Cyclosporine Nephrotoxicity In Rats Research Articles

Effects of Atorvastatin and Carvedilol on Chronic Cyclosporine Nephrotoxicity in Rats

Background: To evaluate possible renoprotective effects of carvedilol and atorvastatin in CsA induced kidney injury in a rat model. Methods: Twenty-two rats were divided into five groups. Control, CsA, CsA+karvedilol, CsA+Atorvastatin, CsA +karvedilol+atorvastatin groups. Drugs were given for 21 days. Results: Serum BUN and creatinine were significantly higher in the CsA group compared to control and were significantly lower in the CsA+carvedilol+atorvastatin group compared to CsA group. Tissue MDA levels were found to be lower in group 3, 4 and 5 than in CsA group. Tissue SOD in CsA +carvedilol and CsA+atorvastatin groups were found higher than the control and CsA groups. Tissue NO levels were found to be higher only in CsA+carvedilol+atorvastatin group compared to CsA group. Apical budding, hyaline casts and apoptosis in the tubular system was significantly higher in the CsA group than in the treatment groups. Osteopontin showed strong positivity especially in the CsA group. Osteopontin density was lower in the treatment group. Conclusions: This is the first study to evaluate MDA, SOD and NO at tissue level for carvedilol in cyclosporine nephrotoxicity. Carvedilol and atorvastatin may contribute to the reduction of renal injury in chronic CsA nehropathy. These agents have provided a protective effect on renal functions especially in combined treatment.

Molecular Mechanisms of the Protective Effects of Calcium Channel Blockers against Cyclosporine Nephrotoxicity in Rats

Nephrotoxicity is a serious side effect for the immunosuppressant drug cyclosporine (CSA). In this study, we tested the hypothesis that the concurrent administration of calcium channel blockers such as verapamil or nifedipine guard against renal dysfunction induced by CSA in rats. Studies were then extended to investigate the roles of inflammatory, oxidative, and fibrotic pathways in the interaction. Six groups of male rats (n=6 each) were employed and allocated to receive one of the following treatments for 7 consecutive days: vehicle (cremophor), CSA (25 mg.kg−1.day−1), verapamil (2 mg.kg−1.day−1), nifedipine (3 mg.kg−1.day−1), CSA plus verapamil, or CSA plus nifedipine. Biochemical and histomorphometric analyses showed that compared with respective vehicle values, rats treated with CSA exhibited clear signs of nephrotoxicity that included: (i) proteinuria and elevations in serum creatinine and blood urea nitrogen, (ii) deposition of collagen IV in glomerular and tubular areas, and (iii) increases in the glomerulosclerosis index. While the single administration of nifedipine or verapamil caused no specific renal effects, the concurrent use of either calcium channel blocker significantly and equipotently ameliorated the biochemical and morphological derangements caused by CSA. We also report that the inflammatory (NF‐κB expression), oxidative (NADPH‐oxidase activity, reactive oxygen species production), and fibrotic (TGF‐β1 expression) manifestations of renal toxicity induced by CSA were eliminated upon concurrent administration of nifedipine or verapamil. Together, these results highlight the usefulness of calcium channel blocking agents in negating the CSA‐induced nephrotoxicity and predisposing biochemical and molecular machinaries.Support or Funding InformationSupported by the Lebanese National Council for Scientific Research (CNRS) Grant Award for Lebanese graduates (Awardee: Safaa Hammoud)

The effects of valsartan on renal glutathione peroxidase expression in alleviation of cyclosporine nephrotoxicity in rats.

