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Rosiglitazone reverses tenofovir-induced nephrotoxicity
2008; Elsevier BV; Volume: 74; Issue: 7 Linguagem: Inglês
10.1038/ki.2008.252
ISSN1523-1755
AutoresAlexandre Braga Libório, Lúcia Andrade, Leonardo Victor Barbosa Pereira, Talita Rojas Sanches, Maria Heloísa Massola Shimizu, Antônio Carlos Seguro,
Tópico(s)Metabolism and Genetic Disorders
ResumoTenofovir disoproxil fumarate (TDF) is a first-line drug used in patients with highly active retroviral disease; however, it can cause renal failure associated with many tubular anomalies that may be due to down regulation of a variety of ion transporters. Because rosiglitazone, a peroxisome proliferator-activated receptor-γ agonist induces the expression of many of these same transporters, we tested if the nephrotoxicity can be ameliorated by its use. High doses of TDF caused severe renal failure in rats accompanied by a reduction in endothelial nitric-oxide synthase and intense renal vasoconstriction; all of which were significantly improved by rosiglitazone treatment. Low-dose TDF did not alter glomerular filtration rate but produced significant phosphaturia, proximal tubular acidosis, polyuria and a reduced urinary concentrating ability. These alterations were caused by specific downregulation of the sodium-phosphorus cotransporter, sodium/hydrogen exchanger 3 and aquaporin 2. A Fanconi's-like syndrome was ruled out as there was no proteinuria or glycosuria. Rosiglitazone reversed TDF-induced tubular nephrotoxicity, normalized urinary biochemical parameters and membrane transporter protein expression. These studies suggest that rosiglitazone treatment might be useful in patients presenting with TFV-induced nephrotoxicity especially in those with hypophosphatemia or reduced glomerular filtration rate. Tenofovir disoproxil fumarate (TDF) is a first-line drug used in patients with highly active retroviral disease; however, it can cause renal failure associated with many tubular anomalies that may be due to down regulation of a variety of ion transporters. Because rosiglitazone, a peroxisome proliferator-activated receptor-γ agonist induces the expression of many of these same transporters, we tested if the nephrotoxicity can be ameliorated by its use. High doses of TDF caused severe renal failure in rats accompanied by a reduction in endothelial nitric-oxide synthase and intense renal vasoconstriction; all of which were significantly improved by rosiglitazone treatment. Low-dose TDF did not alter glomerular filtration rate but produced significant phosphaturia, proximal tubular acidosis, polyuria and a reduced urinary concentrating ability. These alterations were caused by specific downregulation of the sodium-phosphorus cotransporter, sodium/hydrogen exchanger 3 and aquaporin 2. A Fanconi's-like syndrome was ruled out as there was no proteinuria or glycosuria. Rosiglitazone reversed TDF-induced tubular nephrotoxicity, normalized urinary biochemical parameters and membrane transporter protein expression. These studies suggest that rosiglitazone treatment might be useful in patients presenting with TFV-induced nephrotoxicity especially in those with hypophosphatemia or reduced glomerular filtration rate. Tenofovir disoproxil fumarate (TDF) is the first nucleotide reverse transcriptase inhibitor approved for the treatment of HIV.1.Fung H.B. Stone E.A. Piacenti F.J. Tenofovir disoproxil fumarate.Clin Ther. 2002; 24: 1515-1548Abstract Full Text PDF PubMed Scopus (104) Google Scholar Presenting low protein binding, TDF is freely filtered through the glomerulus without being degraded in the body. Similar to cidofovir and adefovir, TDF is eliminated through active tubular secretion via the human organic anion transporter 1.2.Kearney B.P. Flaherty J.F. Shah J. Tenofovir disoproxil fumarate: clinical pharmacology and pharmacokinetics.Clin Pharmacokinet. 