Tubular Necrosis In Rats Research Articles

Histopathological nephrotoxic features of high oral doses of ubiquinone in rats

Abstract Co-enzyme Q10 (Co-Q10) plays a key role in the cellular respiration for the production of ATP. The toxicity of quinones to the kidney appears to depend on variety of factors, including genetic polymorphisms and the individual’s comorbidites. The aim of the present study was to assess histologically the nephrotoxic effects of 6 weeks daily oral intake of Co-Q10 in experimental animals. Twenty-five Wistar rats weighing between 220-270 g were randomly divided into two groups: experimental “treated” and control “untreated” groups (n=15, n=10, respectively). The animals of the experimental group received 300 mg/kg daily dose of gelatinous capsules of Co-Q10 by oral gavage for six weeks. At the end of the study, all animals were sacrificed under general anesthesia and samples of the kidneys were excised for microscopic histopathological assessment of renal tissue using stain. The experimental group showed a range of mild to severe dilatation of Bowman’s space, with a mean corpuscular diameter of 294±38 µm that was significantly higher (p <0.05) than that of the untreated control group 208±31 µm. Shrinkage to complete destruction of the glomeruli was observed in the experimental group only. The long-term use of high doses of Co-Q10 had revealed a selective nephrotoxicity towards podocytes. This might be a risk factor leading to renal proximal tubular necrosis in rats and the subsequent renal function deterioration.

The attenuative effects of oral resveratrol on renal changes induced by vanadium injection in rats

Introduction: Resveratrol (trans-3,5,4’-trihydroxystilbene) as a polyphenol with potential antioxidant and anti-inflammatory properties is known as an effective herbal medicine in different disorders in rats. Objectives: The present study was carried out to investigate the protective effects of oral consumption of resveratrol on vanadium induced renal injury in male Wistar rats. Materials and Methods: Animals received either ammonium metavanadate (AMV, 5 mg/ kg/d, (intraperitoneally; 14 consecutive days) or resveratrol solution (10 mg/kg and 50 mg/kg, gastric gavage) along with AMV treatment. The last group received resveratrol alone (50 mg/ kg, gastric gavage) for 4 weeks. Results: AMV injection caused progressive tubular damages resembling acute tubular necrosis. Microscopic views revealed tubular attenuation and blebbing. In addition, progressive peritubular congestion of the capillaries observed while no evidence of renal fibrosis was present in trichrome staining. Further, levels of the renal transforming growth factor β1 (TGF-β1) as an index of fibrosis had no difference in treated animals as compared with the control (13.4±1.2 versus 11.24±0.93 pg/mg protein) at the P<0.05. However, in AMVtreated animals receiving the higher dose of resveratrol (50 mg/kg), the renal superoxide dismutase (SOD) activity, showed no difference as compared with the saline-treated rats (42±1.3 versus 51±1.4). Conclusions It is evident that AMV injection had no ability to induce renal fibrosis in rats while it evokes renal destructive lesions based on pathological results and enzyme levels. Moreover, our preliminary results suggest that resveratrol in high dose (50 mg/kg) could confer a minor role against AMV induced renal tubular necrosis in rats due to pathological results.

Evaluation of Renal Protection From High Doses of Melatonin in an Experimental Model of Renal Ischemia and Reperfusion in Hyperglycemic Rats

