The Comparative Effects of Inulin and Bacillus clausii on LPS-Induced Endotoxemic Rat Liver.

BACKGROUND: This paper sought to investigate the modifies of inulin and Bacillus clausii on the lipopolysaccharides (LPS) inducing oxidative stress signaling pathway in the endotoxemic rat model. METHODS: Wistar albino male rats (n = 36), divided into six groups, were formed randomly in the following stages: the control group; the prebiotic group (Inulin; 500 mg/kg); the probiotic group (Bacillus clausii; 1x109 CFU); the LPS group (1.5 mg/kg) as the endotoxemic model; the prebiotic group + LPS; and the probiotic group + LPS as treatment groups. RESULTS: The reactive oxygen species (ROS), advanced oxidation products of protein (AOPP), thiobarbituric acid reactive substances (TBARS), total oxidant status (TOS), oxidative stress index (OSI), and myeloperoxidase activity (MPO) levels increased in LPS-induced toxicity. Prebiotic treatment decreased LPS-induced hepatotoxicity on rat liver as observed in the decrease in the levels of oxidative stress parameters, such as ROS, TBARS, TOS, and OSI. The effect of the probiotic treatment on the ROS, AOPP, TOS, OSI levels was not statistically significant. However, it was determined that probiotic application was effective in the TBARS, TAS, and GSH levels. When the biochemical results of the prebiotic and probiotic treatment applications were compared, it was found that the prebiotic treatment was more effective on oxidative stress parameters (ROS, TBARS, TOS, and OSI). In addition, the histological damage score and MPO-staining results of the prebiotic treatment group were found to be more effective than the probiotic group. CONCLUSION: In this first study, where inulin and Bacillus clausii spores are used against liver damage caused by LPS, inulin provides much more effective protection than Bacillus clausii spores.

The Effects of Nitric Oxide and Inhibitor, and Combination of Albendazole and Praziquantel On Liver in Mice Injected with Echinococcus granulosus Larvae.

In this study, the role of nitric oxide (NO) in the pathogenesis of hydatidosis and the interaction with effects of anthelmintic drugs, albendazole and praziquantel, were examined in larval infection caused by protoscolices obtained from hydatid cysts of sheep liver in Albino Balb/c mice. Animals were divided into ten groups including controls and infected groups. Larval infection was established with intraperitoneal injection of protoscolices. Eight months after infection with protoscolices, the infected animals were divided into 6 groups. The infected animals were given a selective inhibitor of inducible nitric oxide synthase (iNOS) L-N6-(1-Iminoethyl) lysine-hydrochloride (L-NIL), NO donor sodium nitroprusside (SNP), albendazole and praziquantel as anthelmintic drugs for 7 days. In addition, control groups were composed of intact group, control, anthelmintic drugs + L-NIL, and anthelmintic drugs + SNP. The liver and blood samples were taken for cytological, histological, immunohistochemical and biochemical analyses 7 days after treatments at the end of experiment. The animals injected with protoscolices showed histopathological changes including inflammation areas, infiltration and accumulation of leukocytes, dilation of sinusoids, and damage in endothelial cells and hepatocytes at light microscopy. Electron microscopy were revealed severe damage in sinusoidal endothelial cells, leukocytes especially eosinophils in sinusoid lumens and disorganization in endoplasmic reticulum and nuclear membrane. Endothelial nitric oxide synthase (eNOS) and iNOS reactions were increased in the tissue. Anthelmintic drugs decreased inflammation areas and damages; however, it did not change NOS reactions in the animals given protoscolices. L-NIL and SNP diminished both iNOS and eNOS reactions. Unlike the group administered the inhibitor, SNP treated group exhibited less inflammation areas. Combination of these substances and drugs resulted in decreased inflammation areas. eNOS and iNOS reactions decreased in the drugs and SNP administered group, while only iNOS reaction was decreased in L-NIL given infection group. In addition, the infected groups which received SNP displayed expanded sinusoids and hepatocytes with vacuoles, intriguingly. While levels of serum nitrite/nitrate elevated only in the infection group given drugs and SNP, it decreased in the L-NIL administered group. Tissue level of malondialdehyde increased in infection groups with drugs and SNP. In conclusion, the results indicated that NO plays an important role in the pathogenesis of hydatidosis.

Resuscitation with PEGylated carboxyhemoglobin preserves renal cortical oxygenation and improves skeletal muscle microcirculatory flow during endotoxemia.

