The role of nitric oxide pathway in the renal ischemia-reperfusion injury in rats

INTRODUCTION: Nitric oxide (NO), synthesized from L-arginine by the enzyme nitric oxide synthase (NOS), seems to play an ambiguous role during tissue ischemia-reperfusion injury. Our objective was to evaluate the effects of L-arginine, a NO donor, and N(G)-nitro-L-arginine-methylester (L-NAME), a NOS inhibitor, on oxidative stress, renal dysfunction, histologic alterations and surgical mortality rate induced by renal ischemia-reperfusion (RIR) in uninephrectomized rats. MATERIALS AND METHODS: One-hundred and ninety-seven Wistar rats were randomized into five experimental groups. Group 1: sham operation; group 2: right uninephrectomy (UNI); group 3: UNI + RIR in the contralateral kidney; group 4: UNI + L-NAME (20 mg/kg; intraperitoneally) + RIR; and group 5: UNI + L-arginine + RIR. The effect of the drugs was evaluated by lipid peroxidation measured by the renal malondialdehyde (MD) content and chemiluminescence (CL) levels, serum creatinine (Cr) levels, urinary volume, tubular necrosis and athrophy, inflammatory infiltrate, interstitial fibrosis as histologic evaluation and surgical mortality rate after the procedures. A P value less than 0.05 was considered significant. RESULTS: Right uninephrectomy did not alter the renal parameters. RIR increased Cr levels (at 24 and 96 h of reperfusion), index of lipid peroxidation (both MD and QL levels), and worsened the histologic aspects. Pretreatment with L-arginine reduced the kidney levels of QL when compared with the non-treated group (5574 +/- 909 vs. 13 660 +/- 1104 cps/mg of protein; P < 0.05) but increased the MD levels (0.97 +/- 0.24 vs. 0.79 +/- 0.06 nmol/mg of protein; P < 0.05). Moreover, L-arginine attenuated the increment of Cr levels, inflammatory infiltrate and tubular athrophy in rats subjected to RIR (P < 0.05). On the other hand, pretreatment with L-NAME increased both CL (17 482 +/- 4397 vs. 13 660 +/- 1104 cps/mg of protein; P < 0.05) and MD levels (1.16 +/- 0.11 vs. 0.79 +/- 0.06 nmol/mg of protein; P < 0.05). Furthermore, L-NAME worsened the renal dysfunction (P < 0.05) at 192 h after the RIR, and surgical mortality rates were similar (P > 0.05). CONCLUSION: L-arginine has a tendency to exert a beneficial effect on renal damage during RIR in rats. Moreover, L-NAME seems to worsen the renal damage by increasing the kidney-levels of CL and impairment of renal function probably due to reduction of NO production.     

Renal protective effect of molsidomine and L-arginine in ischemia-reperfusion induced injury in rats

BACKGROUND: Nitric oxide (NO), synthesized from L-arginine by the enzyme NO synthase (NOS) seems to play an ambiguous role during tissue ischemia-reperfusion (I/R) injury. This study was designed to investigate the effects of molsidomine, a NO donor and L-arginine in I/R induced renal failure in rats METHODS: The protective effect of molsidomine and L-arginine against the damage inflicted by I/R was investigated in Sprague-Dawley rats. In one set of experiments animals were unilaterally nephrectomized, and subjected to 45 min of left renal pedicle occlusion and in another set both the renal pedicles were occluded for 45 min followed by 24 h of reperfusion. Molsidomine (10 mg/kg, p.o.) was administered twice, 30 min before ischemia and 12 h after the reperfusion period, while L-arginine was administered once, 30 min before ischemia. At the end of the reperfusion period, rats were sacrificed. Tissue and urine nitrite levels were measured to assess the total NO levels. Thiobarbituric acid reactive substances (TBARS), reduced glutathione (GSH) levels, catalase (CAT), and superoxide dismutase (SOD) activities were determined in renal tissue. Serum creatinine and BUN concentrations were measured for the evaluation of renal function. RESULTS: Ischemic control animals demonstrated severe deterioration of renal function, renal morphology, reduced levels of tissue, and urine NO levels and a significant renal oxidative stress. Pretreatment of animals with molsidomine and L-arginine markedly attenuated renal dysfunction, morphological alterations, improved the tissue as well as urine NO contents, reduced elevated TBAR levels and restored the depleted renal antioxidant enzymes. CONCLUSIONS: The findings imply that NO play a causal role in I/R induced renal injury.     

