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. 2014 May 16;9(5):e97824. doi: 10.1371/journal.pone.0097824

Figure 3. APN Induces FoxO1 and JNK Phosphorylation in RANKL-Treated RAW264.7 Cells.

Figure 3

(A) qRT-PCR of FoxO1 mRNA levels in RAW264.7 cells treated with 50 ng/ml RANKL in the absence (control) and presence of 0.5 µg/ml APN treatment for 24 hours. The mRNA level was normalized with those of GAPDH. (B) Western blot for phosphorylated JNK in RAW264.7 cells treated with 50 ng/ml RANKL in the absence or presence of 0.5 µg/ml APN for 15 or 30 minutes. JNK was detected as the loading control. Data are shown as mean ± SD of three independent experiments. *P<0.05. (C) Western blot of FoxO1 nuclear protein extracted from RAW264.7 cells that were treated with 50 ng/ml RANKL in the presence and absence of 0.5 µg/ml APN, with or without prior treatment with the JNK inhibitor SP600125 for 2 hours. DMSO was used to dissolve SP600125. Nuclear Lamin B1 was used as the loading control. Data are shown as mean ± SD of three independent experiments. *P<0.05.