2004; 43: 595-612Crossref PubMed Scopus (326) Google Scholar High intracellular concentrations of TDF in tubular cells can interfere with cell function.3.Birkus G. Hitchcock M. Cihlar T. Assessment of mitochondrial toxicity in human cells treated with tenofovir: comparison with other nucleoside reverse transcriptase inhibitors.Antimicrob Agents Chemother. 2002; 46: 716-723Crossref PubMed Scopus (374) Google Scholar Currently, TDF is considered as a first-line drug for patients who are highly active antiretroviral therapy (HAART)-naïve and is an option in many HAART combinations.4.Panel on Clinical Practices for Treatment of HIV Infection Guidelines for the use of antiretroviral agents in HIV-1-infected adults and adolescents. Department of Health and Human Services, Washington, DCMay 04, 2006http://AIDSinfo.nih.govGoogle Scholar It has recently been shown that, in such antiretroviral-naïve patients, the TDF–emtricitabine–efavirenz combination is superior to the zidovudine–lamivudine–efavirenz combination in terms of virological suppression and CD4 response.5.Gallant J.E. DeJesus E. Arribas J.R. et al.Tenofovir DF, emtricitabine, and efavirenz vs zidovudine, lamivudine, and efavirenz for HIV.N Engl J Med. 2006; 354: 251-260Crossref PubMed Scopus (780) Google Scholar In HIV-infected patients, treatment with TDF has been associated with good virological suppression and favorable clinical outcomes. However, TDF has serious side effects, especially with long-term use. Among the principal side effects associated with TDF use are hypophosphatemia,6.Breton G. Alexandre M. Duval X. et al.Tubulopathy consecutive to tenofovir-containing antiretroviral therapy in two patients infected with human immunodeficiency virus-1.Scand J Infect Dis. 2003; 36: 527-528Crossref Scopus (28) Google Scholar,7.Karras A. Lafaurie M. Furco A. et al.Tenofovir-related nephrotoxicity in human immunodeficiency virus-infected patients: three cases of renal failure, Fanconi syndrome, and nephrogenic diabetes insipidus.Clin Infect Dis. 2003; 36: 1070-1073Crossref PubMed Scopus (319) Google Scholar,8.Badiou S. De Boever C.M. Terrier N. et al.Is tenofovir involved in hypophosphatemia and decrease of tubular phosphate reabsorption in HIV-positive adults?.J Infect. 2006; 52: 335-338Abstract Full Text Full Text PDF PubMed Scopus (32) Google Scholar renal failure,9.Antoniou T. Raboud J. Chirhin S. et al.Incidence of and risk factors for tenofovir-induced nephrotoxicity: a retrospective cohort study.HIV Med. 2005; 6: 284-290Crossref PubMed Scopus (77) Google Scholar,10.El Sahly H.M. Teeter L. Zerai T. et al.Serum creatinine changes in HIV-seropositive patients receiving tenofovir.AIDS. 2006; 20: 786-787Crossref PubMed Scopus (12) Google Scholar and tubular toxicity. The tubular toxicity induced by TDF can manifest as renal tubular acidosis,11.Earle K.E. Seneviratne T. Shaker J. et al.Fanconi's syndrome in HIV+ adults: report of three cases and literature review.J Bone Miner Res. 2004; 19: 714-721Crossref PubMed Scopus (117) Google Scholar Fanconi syndrome,7.Karras A. Lafaurie M. Furco A. et al.Tenofovir-related nephrotoxicity in human immunodeficiency virus-infected patients: three cases of renal failure, Fanconi syndrome, and nephrogenic diabetes insipidus.Clin Infect Dis. 2003; 36: 1070-1073Crossref PubMed Scopus (319) Google Scholar,12.Izzedine H. Isnard-Bagnis C. Hulot J.S. et al.Renal safety of tenofovir in HIV treatment-experienced patients.AIDS. 2004; 18: 1074-1076Crossref PubMed Scopus (156) Google Scholar,13.Verhelst D. Monge M. Meynard J.L. et al.Fanconi syndrome and renal failure induced by tenofovir: a first