YTHDF2 Regulates Macrophage Polarization through NF-κB and MAPK Signaling Pathway Inhibition or p53 Degradation

Figure 5

Role of YTHDF2 silencing in activation of NF-κB and MAPK signalings within M1 macrophages. M0 macrophages were subject to transfection using YTHDF2 siRNAs or NC-siRNA for 24 h before 0-, 15-, 30-, 60-, and 120-min LPS+ INF-γ treatment. (a) p65, IκBα, and IKKα/β phosphorylation expression within NF-κB pathway were analyzed through WB, with VINCULIN being the endogenous reference. (b) Quantification of p65, IκBα, and IKKα/β phosphorylation levels in comparison with control. (c) p38, ERK, and JNK phosphorylation levels within MAPK pathway were analyzed through western blotting. (d) Quantification of p38, JNK, and ERK phosphorylation levels in comparison with control. (e, f) M0 macrophages subject to transfection using YTHDF2 siRNA or NC-siRNA were further exposed to BAY 11-7082, SB203580, or SP600125 (inhibitors for NF-κB, p38, and JNK pathways, separately) for a 2-h period, while non-treated cells served as blank control. Thereafter, LPS/IFN-γ was added to stimulate cells for a 6-h period. IL-6 and TNF-α mRNA expression were determined through qRT-PCR, with GAPDH being an endogenous reference. Data are denoted by (). ; .

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