Endotoxin-induced acute renal failure in rats: effects of L-arginine and nitric oxide synthase inhibition on renal function

BACKGROUND: The regulation of renal hemodynamics is closely related to the L-arginine (L-Arg)/nitric oxide (NO) pathway. NO - metabolized from L-Arg - is capable of improving renal function in ischemic and toxic acute renal failure (ARF), while NO synthase (NOS) inhibition induces deterioration in renal function. The mortality rate in patients with septic shock is increased when treated with a non-selective NOS inhibitor, while the incidence of ARF requiring renal replacement therapy is unaffected. To date, there are no studies on the impact of NOS substrate (L-Arg) and inhibitor (L-NMMA) on renal function in early lipopolysaccharide (LPS)-induced ARF. METHODS: ARF was induced by intravenous (i.v.) LPS. Animals were treated with L-Arg, L-NMMA (NOS substrate and inhibitor), a combination of both or saline. Glomerular filtration rate (GFR), urine flow, fractional sodium excretion, excretion of NO metabolism stable end products and blood pressure (BP) were recorded at baseline, after ARF induction, during drug infusion and thereafter. RESULTS: L-Arg induced better GFR during infusion. Excretion of the NO metabolism end products was highest in the L-Arg group and lowest in the NOS inhibitor group. L-Arg administration had no influence on BP, while L-NMMA induced a slight elevation. CONCLUSIONS: We conclude that exogenous L-Arg exerts beneficial effects in early LPS-induced ARF in rats during drug infusion, while NOS inhibition has no influence on GFR. Subcellular compartmentalization of the L-Arg pool in cytoplasma and the rapid utilization of exogenous L-Arg in such a micro-environment could explain this effect, which has been observed in other ARF models and was called the "L-Arg paradox". In further studies the effects of early and prolonged administration of L-Arg in endotoxinemia should be investigated.     

Ischemic acute renal failure in the rat: effects of L-arginine and superoxide dismutase on renal function

BACKGROUND: Regulation of renal hemodynamics -- especially intraglomerular hemodynamics -- is closely related to the L-arginine (L-Arg)/nitric oxide (NO) pathway, both under basal conditions and in acute renal failure (ARF). Also, superoxide anions -- which may react with NO -- play a role in ischemic ARF. L-Arg not only has beneficial effects on glomerular filtration rate (GFR) but also reduces O2(-) production and prevents NO synthase isoform I up-regulation. Thus, it is of interest to elucidate whether the potential beneficial effects of L-Arg in reperfusion can be augmented by additional treatment with superoxide dismutase (SOD). METHODS: ARF was induced by renal artery clamping for 40 minutes. Animals were treated with either L-Arg, SOD, a combination of both, or saline. GFR, renal plasma flow (RPF), filtration fraction (FF) and blood pressure were recorded at baseline, after induction of ARF, during drug infusion and thereafter. RESULTS: Renal artery clamping induces a severe drop of GFR, RPF and FF, which all are improved by L-Arg and SOD. Increasing GFR is mainly due to better renal perfusion. FF fell after reperfusion and increased with L-Arg and SOD, indicating improvement of disturbed intrarenal hemodynamics. Combined administration of L-Arg and SOD showed similar effects in comparison with each substance alone, but did not induce additional effects on GFR and RPF. CONCLUSIONS: L-Arg and SOD exert beneficial effects in ischemic ARF. Probably, improvements in reducing NO availability and in enhancing O2(-) formation are both playing a mediating role. The underlying mechanisms regulating the interplay between NO availability and O2(-) formation need to be elucidated in further studies using -- aside from other means -- selective NOS inhibitors, intervention in different experimental phases and treatment for a longer period.     

