Research Paper Volume 13, Issue 6 pp 8380—8395

Figure 5. Extracellular HMGB1 upregulates miR-21 expression via activating Rage/JNK signaling pathway. (A) Immunoblot analysis indicates rhHMGB1 promotes CD44 expression by activating Rage/JNK signaling pathway. HepG2 and HCCLM3 cells were treated with negative control, Rage siRNA, JNK inhibitor(SP600125,SP,20μm) and rhHMGB1 (1μg/ml) for 24h. (B) Q-PCR analysis indicates inactivating Rage/JNK signaling downregulates miR-21 expression caused by rhHMGB1. HepG2 and HCCLM3 cells were treated with negative control, Rage siRNA, SP(20μm) and rhHMGB1 (1μg/ml) for 24h. (C–E) Rage/JNK pathway accounts for HCC sphere formations and invasion caused by rhHMGB1. Number and size of spheres and invaded cells were photographed and analyzed. HepG2 cells were treated with negative control, Rage siRNA, SP(20μm) and rhHMGB1 (1μg/ml) for 24h. (F) Immunoblot analysis indicates miR-21 mimic, Rage siRNA and SP all decrease CD44 expression and OCT4 nuclear translocation caused by rhHMGB1. HepG2 cells were treated with negative control, Rage siRNA, SP(20μm), miR-21 mimic and rhHMGB1 (1μg/ml) for 24h. Data are means ± SEM, * means p<0.05, ** means p<0.01, *** means p<0.001 by unpaired student T test.