Figure 2

Tubular apoptosis and injury is increased in NOX4 KO mice compared to WT after IRI (A–E).

(A) Representative images of renal cortex TUNEL/DAPI staining and cortico-medullary Haematoxylin Eosin staining depicting tubular injury in WT and NOX4 KO mice 24 hours after IR are shown. Scale bar 50 μm. (B) Quantification of renal cortico-medullary junction injury score in NOX4 KO mice compared to WT (WT, n = 10; NOX4 KO, n = 9) after ischemia reperfusion. Results are expressed as the mean of arbitrary values ± SEM, ns p > 0.05, *p < 0.05. (C) Quantification of TUNEL staining of apoptotic cells in renal cortex of NOX4 KO mice compared to those of their WT littermates (WT, n = 10; NOX4, n = 9). Results are expressed as the mean ratio of the density of TUNEL positive cells measured in each sample over the mean density of TUNEL positive cells measured in WT ± SEM, ns p > 0.05, *p < 0.05. (D) Serum creatinine measurement performed on WT and NOX4 KO mice (WT, n = 10; NOX4 KO, n = 9) after IRI. Results are expressed as relative values of serum creatinine of NOX4 KO versus WT in different experiments in mean ± SEM, ns p > 0.05, *p < 0.05. (E) AnnexinV/PI assay analysis of cell apoptosis in MEF cells derived from WT and NOX4 KO mice. Results are expressed as mean percentage of late apoptotic cells (cells positive for both Annexin V and PI ± SEM, (n = 3), ns p > 0.05, *p < 0.05. (F) PARP and Cleaved PARP Western blot analysis performed on freshly isolated renal cortical tubule suspensions protein samples from WT and NOX4 KO. Representative picture and Western blot densitometric quantification are shown (WT n = 3; KO n = 3). Results are expressed as the mean of individual densitometric values over the mean densitometric value obtained in WT animals ± SE, ns p > 0.05 and *p < 0.05.