology Research / 2018 / Article / Fig 2

Research Article

Activation of PAR4 Upregulates p16 through Inhibition of DNMT1 and HDAC2 Expression via MAPK Signals in Esophageal Squamous Cell Carcinoma Cells

Figure 2

Effect of MAPK on DNMT1, HDAC2, and p16 expression by PAR4-AP in human ESCC cells. (a) Western blotting showing p16 protein levels in ESCC cell lines (EC109 and TE-1) with PAR4-AP treatment at 0, 1, and 2 h, respectively. (b) RT-PCR analysis showing p16 gene levels in ESCC cell lines (EC109 and TE-1) with PAR4-AP treatment at 0, 1, and 2 h, respectively. The results were calculated by normalizing to β-actin in the same sample with the ΔCt method. The changes in the relative mRNA levels are expressed as fold changes compared with the controls. (c) Western blotting showing p-ERK1/2 and p-p38 expression in ESCC cells with PAR4-AP treatment at 0, 1, 2, 6, 12, and 24 h, respectively. (d) Western blotting analyses for DNMT1, HDAC2, and p16 protein expressions in ESCC cells following treatment with PAR4-AP for 2 h and pretreatment with U-46619 (an activator of ERK1/2 and p38), tBHQ (an activator of ERK1/2), PD98059 (an inhibitor of ERK1/2), or SB203580 (an inhibitor of p38) for 60 min. The mean optic densities of the proteins were calculated by normalizing to GAPDH. All values are expressed as the means ± SEMs (). versus controls; versus PAR4-AP-only groups.
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