case report.Am J Kidney Dis. 2002; 40: 1331-1333Abstract Full Text Full Text PDF PubMed Scopus (297) Google Scholar or nephrogenic diabetes insipidus (NDI).7.Karras A. Lafaurie M. Furco A. et al.Tenofovir-related nephrotoxicity in human immunodeficiency virus-infected patients: three cases of renal failure, Fanconi syndrome, and nephrogenic diabetes insipidus.Clin Infect Dis. 2003; 36: 1070-1073Crossref PubMed Scopus (319) Google Scholar,13.Verhelst D. Monge M. Meynard J.L. et al.Fanconi syndrome and renal failure induced by tenofovir: a first case report.Am J Kidney Dis. 2002; 40: 1331-1333Abstract Full Text Full Text PDF PubMed Scopus (297) Google Scholar,14.Rollot F. Nazal E.M. Chauvelot-Moachon L. et al.Tenofovir-related Fanconi syndrome with nephrogenic diabetes insipidus in a patient with acquired immunodeficiency syndrome: the role of lopinavir–ritonavir–didanosine.Clin Infect Dis. 2003; 37: E174-E176Crossref PubMed Scopus (131) Google Scholar Another significant side effect is reduced bone mineral density.11.Earle K.E. Seneviratne T. Shaker J. et al.Fanconi's syndrome in HIV+ adults: report of three cases and literature review.J Bone Miner Res. 2004; 19: 714-721Crossref PubMed Scopus (117) Google Scholar,15.Parsonage M.J. Wilkins E.G. Snowden N. et al.The development of hypophosphataemic osteomalacia with myopathy in two patients with HIV infection receiving tenofovir therapy.HIV Med. 2005; 6: 341-346Crossref PubMed Scopus (105) Google Scholar Through bone histology in animals, it has been demonstrated that TDF treatment causes osteomalacia,16.Van Rompay K.K. Brignolo L.L. Meyer D.J. et al.Biological effects of short-term or prolonged administration of 9-[2-(phosphonomethoxy) propyl]adenine (tenofovir) to newborn and infant rhesus macaques.Antimicrob Agents Chemother. 2004; 48: 1469-1487Crossref PubMed Scopus (122) Google Scholar possibly by reducing serum phosphorus levels. Although the incidence of adverse effects is low, the number of patients presenting with such side effects is likely to grow in parallel with the increasing use of TDF. The peroxisome proliferator-activated receptor-γ (PPAR-γ) is a member of the nuclear receptor superfamily of ligand-activated transcription factors. To regulate the transcription of numerous target genes, ligand-activated PPAR-γ binds to a specific DNA site known as the peroxisome proliferator response element.17.Michalik L. Wahli W. Peroxisome proliferator-activated receptors: three isotypes for a multitude of functions.Curr Opin Biotechnol. 1999; 6: 590-595Google Scholar,18.Boulanger H. Mansouri R. Gautier J.F. et al.Are peroxisome proliferator-activated receptors new therapeutic targets in diabetic and non-diabetic nephropathies?.Nephrol Dial Transplant. 2006; 21: 2696-2702Crossref PubMed Scopus (9) Google Scholar The PPAR-γ is widely distributed throughout the tissues, especially in adipose tissue. In renal tissue, PPAR-γ is expressed in the medullary collecting ducts, proximal tubules, mesangial cells, and renal microvasculature.19.Guan Y. Peroxisome proliferator-activated receptor family and its relationship to renal complications of the metabolic syndrome.J Am Soc Nephrol. 2004; 15: 2801-2815Crossref PubMed Scopus (145) Google Scholar Song et al.20.Song J. Knepper M.A. Hu X. et al.Rosiglitazone activates renal sodium and water-reabsorptive pathways and lowers blood pressure in normal rats.J Pharmacol Exp Ther. 2004; 308: 426-433Crossref PubMed Scopus (112) Google Scholar demonstrated the effects that a PPAR-γ agonist has on transporters in the renal tubular epithelium. The authors found that, in normal rats, the administration of the PPAR-γ agonist rosiglitazone (RSG) was associated with increased expression of the bumetanide-sensitive Na–K–2Cl cotransporter (NKCC2), the sodium/hydrogen exchanger 3 (NHE3), and the sodium-phosphate cotransporter subtype IIa (NaPi-IIa), as well as aquaporin 2 (AQP2). In addition, RSG administration increased the expression of endothelial nitric-oxide synthase (eNOS). In this study, we investigated the mechanisms of TDF-induced nephrotoxicity, focusing on its tubular effects. We also tested the hypothesis that RSG protects against TDF-induced renal impairment. Three sets of experiments were performed. The first was designated as the high-TDF (Hi-TDF) set. The Hi-TDF experiments involved two groups: Hi-TDF (rats fed with a diet containing 300 mg of TDF/kg of food for 30 days); and Hi-TDF+RSG (rats fed with a diet containing 300 mg of TDF/kg of food for the first 15 days and, for the subsequent 15 days, 300 mg of TDF plus 92 mg of RSG/kg of food). The second set of experiments was designated as the low-TDF (Lo-TDF) set. The Lo-TDF experiments involved two groups: Lo-TDF (rats fed with a diet containing 50 mg of TDF/kg of food for 30 days); and Lo-TDF+RSG (rats fed with a diet containing 50 mg of TDF/kg of food for the first 15 days and, for the subsequent 15 days, 50 mg of TDF plus 92 mg of RSG/kg of food). For the Hi-TDF and Lo-TDF experiments, we included a control group (rats fed with a normal diet for 30 days). The third set of experiments was designated as the RSG-only set and involved a group designated as RSG (rats fed with a diet containing 92 mg of RSG/kg of food for 15 days). For the RSG-only experiments, we included an additional control group (rats fed with a normal diet for 15 days). In all three sets of experiments, each rat was provided with 25 g of food per day, and all presented with similar ingestion (≈23 g). As can be seen in Tables 1 and 2, the TDF-induced reduction in creatinine clearance was dose dependent. In the Lo-TDF-treated rats, there were only minimal alterations in renal blood flow (RBF), which were not accompanied by any significant changes in creatinine clearance. Conversely, creatinine clearance was significantly lower in the Hi-TDF group rats than in the corresponding control rats.Table 1Renal function and hemodynamic measurements in normal (control) rats, in rats treated for 30 days with a high dose of TDF (300 mg/kg of food; Hi-TDF group), and in rats treated for 30 days with the same high dose of TDF, to which RSG (92 mg/kg of food) was added on day 15 and continued throughout (Hi-TDF+RSG group)GroupBWUOCrClMAPRBFRVRControl (n=4)272±1811.5±1.10.83±0.07117±3.86.1±0.1519.4±0.8Hi-TDF (n=5)259±1513.9±2.10.19±0.01aP<0.001 vs other groups.140±4.2bP<0.05 vs other groups.2.3±0.20c,P<0.001 vs control.dP<0.05 vs Hi-TDF+RSG.60.9±1.1aP<0.001 vs other groups.Hi-TDF+RSG (n=5)267±811.8±1.30.58±0.04eP<0.01 vs control by analysis of variance and Tukey's multiple comparison test.125±3.54.3±0.35eP<0.01 vs control by analysis of variance and Tukey's multiple comparison test.29.7±0.9eP<0.01 vs control by analysis of variance and Tukey's multiple comparison test.BW, body weight (g); CrCl, creatinine clearance (ml/min/100 g); MAP, mean arterial pressure (mm Hg); RBF, renal blood flow (ml/min); RSG, rosiglitazone; RVR, renal vascular resistance (mm Hg/ml/min); TDF, tenofovir disoproxil fumarate; UO, urine output (ml/day).Data are expressed as mean±s.e.m.a P<0.001 vs other groups.b P<0.05 vs other groups.c P<0.001 vs control.d P<0.05 vs Hi-TDF+RSG.e P<0.01 vs control by analysis of variance and Tukey's multiple comparison test. Open table in a new tab Table 2Renal function and hemodynamic measurements in normal (control) rats, in rats treated for 30 days with a low dose of TDF (50 mg/kg of food; Lo-TDF group), and in rats treated for 30 days with the same low dose of TDF, to which RSG (92 mg/kg of food) was added on day 15 and