L-arginine as a therapeutic tool in kidney disease

Infusion of L-arginine in experimental animals increases renal plasma flow (RPF) and glomerular filtration rate (GFR). It is likely that a component of these hemodynamic changes are mediated by nitric oxide (NO) as suggested by studies with specific antagonists of L-arginine metabolism. L-arginine administration ameliorates the infiltration of the renal parenchyma by macrophages in rats with obstructive nephropathy or rats with puromycin-induced nephrotic syndrome. L-arginine administration also blunts the increase in interstitial volume, collagen IV, and alpha-smooth muscle actin. Rats with a remnant kidney given 1% L-arginine in the drinking water had a greater GFR and RPF. L-arginine administration also decreased proteinuria. Diabetic rats given L-arginine had significantly lower excretion of protein and cyclic guanosine monophosphate than diabetic rats not receiving L-arginine. Despite persistent hyperglycemia, the administration of L-arginine prevented the development of hyperfiltration and ameliorated proteinuria in diabetic rats. In the setting of ischemic acute renal failure, the administration of L-arginine had a beneficial effect on GFR and RPF, decreased O2- production, diminished up-regulation of soluble guanylate cyclase, and prevented up-regulation of inducible NO synthase (iNOS). The pharmacokinetics of L-arginine indicate that side effects are rare and mostly mild and dose dependent.     

Renal and metabolic effects of L-arginine infusion in kidney transplant recipients.

AIM AND METHODS: In order to investigate the role of kidney damage on renal response to L-arginine (L-Arg) infusion in transplant patients receiving cyclosporine A (CsA) treatment, we assessed systemic and glomerular hemodynamic variables, the fraction excretion of urinary sodium, albumin, cyclic GMP (as an index of nitric oxide (NO) production from L-Arg) and urea excretion (as an index of ureagenesis), and glucoregulatory hormone levels in five normal volunteers and 21 renal allograft recipients (aged 10-20 years) treated with CsA, 10 with normal renal function and 11 with chronic renal insufficiency. RESULTS: In the normal subjects, L-Arg infusion (290 mg/min/1.73 m2 for 1 h) significantly reduced mean arterial pressure (MAP) (76+/-7 to 70+/-5 mmHg) and renal vascular resistance (RVR), and increased GFR (103+/-9 to 122+/-7 min/1.73 m2), RPF, urinary cyclic GMP excretion (0.40+/-0.1 to 0.60+/-0.1 nmol/100 ml glomerular filtrate (GF)), and sodium and albumin excretion. Neither the patients with chronic graft dysfunction nor those with a normal graft responded to L-Arg infusion: RVR remained high, and MAP, GFR, RPF, fractional excretion of sodium and urinary excretion of albumin and cyclic GMP were unchanged in both groups of patients. Glucagon, insulin and urinary urea excretion rose significantly in controls and both patient groups. CONCLUSION: The hemodynamic effects of L-Arg infusion were inhibited in the patients, regardless of their degree of renal function, possibly because L-Arg-NO production was blunted.     

Haemodynamic effects of valsartan in acute renal ischaemia/reperfusion injury.

BACKGROUND: Acute deterioration of renal function is an important side-effect of angiotensin-converting enzyme (ACE) inhibitors, especially if accompanied by other nephrotoxic events. Angiotensin II receptor(1) blockers (ARB) are thought to have fewer side-effects on renal perfusion and function. We examined the effects of valsartan (VAL) on kidney function as well as the contribution of the nitric oxide (NO) system in a rat model of ischaemic acute renal failure (ARF). METHODS: ARF was induced by 40 min of clamping of both renal arteries in female Sprague-Dawley rats. Renal haemodynamic and tubular parameters were determined during post-ischaemic infusion of vehicle, VAL, VAL and the NO-synthase substrate L-arginine, and VAL together with inhibition of NO synthases (NOS) by L-NMMA. RESULTS: Clamping induced acute renal failure with marked decreases in glomerular filtration rate (GFR) and renal plasma flow (RPF) accompanied by a rise in renal vascular resistance (RVR) and fractional sodium excretion. Valsartan caused a slight but significant improvement of GFR and RPF without full recovery of renal function and caused a lowering of RVR and tubular sodium loss. L-arginine-co-administration had no additive beneficial effect. Valsartan-induced changes were not significantly depressed by unspecific inhibition of NOS. CONCLUSIONS: Inhibition of the angiotensin II-receptor(1) diminishes the deleterious effects of ischaemia and reperfusion on glomerular function and on the renal microcirculation. An involvement of the NO system could not be demonstrated.     

