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 (O(2) (-)) 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 O(2) (-) and NO generation, lipid peroxidation (MDA) and monocyte/macrophage infiltration were found in nephrotic animals. Renal oxidative stress (O(2) (-), 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.

Increased apoptosis in acute puromycin aminonucleoside nephrosis.

Acute puromycin aminonucleoside nephrosis (PAN) in rats is characterized by heavy proteinuria associated with renal hypercellularity. The role of apoptosis in the resolution of renal hypercellularity was investigated in PAN. To study the participation of apoptosis in PAN, renal tissues were collected from nephrotic and control rats on weeks 1, 2 and 7 after a single puromycin aminonucleoside injection. Apoptosis was evaluated by light and electron microscopy. Apoptotic DNA fragmentation was detected by TUNEL staining. Renal tissues were also evaluated by the presence of leukocyte common antigen (LCA), ED1 (macrophages) and proliferating cell nuclear antigen (PCNA) with corresponding monoclonal antibodies. An increased number of apoptotic (TUNEL+) cells was observed in the glomerulus at week 1. Electron microscopy analysis showed glomerular apoptosis mainly in endothelial cells. In the interstitium and tubules, increased apoptosis was observed at weeks 1 and 2. Increased apoptosis was accompanied with increased LCA+, ED1+ and PCNA+ cells in the interstitium and with increased PCNA+ cells in tubules. There was a high significant correlation between the number of apoptotic cells and the number of interstitial LCA+, ED1+ and PCNA+ cells. Tubular PCNA expression was correlated with tubular apoptosis. We also observed significant correlation between glomerular, interstitial and tubular apoptosis with proteinuria during the nephrosis. Double staining analysis showed that about 13% of interstitial or tubular apoptotic cells were positive for PCNA. All these values returned to normal by week 7. These results indicate that apoptosis is involved in the repairing process of this disease model.

Distal nephron sodium handling by normal, nephrotic, pyelonephritic and partially nephrectomized rats.

The renal tubular handling of sodium was evaluated in 10 normal (N), 9 edematous nephrotoxic serum nephrotic (Ne), 9 pyelonephritic (Py) and 12 partially nephrectomized (Nx) Wistar rats during maximal free-water clearance induced by intravenous infusion of hypotonic sodium chloride solutions. Fractional deliveries of sodium to distal nephron [(CH2O + CNa)/GFR%] were similar (P greater than 0.05) in N (23.2 +/- 0.69%), Ne (24.3 +/- 1.30%) and Nx (26.9 +/- 2.6%) rats. However, the fractional distal sodium reabsorption [CH2O/(CH2O + CNa)%] was significantly higher (P less than 0.05) in Ne (76.8 +/- 1.80%) and lower (P less than 0.05) in Nx (33.3 +/- 3.1%) rats when compared to normal rats (50.3 +/- 1.80%). Py rats exhibited a higher fractional delivery of sodium to distal nephron (33.5 +/- 3.4%) and a lower fractional distal sodium reabsorption (35.9 +/- 5.9%) when compared to normal rats (P less than 0.05). These data suggest that the distal nephron, by altering sodium reabsorption, contributes to the maintenance of sodium balance in Nx rats, whereas in Ne rats the distal nephron generates a positive sodium balance and edema. At very low glomerular filtration rates (GFR), as is the case for Py rats, sodium balance is maintained both by the proximal and distal portions of the nephron.