PEGylated carboxyhemoglobin (PEGHbCO), which has carbon monoxide-releasing properties and plasma expansion and oxygen-carrying properties, may improve both skeletal microcirculatory flow and renal cortical microcirculatory Po2 (CµPo2) and, subsequently, limit endotoxemia-induced acute kidney injury. Anesthetized, ventilated Wistar albino rats (n = 44) underwent endotoxemic shock. CµPo2 was measured in exposed kidneys using a phosphorescence-quenching method. Rats were randomly assigned to the following five groups: 1) unresuscitated lipopolysaccharide (LPS), 2) LPS + Ringer's acetate (RA), 3) LPS + RA + 0.5 µg·kg·-1min-1 norepinephrine (NE), 4) LPS + RA + 320 mg/kg PEGHbCO, and 5) LPS + RA + PEGHbCO + NE. The total volume was 30 mL/kg in each group. A time control animal group was used. Skeletal muscle microcirculation was assessed by handheld intravital microscopy. Kidney immunohistochemistry and myeloperoxidase-stained leukocytes in glomerular and peritubular areas were analyzed. Endotoxemia-induced histological damage was assessed. Plasma levels of IL-6, heme oxygenase-1, malondialdehyde, and syndecan-1 were assessed by ELISA. CµPo2 was higher in the LPS + RA + PEGHbCO-resuscitated group, at 35 ± 6mmHg compared with 21 ± 12 mmHg for the LPS+RA group [mean difference: -13.53, 95% confidence interval: (-26.35; -0.7156), P = 0.035]. The number of nonflowing, intermittent, or sluggish capillaries was smaller in groups infused with PEGHbCO compared with RA alone (P < 0.05), while the number of normally perfused vessels was greater (P < 0.05). The addition of NE did not further improve CµPo2 or microcirculatory parameters. Endotoxemia-induced kidney immunohistochemistry and histological alterations were not mitigated by PEGHbCO 1 h after resuscitation. Renal leukocyte infiltration and plasma levels of biomarkers were similar across groups. PEGHbCO enhanced CµPo2 while restoring skeletal muscle microcirculatory flow in previously nonflowing capillaries. PEGHbCO should be further evaluated as a resuscitation fluid in mid- to long-term models of sepsis-induced acute kidney injury.

Mycophenolate mofetil improves renal haemodynamics, microvascular oxygenation, and inflammation in a rat model of supra-renal aortic clamping-mediated renal ischaemia reperfusion injury.

Ischaemia/reperfusion (I/R) is one of the main causes of acute kidney injury (AKI), which is characterized by sterile inflammation and oxidative stress. Immune cell activation can provoke overproduction of inflammatory mediators and reactive oxygen species (ROS), leading to perturbation of the microcirculation and tissue oxygenation associated with local and remote tissue injury. This study investigated whether the clinically employed immunosuppressant mycophenolate mofetil (MMF) was able to reduce I/R-induced renal oxygenation defects and oxidative stress by preventing sterile inflammation. Rats were divided into three groups (n=6/group): (1) a sham-operated control group; (2) a group subjected to renal I/R alone (I/R); and (3) a group subjected to I/R and MMF treatment (20 mg/kg prior to I/R) (I/R+MMF). Ischaemia was induced by a vascular occluder placed on the abdominal aorta for 30 minutes, followed by 120 minutes of reperfusion. Renal I/R deteriorated renal oxygenation (P<.001) and oxygen delivery (P<.01), reduced creatinine clearance (P<.01) and tubular sodium reabsorption (P<.001), and increased iNOS, renal tissue injury markers (P<.001), and IL-6 (P<.001). Oral MMF administration prior to insult restored renal cortical oxygenation (P<.05) and iNOS, renal injury markers, and inflammation parameters (P<.001) to near-baseline levels without affecting renal function. MMF exerted a prophylactic effect on renal microvascular oxygenation and abrogated tissue inflammation and renal injury following lower body I/R-induced AKI. These findings may have clinical implications during major vascular or renal transplant surgery.

Effects of a human recombinant alkaline phosphatase on renal hemodynamics, oxygenation and inflammation in two models of acute kidney injury.

Two small clinical trials indicated that administration of bovine intestinal alkaline phosphatase (AP) improves renal function in critically ill patients with sepsis-associated acute kidney injury (AKI), for which the mechanism of action is not completely understood. Here, we investigated the effects of a newly developed human recombinant AP (recAP) on renal oxygenation and hemodynamics and prevention of kidney damage and inflammation in two in vivo AKI models. To induce AKI, male Wistar rats (n=18) were subjected to renal ischemia (30min) and reperfusion (I/R), or sham-operated. In a second model, rats (n=18) received a 30min infusion of lipopolysaccharide (LPS; 2.5mg/kg), or saline, and fluid resuscitation. In both models, recAP (1000U/kg) was administered intravenously (15min before reperfusion, or 90min after LPS). Following recAP treatment, I/R-induced changes in renal blood flow, renal vascular resistance and oxygen delivery at early, and cortical microvascular oxygen tension at late reperfusion were no longer significantly affected. RecAP did not influence I/R-induced effects on mean arterial pressure. During endotoxemia, recAP treatment did not modulate the LPS-induced changes in systemic hemodynamics and renal oxygenation. In both models, recAP did exert a clear renal protective anti-inflammatory effect, demonstrated by attenuated immunostaining of inflammatory, tubular injury and pro-apoptosis markers. Whether this renal protective effect is sufficient to improve outcome of patients suffering from sepsis-associated AKI is being investigated in a large clinical trial.

