CD38 Deficiency Protects the Heart from Ischemia/Reperfusion Injury through Activating SIRT1/FOXOs-Mediated Antioxidative Stress Pathway

Figure 4

CD38 knockdown protects cells from H/R-induced injury in H9c2 cells through activating FOXOs signaling pathway. (a–c) The protein expressions of FOXO1 and FOXO3 in H9c2 cells. The FOXO proteins were examined by western blot in CD38 knockdown H9c2 cells (a) and the quantitative analysis of the protein expressions of FOXO1 (b) and FOXO3 (c) was performed. (d) The immunofluorescence images of nuclear localization of FOXO1. The H9c2 cells were subjected to immunostaining with anti-FOXO1 antibody (green) and DAPI (blue) (magnification, ×400 scale bars, 20 μm). (e–j) Expressions of SOD2 and catalase (CAT). The mRNA and protein expressions of FOXO1 targeted genes SOD2 and CAT were determined by qPCR (e, h) and western blot with SOD2 (f, g) and CAT (i, j) antibodies, respectively. GAPDH was used as loading control and the data were presented as mean ± SD, , and and . (k) Cell viabilities after H/R stimulation with various reagents. The cell viabilities were determined by CCK8 assay after treatment with SIRT1 specific inhibitor EX527 and PI3K inhibiter LY294002 followed with H/R stimulation in H9c2 cells.

(a)

(b)

(c)

(d)

(e)

(f)

(g)

(h)

(i)

(j)

(k)