Increased oxidative stress and apoptosis in acute puromycin aminonucleoside nephrosis

Accumulating evidence demonstrates that oxidative stress is one of the underlying mechanisms to induce apoptosis in different biological systems. The aim of this study was to examine the simultaneous presence and correlation between oxidative stress events, apoptosis, apoptosis-associated proteins and monocyte/macrophage infiltration during the course of acute puromycin aminonucleoside nephrosis (PAN). To induce nephrosis, Sprague-Dawley rats were injected intraperitoneally with puromycin aminonucleoside and killed at weeks 1 and 2 of nephrosis. Controls represent animals injected with 0.9% saline solution. Kidney sections were homogenized to measure nitric oxide (NO), malondialdehyde (MDA), reduced glutathione (GSH), superoxide dismutase (SOD) and catalase activities by appropriate enzymatic and biochemical methods. Renal frozen sections were studied for superoxide anion (O₂^-) by a histochemical method, for apoptosis by TUNEL (terminal-deoxynucleotidyl- transferase-mediated dUTP-digoxigenin nick end labelling) and for apoptosis-associated protein expression and monocyte/macrophage infiltration by monoclonal antibodies. Increased renal apoptosis, p53, Bax, Bcl-2 accompanied by increased O₂^- and NO generation, lipid peroxidation (MDA) and monocyte/macrophage infiltration were found in nephrotic animals. Renal oxidative stress (O₂^-, NO and MDA) was correlated with apoptosis, p53 expression, monocyte/macrophage cells and proteinuria. Anti-oxidant molecules (SOD and GSH) remained unchanged apart from a decreased activity of catalase which correlated with glomerular apoptosis. In conclusion, the close correlation between the presence of apoptosis and oxidative events confirms the role of oxidative stress in the apoptosis observed during PAN.