Effects of N-acetylcysteine (NAC) supplementation in resuscitation fluids on renal microcirculatory oxygenation, inflammation, and function in a rat model of endotoxemia.

BACKGROUND: Modulation of inflammation and oxidative stress appears to limit sepsis-induced damage in experimental models. The kidney is one of the most sensitive organs to injury during septic shock. In this study, we evaluated the effect of N-acetylcysteine (NAC) administration in conjunction with fluid resuscitation on renal oxygenation and function. We hypothesized that reducing inflammation would improve the microcirculatory oxygenation in the kidney and limit the onset of acute kidney injury (AKI). METHODS: Rats were randomized into five groups (n = 8 per group): (1) control group, (2) control + NAC, (3) endotoxemic shock with lipopolysaccharide (LPS) without fluids, (4) LPS + fluid resuscitation, and (5) LPS + fluid resuscitation + NAC (150 mg/kg/h). Fluid resuscitation was initiated at 120 min and maintained at fixed volume for 2 h with hydroxyethyl starch (HES 130/0.4) dissolved in acetate-balanced Ringer's solution (Volulyte) with or without supplementation with NAC (150 mg/kg/h). Oxygen tension in the renal cortex (CµPO2), outer medulla (MµPO2), and renal vein was measured using phosphorimetry. Biomarkers of renal injury, inflammation, and oxidative stress were assessed in kidney tissues. RESULTS: Fluid resuscitation significantly improved the systemic and renal macrohemodynamic parameters after LPS. However, the addition of NAC further improved cortical renal oxygenation, oxygen delivery, and oxygen consumption (p < 0.05). NAC supplementation dampened the accumulation of NGAL or L-FABP, hyaluronic acid, and nitric oxide in kidney tissue (p < 0.01). CONCLUSION: The addition of NAC to fluid resuscitation may improve renal oxygenation and attenuate microvascular dysfunction and AKI. Decreases in renal NO and hyaluronic acid levels may be involved in this beneficial effect. A therapeutic strategy combining initial fluid resuscitation with antioxidant therapies may prevent sepsis-induced AKI.

Fully Balanced Fluids do not Improve Microvascular Oxygenation, Acidosis and Renal Function in a Rat Model of Endotoxemia.

The expectation of fluid therapy in patients with septic shock is that it corrects hypovolemia, with the aim of restoring tissue perfusion and oxygenation and organ function. This study investigated whether different types of resuscitation fluids were effective in improving renal microcirculatory oxygenation, acidosis, oxidative stress, and renal function in a rat model of endotoxemic shock. Five groups of rats were used: a sham group, a lipopolysaccharide (LPS) group, and three LPS groups that received 30 mL/kg/h of 0.9% sodium chloride (0.9% NaCl), a new bicarbonate buffered crystalloid solution closely resembling the composition of plasma (FB-Cxt) or a hydroxyethyl starch-ringer acetate solution. Systemic hemodynamic variables, renal blood flow, microvascular oxygenation, oxidative/nitrosative stress, and renal function were measured. LPS-induced shock was only partially resolved by fluid administration. Animals became arterially hypotensive despite adequate central venous pressure. Hydroxyethyl starch-ringer acetate was more effective at improving arterial pressures and renal blood flow than 0.9% NaCl or FB-Cxt. Fluids had marginal effects on pH and HCO3 levels irrespective of the buffer, or on renal µPO2 and dysfunction. Colloids increased the markers of renal oxidative stress (P < 0.001), whereas unbalanced crystalloids increased the markers of nitrosative stress during sepsis (P < 0.01). Endotoxemia-induced acidosis and decreases in renal µPO2 or renal injury were not corrected solely by fluid resuscitation, irrespective of the buffer of the fluid. Our study supported the idea that fluids must be supplemented by other compounds that specifically correct renal inflammation and oxygenation to be effective in resolving septic shock-induced renal failure.

Scavenging reactive oxygen species using tempol in the acute phase of renal ischemia/reperfusion and its effects on kidney oxygenation and nitric oxide levels.

