Research Paper Volume 13, Issue 18 pp 22556—22570

Figure 3. Knockdown of NOX2 and NOX4 expression could reduce DOX-induced ROS production and expression of apoptotic proteins in H9c2 cells. (A) Fluorescence microscope observation of H9c2 cells after transfecting with Negative siRNA, NOX2 siRNA and NOX4 siRNA (×100). (B) Three distinct NOX2 siRNAs and one negative siRNA were designed, and Nox2-rat-663 was selected as the best interference fragment by PCR. (C) Three distinct NOX4 siRNAs and one negative siRNA were designed, and Nox4-rat-576 was selected as the best interference fragment by PCR. (D) Western blot analysis of NOX2 siRNA and DOX-treatment on NOX2, caspase-3, cleaved caspase-9 protein levels. (E) Densitometry analysis of the protein bands of NOX2, caspase-3, cleaved caspase-9 proteins. (F) Western blot analysis of NOX4 siRNA and DOX-treatment on NOX4, caspase-3, cleaved caspase-9 protein levels. (G) Densitometry analysis of the protein bands of NOX4, caspase-3, cleaved caspase-9 proteins. (H) Effect of NOX2 siRNA, NOX4 siRNA and DOX-treatment on ROS levels at 12 h time point in H9c2 cells. (I) Microscopic analysis of NOX2 siRNA, NOX4 siRNA and DOX-treatment on ROS levels by DCF Fluorescence. *P<0.05 vs. Control; †P<0.05 vs. Dox. DOX, doxorubicin; siRNA, small interfering RNA.