Introduction: Nephrotoxicity as a side effect caused by the immunosuppressive drug, cyclosporine-A (CsA), can be a major problem in transplant medicine. Oxidative stress may play an important role in the CsA-induced nephrotoxicity. It has been shown that the antihypertensive drug, valsartan (Val), has also renoprotective effects but, its molecular mechanism is largely unknown. In the present study, it was aimed to evaluate the Val effect in the alleviation of CsA nephrotoxicity via probable renal glutathione peroxidase (GPx) upregulation and oxidative stress decrease. Methods: Thirty-two Sprague-Dawley rats were divided into four groups based on CsA and/or Val administration: group A (Control, 1 mL/kg/day of olive oil as vehicle), group B (CsA, 30 mg/kg/day), group C (CsA+Val, 30+30 mg/kg/day), and group D (Val, 30 mg/kg/day). After the administration period (six weeks), renal GPx expression was evaluated by real-time polymerase chain reaction (PCR). Plasma levels of GPx and 8-Hydroxydeoxyguanosine (8-OHdG) were measured by enzyme-linked immunosorbent assay (ELISA). Malondialdehyde (MDA) and protein carbonyl groups (PCG) were measured by spectrophotometer. Plasma levels of urea and creatinine were measured by an autoanalyzer. Results: CsA treatment led to the decrease in renal expression and plasma levels of GPx in comparison to other study groups. Rats received CsA were detected to have significantly (p<0.05) higher plasma 8-OHdG, MDA, PCG, urea, and creatinine levels in comparison to other groups. Plasma urea and creatinine levels were negatively correlated with renal GPx expression and positively correlated with the oxidative stress markers. Conclusion:Administration of Val may result in attenuating the nephrotoxic side effect of CsA via probable renal GPx upregulation, and subsequently oxidative stress decrease.

Retraction Note: The effects of valsartan on renal glutathione peroxidase expression in alleviation of cyclosporine nephrotoxicity in rats.

[This retracts the article DOI: 10.15171/bi.2016.18.].

Beneficial effect of shallot (Allium ascalonicum L.) extract on cyclosporine nephrotoxicity in rats

The clinical use of an immunosuppressive cyclosporine A (CsA) is limited by its serious nephrotoxic effect. Evidences have suggested the role of oxidative stress in its pathogenesis. Shallot (Allium ascalonicum L.) has recently been shown to possess antioxidative and free radical scavenging abilities. The present study was undertaken to investigate the possible beneficial effect of shallot extract on renal injury caused by CsA. Male Wistar rats were treated orally with vehicle, CsA (25mg/kg), shallot extract (1g/kg), and CsA plus shallot extract for 21 days. Renal function, histopathology, tissue malondialdehyde (MDA) and glutathione (GSH) levels were evaluated 24h after the last treatment. CsA-induced nephrotoxicity was evidenced by increased blood urea nitrogen and serum creatinine, but decreased urea and creatinine clearance. The kidney of CsA treated rats exhibited severe vacuolations and tubular necrosis. CsA also induced oxidative stress, as indicated by increased renal MDA and reduced GSH concentrations. Administration of shallot extract along with CsA counteracted the deleterious effects of CsA on renal dysfunction, oxidative stress markers, and morphological changes. These data indicate the protective potential of shallot extract against CsA nephrotoxicity and suggest a significant contribution of its antioxidant property to this beneficial effect.

In vivo effect of the natural antioxidant hydroxytyrosol on cyclosporine nephrotoxicity in rats