BackgroundMelatonin is a free radical scavenger with important actions in the study of renal ischemia and reperfusion (I/R). This study evaluated possible renal protection of high doses of melatonin in an experimental model of I/R in which rats were submitted to acute hyperglycemia under anesthesia with isoflurane. MethodForty-four male Wistar rats, weighing more than 300 g, were randomly divided into 5 groups: G1, sham (n = 10); G2, melatonin (n = 10; 50 mg.kg−1); G3, hyperglycemia (n = 9; glucose 2.5 g.kg−1); G4, hyperglycemia/melatonin (n = 10; 2.5 g.kg−1 glucose + melatonin 50 mg.kg−1); and G5, I/R (n = 5). In all groups, anesthesia was induced with 4% isoflurane and maintained with 1.5% to 2.0% isoflurane. Intraperitoneal injection of melatonin (G1, G4), glucose (G3, G4), or saline (G1, G5) was performed 40 minutes before left renal ischemia. Serum plasma values for creatinine and glucose were determined at baseline (M1), immediately following reperfusion (M2), and 24 hours after completion of the experiment (M3). Histological analysis was performed to evaluate tubular necrosis (0–5). ResultsSerum glucose was higher at M2 in the groups supplemented with glucose, hyperglycemia (356.00 ± 107.83), and hyperglycemia/melatonin (445.3 ± 148.32). Creatinine values were higher at T3 (P = .0001) for I/R (3.6 ± 0.37), hyperglycemia/melatonin (3.9 ± 0.46), and hyperglycemia (3.71 ± 0.69) and lower in the sham (0.79 ± 0.16) and melatonin (2.01 ± 1.01) groups, P < .05. Histology showed no necrosis injury in the G1, lesion grade 2 in the G2, and severe acute tubular necrosis in the G3: (grade 4), G4: (grade 5) and G5: (grade 4) groups (P < .0001). DiscussionMelatonin protected the kidneys submitted to I/R in rats without hyperglycemia; however, this did not occur when the I/R lesion was associated with hyperglycemia. ConclusionsDue to its antioxidant and antiapoptotic action, melatonin was able to mitigate, but not prevent acute tubular necrosis in rats with hyperglycemia under anesthesia by isoflurane.

Renoprotective effects of the crude extract and solvent fractions of the leaves of Euclea divinorum Hierns against gentamicin-induced nephrotoxicity in rats

Ethnopharmacological relevanceEuclea divinorum Hierns (Ebenaceae) is used in Ethiopian folklore medicine to treat scabies, inflammation of the skin, eczema, abdominal pain, gonorrhea, and kidney problems. However, the claim has not been scientifically validated. Aim of the studyTo assess the renoprotective effects of the crude extract and solvent fractions of E. divinorum leaves against gentamicin-induced nephrotoxicity in rats. Materials and methodRats of either sex were divided into seven experimental groups, each comprising six animals. Group I served as control and given vehicle (Tween 80, 2%, v/v in water) and Group II were treated with gentamicin intraperitoneally (100mg/kg/day) for eight days. Group III–V received crude extract at three different doses 100mg/kg, 150mg/kg and 200mg/kg, respectively. Group VI received 100mg/kg of the methanolic fraction and Group VII 100mg/kg of the aqueous fraction. The extract was administered orally two days before and eight days concomitantly with gentamicin. Following treatment, blood and renal tissue were used to assess creatinine, blood urea nitrogen (BUN), malondialdehyde (MDA), antioxidant enzymes and tubular necrosis using recommended procedures. In addition, free radical scavenging activity was determined using in vitro methods. ResultsGentamicin significantly increased serum creatinine and BUN, MDA and tubular necrosis in rats. It also decreased activity of catalase and superoxide dismutase as well as levels of gluthatione. Pre- and co-treatment with the crude extract and solvent fractions of Euclea divinorum leaves reversed gentamicin-induced alterations as evidenced by a decrease in tubular necrosis, serum and oxidant markers as well as by an increase in antioxidant molecules. Effect was found to decrease with dose when the crude extract was used and maximum protection was conferred by 100mg/kg of the methanolic fraction in both in vivo and in vitro studies. ConclusionsEuclea divinorum reversed gentamicin-induced nephrotoxicity, probably via its antioxidant activity. The fact that the methanol fraction conferred maximum protection suggests that semi-polar antioxidant principles might be responsible for the observed effect.

Effects of hypercholesterolemia on contrast media-induced nephrotoxicity in rats

To explore the effects of short- and long-term dietary hypercholesterolemia on contrast media-induced nephrotoxicity in rats. The male Wistar rats were fed either a normal rodent diet or a high cholesterol diet. At the end of 2 and 8 weeks, 8 rats from each group received a tail vein injection of either Iohexol injection (groups NC and HC) or vehicle (groups N and H). Blood lipid, renal function, renal hemodynamics, renal and urinary prostaglandin E2 (PGE2) and thromboxane B2 (TXB2), renal nitric oxide and malondialdehyde (MDA) were determined at Day 1 following the administration of contrast media. The dosing of contrast media induced obviously increased serum creatinine compared with normal rats ((185 ± 28) vs (53 ± 3) µmol/L, P < 0.01) and severe renal tubular necrosis in rats with a high cholesterol diet for 8 weeks but did not in normal-diet rats or rats with a high cholesterol diet for 2 weeks. The renal and urinary levels of PGE2 and TXB2 increased significantly in rats of groups H and HC at the end of 8 weeks. The renal production of nitric oxide decreased while the concentration of MDA increased markedly in groups HC and H at the end of 8 weeks. Long-term hypercholesterolemia appears to be a risk factor of contrast media-induced acute renal failure. And it may be associated with the disorder of intrarenal prostaglandins and the abnormality of renal nitric oxide system as induced by lipid peroxidation.