Renal hemodynamic effects of L-arginine and sodium nitroprusside in heart transplant recipients

BACKGROUND: Long-term treatment with cyclosporine A (CsA) induces vasoconstriction in the kidney and causes renal impairment. An altered L-arginine (L-Arg)/nitric oxide (NO) pathway may play a key role in CsA nephrotoxicity. METHODS: We studied the effect of L-Arg (dosage, 17 mg/kg/min over 30 min), the precursor of NO synthesis, and sodium nitroprusside (SNP; dosage, 1.0 microgram/kg/min over 30 min) on renal hemodynamics in a double-blind, placebo-controlled, randomized, three-way cross-over study comprising 12 stable cardiac transplant recipients on long-term CsA treatment, 10 patients with chronic nephropathy not receiving CsA, and 13 healthy controls. Renal plasma flow (RPF) and glomerular filtration rate (GFR) were measured by paraaminohippurate (PAH) and the inulin clearance method, respectively. RESULTS: In healthy subjects, L-Arg induced an increase in RPF (P = 0.009) and GFR (P = 0.001). By contrast, L-Arg did not induce renal hemodynamic effects in heart transplant patients or patients with chronic nephropathy. SNP reduced RPF (P = 0.050) and GFR (P = 0.005) in patients with chronic nephropathy but did not affect renal hemodynamics in heart transplant recipients or in healthy subjects. CONCLUSIONS: These data indicate that L-Arg cannot be used to reverse CsA-induced renal vasoconstriction in heart transplant recipients under long-term CsA treatment, although these patients have a normal renal response to SNP.     

Effect of L-arginine infusion in normotensive subjects with and without a family history of hypertension

BACKGROUND: Experimental studies have shown that nitric oxide (NO) generation in the kidney from L-arginine participates in the regulation of renal function. Our purpose was to study the effect of infusion of L-arginine (1, 5, and 10 mg/kg/min) on blood pressure (BP), renal hemodynamics, and urinary excretion of sodium and albumin in normotensive subjects with a family history of either severe hypertension (FHSH, N = 17) or mild hypertension (FHMH, N = 20) and in control subjects (N = 18) without a hereditary predisposition for hypertension. METHODS: The glomerular filtration rate (GFR) and renal plasma flow (RPF) were measured by renal clearances of Cr51 ethylenediaminetetraacetic acid and paramino-hippurate. Renal tubular reabsorption of sodium was estimated by lithium clearance. To evaluate the effect of L-arginine infusion on the L-arginine/NO pathway, we measured the NO-metabolite nitrate in plasma, and urinary excretion of cGMP, the second messenger of NO. The derivative at an L-arginine dose of 7.5 mg/kg/min was used as a measure of sensitivity to L-arginine. RESULTS: There was no difference in baseline systolic BP between the groups, but diastolic BP was significantly higher in FHSH compared with control subjects (P < 0.05). L-arginine caused a significant increase in urine flow, urinary excretion of albumin and sodium, and lithium clearance in all groups. FHSH showed a significantly decreased sensitivity to L-arginine with respect to urine flow rate (P = 0029) compared with FHMH and control subjects. L-arginine caused a significant decrease in the GFR in FHSH (P < 0.02) and control subjects (P < 0.001), but in FHMH, the decrease did not reach statistical significance (P = 0.097). There was no difference in sensitivity to L-arginine with respect to BP, RPF, or GFR between the three groups. In all patients, there was a significant positive relationship between Delta urine flow rate or Delta urinary sodium excretion and Delta GFR during infusion of L-arginine (P = 0.003 and P = 0.03, respectively). Plasma nitrate and urinary cGMP decreased in all groups during the L-arginine infusion. CONCLUSION: L-Arginine infusion in normotensive subjects caused an enhanced urine flow rate and urinary sodium and albumin excretion and a slight reduction in GFR. The effect of L-arginine on the urine flow rate was significantly less pronounced in subjects with a family history of severe hypertension, which may indicate a tubular disturbance in hypertension.     