Cyclosporine A (CsA) is the first-line immunosuppressant used in transplant patients and in auto- immune diseases. Nephrotoxicity is the major limitation of CsA use. Although the mechanisms of nephrotoxicity have not been completely defined, some evidence suggests that reactive oxygen species (ROS) play a causal role. The present study was designed to investigate in vivo effects of hydroxytyrosol (DOPET), a natural olive oil antioxidant, on oxidative stress, renal histology and haemodynamic alterations induced in rats by CsA treatment. Adult Sprague-Dawley rats were treated i.p. with CsA (15 mg/kg) alone or in combination with DOPET (20 mg/kg) for 3 weeks. At the end of the treatment, superoxide concentration within the cells of the abdominal aorta and renal artery was quantified from the oxidation of dihydroethidium (DHE) using fluorescence microscopic imaging analysis. In kidney tissues, lipid peroxidation was measured by thiobarbituric acid-reacting substances (TBARS) assay, glutathione level was assessed enzymatically and the expression of haem oxygenase-1 (HO-1) gene was evaluated by semiquantitative RT-PCR. Renal morphology was studied by classical histological techniques, while the glomerular filtration rate (GFR) was estimated by inulin clearance. Systemic blood pressure was monitored by the tail method and through the catheterization of the carotid artery. CsA administration increased superoxide concentration both in the aorta and in the renal artery, while DOPET completely prevented this effect. Higher levels of TBARS, a significant decrease in GSH and an upregulation of HO-1 mRNA were observed in the kidneys of CsA-treated rats. DOPET treatment reversed quantitatively these effects. However, CsA-dependent changes in renal histology were only partially reversed by DOPET. Finally, CsA induced a severe reduction in GFR and a significant increase in both systolic and diastolic blood pressure; the DOPET treatment had no significant effect on these haemodynamic alterations. The reported data indicate that effective DOPET protection from CsA-induced oxidative stress is associated with a mild effect on histological damages and does not affect the altered glomerular function and the hypertension, thus indicating that kidney injury by CsA is only in part dependent on oxidative stress.

Pioglitazone limits cyclosporine nephrotoxicity in rats

Chronic cyclosporine (CsA) nephrotoxicity is a relevant factor in the pathogenesis of chronic allograft nephropathy. Pioglitazone is an agonist of PPARγ, capable of reducing chronic inflammation. We investigated the capacity of pioglitazone in preventing renal dysfunction. Adult male Wistar rats were assigned to: Vehicle (olive oil 1 ml/kg/day), CsA (10 mg/kg/day) alone and with pioglitazone (5 or 10 mg/kg/day). The animals were sacrificed at 28 days, where blood (serum creatinine ratio, CR) and kidney samples (arteriolopathy analyses) were collected. The mRNA transcripts of TGF-β1, PAI-1, Smad3 and 7 were evaluated by real-time PCR. As expected, CsA treatment significantly decreased renal function that peaked at day 28, compared with vehicle (CR = 1.29 ± 0.03 vs. 0.95 ± 0.14, p < 0.05). In contrast, the administration of pioglitazone 5 or 10 mg/kg combined with CsA resulted in better renal function (CR = 1.09 ± 0.05 and 1.14 ± 0.14, respectively, p < 0.05). Animals treated with CSA showed relevant arteriolopathy (49.5 ± 2.86%) and pioglitazone administration significantly limited it (37.0 ± 3.59% and 36.6 ± 1.72%, respectively, 5 or 10 mg/kg, p < 0.05). In CsA-treated animals (alone and with pioglitazone), TGF-β1 and Smad3 increased significantly. In animals treated with CsA and pioglitazone (5 mg/kg), PAI-1 was significantly lower than CsA alone (3.96 ± 0.92 vs. 7.53 ± 1.38, p < 0.05). Interestingly, the administration of pioglitazone 5 or 10 mg/kg was associated with an increase in Smad7 (1.79 ± 0.25 and 1.75 ± 0.19, respectively), compared to vehicle and to CsA-treated groups (1.08 ± 0.17 and 1.17 ± 0.19, respectively, p < 0.05). These data provide evidence that pioglitazone acts through down regulation of pro-fibrotic cytokine PAI-1 and overexpression of the regulatory Smad7.

