Figure 5

Representative immunoblotting and immunofluorescent analyses for selected differentially expressed proteins identified by proteomic analysis of H9C2 cells in response to doxorubicin treatment and pre-treatment with quercetin. The levels of identified proteins, (A) Aconitase, (B) ATP synthase, (C) Carbonic anhydrase, (D) GRP78, (E) HSP27, (F) HSP60, (G) Peroxiredoxin 6, (H) Tropomyosin 4, (I) Vimmentin, in H9C2 cells in response to doxorubicin treatment and pre-treatment with quercetin were confirmed by immunoblot, while GAPDH was used as loading controls (left panels). The protein expression maps and two-dimensional spot images were shown in right panels and middle panels, respectively. (J) H9C2 cells in different treatment conditions were fixed and incubated with anti-Cofilin-1 antibody (Red) and stained with Phalloidin (Green). Nucleus were stained with DAPI (Blue). Each set of three fields was taken using the same exposure, and images are representative of three different fields. In this validation experiment, H9C2 cells were left untreated, treated with 0.45 μM doxorubicin for 24 h or pretreated with 100 μM quercetin for 4 h followed by 0.45 μM doxorubicin for further 24 h.