continued throughout (Lo-TDF+RSG group)GroupBWUOCrClMAPRBFRVRControl (n=4)272±1811.5±1.10.83±0.07117±3.86.1±0.1519.4±0.8Lo-TDF (n=6)274±1223.4±1.6aP<0.01 vs other groups.0.72±0.08128±3.15.0±0.19bP<0.05 vs other groups.25.6±1.3cP<0.01 vs other groups by analysis of variance and Tukey's multiple comparison test.Lo-TDF+RSG (n=5)262±68.4±1.10.74±0.01117±2.85.9±0.3519.8±0.9BW, body weight (g); CrCl, creatinine clearance (ml/min/100 g); MAP, mean arterial pressure (mm Hg); RBF, renal blood flow (ml/min); RSG, rosiglitazone; RVR, renal vascular resistance (mm Hg/ml/min); TDF, tenofovir disoproxil fumarate; UO, urine output (ml/day).Data are expressed as mean±s.e.m.a P<0.01 vs other groups.b P<0.05 vs other groups.c P<0.01 vs other groups by analysis of variance and Tukey's multiple comparison test. Open table in a new tab BW, body weight (g); CrCl, creatinine clearance (ml/min/100 g); MAP, mean arterial pressure (mm Hg); RBF, renal blood flow (ml/min); RSG, rosiglitazone; RVR, renal vascular resistance (mm Hg/ml/min); TDF, tenofovir disoproxil fumarate; UO, urine output (ml/day). Data are expressed as mean±s.e.m. BW, body weight (g); CrCl, creatinine clearance (ml/min/100 g); MAP, mean arterial pressure (mm Hg); RBF, renal blood flow (ml/min); RSG, rosiglitazone; RVR, renal vascular resistance (mm Hg/ml/min); TDF, tenofovir disoproxil fumarate; UO, urine output (ml/day). Data are expressed as mean±s.e.m. The results of clearance studies and RBF measurements in the Hi-TDF experiments are shown in Table 1. There were no differences among the three groups in terms of body weight. The rats in the Hi-TDF group presented with higher blood pressure and significantly impaired renal function. These alterations were accompanied by intense renal vasoconstriction (reduced RBF and increased renal vascular resistance). In addition, the Hi-TDF group rats presented with markedly lower eNOS expression than the corresponding control rats (Figure 1). Administration of RSG improved renal function, as evidenced by the fact that creatinine clearance was significantly greater in the Hi-TDF+RSG group than in the Hi-TDF group. There was no difference in creatinine clearance between the rats in the RSG group and those in the corresponding control group (0.58±0.03 vs 0.68±0.05 ml/min/100 g body weight). Renal vasoconstriction was lower in the Hi-TDF+RSG group than in the Hi-TDF group (Table 1). In addition, RSG inhibited the TDF-induced downregulation of eNOS expression (Figure 1). In terms of creatinine clearance, there was no significant difference between the Lo-TDF group rats and the corresponding control group rats (Table 2). In the Lo-TDF group rats, urine output was significantly greater than that observed in the control rats (Table 3). The Lo-TDF group rats also presented with markedly lower urine osmolality than did the control rats. Proximal tubular function was impaired in Lo-TDF-treated rats, as evidenced by increased urinary phosphorus excretion and a considerable reduction in serum bicarbonate. Other markers of proximal tubular dysfunction, such as glycosuria and proteinuria, were absent (data not shown). No differences were observed in urinary sodium excretion or urinary potassium excretion. In addition, we observed no differences between the Lo-TDF-treated rats and the corresponding control rats in terms of urinary calcium excretion or urinary magnesium excretion. Administration of RSG reversed the hyperphosphaturia presented by the rats in the Lo-TDF group. In Lo-TDF+RSG-treated rats, urinary phosphorus excretion returned to normal levels by the end of the experiment (Table 3). The acidosis presented by rats in the Lo-TDF group was not observed in the Lo-TDF+RSG group. The Lo-TDF group rats presented with partial NDI, by which