Endogenous nitric oxide and exogenous nitric oxide supplementation in hepatic ischemia-reperfusion injury in the rat

BACKGROUND: Although nitric oxide (NO) is thought to be beneficial in hepatic ischemia-reperfusion (I/R), the mechanisms for this effect are not well established. METHODS: To investigate the effects of endogenous NO and exogenous NO supplementation on hepatic I/R injury and their pathogenic mechanisms, serum ALT and hyaluronic acid (endothelial cell damage), and hepatic malondialdehyde and H2O2 (oxidative stress), myeloperoxidase activity (leukocyte accumulation), and endothelin (vasoconstrictor peptide opposite to NO) were determined at different reperfusion periods in untreated rats and rats receiving L-NAME, L-NAME+L-arginine, and spermine NONOate (exogenous NO donor). RESULTS: After reperfusion every parameter increased in untreated animals. Endogenous NO synthesis inhibition by L-NAME increased hepatocyte and endothelial damage as compared to untreated rats, which was reverted and even improved by the addition of L-arginine. Spermine NONOate also improved this damage. However, different mechanisms account for the beneficial effect of endogenous and exogenous NO. Oxidative stress decreased by both L-NAME and L-NAME+L-arginine, but remained unmodified by spermine NONOate. Myeloperoxidase increased by L-NAME and this effect was reverted by the addition of L-arginine, whereas no change was observed with spermine NONOate. Endothelin levels were not modified by L-NAME and L-NAME+L-arginine, but decreased with spermine NONOate. CONCLUSIONS: These results suggest that, although both endogenous and exogenous NO exert a protective role in experimental hepatic I/R injury, the mechanisms of the beneficial effect of the two sources of NO are different.     

Effects of L-arginine on the kidney levels of malondialdehyde in rats submitted to renal ischaemia-reperfusion

OBJECTIVE: To evaluate the effects of L-arginine, a nitric oxide donor, on kidney levels of malondialdehyde (MDA, a product of cellular lipid peroxidation), serum creatinine levels, and urinary volume in rats undergoing unilateral renal ischaemia-reperfusion. MATERIALS AND METHODS: Wistar rats (117) were randomly distributed into three experimental groups (of four subgroups each) in which were assessed renal cell-lipid peroxidation (kidney levels of MDA), serum creatinine levels and urinary volume. The rats underwent unilateral nephrectomy followed by contralateral renal ischaemia-reperfusion with or with no pretreatment with L-arginine (200 mg/kg) given intraperitoneally. RESULTS: Pretreatment with L-arginine caused significantly higher kidney levels of MDA than in the untreated group (P < 0.05). Furthermore, L-arginine given before surgery attenuated the increase in serum creatinine and significantly increased urinary volume in rats subjected to renal ischaemia-reperfusion (P < 0.05). CONCLUSION: L-arginine tended to be of benefit for renal function during renal ischaemia-reperfusion in rats. Pretreatment with L-arginine (200 mg/kg intraperitoneally) seems to increase the renal damage by increasing kidney levels of MDA.     

L-arginine reduces tubular cell injury in acute post-ischaemic renal failure.

BACKGROUND: The pathophysiology of renal ischaemia, resulting in tubular cell injury and leading to acute renal failure (ARF), remains unclear. An ever-increasing number of investigations focus on a possible role of nitric oxide (NO) in regulating circulation during ARF. In this context, we investigated the influence of chronic stimulation or inhibition of NO synthesis, or both, on haemodynamic parameters, histology and plasma renin activity (PRA) after ischaemia-reperfusion injury of rat kidneys. METHODS: Experiments were performed on adult, male Wistar rats. Before induction of ARF, a group of animals was treated with a NO synthesis inhibitor (L-NAME) and another group was treated with a precursor of NO synthesis (L-arginine). The animals received those substances for 4 weeks. Control groups received the same amount of tap water for 4 or 8 weeks and were divided into groups with ARF (4 weeks--ARF group and 8 weeks ARF group) and a sham-operated group. Another group of rats was treated first with L-NAME and then with L-arginine in their drinking water, for 4 weeks for each of these two substances. All parameters were evaluated 24 h after the induction of ischaemic ARF or the sham operation. RESULTS: Our results show that such long-term stimulation of NO release by L-arginine improved renal haemodynamics in the ischaemic form of ARF. Renal blood flow (RBF) increased by 96% in the L-arginine-treated rats with ARF compared with the group with ARF alone. Inhibition of NO synthesis worsens renal haemodynamics after ARF. However, this aggravation can be reversed by L-arginine. The rate of water reabsorption was reduced in all groups with ARF, but this reduction was least in the group treated with L-arginine. The rate of Na+ reabsorption was reduced in all groups 24 h after renal ischaemia, but a significant decrease was observed after the inhibition of NO synthesis. Histological examination of the kidney specimens showed that morphological changes were least in the rats treated with L-arginine, when compared with all other groups with ARF. Nevertheless, the lesions were most prominent in the L-NAME+ARF group. In this group, the areas of corticomedullar necrosis were more widespread in comparison with other groups, especially the L-arginine group where only swelling of the proximal tubular cells was observed. Treatment with L-NAME was not accompanied by any significant alteration in the plasma concentration of angiotensin I (ANG I), while in the group treated with L-arginine ANG I had a tendency to decrease. CONCLUSIONS: Acute post-ischaemic renal failure may be alleviated by administering the NO substrate (L-arginine). NO acts cytoprotectively on tubular epithelial cells in ischaemia--reperfusion injury of rat kidney. Evidence of this comes from both histopathological findings and increased tubular water and sodium reabsorption. However, inhibition of NO synthesis (provoked by L-NAME) worsens renal haemodynamics and aggravates morphological changes after ARF. These aggravations can, however, be reversed by L-arginine.     