L-Arginine counteracts nitric oxide deficiency and improves the recovery phase of ischemic acute renal failure in rats

BACKGROUND: In ischemic acute renal failure (ARF), nitric oxide-dependent regulation of renal hemodynamics and glomerular function is disturbed. Previous studies indicate that the nitric oxide precursor l-arginine (l-Arg) has beneficial effects on renal function. Here we further analyzed the impact of l-Arg on functional and biochemical parameters of nitric oxide signaling during the course of ischemic ARF. METHODS: Ischemic ARF was induced in rats by bilateral clamping of renal arteries for 45 minutes. l-Arg was applied intraperitoneally during clamping, and orally during 14 days of follow-up. Glomerular filtration rate (GFR) and renal plasma flow (RPF) were measured, and biochemical parameters analyzed by protein immunoblots. RESULTS: Clamping resulted in 70% to 90% reduction of GFR and RPF, with a gradual recovery by day 14. Using an in situ assay with the oxidative fluorescent dye hydroethidine, increased tubular generation of O2- was detected in the early course of ischemic ARF, indicating enhanced oxidative stress. These findings were accompanied by up-regulation of the nitric oxide receptor, soluble guanylate cyclase, and by significant regulatory changes of inducible nitric oxide synthase (iNOS) and endothelial NOS expression. l-Arg had a beneficial effect on GFR and RPF, decreased O2- production, diminished up-regulation of soluble guanylate cyclase, and prevented up-regulation of iNOS. CONCLUSION: Ischemic ARF is accompanied by marked alterations in the expression of key enzymes of the nitric oxide pathway, indicative for deficiency of constitutive NOS activity. l-Arg supplementation reduces O2- generation and significantly improves the expression of nitric oxide signaling proteins as well as the recovery phase of ischemic ARF.     

Can L-arginine manipulation reduce renal disease?

The administration of L-arginine to normal animals leads to an increase in renal plasma flow and glomerular filtration rate (GFR). Administration on a chronic basis of N-nitro-L-arginine methylester (L-NAME), an antagonist of L-arginine, increases blood pressure and reduces the ultrafiltration coefficient. In rats with ureteral obstruction, the administration of L-arginine increases GFR and renal blood flow in the postobstructive kidney. Administration of L-arginine decreased the macrophage infiltration of the renal parenchyma that occurs in this model. L-arginine administration also blunted the increases in interstitial volume, collagen deposition, and expression of alpha-smooth muscle actin in the obstructed kidney. L-arginine administration to rats with subtotal nephrectomy reduced proteinuria and the number of abnormal glomeruli. Some of these effects may be mediated by nitric oxide (NO). In rats with diabetes, administration of L-arginine decreased hyperfiltration and proteinuria. The role of arginine and NO in glomerular diseases is controversial. In general most of the evidence indicates a beneficial change in the renal pathology and function in animals with glomerulonephritis receiving L-arginine. Most of the evidence indicates that the L-arginine-NO pathway has an important role in ameliorating hypertension, renal disease, inflammation and atherosclerosis.     

Paradoxical increase in nitric oxide synthase activity in hypercholesterolaemic rats with impaired renal function and decreased activity of nitric oxide.

BACKGROUND: We have shown that acute exposure of oxidized low-density lipoprotein (OX-LDL) induces vasoconstriction in renal vessels and reduces glomerular filtration rate (GFR) in an isolated perfused rat kidney model by decreasing the activity of nitric oxide (NO). L-arginine has a protective role against OX-LDl-induced vasoconstriction. Micropuncture studies have demonstrated that short-term diet-induced hypercholesterolaemia is associated with decreased GFR and renal blood flow and increased glomerular capillary pressure. This may be mediated by decreased activity of NO. METHODS: Rats were made hypercholesterolaemic by supplementing the standard chow with 4% cholesterol and 1% sodium cholate. A group of rats on hypercholesterolaemic diet also received L-arginine in the drinking water. After 4 and 6 weeks, blood samples and 24-h urine samples were collected for the measurement of biochemical parameters. After 6 weeks, all rats were subjected to isolated perfusion of kidneys at a constant pressure of 100 mmHg. During isolated perfusion, the unused contralateral kidney was taken for morphological studies and for assessing the activity of nitric oxide synthase enzyme by beta-NADPH diaphorase histochemistry. RESULTS: Rats fed a high-cholesterol diet had LDL levels 3-6 times greater than the rats fed standard chow. Rats that received L-arginine in the drinking water had serum L-arginine levels 5-6 times greater than control rats. At 6 weeks, creatinine clearance was significantly lower in the rats on the high-cholesterol diet compared to the rats on standard chow and rats on high-cholesterol diet plus L-arginine. Twenty-four-hour urinary total nitrate and nitrite excretion in the hypercholesterolaemic rats was 1.5-2 times greater than that of control rats. Twenty-four-hour urinary cGMP excretion was significantly lower in the rats on a high-cholesterol diet, but in the rats on high-cholesterol diet and L-arginine, 24-h urinary cGMP excretion was not significantly different from that of control rats. During isolated perfusion of kidneys, renal perfusate flow was found to be significantly reduced in the kidneys taken from the rats on a high-fat diet compared to controls. L-arginine supplementation in the drinking water almost completely reversed the effect of a high-fat diet. Inulin clearance was also significantly reduced in kidneys on a high-fat diet in contrast to controls but not in kidneys on high fat-diet and L-arginine. Basal cGMP excretion in urine was significantly lower in the kidneys taken from the rats on a high-fat diet compared to controls. L-arginine supplementation restored the basal cGMP excretion in these kidneys. NO synthase (NOS) enzyme activity as assessed by NADPH diaphorase activity showed that kidney sections taken from the rats on a high-fat diet showed more intense staining, indicating increased activity compared to the kidney sections taken from the rats on a normal diet. CONCLUSION: Though activity of NO is diminished in hypercholesterolaemic rats with impaired renal function, there is a paradoxical increase in NO production and NOS activity. L-arginine reverses the effects of a high-fat diet.     