Systemic administration of recombinant human relaxin (RHRLX) ameliorates the acute cyclosporine nephrotoxicity in rats

Relaxin: a 6KD peptide hormone of pregnancy belonging to insulin family, has three major biological actions: antifibrotic, angiogenic and vasodilatory. Previous publications have shown that treatment with recombinant human relaxin (rhRlx) elicits potent renal vasodilation and hyperfiltration in conscious rats in a nitric oxide (NO)-dependent manner, thereby mimicking pregnancy. The objective of this study was to evaluate the effect of rhRlx treatment in acute cyclosporine nephrotoxicity in rats. A total of 124 rats female Sprague Dawley rats were treated with either Cyclosporine (30 mg/kg po day 0-10) or vehicle (olive oil) for 10 days. Recombinant human relaxin (rhRlx) at a dose of either 0.1 mg/kg/day or 0.5 mg/kg/day was administered to rats subcutaneously via Alzet pumps (Alza Corp., Mountain view, CA) from day 0-10. On day 5 and day 10 of treatment, the kidney function (urine flow rate [UFR] and glomeruli filtration rate [GFR]) measured by metabolic cages was recorded. On day 10, the animals were euthanized and the kidneys fixed for were subjected to histological evaluation. Consistent with previously published results, systemic administration of rhRlx (0.5 mg/kg) to normal rats enhanced both GFR and UFR by day 5 (147% & 168% compared to the baseline, respectively). CsA treatment alone significantly reduced GFR and UFR on both day 5 and day 10 compared to baseline by 30-35% (p<0.05). The GFR and UFR in animals treated with both CsA and rhRlx at either a dose of 0.1mg/kg or 0.5 mg/kg were significantly improved compare to those of animals treated with CsA alone (p<0.05). Histological evaluation of cross-sections of the kidneys from rats at day 10 revealed marked improvement on tubulopathy and arteriolopathy. The data from this study demonstrated that systemic administration of rhRlx enhanced kidney functions in normal rats and ameliorated cyclosporine induced acute nephrotoxicity in rats. The results indicate that systemic administration of rhRlx may have potential benefit in preventing cyclosporine-induced nephrotoxicity.

Individual or combined effects of enalapril and verapamil on chronic cyclosporine nephrotoxicity in rats.

Previous studies have demonstrated that enalapril and verapamil seem to attenuate the cyclosporine nephrotoxicity. However, the mechanisms have not been completely understood, especially on molecular events. The aim of this study was to examine the effect of individual or combined treatment on osteopontin, TGF-beta, endothelin-1 and procollagen alpha 1(I) mRNA expressions. Enalapril (50 mg/L in drinking water) and verapamil (0.5 mg/kg/day, subcutaneously), alone or in combination, were administered to rats with chronic cyclosporine nephrotoxicity (cyclosporine, 25 mg/kg/day, subcutaneously) (n = 5 each). Five rats treated with olive oil vehicle were used as control. After 4 weeks, biochemical parameters were measured, and renal cortical mRNA levels were evaluated by Northern blot analysis. Cyclosporine reduced renal creatinine clearance significantly and induced renal cortical osteopontin, TGF-beta, endothelin-1 and procollagen alpha 1(I) gene expressions around 13.5 +/- 1.3, 2.4 +/- 0.2, 1.5 +/- 0.1, 1.9 +/- 0.1 folds, respectively. Individual treatment with enalapril or verapamil significantly suppressed the osteopontin and TGF-beta mRNA expression, but not endothelin-1 and procollagen alpha 1(I). Combined treatment also inhibited the osteopontin and TGF-beta mRNA expression but there was no difference between combined and individual treatment. In conclusion, enalapril or verapamil significantly blunted the cyclosporine-induced osteopontin and TGF-beta gene expressions. However, combined treatment did not show any additive effect.

Oral Supplementation of L-Arginine Prevents Chronic Cyclosporine Nephrotoxicity in Rats