Real-time monitoring of nitric oxide in ischemia-reperfusion rat kidney

In this study we attempted to clarify the release of nitric oxide (NO) and its role in the ischemia-reperfusion rat kidney. After right nephrectomy, male Wistar rats were divided into four groups: one sham operated and three groups who underwent ischemia (30 min) and reperfusion of the left renal artery. Thirty minutes prior to ischemia-reperfusion, two groups were injected intraperitoneally with 10 and 30 mg/kg of NG-nitro-L-arginine methylester (L-NAME). Real-time monitoring of blood flow and NO release in the rat kidney was measured with a laser Doppler flowmeter and an NO-selective electrode, respectively. Serum creatinine and blood urea nitrogen (BUN) levels were measured 1 and 7 days after the induction of ischemia-reperfusion. Clamping of the renal artery decreased blood flow to 1-5% of the basal level measured before clamping. After removal of the clip, the blood flow of the 30 mg/kg L-NAME rats was significantly lower than that of the controls. Immediately following the clipping of the renal artery, NO release rapidly increased. After removing the clip, NO release immediately returned to three-quarters of the basal level. Serum creatinine and BUN levels of the ischemia-reperfusion rats were slightly but not significantly higher and those of 30 mg L-NAME rats were significantly higher than those of the control or ischemia-reperfusion rats 1 day and 7 days after ischemia-reperfusion. Our data suggest that NO acts as a cytoprotective agent in ischemia-reperfusion injury of the rat kidney.     

Taurine protects against early and late skeletal muscle dysfunction secondary to ischaemia reperfusion injury.

OBJECTIVE: To assess the efficacy of the cytoprotective B-amino acid taurine in prevention of skeletal muscle dysfunction secondary to ischaemia-reperfusion (IR) injury. DESIGN: Randomised controlled animal study. SETTING: Biomedical research laboratory, teaching hospital, Republic of Ireland. ANIMALS: 96 Sprague Dawley rats. INTERVENTIONS: Rats were randomised into three groups (n = 24/group): control (sham); ischaemia-reperfusion (untreated); and ischaemia-reperfusion + taurine. A further 24 rats were given taurine alone. The rat cremaster skeletal muscle model of four hours of ischaemia followed by reperfusion was used. Taurine 4%wt/vol was given in the animals' water throughout the experiment, beginning 48 hours before the ischaemia was initiated. OUTCOME MEASURES: 8 rats were killed from each group and muscle contractile function was assessed using electrical field stimulation in a tissue bath at 24 hrs, 48 hrs and 7 days. RESULTS: Ischaemia followed by 24 hours, 48 hours or 7 days of reperfusion resulted in significant reduction in both muscle twitch and tetanic contractile function (p < 0.05). This was reversed by taurine, which resulted in significant preservation of twitch and tetanic contractility at all time points including one week of reperfusion (p < 0.05). There was no difference in muscle function between muscle treated with taurine after ischaemia-reperfusion and control muscle, with the same operation but without ischaemia, from 48 hours onwards. CONCLUSIONS: These data show that pharmaceutical use of the endogenous amino acid taurine, unlike many-other agents, protects electrophysiological function in skeletal muscle against both early and late ischaemia-reperfusion injury.     