time urine output and osmolality had normalized in the Lo-TDF+RSG-treated rats (Table 3).Table 3Functional parameters in normal (control) rats, in rats treated for 30 days with a low dose of TDF (50 mg/kg of food; Lo-TDF group), and in rats treated for 30 days with the same low dose of TDF, to which RSG (92 mg/kg of food) was added on day 15 and continued throughout (Lo-TDF+RSG group)ControlLo-TDFLo-TDF+RSGUrine output (ml/day)11.5±0.523.4±1.6aP<0.001 vs control and Lo-TDF+RSG.8.4±1.1UOsm (mOsm/kg H2O)988±101520±85b,P<0.001 vs Lo-TDF+RSG.cP<0.05 vs control.1360±138cP<0.05 vs control.UPV (μmol/day)316±33648±41aP<0.001 vs control and Lo-TDF+RSG.351±36UNaV (mEq./day)1.30±0.221.45±0.141.46±0.15UKV (mEq./day)1.16±0.211.65±0.381.48±0.18UMgV (mg/day)0.96±0.260.92±0.281.04±0.13UCaV (μg/day)354±25298±22362±32PH7.42±0.017.31±0.02aP<0.001 vs control and Lo-TDF+RSG.7.42±0.02Serum bicarbonate (mEq./l)24.6±1.218.3±0.8dP<0.05 vs control and Lo-TDF+RSG.23.7±1.0Serum anion gapeAnion gap is calculated as sodium-chloride-bicarbonate.18.3±1.817.9±2.018.5±1.6RSG, rosiglitazone; TDF, tenofovir disoproxil fumarate; UOsm, urinary osmolality; UNaV, urinary sodium; UKV, urinary potassium; UPV, urinary phosphorus; UMgV, urinary magnesium; UCaV, urinary calcium.a P<0.001 vs control and Lo-TDF+RSG.b P<0.001 vs Lo-TDF+RSG.c P<0.05 vs control.d P<0.05 vs control and Lo-TDF+RSG.e Anion gap is calculated as sodium-chloride-bicarbonate. Open table in a new tab RSG, rosiglitazone; TDF, tenofovir disoproxil fumarate; UOsm, urinary osmolality; UNaV, urinary sodium; UKV, urinary potassium; UPV, urinary phosphorus; UMgV, urinary magnesium; UCaV, urinary calcium. Interestingly, compared with the corresponding control group rats, the RSG group rats presented with an increase in body weight, as well as an increase in urinary osmolality. However, there was no difference between the two groups in terms of urine volume. Surprisingly, the RSG group rats also presented significantly greater urinary excretions than those in the corresponding control group, including urinary excretion of sodium, potassium, and phosphorus (Table 4).Table 4Functional parameters in normal (control) rats and in rats treated for 15 days with RSG (92 mg/kg of food, RSG group)BW (g)UOsm (mOsm/kg H2O)Urine output (ml/day)UNaV (mEq./day)UKV (mEq./day)UPV (μmol/day)Control group252±7410±7326±3.20.6±0.051.2±0.04524±25RSG group293±6aP<0.001 vs control.627±55bP<0.05 vs control.22±3.11.48±0.15aP<0.001 vs control.1.4±0.08bP<0.05 vs control.848±35.4aP<0.001 vs control.BW, body weight, RSG, rosiglitazone; UOsm, urinary osmolality; UNaV, urinary sodium; UKV, urinary potassium; UPV, urinary phosphorus.a P<0.001 vs control.b P<0.05 vs control. Open table in a new tab BW, body weight, RSG, rosiglitazone; UOsm, urinary osmolality; UNaV, urinary sodium; UKV, urinary potassium; UPV, urinary phosphorus. Tubular reabsorption of phosphorus is largely performed in the proximal tubules via NaPi-IIa. To investigate the cause of hyperphosphaturia in TDF-treated rats, we examined NaPi-IIa expression in the renal cortex. As shown in Figure 2a, NaPi-IIa protein expression in TDF-treated rats was significantly lower than that seen in control rats (67.7±1.4 vs 99.5±0.5%, P<0.01). Although rats treated with TDF presented with low serum bicarbonate levels and low serum pH, urine pH also remained low (Table 3), suggesting proximal tubular dysfunction. To thoroughly investigate the cause of the serum acidosis, we examined NHE3 protein expression. In comparison with control rats, those treated with TDF presented with lower NHE3 protein expression (100±1 vs 59±2.1%, P<0.01; Figure 2b), which is consistent with the proximal type of renal tubular acidosis. Figure 3 shows representative immunoblots of renal cortex