BACKGROUND: Renal ischemia/reperfusion (I/R) injury is commonly seen in kidney transplantation and affects the allograft survival rates. We aimed to test our hypothesis that scavenging reactive oxygen species (ROS) with tempol would protect renal oxygenation and nitric oxide (NO) levels in the acute phase of renal I/R. METHODS: Rats were randomly divided: (1) no I/R, no tempol; (2) no I/R, but with tempol; (3) I/R without tempol; and (4) I/R with tempol. I/R was induced by 30-min clamping of the renal artery. Tempol (200 µmol/kg/h/i.v) was administered 15 min prior to I/R. RESULTS: I/R without tempol led to a significant decrease in renal oxygen delivery and microvascular oxygenation. Tempol, however, protected renal oxygenation after I/R. At R90, the creatinine clearance rate was lower in the I/R-subjected group that did not receive tempol compared to that in the other groups. I/R injury without tempol treatment led to a significant increase in tissue malondialdehyde levels and a significant decrease in tissue NO levels. Tempol administration before I/R could prevent oxidative stress and altered tissue NO levels. CONCLUSIONS: This underscores that unbalance between oxygen, NO, and ROS forms an important component of the pathogenesis of I/R-induced AKI and should therefore be taken into account when designing a prevention/treatment strategy for renal I/R injury in transplantation.

TEMPOL has limited protective effects on renal oxygenation and hemodynamics but reduces kidney damage and inflammation in a rat model of renal ischemia/reperfusion by aortic clamping.

BACKGROUND: Renal ischemia-reperfusion (I/R) is a common clinical complication in critically ill patients that is associated with considerable morbidity and mortality. Renal I/R is a major cause of acute kidney injury (AKI) resulting from I/R-induced oxidative stress, sterile inflammation, and microcirculatory perfusion defects, which can be ameliorated with the superoxide scavenger TEMPOL. The most common cause of AKI in the clinical setting is aortic surgery with suprarenal aortic clamping. The protective effect of TEMPOL in aortic clamping-induced renal I/R has not been studied before. AIM: To evaluate the protective effects of TEMPOL on oxidative stress, inflammation, tissue injury, and renal hemodynamics and oxygenation in a clinically representative rat model of I/R using aortic cross-clamping. METHODS: Animals (N = 24) were either sham-operated or subjected to ischemia (30 min) and 90-min reperfusion, with or without TEMPOL treatment (15 min before ischemia and during entire reperfusion phase, 200 µmol/kg/h). Systemic and renal hemodynamics, renal oxygenation, and blood gas values were determined at 15 min and 90 min of reperfusion. At 90-min reperfusion, iNOS, inflammation (IL-6, MPO), oxidative stress (MDA), and tissue damage (NGAL, L-FABP) were determined in tissue biopsies. RESULTS: TEMPOL administration at a cumulative dose of 400 µmol/kg conferred a protective effect on AKI in terms of reducing renal damage, inflammation, and iNOS activation. With respect to renal hemodynamics and oxygenation, TEMPOL only reduced renal vascular resistance to near-baseline levels at both reperfusion time points and partially ameliorated the I/R-induced drop microvascular partial tension of oxygen at 90 min reperfusion. Also, TEMPOL alleviated the I/R-induced metabolic acidosis. However, TEMPOL exerted no restorative effect in terms of the severely reduced mean arterial pressure, renal blood flow, and renal oxygen delivery and consumption. The renal oxygen extraction ratio remained unchanged during the 90-min reperfusion phase. Kidneys in all groups were anuric throughout the experiment. CONCLUSIONS: This clinically representative renal I/R model, which entails both renal I/R and hind limb I/R as opposed to the standardly used renal I/R model that employs renal artery clamping, resulted in relatively moderate direct AKI. The damage was exacerbated by the perturbed systemic hemodynamics and metabolic acidosis as a result of the hind limb I/R. TEMPOL partially intervened in the factors that led to AKI as well as renal microvascular partial tension of oxygen and metabolic acidosis. However, more effective interventions should be devised for the mean arterial pressure drop (i.e., anuria) associated with aortic clamping and for restoring other critical renal hemodynamic and oxygenation parameters in order to improve post-I/R renal function. RELEVANCE FOR PATIENTS: TEMPOL is a promising compound that has been shown to protect kidneys from I/R damage, which is relevant in kidney transplantation, pancreas transplantation, and aortic aneurysm repair in kidney transplant patients. This study suggests that intervening with TEMPOL is not sufficient to ensure optimal clinical outcome in patients that have undergone aortic clamping and that more effective interventions should be investigated.

Ascorbic acid improves renal microcirculatory oxygenation in a rat model of renal I/R injury.