Effect of irbesartan on the antioxidant defence system and nitric oxide release in diabetic rat kidney

BACKGROUND/AIMS: Increased oxidative stress is involved in the aetiology of diabetic nephropathy, and angiotensin II is reported to play a considerable role in the development of renal damage in diabetic kidney. Angiotensin antagonism can slow the progression of renal impairment in diabetes. The present study was thus designed to examine the effect of an angiotensin II type 1 (AT1) receptor antagonist, irbesartan on renal function, oxidative stress and nitric oxide (NO) release in streptozotocin (STZ)-induced diabetic rats. METHODS: Diabetes was induced by a single intraperitoneal injection of streptozotocin (65 mg/kg) in rats. After 4 weeks of STZ injection, rats were divided into four groups: the control rats, diabetic rats and diabetic rats treated with irbesartan (25 and 50 mg/kg, orally) respectively till 8 weeks starting from 4 weeks after STZ injection. Renal function was assessed by creatinine, blood urea nitrogen, creatinine clearance and urea clearance. Oxidative stress was measured by renal malondialdehyde (MDA), reduced glutathione (GSH), superoxide dismutase (SOD) and catalase. We also measured renal nitrite levels. RESULTS: At the end of the 8th week, diabetic rats exhibited renal dysfunction as evidenced by reduced creatinine and urea clearance along with enhanced albumin excretion rate as compared with control rats. Biochemical analysis of kidneys revealed a marked increase in oxidative stress demonstrated by increased lipid peroxidation and decreased activities of key antioxidant enzymes, GSH, SOD and catalase in diabetic rats. NO release was also significantly higher in diabetic rats than controls. Chronic treatment with irbesartan in diabetic rats significantly attenuated both renal dysfunction and oxidative stress along with increased NO levels as compared with untreated diabetic rats. The kidneys of diabetic rats showed morphological changes such as hyaline casts, glomerular thickening and moderate interstitial fibrosis and arteriolopathy, whereas irbesartan administration markedly prevented diabetic-induced renal morphological alterations. CONCLUSIONS: The present study suggests that oxidative stress/nitrosative stress is increased in the diabetic kidney and AT1 receptor blockade can prevent these changes. The results also suggest that in STZ-induced diabetic rats, the protective action of irbesartan might be mediated, at least in part, by its effect on tissue oxidant/antioxidant status.     

Diltiazem attenuates oxidative stress in diabetic rats

Diabetic nephropathy is the main cause of end stage renal damage. Oxidative stress is involved in the etiology of diabetic nephropathy and intracellular calcium is reported to play a considerable role in the development of renal damage in the diabetic kidney. Calcium antagonism can slow the progression of renal impairment in diabetes. The present study was thus designed to examine the effect of a nondihydropyridine calcium channel blocker, diltiazem, on renal function, oxidative stress, and nitric oxide (NO) release in streptozotocin (STZ)-induced diabetic rats. Diabetes was induced by a single intraperitoneal injection of STZ (65 mg/kg) in rats. After 4 weeks of STZ injection, the rats were divided in to four groups: control rats, diabetic rats treated with saline, and two groups of diabetic rats treated with diltiazem (5 and 10 mg/kg, i.p, respectively) for 8 weeks starting from 4 weeks after STZ injection. Renal function was assessed by creatinine, blood urea nitrogen, creatinine clearance, and urea clearance. Oxidative stress was measured by renal malondialdehyde (MDA), reduced glutathione (GSH), superoxide dismutase (SOD), and catalase. We also measured renal nitrite levels. At the end of the 8 weeks, diabetic rats exhibited renal dysfunction as evidenced by reduced creatinine and urea clearance along with enhanced albumin excretion rate as compared with control rats. Biochemical analysis of kidneys revealed a marked increase in oxidative stress demonstrated by increased lipid peroxidation and decreased activities of key antioxidant enzymes, GSH, SOD, and catalase in diabetic rats. Release of NO also significantly higher in diabetic rats than controls. Chronic treatment with diltiazem in diabetic rats significantly attenuated both renal dysfunction and oxidative stress along with increased NO levels as compared with untreated diabetic rats. The kidneys of diabetic rats showed morphological changes such as hyaline casts, glomerular thickening, and moderate interstitial fibrosis and arteriolopathy, whereas diltiazem administration markedly prevented diabetic-induced renal morphological alterations. The present study suggests that oxidative stress/nitrosative stress is increased in the diabetic kidney and calcium channel blockage can prevent these changes. The results also suggest that in STZ-induced diabetic rats, the protective action of diltiazem might be mediated, at least in part, by its effect on tissue oxidant/antioxidant status.     