Nephroprotective effects of subcapsular transplantation of metanephric mesenchymal cells on gentamicin-induced acute tubular necrosis in rats

The subcapsular transplantation of metanephric mesenchymal cells (MMCs) may be a new therapeutic approach for the treatment of acute tubular necrosis (ATN). To investigate this hypothesis and provide evidence for its possible use in the clinic, we evaluated the nephroprotective effects of transplanting MMCs into the renal subcaspsule of rats with ATN induced by gentamicin. MMCs were expanded in culture. After gentamicin-induced ATN was established, fluorescently-labeled cells were transplanted and traced in kidney tissues by fluorescence microscopy. Serum creatinine (Cr), urea nitrogen (BUN), and N-acetyl-b-D-glucosaminidase (NAG) levels were determined at different time points. Kidney pathology was studied by hematoxylin-eosin staining. Apoptosis was examined by the TUNEL assay. In the MMCs-treated group, the mortality rate decreased; BUN, Cr, and NAG levels peaked at 8 days, and were significantly lower than those in the other groups at 11 and 14 days. RIMM-18 cells locally recruited through precise tropism to sites of injury had the ability to migrate into the tubuli from the renal subcapsule. Damage to the cell-treated kidneys was reduced. The pathologic lesion scores of tubular damage reached the highest values at 8 days in the treated kidneys and 11 days in the untreated ones. The apoptotic index showed that the peaks of apoptosis occurred at earlier stages of the injury process in cell-treated than in untreated kidney and thereafter declined in a time-dependent manner. The subcapsular transplantation of MMCs could ameliorate renal function and repair kidney injury.

Effect of Stem Cell Factor and Granulocyte-Macrophage Colony-Stimulating Factor-Induced Bone Marrow Stem Cell Mobilization on Recovery from Acute Tubular Necrosis in Rats

Background: Acute tubular necrosis (ATN) is the most common reason for acute kidney injury (AKI), and there is still an absence of effective therapies. Objective: To assess the value of bone marrow cell mobilization by stem cell factor (SCF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) therapy in rats with gentamicin-induced ATN. Methods: ATN was induced in male Sprague–Dawley (SD) rats with five daily high-dose intraperitoneal injections of gentamicin. Subcutaneous injections of SCF and GM-CSF were administered simultaneously and these cytokines were observed on days 2, 5, 10, 17, 24, and 31. Peripheral blood and renal tissue CD34+ cell count, mortality rate, blood urea nitrogen (BUN), serum creatinine (SCr), creatinine clearance rate (CCr), and histopathologic lesion scores were determined. Twelve hours after bone marrow ablation (BMA) by lethal X-ray radiation, specific pathogen-free (SPF) ATN rats were given five daily injections of SCF and GM-CSF. BUN, SCr, and histopathologic lesion scores were evaluated on days 2, 5, and 10. Results: Peripheral blood CD34+ cell count increased significantly in ATN rats between 2 and 10 days after SCF and GM-CSF injection. Mortality was reduced from 34.7% in the ATN group to 18.6% in the ATN+CSF. In addition, cytokines administration significantly decreased SCr and BUN. Moreover, cytokines rapidly ameliorated tubular injury. There was no significant effect on ATN rats after BMA. Conclusions: This study demonstrated that SCF and GM-CSF effectively mobilized bone marrow cells in ATN rats, and cytokines administration partially prevented gentamicin-induced ATN. These results suggest that bone marrow stem cell (BMSC) mobilization may be an effective therapy for ATN.