Symmetrical dimethylarginine: a new combined parameter for renal function and extent of coronary artery disease

Symmetrical dimethylarginine (SDMA) is the structural isomer of the endogenous nitric oxide synthase (NOS) inhibitor asymmetric dimethylarginine. Whereas the major route of asymmetric dimethylarginine elimination is the hydrolytic degradation by dimethylarginine dimethylaminohydrolase, SDMA is eliminated by renal excretion. SDMA does not directly inhibit NOS but is a competitor of arginine transport. This study showed for the first time that measurement of SDMA can be a marker of estimated GFR and extent of coronary artery disease (CAD). In 97 patients with CAD, SDMA was a marker of estimated GFR. On multiple regression analysis of the CAD parameter stenosis score, SDMA was the only parameter retained. In addition, endothelial cells from the third passage were cultured in medium that contained 70 micromol/L arginine and was incubated for 24 h in the presence of various concentration of SDMA (0, 2, 5, 10, and 100 micromol/L). The levels of nitrate and nitrite in conditioned media, the protein expression of NOS, and the content of reactive oxygen species in endothelial cells were determined. SDMA inhibited dose dependently the NO synthesis in intact endothelial cells, whereas it had no effect on protein expression of NOS. This effect was associated with an increase in reactive oxygen species. Co-incubation with L-arginine but not D-arginine reversed the effect of SDMA on NOS pathway. Our data suggest that SDMA reduced the endothelial NO synthesis, probably by limiting L-arginine supply to NOS. It is concluded that SDMA might be a useful parameter for detecting patients in very early stages of chronic kidney disease and for determining their risk for developing cardiovascular disease.     

肾脂肪囊内注射黄芪和多巴胺对大鼠内毒素休克所致急性肾衰竭的防治作用

目的:探讨肾脂肪囊内联合应用黄芪和多巴胺对大鼠内毒素休克所致急性肾衰竭的防治作用及其机制.方法:SD大鼠随机分为对照组、内毒素组、静脉应用与肾脂肪囊内应用黄芪和多巴胺组,分别测定平均动脉压(MAp)、肾血浆流量(RPF)、肾小球滤过率(GFR)、尿量(UO)、滤过钠排泄分数(FENa)以及肾组织一氧化氮(NO)和内皮素(ET)浓度,光镜及电镜观察病理改变.结果:内毒素组MAp和RPF、GFR、UO显著降低,FENa初4 h极度减少,随后增高,至12 h后接近或大于2%,肾组织NO和ET浓度明显升高;黄芪和多巴胺联合应用可稳定MAp,提高RPF和GFR,增加UO,改善FENa,降低肾组织NO和ET水平,减轻组织病理损害,尤以肾脂肪囊内给药组肾脏保护效果显著,实验后期作用更为明显.结论:黄芪和多巴胺联合应用可纠正大鼠内毒素休克低血压状态,改善肾脏功能,减轻病理损害,可能与作用于NO和ET系统有关.     