homogenates probed with the antibody against the α-subunit of the epithelial sodium channel (α-EnaC). Abundance of α-ENaC was unaffected by TDF treatment (Lo-TDF: 108.3%±1.7; control: 100%, P=NS). Protein expression of NHE3 was reduced in TDF-treated rats. As no increase in urinary sodium excretion was observed in the treated rats, we attempted to determine whether distal segments of the nephron play a compensatory role by reabsorbing the excess sodium delivered. It is well known that apical NKCC2 is the major transporter for sodium reabsorption by the thick ascending limb. As indicated in Figure 4a, NKCC2 protein expression was significantly higher in the medullae of Lo-TDF group rats than in those of control rats (136±2.2 vs 99±1.0%, P<0.001). As can be seen in Figure 4b, treatment with TDF reduced AQP2 abundance significantly (67.5±2.5 vs 99±1, P<0.01). This can partially explain the reduced urinary concentrating ability. In rats receiving low-dose TDF, RSG restored NaPi-IIa expression (Figure 2a), explaining the normalization of phosphaturia observed in the Lo-TDF+RSG group. There was no significant difference between the RSG group rats and the controls in terms of NaPi-IIa expression (RSG: 89.3±2.3%). In the Lo-TDF+RSG-treated rats, NHE3 protein expression was higher than that seen in the Lo-TDF group rats (93.3±4.4 vs 59±2.1, P<0.01; Figure 2b), and NHE3 protein abundance was comparable with that observed in control rats (93.3±4.4 vs 100±1%, P=NS). The RSG group rats presented with no significant difference in serum pH or NHE3 expression in comparison with the control group rats (RSG: 108±7.3%). Figure 3 shows that α-ENaC expression was greater in the Lo-TDF+RSG-treated rats than in the Lo-TDF-treated rats (145±2.9 vs 108.3±1.7%, P<0.001). In addition, α-ENaC expression was greater in the Lo-TDF+RSG-treated rats than in control rats (145±2.9 vs 100%, P<0.001). In the RSG group, α-ENaC expression was increased in comparison with the control group (RSG: 155±2.9%, P<0.001). In addition, there was a significant difference between the RSG group and the Lo-TDF+RSG group in terms of α-ENaC expression, which was greater in the RSG group (P<0.05). Figure 4a shows that Lo-TDF+RSG rats presented with levels of NKCC2 expression comparable with those seen in Lo-TDF group rats (145±2.9 vs 136±2.2%, P=NS). Expression of NKCC2 was significantly higher in the Lo-TDF+RSG and Lo-TDF group rats than in the control rats and in the RSG rats (P<0.001). In the RSG group, NKCC2 expression did not differ significantly from that observed for the controls (RSG: 111±5.2%). Figure 4b shows that Lo-TDF+RSG rats presented greater AQP2 expression than that seen in the Lo-TDF group rats (143.3±7.2 vs 67.5±2.5%, P<0.001). In addition, AQP2 abundance was greater in Lo-TDF+RSG rats than in control rats (143.3±7.2 vs 99±1%, P<0.001). In the RSG group, AQP2 expression increased in comparison with that observed in the control group (RSG group: 165±5.4%, P<0.001). In addition, there was a significant difference between the RSG group and the Lo-TDF+RSG group in terms of AQP2 expression, which was higher in the RSG group (P<0.05). Treatment with TDF induced a broad spectrum of nephrotoxicity, including renal failure, proximal tubular injury (hyperphosphaturia and renal tubular acidosis), and reduced urinary concentrating ability. The dose-dependent deterioration in renal function was associated with renal vasoconstriction and reduced abundance of eNOS. We also demonstrated downregulation of membrane transporters (NaPi-IIa, NHE3, and AQP2). Most importantly, RSG reversed all TDF-induced renal alterations. It has been demonstrated that changes in the expression of membrane protein transporters can occur in many forms of acute kidney injury, including those caused by ischemia or nephrotoxicity.21.Andrade L. Rebouças N.A. Seguro A.C. Down-regulation of Na+ transporters and AQP2 is responsible for acyclovir-induced polyuria and hypophosphatemia.Kidney Int. 2004; 65: 175-183Abstract Full Text Full Text PDF PubMed Scopus (13) Google Scholar,22.Kwon T.H. Frøkiaer J. Han J.S. et al.Decreased abundance of major Na+ transporters in kidneys of rats with ischemia-induced acute renal failure.Am J Physiol Renal Physiol. 