Effects of Short- and Long-Term Hypercholesterolemia on Contrast-Induced Acute Kidney Injury

Background: Whether hypercholesterolemia is a risk factor for contrast-induced acute kidney injury (CI-AKI) remains unclear. In the present study, the effects of short- and long-term dietary hypercholesterolemia on contrast media-induced nephrotoxicity were evaluated. Methods: Rats were fed either a normal rodent diet (N) or high-cholesterol diet (H). At the end of 2 and 8 weeks, 8 rats from each diet group were given a tail vein injection of either iohexol (group NC and group HC) or vehicle (group N and group H). Blood lipids, renal function and renal hemodynamics were evaluated 1 day after contrast media administration. Renal and urinary prostaglandin E₂ (PGE₂) and thromboxane B₂ (TXB₂) were detected by radioimmunoassay. Renal nitric oxide and malondialdehyde (MDA) were measured by the Griess reaction and thiobarbituric acid method, respectively. Results: Contrast media administration increased serum creatinine levels and induced severe renal tubular necrosis in rats fed the high-cholesterol diet for 8 weeks but not in rats fed the normal diet or high-cholesterol diet for 2 weeks. The renal and urinary PGE₂ and TXB₂ levels increased significantly in rats in group H and group HC at the end of 8 weeks. Renal nitric oxide production decreased, and MDA levels increased markedly in group HC and group H at the end of 8 weeks. Conclusions: We conclude that long-term hypercholesterolemia appeared to be a risk factor for CI-AKI, which might be associated with disorders in intrarenal prostaglandins and abnormalities in renal nitric oxide system induced by lipid peroxidation.

Integrated transcriptomic and proteomic evaluation of gentamicin nephrotoxicity in rats

Gentamicin is an aminoglycoside antibiotic, which induces renal tubular necrosis in rats. In the context of the European InnoMed PredTox project, transcriptomic and proteomic studies were performed to provide new insights into the molecular mechanisms of gentamicin-induced nephrotoxicity. Male Wistar rats were treated with 25 and 75 mg/kg/day subcutaneously for 1, 3 and 14 days. Histopathology observations showed mild tubular degeneration/necrosis and regeneration and moderate mononuclear cell infiltrate after long-term treatment. Transcriptomic data indicated a strong treatment-related gene expression modulation in kidney and blood cells at the high dose after 14 days of treatment, with the regulation of 463 and 3241 genes, respectively. Of note, the induction of NF-kappa B pathway via the p38 MAPK cascade in the kidney, together with the activation of T-cell receptor signaling in blood cells were suggestive of inflammatory processes in relation with the recruitment of mononuclear cells in the kidney. Proteomic results showed a regulation of 163 proteins in kidney at the high dose after 14 days of treatment. These protein modulations were suggestive of a mitochondrial dysfunction with impairment of cellular energy production, induction of oxidative stress, an effect on protein biosynthesis and on cellular assembly and organization. Proteomic results also provided clues for potential nephrotoxicity biomarkers such as AGAT and PRBP4 which were strongly modulated in the kidney. Transcriptomic and proteomic data turned out to be complementary and their integration gave a more comprehensive insight into the putative mode of nephrotoxicity of gentamicin which was in accordance with histopathological findings.

Experimental study of the prevention and treatment of acute renal tubular necrosis with erythropoietin in rats

Objective To study the preventive effects of erythropoietin （EPO） on acute tubular necrosis of kidney in rats. Method The rat models of acute renal tubular necrosis were established with injecting glycerol in dose of 10 mL/kg. Thirty Wistar rats were randomly （random number） divided into control group, model group and EPO treatment group. EPO was administered intravenously into rats of treatment group in a dose of 1000IU/kg. Levels of blood urea nitrogen （BUN） and serum creatinine （Scr）, urine osmolality, urine N-acetyl-D-glucosaminidase （NAG）, urine β2-microglobulin （β2-MG）, tissue MDA and SOD of rats in the three groups were assayed after the experiment. Renal histological examination was also performed. Results Compared with model group, the levels of BUN and Scr, urine osmolality, NAG,β2-MG and tissue MDA in EPO treament group were significantly lower, but urine osmolality and tissue SOD of rats remarkably increased in comparison with model group. EPO also lessened the histological changes in treatment group. Conclusions EPO has some protective effects on acute renal tubular necrosis in rats, which is probably through preventing oxygen free radical damage and elevating endogenous antioxidation potential. Key words: Erythropoietin; Acute tubular necrosis; Oxygen free radicals

Spirulina Platensis Protects Against Renal Injury in Rats with Gentamicin-Induced Acute Tubular Necrosis