Effects of nitric oxide on gentamicin toxicity in isolated perfused rat kidneys

BACKGROUND: There have been many studies in recent years concerning the role of nitric oxide (NO) in acute renal failure (ARF). In this study, the effects of the inhibition or the induction of NO synthase (NOS) on gentamicin-induced ARF was investigated in isolated perfused rat kidneys. METHODS: Kidneys from male Sprague-Dawley rats were perfused in situ for 90 min. Perfusion was conducted in the presence of inulin (60 mg/dL in perfusion buffer) as a glomerular filtration rate (GFR) marker. Six groups (total: 42 rats) were studied: group 1, controls with no treatment; group 2, L-arginine (2 mM in perfusate); group 3, L-nitro-arginine-methyl ester (L-NAME, 0.1 mM in perfusate); group 4, gentamicin (GM, 0.5 mg/mL in perfusate); group 5, GM + L-arginine (same dose as groups 2 and 4) and; group 6, GM + L-NAME (same dose as groups 3 and 4). Cell injury was assessed by measuring N-acetyl-beta-D-glucosaminidase (NAG), lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) activity in urine. RESULTS: L-arginine prevented, whereas L-NAME enhanced, GM-induced enzyme release and GFR reduction. Histological studies showed that GM-treated kidneys had clear signs of tubular damage and this damage was increased by simultaneous L-NAME and GM administration. CONCLUSION: This study suggests that NO formation could prevent the GM-induced nephrotoxicity in this ARF model.     

Theophylline in rats during maintenance phase of post-ischemic acute renal failure

We have shown previously that theophylline increases both renal plasma flow (RPF) and glomerular filtration rate (GFR) during the initiation phase of post-ischemic acute renal failure (ARF) in rats. The purpose of the present experiments was to determine the effects of theophylline during the maintenance phase of ARF, five days after initiation. Clearance techniques were used to measure renal function in a control group of pentobarbital anesthetized rats (group C) and in three experimental groups, five days after subjecting the left kidney to a thirty-minute period of complete ischemia. Group SS received saline during both the ischemic episode and the clearance measurements; group ST received saline during ischemia and theophylline, acutely, during the clearance measurements; group TS received theophylline during ischemia and saline during the clearance measurements. In comparison with the values for the control group (group C), RPF and GFR of the post-ischemic left kidneys of group SS were approximately half normal. In groups ST and TS, RPF and GFR of the left kidneys were higher than in group SS. Collectively, these results demonstrate that pretreatment with theophylline during the initiation phase of ischemia-induced ARF leads to increased RPF and GFR during the maintenance phase, and that acute theophylline treatment during the maintenance phase acutely increases RPF and GFR. Since increases in GFR were associated with increases in RPF, and since theophylline is an adenosine receptor antagonist, these results are consistent with the hypothesis that adenosine-mediated hemodynamic changes play a pathogenic role in ischemia-induced ARF in rats.     

Impaired basal NO activity in patients with glomerular disease and the influence of oxidative stress

Endothelial dysfunction has been found to be linked to and predictive of cardiovascular events. Whether endothelial function of the renal vasculature is impaired in patients with chronic glomerular disease and whether oxidative stress is of importance in this setting has not yet been determined. In this study, endothelial function of the renal vasculature was investigated in 25 patients with chronic glomerular disease and 50 control subjects matched for age and blood pressure. Renal plasma flow (RPF) and glomerular filtration rate were measured by constant infusion input clearance technique at baseline and following infusions of the nitric oxide synthase (NOS) inhibitor N(G)-monomethyl-L-arginine (L-NMMA, 4.25 mg/kg), the substrate of NOS L-arginine (100 mg/kg) and the antioxidant vitamin C (3 g co-infused with L-arginine 100 mg/kg). At baseline, RPF was similar in the two groups. The reduction in RPF in response to L-NMMA was less pronounced in patients with chronic glomerular disease compared to control subjects (-4.6+/-12 vs -9.8+/-9%; P=0.040), indicating reduced basal nitric oxide (NO) activity in chronic glomerular disease. Co-infusion of the antioxidant vitamin C on top of L-arginine induced a more pronounced increase in RPF in patients with chronic glomerular disease than in control subjects (21.7+/-17 vs 10.9+/-22%; P=0.036). Our findings suggest that basal NO activity of the renal vasculature is reduced in patients with chronic glomerular disease compared to age- and blood pressure-matched control subjects. This might be in part related to increased oxidative stress.     