Ischemic Postconditioning Modified Renal Oxidative Stress and Lipid Peroxidation Caused By Ischemic Reperfusion Injury in Rats

Several recent studies have shown that ischemic postconditioning (IPostC) protects hears from ischemic reperfusion insults in various animal models. However, the mechanism of IPostC remains unclear. In the present study, we investigated the hypothesis that PostC protected kidneys against ischemic reperfusion injury by modifying renal oxidative stress and lipid peroxidation. Rats underwent 45 minutes of renal pedicle ligature followed by reperfusion for 1, 3, 6, 12, or 24 hours. IPostC was performed using 6, 10 second cycles of reperfusion and 10 seconds of renal pedicle occlusion at the end of the ischemia. Our data showed that IPostC attenuated renal dysfunction, significantly increasing the activity of antioxidases, including superoxide dismutase (SOD), catalase (CAT), and glutathione perokidase (GSH-Px) in renal homogenates, and concentrations of GSH and SOD expression. The level of malondialdehyde (MDA) and the activity of myeloperoxidase (MPO) were significantly decreased in IPostC rats. These results indicated that the protective effects of IPosC may be related to modification of renal oxidative stress and lipid peroxidation caused by ischemic reperfusion injury in rats.     

Nitrite-derived nitric oxide protects the rat kidney against ischemia/reperfusion injury in vivo: role for xanthine oxidoreductase

In normal conditions, nitric oxide (NO) is oxidized to the anion nitrite, but in hypoxia, this nitrite may be reduced back to NO by the nitrite reductase action of deoxygenated hemoglobin, acidic disproportionation, or xanthine oxidoreductase (XOR). Herein, is investigated the effects of topical sodium nitrite administration in a rat model of renal ischemia/reperfusion (I/R) injury. Rats were subjected to 60 min of bilateral renal ischemia and 6 h of reperfusion in the absence or presence of sodium nitrite (30 nmol) administered topically 1 min before reperfusion. Serum creatinine, serum aspartate aminotransferase, creatinine clearance, fractional excretion of Na(+), and plasma nitrite/nitrate concentrations were measured. The nitrite-derived NO-generating capacity of renal tissue was determined under acidic and hypoxic conditions by ozone chemiluminescence in homogenates of kidneys that were subjected to sham, ischemia-only, and I/R conditions. Nitrite significantly attenuated renal dysfunction and injury, an effect that was abolished by previous treatment of rats with the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazole-1-oxyl-3-oxide (2.5 mumol intravenously 5 min before ischemia and 50 nmol topically 6 min before reperfusion). Renal tissue homogenates produced significant amounts of NO from nitrite, an effect that was attenuated significantly by the xanthine oxidoreductase inhibitor allopurinol. Taken together, these findings demonstrate that topically administered sodium nitrite protects the rat kidney against I/R injury and dysfunction in vivo via the generation, in part, of xanthine oxidoreductase-catalyzed NO production. These observations suggest that nitrite therapy might prove beneficial in protecting kidney function and integrity during periods of I/R such as those encountered in renal transplantation.     

EUK-134 reduces renal dysfunction and injury caused by oxidative and nitrosative stress of the kidney

BACKGROUND/AIMS: Oxidative and nitrosative stress plays important roles in the pathogenesis of renal ischemia/reperfusion (I/R) injury. Here we investigate the effect of EUK-134, a synthetic superoxide dismutase and catalase mimetic, (i) on renal dysfunction and injury caused by I/R in vivo and (ii) on proximal tubular cell (PTC) injury and death caused by oxidative and nitrosative stress. METHODS: Rats, subjected to bilateral renal ischemia (45 min) followed by reperfusion (6 h), were administered EUK-134 (0.3 and 3 mg/kg, i.v.) prior to and during reperfusion, after which biochemical and histological indicators of renal dysfunction and injury were measured. The expression of poly(ADP-ribose) (PAR) and inducible nitric oxide (NO) synthase (iNOS) and nitrotyrosine formation were determined immunohistochemically and used as indicators of oxidative and nitrosative stress. Primary cultures of rat PTCs, isolated and cultured from the kidney cortex, were incubated with hydrogen peroxide (H2O2; 1 mM for 2 h) in the presence of increasing concentrations of EUK-134 (1-100 microM) after which PTC injury and death were measured. The effects of EUK-134 on serum levels of NO in rats subjected to renal I/R or on NO production by PTCs incubated with interferon-gamma (IFN-gamma, 100 IU/ml) and bacterial lipopolysaccharide (LPS, 10 microg/ml) in combination for 24 h were also measured. RESULTS: EUK-134 produced a significant reduction in renal dysfunction and injury caused by I/R. Specifically, serum creatinine levels, an indicator of renal dysfunction, were reduced from 227 +/- 11 (n = 12, I/R only) to 146 +/- 9 microM (n = 12, I/R +3 mg/kg EUK-134). Urinary N-acetyl-beta-D-glucosaminidase activity, an indicator of tubular damage, was reduced from 42 +/- 5 (n = 12, I/R only) to 22 +/- 3 IU/l (n = 12, I/R +3 mg/kg EUK-134). EUK-134 significantly reduced renal injury caused by oxidative stress in vivo (reduction in PAR formation), and in vitro EUK-134 reduced PTC injury and death caused by H2O2. However, EUK-134 also reduced nitrosative stress caused by I/R in vivo (reduction of iNOS expression and nitrotyrosine formation), which was reflected by a significant reduction in serum NO levels in rats subjected to renal I/R. Specifically, serum NO levels were reduced from 57 +/- 12 (n = 12, I/R only) to 23 +/- 3 mM (n = 12, I/R +3 mg/kg EUK-134). In vitro, EUK-134 significantly reduced NO production by PTCs incubated with IFN-gamma/LPS. CONCLUSION: We propose that EUK-134 reduces renal I/R injury not only via reduction of oxidative stress, but also by reducing nitrosative stress caused by renal I/R.     