The present study was carried out to evaluate the renoprotective antioxidant effect of Spirulina platensis on gentamicin-induced acute tubular necrosis in rats. Albino-Wistar rats, (9male and 9 female), weighing approximately 250 g, were used for this study. Rats were randomly assigned to three equal groups. Control group received 0,9 % sodium chloride intraperitoneally for 7 days at the same volume as gentamicin group. Gentamicin group was treated intraperitoneally with gentamicin, 80 mg/kg daily for 7 days. Gentamicin+spirulina group received Spirulina platensis 1000 mg/kg orally 2 days before and 7 days concurrently with gentamicin (80 mg/kg i.p.). Nephrotoxicity was assessed by measuring plasma nitrite concentration, stabile metabolic product of nitric oxide with oxygen. Plasma nitrite concentration was determined by colorimetric method using Griess reaction. For histological analysis kidney specimens were stained with hematoxylin-eosin (HE) and periodic acid-Schiff (PAS) stain. Plasma nitrite concentration and the level of kidney damage were significantly higher in gentamicin group in comparison both to the control and gentamicin+spirulina group. Spirulina platensis significantly lowered the plasma nitrite level and attenuated histomorphological changes related to renal injury caused by gentamicin. Thus, the results from present study suggest that Spirulina platensis has renoprotective potential in gentamicin-induced acute tubular necrosis possibly due to its antioxidant properties.

Validation of putative genomic biomarkers of nephrotoxicity in rats

Drug-induced renal injury is a common finding in the early preclinical phase of drug development. But the specific genes responding to renal injury remain poorly defined. Identification of drug-induced gene changes is critical to provide insights into molecular mechanisms and detection of renal damage. To identify genes associated with the development of drug-induced nephrotoxicity, a literature survey was conducted and a panel of 48 genes was selected based on gene expression changes in multiple published studies. Male Sprague–Dawley rats were dosed daily for 1, 3 or 5 days to the known nephrotoxicants gentamicin, bacitracin, vancomycin and cisplatin, or the known hepatotoxicants ketoconazole, 1-naphthyl isothiocyanate and 4,4-diaminodiphenylmethane. Histopathological evaluation and clinical chemistry revealed renal proximal tubular necrosis in rats treated with the nephrotoxicants, but not from those treated with the hepatotoxicants. RNA was extracted from the kidney, and RT-PCR was performed to evaluate expression profiles of the selected genes. Among the genes examined, 24 genes are confirmed to be highly induced or repressed in rats treated with nephrotoxicants; further investigation identified that 5 of the 24 genes were also altered by hepatotoxicants. These data led to the identification of a set of genomic biomarker candidates whose expression in kidney is selectively regulated only by nephrotoxicants. Among those genes displaying the highest expression changes specifically in nephrotoxicant-treated rats were kidney injury molecule 1 (Kim1), lipocalin 2 (Lcn2), and osteopontin (Spp1). The establishment of such a genomic marker set offers a new tool in our ongoing quest to monitor nephrotoxicity.

The Effects of Inducible Nitric Oxide Synthase Inhibitor L-N6-(1-Iminoethyl) Lysine in Gentamicin-Induced Acute Tubular Necrosis in Rats

The aim of this study was to investigate the role of inducible nitric oxide synthase (iNOS) in gentamicin-induced acute tubular necrosis in rats using the iNOS inhibitor L-N6-(1-iminoethyl) lysine (L-NIL). Wistar rats, both sexes (n=18), were equally divided into three groups. Gentamicin group received intraperitoneally (i.p.) gentamicin in 0.9 % NaCl at a dose of 80 mg/kg/day for five consecutive days. L-NIL+gentamicin group received L-NIL at a dose of 3 mg/kg i.p. 36, 24 and 12 h before first dose of gentamicin. Control group received 0.9 % NaCl i.p. for five consecutive days at the equal volume as gentamicin group. Griess reaction was used for determination plasma level of NO. Semiquantitative histological analysis was used for the evaluation of kidney damage level. The plasma NO level and the level of kidney damage were statistically higher in gentamicin group in comparison to the control group (p=0.046). Application of L-NIL prior to gentamicin led to certain decrease in the plasma level of NO as well as in the level of kidney damage. Application of L-NIL, prior to gentamicin administration, did not provide complete protective effects of L-NIL on the kidney, which was demonstrated on kidney sections. The lack of anticipated protective effect of L-NIL on kidney tissue might be explained with the fact that we have used L-NIL prior but not during/after gentamicin administration. It would be necessary to examine the effects of L-NIL administration not only before, but as well during and possibly after the administration of gentamicin.