Oral sorbent AST-120 increases renal NO synthesis in uremic rats.

OBJECTIVE: The urine level of nitric oxide (NO) metabolites, i.e., nitrates/nitrites (NOx), in chronic renal failure (CRF) is decreased because of reduced renal synthesis of NO. We determined whether the administration of an oral sorbent, AST-120, increases the urine level of NOx and the renal expression of nitric oxide synthase (NOS) isoforms in CRF rats. METHODS: Chronic renal failure rats were produced by 4/5 nephrectomy. Rats were randomized into two groups: CRF control rats, and AST-120-treated CRF rats. The AST-120 was administered to the rats at a dose of 4 g/kg with powder chow for 16 weeks, whereas powder chow alone was administered to control rats. The urine levels of NOx were measured by using a NOx colorimetric assay kit. The expression of endothelial NOS (eNOS), inducible NOS (iNOS), and neuronal NOS (nNOS) in the kidney was determined by immunohistochemistry. Serum and urine levels of indoxyl sulfate were determined by high-performance liquid chromatography. RESULTS: Urine levels of NOx and the expression of glomerular eNOS and tubulointerstitial nNOS were significantly decreased in CRF rats compared with normal rats. The administration of AST-120 to CRF rats significantly increased urine levels of NOx and the expression of glomerular eNOS and tubulointerstitial nNOS. The administration of AST-120 to CRF rats significantly decreased urine and serum levels of indoxyl sulfate. CONCLUSIONS: The oral sorbent AST-120 increases NO synthesis in the kidneys of uremic rats by increasing the renal expression of eNOS and nNOS, through alleviation of indoxyl sulfate overload on the kidney.     

Nitric oxide synthase isoforms and glomerular hyperfiltration in early diabetic nephropathy

This study tested the hypothesis that nitric oxide (NO)-mediated renal vasodilation due to the activity of the inducible nitric oxide synthase (iNOS) contributes to glomerular hyperfiltration in diabetic rats. Two weeks after induction of diabetes mellitus by streptozotocin, mean arterial BP (MAP), GFR (inulin clearance), and renal plasma flow (RPF) (para-aminohippurate clearance) were measured in conscious instrumented rats. Diabetic rats had elevated GFR (3129 +/- 309 microl/min versus 2297 +/- 264 microl/min in untreated control rats, P < 0.05) and RPF (10526 +/- 679 microl/min versus 8005 +/- 534 microl/min), which was prevented by chronic insulin treatment. Intravenous administration of 0.1 and 1 mg of L-imino-ethyl-lysine (L-NIL), an inhibitor of iNOS, did not affect MAP, GFR, or RPF, either in diabetic or control rats. A higher L-NIL dose (10 mg) increased MAP and decreased RPF in diabetic rats significantly (n = 6, P < 0.05), but not in controls (n = 6). In addition, 0.1 mg of NG-nitro-L-arginine methyl ester (L-NAME), a nonselective blocker of NOS isoforms, decreased GFR (2389 +/- 478 microl/min) and RPF (7691 +/- 402 microl/min) in diabetic animals to control levels, while renal hemodynamics in normoglycemic rats were not altered. Higher L-NAME doses (1 and 10 mg) reduced GFR and RPF in diabetic and control rats to identical levels. In glomeruli isolated from diabetic and control rats, neither iNOS mRNA nor iNOS protein expression was detected. In contrast, increased protein levels of endothelial constitutive NOS (ecNOS) were found in glomeruli of diabetic rats compared with controls. By immunohistochemistry, ecNOS but not iNOS staining was observed in the endothelium of preglomerular vessels and in diabetic glomeruli. These results support the notion that increased NO availability due to greater abundance of ecNOS contributes to the pathogenesis of glomerular hyperfiltration in early experimental diabetic nephropathy. In contrast, we found no functional or molecular evidence for increased glomerular expression and activity of iNOS in diabetic rats.     

Hyperhomocysteinemia induces renal hemodynamic dysfunction: is nitric oxide involved?

