Fig. 2

Knockdown of PIAS1 aggravates injury after H/R via activating NF-κB pathways. a Western blot analysis of PIAS1 proteins in H9C2 cells transfected with si-NC (non-specific siRNA) or si-PIAS1 (against PIAS1 siRNA). b TUNEL staining of apoptotic H9C2 cells transfected with si-NC or si-PIAS1 under normoxic condition or H/R treatments. Scale bars, 100um. c Quantification of apoptotic rates based on TUNEL staining (n = 5). d RT-PCR analysis of IL-1β, IL-6 and TNFα in H9C2 cells transfected with si-NC or si-PIAS1 under normoxic condition or H/R treatment (n = 3). e Western blot analysis of phosphorylated IκBα proteins in H9C2 cells transfected with si-NC or si-PIAS1 under normoxic condition or H/R treatments. Quantification of the densitometry of the western blot band is shown above (n = 3). f Representative immunofluorescence images of p65 proteins (Green) in H9C2 cells transfected with si-NC or si-PIAS1 under normoxic condition or H/R treatments. DAPI indicates cell nucleus. Scale bars, 100um. g Quantification of nuclear p65-positive H9C2 cells based on TUNEL staining (n = 5). Experiments in a, d and e were performed three times, and experiments in b, c, f and g were performed five times. Data presented are means ± SD, ^#P < 0.05 vs. H/R + si-NC group, ^**P < 0.01 vs. sham group, and ^##P < 0.01 vs. H/R + si-NC group