2000; 278: F925-F939PubMed Google Scholar However, it remains unknown whether these alterations are specific to injury caused by TDF administration or are simply characteristic of how renal tubular cells react under stress (for example, ischemia) Nevertheless, in this study, alterations of this type occurred in rats treated with tenofovir. In experimental studies involving isolated human proximal tubular cells, TDF presented with lower cytotoxicity than other nucleoside reverse transcriptase inhibitors.23.Birkus G. Hitchcock M.J. Cihlar T. Assessment of mitochondrial toxicity in human cells treated with tenofovir: comparison with other nucleoside reverse transcriptase inhibitors.Antimicrob Agents Chemother. 2002; 46: 716-723Crossref PubMed Scopus (309) Google Scholar However, there are studies showing that patients using TDF-containing HAART regimens present with renal dysfunction, whereas control HIV patients do not.24.Mauss S. Berger F. Schmutz G. Antiretroviral therapy with tenofovir is associated with mild renal dysfunction.AIDS. 2005; 19: 1993-1995Google Scholar In other reports, the incidence of renal failure associated with TDF-containing HAART regimens is no different from that associated with other HAART regimens.25.Gallant J.E. Staszewski S. Pozniak A.L. et al.Efficacy and safety of tenofovir DF vs stavudine in combination therapy in antiretroviral-naive patients: a 3-year randomized trial.JAMA. 2004; 292: 191-201Crossref PubMed Scopus (1264) Google Scholar,26.Izzedine H. Hulot J.S. Vittecoq D. et al.Long-term renal safety of tenofovir disoproxil fumarate in antiretroviral-naive HIV-1-infected patients: data from a double-blind randomized active-controlled multicentre study.Nephrol Dial Transplant. 2005; 20: 743-746Crossref PubMed Scopus (164) Google Scholar For treatment-naïve patients, as well as for treatment-experienced patients, TDF has rapidly become a favored nucleoside component of antiretroviral regimens. In this study, rats receiving high-dose TDF developed severe renal failure accompanied by intense renal vasoconstriction, neither of which was observed in those receiving the low dose. It has been suggested that TDF causes direct proximal tubular damage, leading to renal failure.27.Izzedine H. Launay-Vacher V. Deray G. Antiviral drug-induced nephrotoxicity.Am J Kidney Dis. 2005; 45: 804-817Abstract Full Text Full Text PDF PubMed Scopus (301) Google Scholar We have demonstrated herein that another mechanism contributes to TDF-induced renal failure, namely diminished production of NO. In previous studies conducted by our group in patients and in animal models, we found that indinavir also decreases NO production.28.Eira M. Araujo M. Seguro A.C. Urinary NO3 excretion and renal failure in indinavir-treated patients.Braz J Med Biol Res. 2006; 39: 1065-1070Crossref PubMed Scopus (9) Google Scholar,29.de Araújo M. Seguro A.C. Vasodilator agents protect against indinavir nephrotoxicity.Antivir Ther. 2003; 8: 295-299PubMed Google Scholar Various studies have shown that PPAR-γ agonists are efficacious in slowing the progression of glomerulosclerosis (diabetic and nondiabetic), ischemia–reperfusion injury, and cisplatin nephrotoxicity.19.Guan Y. Peroxisome proliferator-activated receptor family
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