BACKGROUND AND OBJECTIVES: Acute kidney injury (AKI) is a clinical condition associated with a degree of morbidity and mortality despite supportive care, and ischemia/reperfusion injury (I/R) is one of the main causes of AKI. The pathophysiology of I/R injury is a complex cascade of events including the release of free oxygen radicals followed by damage to proteins, lipids, mitochondria, and deranged tissue oxygenation. In this study, we investigated whether the antioxidant ascorbic acid would be able to largely prevent oxidative stress and consequently, reduce I/R-related injury to the kidneys in terms of oxygenation, inflammation, and renal failure. MATERIALS AND METHODS: Rats were divided into three groups (n = 6/group): (1) a time control group; (2) a group subjected to renal ischemia for 60 min by high aortic occlusion followed by 2 h of reperfusion (I/R); and (3) a group subjected to I/R and treated with an i.v. 100 mg/kg bolus ascorbic acid 15 min before ischemia and continuous infusion of 50 mg/kg/hour for 2 h during reperfusion (I/R + AA). We measured renal tissue oxidative stress, microvascular oxygenation, renal oxygen delivery and consumption, and renal expression of inflammatory and injury markers. RESULTS: We demonstrated that aortic clamping and release resulted in increased oxidative stress and inflammation that was associated with a significant fall in systemic and renal hemodynamics and oxygenation parameters. The treatment of ascorbic acid completely abrogated oxidative stress and inflammatory parameters. However, it only partly improved microcirculatory oxygenation and was without any effect on anuria. CONCLUSION: The ascorbic acid treatment partly improves microcirculatory oxygenation and prevents oxidative stress without restoring urine output in a severe I/R model of AKI.

Erythropoietin pretreatment suppresses seizures and prevents the increase in inflammatory mediators during pentylenetetrazole-induced generalized seizures.

Erythropoietin (EPO) suppresses epileptic seizures, but the mechanism is unclear. The search for novel targets in the therapy of epilepsy has focused recently on brain inflammation since brain inflammation and the associated blood-brain barrier (BBB) damage appears to be an integral part of epilepsy pathophysiology. We examined the effects of EPO on proinflammatory mediators in brain and serum in PTZ-induced generalized seizure model. The inflammation markers (IL-1ß, TNF-α, IL-6, IL-10), BBB and neuron damage markers (S100B, Neuron specific enolase; NSE, respectively) in serum and brain of Sprague-Dawley male rats were examined with the ELISA method. Nitric oxide synthase (NOS) isoforms were investigated immunohistochemically in hippocampus. EPO treatment 4 h and 24 h before PTZ administration had diverse effects. EPO treatment 4 h before PTZ administration elongated the seizure latency, decreased the inflammation and damage markers in serum and brain significantly, whereas EPO treatment 24 h before PTZ administration lowered inflammation and damage markers to control levels and decreased the seizure stage. PTZ-induced seizures increased inducible NOS (iNOS) activity and decreased endothelial NOS (eNOS) activity in hippocampus. Both EPO pretreatments reversed these effects. These findings, i.e., decreased iNOS activity and increased eNOS activity by EPO suggest the first time that the favorable effect of EPO pretreatment on inflammatory mediators triggered by PTZ-induced seizures. This can provide further insight into epilepsy treatment and new prophylactic strategies against epilepsy risk.

Bis maltolato oxovanadium (BMOV) and ischemia/reperfusion-induced acute kidney injury in rats.

BACKGROUND: The aim of the present study was to test the potential protective effects of the organic vanadium salt bis (maltolato) oxovanadium (BMOV; 15 mg/kg) in the context of renal ischemia/reperfusion (30 min of ischemia) and its effects on renal oxygenation and renal function in the acute phase of reperfusion (up to 90 min post-ischemia). METHODS: Ischemia was established in anesthetized and mechanically ventilated male Wistar rats by renal artery clamping. Renal microvascular and venous oxygenation were measured using phosphorimetry. Creatinine clearance rate, sodium reabsorption, and renal oxygen handling efficiency were considered markers for renal function. RESULTS: The main findings were that BMOV did not affect the systemic and renal hemodynamic and oxygenation variables and partially protected renal sodium reabsorption. CONCLUSIONS: Pretreatment with the organic vanadium compound BMOV did not protect the kidney from I/R injury.

The acute effects of acetate-balanced colloid and crystalloid resuscitation on renal oxygenation in a rat model of hemorrhagic shock.