Hyperhomocysteinemia is associated with endothelial dysfunction, although the underlying mechanism is unknown. Previous studies have shown that nitric oxide (NO) plays an important role in the regulation of systemic and renal hemodynamics. This study investigated whether hyperhomocysteinemia induces renal oxidative stress and promotes renal dysfunction involving disturbances of the NO-pathway in Wistar rats. During 8 wk, control (C) and hyperhomocysteinemic (HYC) groups had free access to tap water and homocysteine-thiolactone (HTL, 50 mg/kg per d), respectively. At 8 wk, plasma homocysteine concentration, renal superoxide anion (O(2)), nitrotyrosine, and nitrite+nitrate levels, and renal function were measured. To assess NO involvement, the responses to L-Arginine (L-Arg, 300 mg/kg) and N(G)-nitro-L-arginine-methyl-ester (L-NAME, 20 microg/kg per min for 60 min) were analyzed. The HYC group showed higher homocysteine concentration (7.6 +/- 1.7 versus 4.9 +/- 1.0 micromol/L; P < 0.001), (O(2) production (157.92 +/- 74.46 versus 91.17 +/- 29.03 cpm. 10(3)/mg protein), and nitrite+nitrate levels (33.4 +/- 5.1 versus 11.7 +/- 4.3 micro mol/mg protein; P < 0.001) than the control group. Western blot analyses showed a nitrotyrosine mass 46% higher in the HYC group than in the controls. Furthermore, the HYC group showed lower GFR, renal plasma flow (RPF), and higher renal vascular resistance (RVR) than the controls. After L-Arg administration, the responses of GFR, RPF, and RVR were attenuated by 36%, 40%, and 50%, respectively; after L-NAME, the responses of RPF and RVR were exaggerated by 79% and 112%, respectively. This suggests a reduced NO bioavailability to produce vasodilation and an enhanced sensitivity to NO inhibition. In conclusion, hyperhomocysteinemia induces oxidative stress, NO inactivation, and renal dysfunction involving disturbances on the NO-pathway.     

Nitric oxide, sepsis, and the kidney

Although excess nitric oxide (NO) production plays a major role in the hypotension characteristic of sepsis, concurrent constitutive NO generation in the kidney during sepsis is essential for preservation of renal perfusion and prevention of glomerular thrombosis. The authors have shown that although all nitric oxide synthase (NOS) inhibitors restore normal blood pressure in lipopolysaccharide (LPS) treated rats, only selective inducible NOS (iNOS) inhibition prevents the reductions in glomerular filtration rate (GFR), whereas nonselective inhibition of NOS further decreases GFR. Glomerular endothelial NOS (eNOS) activity was found to be inhibited by LPS. The decrease in eNOS activity was completely prevented by selective iNOS inhibition in vivo and in vitro. The adverse renal outcomes after LPS administration correlated with decreased glomerular eNOS activity rather than elevated NO production. These findings suggest that the decrease in GFR after LPS is caused by local inhibition of eNOS by iNOS possibly via NO autoinhibition. Selective inhibition of iNOS could represent a substantially superior approach for the treatment of the sepsis syndrome.     

Atorvastatin improves the course of ischemic acute renal failure in aging rats

Statins increase the production of nitric oxide (NO) and have beneficial effects on the course of acute renal failure (ARF) in young rats. The effects of a short-term treatment with atorvastatin (ATO) on ischemic ARF in old rats, characterized by a great susceptibility to ischemia, was tested. No difference was found in renal dynamics between young (Y, 3 mo old) and old (O, 18 mo old) rats in normal conditions (CON) or after ATO treatment (12 mg/kg/d for 14 d). Twenty-four hours after clamping of both renal arteries, a more pronounced decrease in GFR was observed in O rats versus Y rats after a greater renal vasoconstriction and hypoperfusion of aging animals. Pretreatment with ATO mitigated renal vasoconstriction in O rats and restored GFR values to Y rats. Nitrate excretion was enhanced in Y rats after ARF but was not further modified by ATO; in O rats, ARF did not increase nitrate excretion, which was raised after ATO treatment. This reflected the increase in endothelial NO synthase (eNOS)-mRNA expression and eNOS protein observed in old ATO-treated animals with ARF. ATO treatment had also a significant protective effect against the cell injury at tubular level in O, but not Y, rats. The Ras system was not influenced by ATO in O rats, whereas the activation of Rho proteins was partially inhibited by ATO. Low-dose treatment with ATO enhances NO availability in aging rats, improving renal dynamics and conferring a peculiar histologic protection at tubular level after ischemia.