Renal ischemia/reperfusion leads to macrophage-mediated increase in pulmonary vascular permeability.

BACKGROUND: Despite the advent of dialysis, survival with acute renal failure when associated with multiorgan failure is poor. The development of lung injury after shock or visceral ischemia has been shown; however, the effects of isolated renal ischemia/reperfusion injury (IRI) on the lungs are unclear. We hypothesized that isolated renal IRI could alter pulmonary vascular permeability (PVP) and that macrophages could be important mediators in this response. METHODS: Rats (N = 5 per group) underwent renal ischemia for 30 minutes, followed by reperfusion. Lung vascular permeability was evaluated by quantitation of Evans blue dye extravasation from vascular space to lung parenchyma at 1, 24, 48, or 96 hours after reperfusion. Serum was collected for blood urea nitrogen and creatinine at each time point. To examine the role of the macrophage, the macrophage pacifant CNI-1493, which inhibits the release of macrophage-derived inflammatory products, was administered in a blinded fashion during renal IRI. RESULTS: PVP was significantly (P < 0.05) increased at 24 hours and peaked at 48 hours after IRI compared with shams as well as baseline levels. PVP after IRI became similar to shams after 96 hours. This correlated with increases in blood urea nitrogen and creatinine at similar time points. At 48 hours, CNI-1493 significantly abrogated the increase in PVP compared with IRI alone. However, CNI-1493 did not alter the course of the acute renal failure. Pulmonary histology demonstrated interstitial edema, alveolar hemorrhage, and red blood cell sludging after renal IRI, which was partially attenuated by CNI-1493. CONCLUSIONS: Increased PVP develops after isolated renal IRI, and macrophage-derived products are mediators in this response. These findings have implications for understanding the mechanisms underlying respiratory dysfunction associated with acute renal failure.     

Role of nitric oxide in resveratrol-induced renal protective effects of ischemic preconditioning

BACKGROUND: Resveratrol, a natural antioxidant and polyphenol found in red wine and grapes, has been found to pharmacologically precondition the heart through upregulation of nitric oxide (NO). This study was designed to explore the involvement of NO in the renoprotective effect of resveratrol in renal ischemic preconditioning in rat kidney. METHODS: Ischemic preconditioning was induced by three cycles 2-minutes of ischemia followed by 5 minutes of reperfusion before 45 minutes of prolonged ischemia. Resveratrol was given 1 hour before the surgical procedures. RESULTS: Ischemic preconditioning and resveratrol treatment significantly improved the renal dysfunction, decrease in total NO levels, and oxidative stress induced by 45 minutes of ischemia followed by 24 hours of reperfusion. Histopatholgic examination of the kidneys of ischemic/reperfusion rats revealed severe renal damage, which was attenuated in both preconditioned and resveratrol-treated animals. Preconditioning and resveratrol administration led to a marked increase in NO levels in kidney. Renoprotective effects of resveratrol were abolished when animals were pretreated with NG-nitro-L-arginine methyl ester, a nonspecific NO synthase inhibitor. CONCLUSIONS: These findings demonstrate an important contributory role of NO in the protection afforded by resveratrol in renal ischemic preconditioning. CLINICAL RELEVANCE: It is now well established that brief periods of ischemia followed by reperfusion render a variety of tissues tolerant to subsequent ischemia/reperfusion-induced injury. This phenomenon, referred to as ischemic preconditioning, was first demonstrated in the dog myocardium. The potential for clinical application of such a powerful protective phenomenon has generated enormous interest in identifying the underlying intracellular signaling pathways, with the ultimate aim of pharmacologically exploiting these mechanisms to develop therapeutic strategies that can enhance tolerance to ischemia/reperfusion injury in patients. This study explored the possible involvement of nitric oxide in renal ischemic preconditioning.     