INTRODUCTION: Fluid resuscitation therapy is the initial step of treatment for hemorrhagic shock. In the present study we aimed to investigate the acute effects of acetate-balanced colloid and crystalloid resuscitation on renal oxygenation in a rat model of hemorrhagic shock. We hypothesized that acetate-balanced solutions would be superior in correcting impaired renal perfusion and oxygenation after severe hemorrhage compared to unbalanced solutions. METHODS: In anesthetized, mechanically ventilated rats, hemorrhagic shock was induced by withdrawing blood from the femoral artery until mean arterial pressure (MAP) was reduced to 30 mmHg. One hour later, animals were resuscitated with either hydroxyethyl starch (HES, 130/0.42 kDa) dissolved in saline (HES-NaCl; n=6) or a acetate-balanced Ringer's solution (HES-RA; n=6), as well as with acetated Ringer's solution (RA; n=6) or 0.9% NaCl alone (NaCl; n=6) until a target MAP of 80 mmHg was reached. Oxygen tension in the renal cortex (CµPO2), outer medulla (MµPO2), and renal vein were measured using phosphorimetry. RESULTS: Hemorrhagic shock (MAP=30 mmHg) significantly decreased renal oxygenation and oxygen consumption. Restoring the MAP to 80 mmHg required 24.8±1.7 ml of NaCl, 21.7±1.4 ml of RA, 5.9±0.5 ml of HES-NaCl (p<0.05 vs. NaCl and RA), and 6.0±0.4 ml of HES-RA (p<0.05 vs. NaCl and RA). NaCl, RA, and HES-NaCl resuscitation led to hyperchloremic acidosis, while HES-RA resuscitation did not. Only HES-RA resuscitation could restore renal blood flow back to ∼85% of baseline level (from 1.9±0.1 ml/min during shock to 5.1 ml±0.2 ml/min 60 min after HES-RA resuscitation) which was associated with an improved renal oxygenation (CµPO2 increased from 24±2 mmHg during shock to 50±2 mmHg 60 min after HES-RA resuscitation) albeit not to baseline level. At the end of the protocol, creatinine clearance was decreased in all groups with no differences between the different resuscitation groups. CONCLUSION: While resuscitation with the NaCl and RA (crystalloid solutions) and the HES-NaCl (unbalanced colloid solution) led to hyperchloremic acidosis, resuscitation with the HES-RA (acetate-balanced colloid solution) did not. The HES-RA was furthermore the only fluid restoring renal blood flow back to ∼85% of baseline level and most prominently improved renal microvascular oxygenation.

Balanced vs unbalanced crystalloid resuscitation in a near-fatal model of hemorrhagic shock and the effects on renal oxygenation, oxidative stress, and inflammation.

BACKGROUND: The aim of the present study was to test the hypothesis that balanced crystalloid resuscitation would be better for the kidney than unbalanced crystalloid resuscitation in a rat hemorrhagic shock model. METHODS: Male Wistar rats were randomly assigned to four groups (n=6/group): (1) time control; (2) hemorrhagic shock control; (3) hemorrhagic shock followed by unbalanced crystalloid resuscitation (0.9% NaCl); and (4) hemorrhagic shock followed by acetate and gluconate-balanced crystalloid resuscitation (Plasma Lyte). We tested the solutions for their effects on renal hemodynamics and microvascular oxygenation, strong-ion difference, systemic and renal markers of inflammation and oxidative stress including glycocalyx degradation as well as their effects on renal function. RESULTS: The main findings of our study were that: (1) both the balanced and unbalanced crystalloid solutions successfully restored the blood pressure, but renal blood flow was only recovered by the balanced solution although this did not lead to improved renal microvascular oxygenation; (2) while unbalanced crystalloid resuscitation induced hyperchloremia and worsened metabolic acidosis in hemorrhaged rats, balanced crystalloid resuscitation prevented hyperchloremia, restored the acid-base balance, and preserved the anion gap and strong ion difference in these animals; (3) in addition balanced crystalloid resuscitation significantly improved renal oxygen consumption (increased VO(2), decreased [Formula: see text] ); and (4) however neither balanced nor unbalanced crystalloid resuscitation could normalize systemic inflammation or oxidative stress. Functional immunohistochemistry biomarkers showed improvement in L-FABP in favor of balanced solutions in comparison to the hemorrhagic group although no such benefit was seen for renal tubular injury (measured by NGAL) by giving either unbalanced or balanced solutions. CONCLUSIONS: Although balanced crystalloid resuscitation seems superior to balanced crystalloid resuscitation in protecting the kidney after hemorrhagic shock and is certainly better than not applying fluid resuscitation, these solutions were not able to correct systemic inflammation or oxidative stress associated with hemorrhagic shock.

The role of renal hypoperfusion in development of renal microcirculatory dysfunction in endotoxemic rats.