This study was performed to evaluate the effect of L-arginine (L-Arg) on the prevention of chronic cyclosporine (CsA) nephrotoxicity in rats. Rats pair-fed a low-salt diet (0.05%) were given CsA (15 mg/kg/day s.c.), CsA and L-Arg (L-Arg group, 1.25 g/l water), CsA and N-nitro-L-arginine methyl ester (L-NAME group, 70 mg/l water) or vehicle. After 28 days, the L-Arg group had a higher glomerular filtration rate compared to the CsA (0.42±0.05 vs. 0.31±0.06 ml/min/100 g, p < 0.05) and the L-NAME groups (vs. 0.19± 0.04 ml/min/100 g, p < 0.05) and a significantly lower serum creatinine level compared to the CsA (0.70±0.06 vs. 0.92±0.12 mg/dl, p < 0.05) and the L-NAME groups (vs. 1.21±0.17 mg/dl, p < 0.05). The L-Arg group had less fibrosis, tubular injury (TI), and arteriolopathy than the CsA (fibrosis 0.39±0.14 vs. 0.74±0.15; TI 1.3±0.3 vs. 2.0±0.1; arteriolopathy 33±7 vs. 48±17, p < 0.05, respectively) and the L-NAME groups (fibrosis vs. 1.67±0.32, TI vs. 2.6±0.3, arteriolopathy vs. 63±10, p < 0.05, respectively). Plasma renin activity in the L-Arg group was less than in the CsA (18±2 vs. 23±3 ng Ang I/ml/h, p < 0.05) and the L-NAME groups (vs. 30±3 ng Ang I/ml/h, p < 0.05). Nitric oxide production in L-Arg group was higher than in the CsA (24.2±1.7 vs. 11.1±1.5 µmol/24 h, p < 0.05) and the L-NAME groups (vs. 8.4±1.0 µmol/24 h, p < 0.05). In conclusion, the nitric oxide pathway is associated with the pathogenesis of chronic CsA nephrotoxicity, and exogenous L-Arg supplementation is effective in reducing chronic CsA nephrotoxicity in rats.

Light microscopic and electron microscopic features of cyclosporine nephrotoxicity in rats.

In order to clarify morphologic changes associated with cyclosporine (CS) nephrotoxicity, CS in ethyl alcohol at 25 mg/kg/day i.p. was administered to male Sprague-Dawley rats for periods of 1 to 8 weeks. Mean systolic BP was slightly increased in the CS group at 4 weeks (p < 0.05), but there was no difference compared to a control group at 8 weeks. Blood urea nitrogen was significantly elevated at 4 weeks and continued to rise (p < 0.005), whereas serum creatinine was elevated at 8 weeks. Microscopic examination of the kidneys from CS-treated rats at one week revealed cytoplasmic vacuolization in all segments of the proximal tubules, tubular inclusion bodies, and peritubular capillary congestion. Ultrastructurally, some vacuoles were neutral fat droplets, while others appeared as single membrane-bound structures due to dilatation of the endoplasmic reticulum. The tubular inclusion bodies were enlarged autolysosomes filled with distorted mitochondrial fragments. At two weeks, tubular regeneration was prominent, in addition to the above mentioned toxic tubulopathy. At four weeks, focal areas of interstitial fibrosis and tubular atrophy associated with cystic dilatation were seen. At 8 weeks, interstitial and intratubular microcalcification were present, in addition to patchy foci of interstitial fibrosis, but vascular lesions were not demonstrated. Although renal tubular changes characterized by vacuolization, inclusion bodies, and microcalcification and interstitial fibrosis are not specific for CS toxicity, these changes are commonly found in both humans and rats at high doses of CS.

Arginine feeding modifies cyclosporine nephrotoxicity in rats.

Glycine (G) infusion causes renal vasodilation mediated by nitric oxide (NO). Cyclosporine A (CsA) nephrotoxicity is characterized by preglomerular vasoconstriction and decreased efferent arteriolar tone probably related to reduced NO and angiotensin II, respectively. L-Arginine (ARG) is a precursor to NO. To test the hypothesis that chronic CsA decreases renal NO activity, we compared the glomerular hemodynamic response to glycine infusion in rats after 8 d of CsA (30 mg/kg per d s.c.), CsA and ARG (1.6 g/kg per d p.o.) (A/CsA), and in two groups of pair-fed controls (CON, A/CON). Single nephron GFR (SNGFR), single nephron plasma flow (SNPF), glomerular capillary hydrostatic pressure gradient (delta P), proximal tubular reabsorption (APR), and kidney tissue angiotensin II (AIIk) were measured before and during G. CsA was associated with baseline decrements in SNGFR, SNPF, delta P, and AIIk, and with a blunted hemodynamic response to G. In CON, ARG did not affect baseline hemodynamics or modify the response to G. In CsA, ARG decreased baseline preglomerular resistance and restored the glomerular hemodynamic response to G. G was associated with a significant increase in AIIk in both CON and CsA. These findings suggest that (a) CsA is associated with decreased AIIk, and (b) CsA may diminish NO activity within the kidney, and that this capacity may be partially restored by arginine feeding.