A Protein Toxin from the Sea Anemone Phyllodiscus semoni Targets the Kidney and Causes a Severe Renal Injury with Predominant Glomerular Endothelial Damage

Envenomation by the sea anemone Phyllodiscus semoni causes fulminant dermatitis and, rarely, acute renal failure in humans. Here, we investigated whether the venom extracted from the nematocysts (PsTX-T) was nephrotoxic when administered intravenously in rats and whether PsTX-T induced activation of the complement system. Although small dose of PsTX-T induced acute tubular necrosis in rats resembling pathology seen in patients, kidneys displayed glomerular injury with glomerular endothelial damage, thrombus formation, mesangiolysis, and partial rupture of glomerular basement membrane, accompanied by severe tubular necrosis at 24 hours after administration of 0.03 mg of PsTX-T per animal, similar to the glomerular findings typical of severe hemolytic uremic syndrome. The early stage injury was accompanied by specific PsTX-T binding, massive complement C3b, and membrane attack complex deposition in glomeruli in the regions of injury and decreased glomerular expression of complement regulators. A pathogenic role for complement was confirmed by demonstrating that systemic complement inhibition reduced renal injury. The isolated nephrotoxic component, a 115-kd protein toxin (PsTX-115), was shown to cause identical renal pathology. The demonstration that PsTX-T and PsTX-115 were highly nephrotoxic acting via induction of complement activation suggests that inhibition of complement might be used to prevent acute renal damage following envenomation by P. semoni.

Effect of inducers of DT-diaphorase on the haemolytic activity and nephrotoxicity of 2-amino-1,4-naphthoquinone in rats

Reduction of naphthoquinones by DT-diaphorase is often described as a detoxification reaction. This is true for some naphthoquinone derivatives, such as alkyl and di-alkyl naphthoquinones, but the situation with other substances, such as 2-hydroxy-1,4-naphthoquinone, is more complex. In the present study, the effect of several substances that are known to increase tissue activities of DT-diaphorase on the toxicity of 2-amino-1,4-naphthoquinone has been investigated. Like 2-hydroxy-1,4-naphthoquinone, the 2-amino-derivative was found to cause both haemolytic anaemia and renal tubular necrosis in rats. Again like 2-hydroxy-1,4-naphthoquinone, the severity of the haemolysis induced by the 2-amino derivative was increased in animals pre-treated with inducers of DT-diaphorase, but the degree of nephrotoxicity was decreased. With these substances, therefore, DT-diaphorase both activates and detoxifies the quinone, depending on the target organ. It is not possible to generalise with regard to the effects of modulation of tissue levels of DT-diaphorase on naphthoquinone toxicity in vivo, since this may change not only the severity of the toxic effects, but also the target organ specificity. In evaluating the possible therapeutic applications of such compounds, the possibility of toxic effects upon the blood and kidney must be borne in mind. In man, renal damage by compounds such as 2-hydroxy- and 2-amino-1,4-naphthoquinone may be a particular problem, because of the low level of DT-diaphorase in human liver.

Nitric Oxide in Gentamicin-induced Acute Tubular Necrosis in Rats

Gentamicin is still widely used in clinical practice in spite of its renal toxicity. The role of nitric oxide (NO) in that process is not completely elucidated. The aim of this study was to investigate the relationship between plasma level of NO and the histopathological changes of kidney in acute tubular necrosis (ATN) induced by gentamicin in rats. Study was carried out in Albino-Wistar rats, both sexes (n=16), average body weight 200-250 g. divided in two equal groups: control and gentamicin group. The control group was injected with 0.9% NaCl i.p. and gentamicin group was injected with gentamicin in the dose of 80 mg/kg/day i.p. in a period of 5 consecutive days. NO plasma level was determined by the production of nitrates and nitrites using classical colorimetrical Griess reaction. Kidney specimens were stained with hematoxylin-eosin (H-E) and Periodic acid-Schiff (PAS) stain. Semiquantitative histological analysis was used for the evaluation of the level of kidney damage. Both, the plasma NO level and the level of kidney damage were statistically higher in rats with gentamicin-induced ATN in comparison to the control group. In spite of that the correlation between plasma NO level and the level of kidney damage was not found. The rise of plasma level NO in gentamicin induced ATN in rats could possibly indicate on the role of NO in renal damage caused by gentamicin.