PURPOSE: To study the role of renal hypoperfusion in development of renal microcirculatory dysfunction in endotoxemic rats. METHODS: Rats were randomized into four groups: a sham group (n = 6), a lipopolysaccharide (LPS) group (n = 6), a group in which LPS administration was followed by immediate fluid resuscitation which prevented the drop of renal blood flow (EARLY group) (n = 6), and a group in which LPS administration was followed by delayed (i.e., a 2-h delay) fluid resuscitation (LATE group) (n = 6). Renal blood flow was measured using a transit-time ultrasound flow probe. Microvascular perfusion and oxygenation distributions in the renal cortex were assessed using laser speckle imaging and phosphorimetry, respectively. Interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α were measured as markers of systemic inflammation. Furthermore, renal tissue samples were stained for leukocyte infiltration and inducible nitric oxide synthase (iNOS) expression in the kidney. RESULTS: LPS infusion worsened both microvascular perfusion and oxygenation distributions. Fluid resuscitation improved perfusion histograms but not oxygenation histograms. Improvement of microvascular perfusion was more pronounced in the EARLY group compared with the LATE group. Serum cytokine levels decreased in the resuscitated groups, with no difference between the EARLY and LATE groups. However, iNOS expression and leukocyte infiltration in glomeruli were lower in the EARLY group compared with the LATE group. CONCLUSIONS: In our model, prevention of endotoxemia-induced systemic hypotension by immediate fluid resuscitation (EARLY group) did not prevent systemic inflammatory activation (IL-6, IL-10, TNF-α) but did reduce renal inflammation (iNOS expression and glomerular leukocyte infiltration). However, it could not prevent reduced renal microvascular oxygenation.

Effects of sepiapterin infusion on renal oxygenation and early acute renal injury after suprarenal aortic clamping in rats.

Acute kidney injury (AKI) can occur after aortic clamping due to microvascular dysfunction leading to renal hypoxia. In this rat study, we have tested the hypothesis that the administration of the precursor of the nitric oxide synthase essential cofactor tetrahydrobiopterin (BH4) could restore renal oxygenation after ischemia reperfusion (I/R) and prevent AKI. We randomly distributed rats into 4 groups: sham group; ischemia-reperfusion group; I/R + sepiapterin, the precursor of BH4; and I/R + sepiapterin + methotrexate, an inhibitor of the pathway generating BH4 from sepiapterin. Cortical and outer medullary microvascular oxygen pressure, renal oxygen delivery, renal oxygen consumption were measured using dual-wavelength oxygen-dependent quenching phosphorescence techniques during ischemia and throughout 3 hours of reperfusion. Kidney injury was assessed using myeloperoxidase staining for leukocyte infiltration and urine neutrophil gelatinase-associated lipocalin levels. Ischemia reperfusion induced a drop in microvascular PO2 (P < 0.01 vs. Sham, both), which was prevented by the infusion of sepiapterin. Sepiapterin partially prevented the rise in renal oxygen extraction (P < 0.001 vs. I/R). Finally, treatment with sepiapterin prevented renal infiltration by inflammatory cells and decreased urine neutrophil gelatinase-associated lipocalin levels indicating a decrease of renal injury. These effects were blunted when adding methotrexate, except for myeloperoxidase. In conclusion, the administration of sepiapterin can prevent renal hypoxia and AKI after suprarenal aortic clamping in rats.

L-NIL prevents renal microvascular hypoxia and increase of renal oxygen consumption after ischemia-reperfusion in rats.

Even though renal hypoxia is believed to play a pivotal role in the development of acute kidney injury, no study has specifically addressed the alterations in renal oxygenation in the early onset of renal ischemia-reperfusion (I/R). Renal oxygenation depends on a balance between oxygen supply and consumption, with the nitric oxide (NO) as a major regulator of microvascular oxygen supply and oxygen consumption. The aim of this study was to investigate whether I/R induces inducible NO synthase (iNOS)-dependent early changes in renal oxygenation and the potential benefit of iNOS inhibitors on such alterations. Anesthetized Sprague-Dawley rats underwent a 30-min suprarenal aortic clamping with or without either the nonselective NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) or the selective iNOS inhibitor L-N(6)-(1-iminoethyl)lysine hydrochloride (L-NIL). Cortical (CmicroPo(2)) and outer medullary (MmicroPo(2)) microvascular oxygen pressure (microPo(2)), renal oxygen delivery (Do(2ren)), renal oxygen consumption (Vo(2)(ren)), and renal oxygen extraction (O(2)ER) were measured by oxygen-dependent quenching phosphorescence techniques throughout 2 h of reperfusion. During reperfusion renal arterial resistance and oxygen shunting increased, whereas renal blood flow, CmicroPo(2), and MmicroPo(2) (-70, -42, and -42%, respectively, P < 0.05), Vo(2)(ren), and Do(2ren) (-70%, P < 0.0001, and -28%, P < 0.05) dropped. Whereas L-NAME further decreased Do(2ren), Vo(2)(ren), CmicroPo(2), and MmicroPo(2) and deteriorated renal function, L-NIL partially prevented the drop of Do(2ren) and microPo(2), increased O(2)ER, restored Vo(2)(ren) and metabolic efficiency, and prevented deterioration of renal function. Our results demonstrate that renal I/R induces early iNOS-dependent microvascular hypoxia in disrupting the balance between microvascular oxygen supply and Vo(2)(ren), whereas endothelial NO synthase activity is compulsory for the maintenance of this balance. L-NIL can prevent ischemic-induced renal microvascular hypoxia.