GW274150, a potent and highly selective inhibitor of iNOS,reduces experimental renal ischemia/reperfusion injury

BACKGROUND: Generation of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) may contribute to renal ischemia/reperfusion (I/R) injury. The aim of this study was to investigate the effects of GW274150, a novel, highly selective, potent and long-acting inhibitor of iNOS activity in rat and mouse models of renal I/R. METHODS: Rats were administered GW274150 (5 mg/kg intravenous bolus administered 30 minutes prior to I/R) and subjected to bilateral renal ischemia (45 minutes) followed by reperfusion (6 hours). Serum and urinary indicators of renal dysfunction, tubular and reperfusion injury were measured, specifically, serum urea, creatinine, aspartate aminotransferase (AST) and N-acetyl-beta-d-glucosaminidase (NAG) enzymuria. In addition, renal sections were used for histologic scoring of renal injury and for immunologic evidence of nitrotyrosine formation and poly [adenosine diphosphate (ADP)-ribose] (PAR). Nitrate levels were measured in rat plasma using the Griess assay. Mice (wild-type, administered 5 mg/kg GW274150, and iNOS-/-) were subjected to bilateral renal ischemia (30 minutes) followed by reperfusion (24 hours) after which renal dysfunction (serum urea, creatinine), renal myeloperoxidase (MPO) activity and malondialdehyde (MDA) levels were measured. RESULTS: GW274150, administered prior to I/R, significantly reduced serum urea, serum creatinine, AST, and NAG indicating reduction of renal dysfunction and injury caused by I/R. GW274150 reduced histologic evidence of tubular injury and markedly reduced immunohistochemical evidence of nitrotyrosine and PAR formation, indicating reduced peroxynitrite formation and poly (ADP-ribose) polymerase (PARP) activation, respectively. GW274150 abolished the rise in the plasma levels of nitrate (indicating reduced NO production). GW274150 also reduced the renal dysfunction in wild-type mice to levels similar to that observed in iNOS-/- mice subjected to I/R. Renal MPO activity and MDA levels were significantly reduced in wild-type mice administered GW274150 and iNOS-/- mice subjected to renal I/R, indicating reduced polymorphonuclear leukocyte (PMN) infiltration and lipid peroxidation. CONCLUSIONS: These results suggest that (1). an enhanced formation of NO by iNOS contributes to the pathophysiology of renal I/R injury and (2). GW274150 reduces I/R injury of the kidney. We propose that selective inhibitors of iNOS activity may be useful against renal dysfunction and injury associated with I/R of the kidney.     

Effect of nitric oxide in ischemia/reperfusion of the pancreas

BACKGROUND: Ischemia/reperfusion injury, and thus graft pancreatitis, remains a major problem in pancreas transplantation. Contradictory results about the role of nitric oxide (NO) in pancreatic ischemia/reperfusion have been reported; however, in none of the reports has a detailed comparison between inhibition of NO synthase and NO supplementation been carried out. METHODS: Vascular isolation of the pancreatic tail was performed in landrace pigs. After splenectomy catheters placed in the distal part of the splenic vessels allowed collection of the venous effluent and perfusion of the pancreatic tail. Three hours of complete warm ischemia was followed by 6 h of reperfusion. The effect of the NO donor sodium nitroprusside (SNP) and L-arginine was compared to a control group and NO synthase inhibition with L-NAME. RESULTS: Lipase in the venous effluent of the pancreas was significantly decreased in the SNP and the L-arginine groups. Vascular resistance was markedly elevated in the L-NAME group and reduced in the NO donor groups. Tissue pO2 after reperfusion was only significantly elevated in the SNP group. Granulocyte infiltration and also overall histological tissue injury were most severe in the control group followed by the L-NAME group, the SNP group, and the L-ARG group. CONCLUSION: The data show that supplementation of nitric oxide is clearly protective in pancreatic ischemia/reperfusion. However, inhibition of NO synthesis does not lead to an equally clear aggravation of tissue injury.     