The protective effect of taurine against gentamicin‐induced acute tubular necrosis in rats

Taurine, which is the major intracellular free beta-amino acid, is known to be an endogenous antioxidant and a membrane-stabilizing agent. In this study, we wished to know whether taurine altered the concentration of gentamicin in kidney tissue and could protect against gentamicin-induced acute proximal tubular injury. Wistar albino rats of both sexes were assigned to three groups, which all received one of the following daily intraperitoneal injections for 8 days: (i) 0.9% sodium chloride (NaCl) alone at the same volume as gentamicin treated rats (group C; n=8); (ii) 100 mg/kg/day gentamicin alone (group G; n=8, four male, four female); or (iii) 100 mg/kg/day gentamicin plus 7.5 ml/kg/day taurine (group G+T; n=9, five male, four female). Urine was collected for 24 h for the determination of urine volume and creatinine. Intracardiac blood was collected for blood urea nitrogen (BUN) and serum creatinine determination. The kidneys were removed, weighed, and the left kidneys were subjected to biochemical analysis for the determination of thiobarbituric acid-reactive substance (TBARS) and lactate levels, and glutathione peroxidase (Gpx) and superoxide dismutase (SOD) activities. The right kidneys were divided vertically in half. The upper halves were used for histopathological examination, by light and electron microscopy. The lower halves were used to detect the gentamicin concentration within the kidney tissue, by high-performance liquid chromatography (HPLC). Changes in body weight and normalized kidney weight were recorded. Taurine treatment reduced gentamicin-induced increases in serum creatinine, 24 h urine volume, BUN and tissue lactate and TBARS levels (0.57+/-0.02 vs 1.06+/-0.08 mg/dl, P<0.001; 9.00+/-1.46 vs 20.9+/-2.73 ml, P<0.001; 25.3+/-1.87 vs 54.1+/-6.99 mg/dl, P<0. 001; 2.56+/-0.10 vs 3.44+/-0.08 micromol/g wet tissue, P<0.001; and 66.4+/-3.41 vs 79.5+/-5.07 nmol/g wet tissue, P>0.05, respectively). Taurine reduced the accumulation of gentamicin within the kidney tissue (233+/-29 vs 494+/-93 microg/g wet tissue, P<0.05). Taurine treatment also prevented body weight loss due to gentamicin administration (17.8+/-1.64 vs -10.0+/-7.08 g, P<0.01) and normalized reduced Gpx and SOD activities (3.46+/-0.16 vs 2.37+/-0. 15 U/g wet tissue, P<0.01; and 15577+/-377 vs 12662+/-577 U/g wet tissue, P<0.01, respectively). Light microscopic examination of the renal tissues from gentamicin-treated rats revealed severe histopathological changes, whereas specimens obtained from taurine-treated rats revealed only mild changes. This finding was supported by electron microscopic examination. Our observations suggest that taurine treatment attenuates the accumulation of gentamicin within kidney tissue and counteracts the deleterious effect of gentamicin on renal tubular function. They may have potentially important clinical implications.

Immunochemical detection of quinol--thioether-derived protein adducts.