The effects of nifedipine on cyclosporine nephrotoxicity in rats

The effects of nifedipine on cyclosporine nephrotoxicity in rats

The effects of nifedipine on cyclosporine nephrotoxicity in rats.

We have investigated the effect of nifedipine on cyclosporine nephrotoxicity in the Sprague-Dawley rat employing a repeatable, single-shot, isotopic technique of measuring the glomerular filtration rate (GFR) and effective renal plasma flow (ERPF). Groups of 10 rats received either cyclosporine 5 mg/kg daily or cremaphor with either nifedipine 0.5 mg/kg daily or its vehicle for 14 days. In the cyclosporine group the GFR (P < 0.001, paired t-test), ERPF and filtration fraction (FF) (P < 0.01) all fell significantly. The cyclosporine plus nifedipine group underwent an increase in the ERPF (P < 0.01), the GFR remained unchanged, and the FF fell significantly (P < 0.0001). In this model, nifedipine completely abolished the renal arteriolar vasospasm produced by cyclosporine. That the FF fell in the cyclosporine plus nifedipine-treated animals indicates that cyclosporine has an effect which is not mediated by arteriolar vasoconstriction. This action may be at the glomerular level and is resistant to calcium channel blockade.

Early renal pathophysiology in an acute model of cyclosporine nephrotoxicity in rats.

We have recently described a rat model of acute cyclosporine nephrotoxicity characterized by rapid onset of reproducible mild to moderate renal failure. In the present studies, we have examined early pathophysiologic events and morphologic changes in this model. Following acute intraperitoneal administration of 60 mg/kg of parenteral cyclosporine, renal blood flow (RBF) fell 24% from baseline. Intraperitoneal administration of an oral cyclosporine preparation (60 mg/kg) also reduced RBF (25%), as did administration of an equivalent volume of parenteral cremophore (23%). Renal vascular resistance (RVR) increased significantly in all these groups. In contrast, intraperitoneal administration of mineral oil or olive oil oral vehicle produced no significant change in RBF (4% fall from baseline), and RVR actually decreased in these control animals. Following 2 daily doses of these agents, RBF remained significantly lower in rats given parenteral cyclosporine (5.10 mL/min vs 8.54 mL/min in cremophore rats and 7.28 mL/min in oil control rats) and renal vascular resistance remained high. Systemic blood pressure was also significantly lower in cyclosporine-treated rats at 2 days, and GFR was depressed. Morphologic studies revealed a correlation at 2 days between tubular vacuolation and renal blood flow and renal vascular resistance in cyclosporine-treated rats.

Cyclosporine nephrotoxicity in rats with an acute reduction of renal function.

The early interactive nephrotoxicity of cyclosporine was studied in rats immediately after either unilateral nephrectomy or a combination of temporary renal ischemia and contralateral nephrectomy. Cyclosporine was administered intramuscularly (IM) for 4 weeks. In comparison to vehicle-treated animals, the rats receiving cyclosporine showed neither the compensatory increase in glomerular filtration rate after unilateral nephrectomy nor the recovery of function after renal ischemia. It is concluded that cyclosporine interferes with the normal compensatory response resulting from an acute loss of renal function. This could be due to an interaction with renal hemodynamics or hormonal factors mediating the compensatory renal response.