The tyrosine kinase inhibitor tyrphostin AG126 reduces renal ischemia/reperfusion injury in the rat

BACKGROUND: We investigate the effects of tyrphostin AG126, an inhibitor of tyrosine kinase activity, on the renal dysfunction and injury caused by ischemia/reperfusion (I/R) of the kidney. METHODS: Tyrphostin AG126 (5 mg/kg intraperitoneally) was administered to male Wistar rats 30 minutes prior to bilateral renal ischemia for 45 minutes followed by reperfusion for up to 48 hours. Biochemical markers of renal dysfunction and injury were measured and renal sections assessed for renal injury. Expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and formation of nitrotyrosine and poly (ADP) ribose (PAR) were assessed using immunohistochemistry. Rat proximal tubular cells (PTCs) were incubated with interferon-gamma (100 IU/mL), bacterial lipopolysaccharide (10 microg/mL), and with increasing concentrations of tyrphostin AG126 (0.0001-1 mmol/L) for 24 hours. Nitric oxide production was measured in both plasma from rats subjected to I/R and in incubation medium from PTCs. RESULTS: After 6 hours of reperfusion, tyrphostin AG126 significantly reduced the increase in serum and urinary indicators of renal dysfunction and injury caused by I/R and reduced histologic evidence of renal injury. Tyrphostin AG126 also improved renal function (after 24 and 48 hours of reperfusion) and reduced the histologic signs of renal injury (after 48 hours of reperfusion). Tyrphostin AG126 reduced the expression of iNOS and nitric oxide levels in both rat plasma and in PTC cultures, as well as expression of COX-2. Tyrphostin AG126 also reduced nitrotyrosine and PAR formation, suggesting reduction of nitrosative stress and poly (ADP-ribose) polymerase (PARP) activation, respectively. CONCLUSION: Taken together, these results show that tyrphostin AG126 significantly reduces the renal dysfunction and injury caused by I/R of the kidney. We propose that inhibition of tyrosine kinase activity may be useful against renal I/R injury.     

Effects of Everolimus on Cytokines,Oxidative Stress,and Renal Histology in Ischemia-Reperfusion Injury of the Kidney

Background. To evaluate the effects of everolimus on renal ischemia-reperfusion injury (IRI). Methods. Wistar albino rats were divided into control, ischemia-reperfusion (IR), and ischemia-reperfusion/everolimus (IR/eve) groups. Everolimus was administered for seven consecutive days to the IR/eve group prior to injury. IR and IR/eve groups underwent forty-five minutes ischemia followed by the application of reperfusion at 2 and 24 hours. Blood samples and kidneys were taken from all animals. Results. Serum blood urea nitrogen and creatinine levels increased at two hours of reperfusion in the IR and IR/eve groups, and decreased at 24 hours of reperfusion in the IR group. In the IR/eve group, we detected significantly high interleukin-6 levels and low tumor necrosis factor-α and malondialdehyde levels at 24 hours. Myeloperoxidase levels increased at two hours of reperfusion in the IR/eve group, but decreased significantly at 24 hours. Everolimus did not improve renal tubular and interstitial injuries in renal IRI. Conclusions. It has been demonstrated that pretreatment with everolimus has beneficial effects on cytokines and oxidative stress in renal IRI. However, these effects are insufficient for the correction of histopathological changes and restoration of normal kidney function.     

Transient Hyperglycemia Affects the Extent of Ischemia-Reperfusion-induced Renal Injury in Rats

Background: Chronic hyperglycemia is known to increase renal injury, particularly during ischemia-reperfusion episodes. The goal of this study was to examine whether transient hyperglycemia during or after renal ischemia-reperfusion increased renal dysfunction.

Methods: Male Lewis rats underwent sham operations or unilateral nephrectomies followed by contralateral renal ischemia-reperfusion. Hyperglycemic rats were given 25% dextrose to induce transient hyperglycemia lasting throughout the duration of ischemia (PI rats) or beginning 2 h after initiation of reperfusion (PR rats). Additional vehicle control rats received saline and underwent ischemia-reperfusion surgery as with PI and PR rats. Twenty-five minutes of mild renal ischemia followed by 24 h of reperfusion was induced by occluding the renal artery and vein.

Results: Terminal serum creatinine concentrations were significantly higher in the PI rats when compared with the PR or vehicle control rats. Histology demonstrated significantly increased necrosis in the PI rats relative to PR and control animals. Tissue analyses demonstrated significantly higher heat shock protein 70, heat shock protein 32, and cleaved caspase-3 protein levels in the PI rats. Oxidative stress generated through the xanthine pathway in the PI group was significantly increased compared with the oxidative stress in the PR and vehicle control rats. In contrast, vascular endothelial growth factor and erythropoietin were significantly decreased in the PI rats compared with the PR rats and controls.