Glutathione (GSH) conjugates of hydroquinone (HQ) and 2-bromohydroquinone (2-BrHQ) produce severe renal proximal tubular necrosis in rats. Since the reactivity of quinones lies, in part, in their ability to alkylate proteins, our goal was to develop an immunochemical method with which to investigate the role of protein adduct formation in quinone-thioether-mediated toxicity. An immunogen was synthesized by coupling 2-bromo-6-(N-acetylcystein-S-yl)hydroquinone (2-BrHQ-NAC) to keyhole-limpet hemocyanin (KLH). Anti-2-BrHQ-NAC-KLH antibodies were raised in rabbits and purified by affinity chromatography. Antibody binding to the 2-BrHQ-NAC epitope was confirmed by competitive enzyme-linked immunosorbent assay (ELISA) with a bovine serum albumin conjugate of 2-BrHQ-NAC. Affinity-purified anti-2-BrHQ-NAC-KLH antibodies recognized adducted proteins in the kidneys of rats treated with HQ, 2-BrHQ, 2-bromo-bis(glutathion-S-yl)hydroquinone, 2-(glutathion-S-yl)hydroquinone, 2, 5-bis(glutathion-S-yl)hydroquinone, and 2,3, 5-tris(glutathion-S-yl)hydroquinone. Immunoreactive proteins were found in all renal subcellular fractions of 2-BrHQ-treated rats, and the distribution of adducts was similiar to that obtained by quantifying 2-Br[14C]HQ covalent adducts. Western blot analysis revealed that three proteins, at 42, 46, and 79 kDa, were adducted by all the compounds examined. The identification of these adducted proteins will be required to assess their significance in quinol-thioether-mediated nephrotoxicity.

Choice of tests in the biochemical assessment of nephrotoxicity in dogs and rats: A study with maleic acid

Urinalysis represents a useful tool in the evaluation of new pharmaceutical agents acting on the kidney, during preclinical toxcological studies. In the present study, we studied the response of urinary creatinine, LDH, AAP, ALP, β-GAL, GGT, NAG and protein excretion to a single intravenous dose of maleic acid (25 mg/kg for dogs and 100 mg/kg for rats). Enzymes were selected based on their association with renal toxicity and their localisation within the renal tubule. They included lysosomal, brush border and cytosolic enzymes. In male dogs, increases in enzyme excretion occurred within 2 h of maleic acid administration, peaked 3 h after dosing, and returned towards, or to, predose value at 24 h. Marked increases in enzyme levels occurred only for LDH and GGT (10-fold or more). Additionally, there was a marked increase in total protein excretion whereas creatinine excretion decreased. In male rats, the only major difference from control was a higher 0–24 h protein excretion rate (approximately 2-fold). Moderate increases were also present for ALP, GGT and LDH (< 2-fold). In female rats, there was a marked increase in urinary excretion of proteins and LDH, GGT, NAG and ALP (8–13-fold when compared to controls). There were only moderate increases (2-fold or less) in β-GAL and AAP. Creatinine was unaffected by the treatment. Histopathological examination of the kidney revealed moderate to severe proximal tubular necrosis in dogs and minimal to moderate tubular necrosis in rats. Overall, urinary GGT, LDH, total protein and creatinine concentrations are the most sensitive biochemical indicators of maleic acid nephrotoxicity in dogs. For rats, in addition to LDH excretion, total proteins, NAG, GGT and ALP can be considered to be sensitive markers of renal injury. In the light of these results and those reported in literature, it was concluded that the response of urinary markers of nephrotoxicity is heavily dependent on the nephrotoxic agent, the dose, species, sex and study design. Consequently, a range of markers should be examined in order to identify the most useful.

Insulin-like growth factor I accelerates recovery from ischemic acute tubular necrosis in the rat.

The effects of administering insulin-like growth factor I (IGF-I) were examined in a model of ischemic acute tubular necrosis in rats. Injury was induced by 75 min of bilateral renal artery occlusion. Compared to rats administered vehicle, rats administered IGF-I (100 micrograms/day via continuous subcutaneous infusion) had significantly lower serum creatinine and blood urea nitrogen levels over the course of 7 days postocclusion. Glomerular filtration rate as determined by inulin clearance was examined on day 2 postocclusion and was significantly increased in IGF-I-treated animals (0.16 +/- 0.02 ml per min per 100 g of body weight) compared to vehicle-treated controls (0.08 +/- 0.02 ml per min per 100 g of body weight). The weight loss that occurred during the course of acute tubular necrosis was ameliorated by IGF-I. Mortality was reduced from 36.7% in vehicle-treated rats to 7.1% in rats administered IGF-I. Histologically, there was much less renal injury evident at day 7 postocclusion in the IGF-I-treated rats compared to vehicle-treated controls. In contrast, growth hormone (200 micrograms administered subcutaneously for 4 days) did not affect recovery of renal function or reduce mortality postreperfusion. This report demonstrates a beneficial effect of IGF-I administration in the setting of acute tubular necrosis. Several properties of IGF-I render it a pharmacological agent with excellent potential for treatment of this condition in humans.