The relationship between nitric oxide and leptin in the lung of rat with streptozotocin-induced diabetes.

Lung structural changes and immunoreactivity of endothelial (eNOS)- and inducible nitric oxide synthase (iNOS) were investigated by light microscopy in lungs of treated and untreated diabetic rats. Diabetes was induced by a single intraperitoneal (i.p.) injection of 65 mg kg(-1) streptozotocin (STZ) in Wistar albino male rats. Diabetic rats received daily i.p. doses of dexamethasone (2 mg kg(-1)), leptin (0.5 microg kg(-1)) and intramuscular insulin (20 U kg(-1)) or a combination of these drugs for 1 week starting 4 weeks after the STZ injections. After treatment, the blood levels of glucose, leptin, insulin and nitrate/nitrite (NO(3) (-)/NO(2) (-)) were measured. Dilatation of alveoli and alveolar ducts, partial alveolar wall thickening and increased eNOS- and iNOS characterized the diabetic rat lungs. High blood glucose and nitrate/nitrite levels as well as low insulin and leptin levels were also present. Treatment with insulin, dexamethasone and a combination of these drugs resulted in improvement of the structural and immunohistochemical abnormalities. The most effective treatment was insulin therapy. Leptin administration resulted in increased relative amounts of extracellular material, which led to noticeable respiratory efficiency in the diabetic rat lungs. All treatments except leptin lowered blood glucose levels. The combination of insulin and dexamethasone increased blood leptin and insulin, while the remaining diabetic rats had blood with low leptin and insulin concentrations. These results suggest that therapy with insulin plus dexamethasone but not therapy with leptin is beneficial for diabetics.

Effects of orlistat and its relationship with nitric oxide in the small intestinal mucosa.

Nitric oxide (NO) is known to be a messenger molecule that plays an important role in physiological and pathological conditions. It is synthesized by an enzyme called nitric oxide synthase (NOS). Inducible NOS (iNOS), one of the three isomers of NOS, has both protective and toxic properties. In this study, the role of NO has been evaluated by gastrointestinal symptoms induced by orlistat which is used in obesity treatment. Orlistat was given to Wistar rats with and without iNOS inhibition. The effects of orlistat and inhibition of NOS were studied. Glucose, urea, alanine transaminase (ALT), and gamma glutamil transpeptidase (GGT) were descreased after short- and long- term orlistat applications. Dexamethasone increased level of these enzymes. Cholesterol and triglyceride were increased in all experimental groups than the controls. This increment was more severe in animals received orlistat and dexamethasone together. Small intestinal tissue also were researched histologically and NADPH-diaphorase (NADPH-d) histochemistrically. Orlistat caused histological damages in brush border membranes, connective tissues of villi, and lymphocyte migration also increased. Dexamethasone treatment prevented these damages partially while orlistat increased the NOS distribution in the tissue sections. Dexamethasone, which is an iNOS inhibitor, decreased NADPH-d histochemistry. There was a similiar NOS distribution both in the control and orlistat+dexamethasone group. Hence, we concluded that long- term trials with orlistat and similar drugs are needed.

In vivo inhibition of inducible nitric oxide and evaluation of the brain tissue damage induced by Toxocara canis larvae in experimentally infected mice.

Nitric oxide (NO) is known to be produced by macrophages, endothelial cells and neurons and synthesized by an enzyme called nitric oxide synthase (NOS). Various effector mechanisms and infections can affect the NO production. Excessive amount of NO will lead to biochemical reactions, which cause toxic effects. In this study the role of NO has been evaluated in larval toxocarosis, which is a systemic parasite infection caused by T. canis larvae. Infection was established in the Balb/c mice with or without inducible NOS (iNOS) inhibition and the effects of infection and NOS inhibition were observed according to the results of SOD and LPx measurements in brain tissue and NADPH-diaphorase (NADP-d) histochemistry. Results of NADPH-d histochemistry indicate that iNOS inhibition has protective effect on the brains of infected mice and that larval T. canis infection could be related to oxidative stress, and NO production and iNOS inhibition can protect the tissue from damage in this infection.