Research Paper Volume 11, Issue 6 pp 1745—1758

H2S restores the cardioprotective effects of ischemic post-conditioning by upregulating HB-EGF/EGFR signaling

Figure 7. Knockdown of EGFR cancels the effect of exogenous H2S on cell damage and related signaling pathways in the aged H9C2 cells. (A) The knockdown of EGFR by EGFR-specific siRNA (EGFR siRNA) in the aged H9C2 cells. The cells were transfected with 50 nM EGFR siRNA or negative control siRNA (control siRNA) for 48 h during P(C) The data are the means ± S.E.M. of 3 determinations. * p<0.05 vs. PC + control siRNA group. (B) The knockdown of EGFR inhibited NaHS-increased expression of the HB-EGF and the activity of phosphorylated EGFR. The graphs represent the optical density of the bands of phosphorylated EGFR (p- EGFR) normalized to the expression of total EGFR (t-EGFR). The graphs represent the optical density of the bands of HB-EGF normalized to the expression of GAPDH signal. All data were from three independent experiments. * p<0.05 vs. PC group; # p<0.05 vs. PC + control siRNA + NaHS group. (C) The knockdown of EGFR cancelled NaHS-decreased cell damage. The cells were cultured in glass-bottom dishes and observed using a general inverted microscope (magnification ×100). (D) The knockdown of EGFR inhibited NaHS-up-regulated the ERK1/2 and PI3K-Akt-GSK-3β pathways. The phosphorylation of ERK1/2, PI3K, Akt and GSK-3β was detected using western blotting analysis. The graphs represent the optical density of the bands of phospho-ERK1/2 (p-ERK1/2), PI3K (p- PI3K), Akt (p-Akt) and GSK-3β (p-GSK-3β) normalized to the expression of total-ERK1/2 (t-ERK1/2) PI3K (t-PI3K), Akt (t-Akt) and GSK-3β (t-GSK-3β). All data were from three independent experiments. * p<0.05 vs. PC group; # p<0.05 vs. PC + control siRNA + NaHS group.