Fenofibrate, a PPARα agonist, has renoprotective effects in mice by enhancing renal lipolysis
2011; Elsevier BV; Volume: 79; Issue: 8 Linguagem: Inglês
10.1038/ki.2010.530
ISSN1523-1755
AutoresYuki Tanaka, Shinji Kume, Shin‐ichi Araki, Keiji Isshiki, Masami Chin‐Kanasaki, Masayoshi Sakaguchi, Toshiro Sugimoto, Daisuke Koya, Masakazu Haneda, Atsunori Kashiwagi, Hiroshi Maegawa, Takashi Uzu,
Tópico(s)Alcohol Consumption and Health Effects
ResumoAs renal lipotoxicity can lead to chronic kidney disease (CKD), we examined the role of peroxisome proliferator-activated receptor (PPAR)-α, a positive regulator of renal lipolysis. Feeding mice a high-fat diet induced glomerular injury, and treating them with fenofibrate, a PPARα agonist, increased the expression of lipolytic enzymes and reduced lipid accumulation and oxidative stress in glomeruli, while inhibiting the development of albuminuria and glomerular fibrosis. In mice given an overload of free fatty acid-bound albumin to induce tubulointerstitial injury, fenofibrate attenuated the development of oxidative stress, macrophage infiltration, and fibrosis, and enhanced lipolysis in the renal interstitium. Fenofibrate inhibited palmitate-induced expression of profibrotic plasminogen activator inhibitor-1 (PAI-1) in cultured mesangial cells, and the expression of both monocyte chemoattractant protein-1 and PAI-1 in proximal tubular cells along with the overexpression of lipolytic enzymes. Thus, fenofibrate can attenuate lipotoxicity-induced glomerular and tubulointerstitial injuries, with enhancement of renal lipolysis. Whether amelioration of renal lipotoxicity by PPARα agonists will turn out to be a useful strategy against CKD will require direct testing. As renal lipotoxicity can lead to chronic kidney disease (CKD), we examined the role of peroxisome proliferator-activated receptor (PPAR)-α, a positive regulator of renal lipolysis. Feeding mice a high-fat diet induced glomerular injury, and treating them with fenofibrate, a PPARα agonist, increased the expression of lipolytic enzymes and reduced lipid accumulation and oxidative stress in glomeruli, while inhibiting the development of albuminuria and glomerular fibrosis. In mice given an overload of free fatty acid-bound albumin to induce tubulointerstitial injury, fenofibrate attenuated the development of oxidative stress, macrophage infiltration, and fibrosis, and enhanced lipolysis in the renal interstitium. Fenofibrate inhibited palmitate-induced expression of profibrotic plasminogen activator inhibitor-1 (PAI-1) in cultured mesangial cells, and the expression of both monocyte chemoattractant protein-1 and PAI-1 in proximal tubular cells along with the overexpression of lipolytic enzymes. Thus, fenofibrate can attenuate lipotoxicity-induced glomerular and tubulointerstitial injuries, with enhancement of renal lipolysis. Whether amelioration of renal lipotoxicity by PPARα agonists will turn out to be a useful strategy against CKD will require direct testing. The prevalence of chronic kidney disease (CKD) and subsequent end-stage kidney disease continues to increase worldwide, despite the application of various intensive therapy programs such as antihypertensive therapy.1.Go A.S. Chertow G.M. Fan D. et al.Chronic kidney disease and the risks of death, cardiovascular events, and hospitalization.N Engl J Med. 2004; 351: 1296-1305Crossref PubMed Scopus (8252) Google Scholar Thus, the establishment of new therapeutic strategies is urgently required. Lipotoxicity is an important pathogenic process in several types and stages of CKD.2.Abrass C.K. Cellular lipid metabolism and the role of lipids in progressive renal disease.Am J Nephrol. 2004; 24: 46-53Crossref PubMed Scopus (194) Google Scholar, 3.Bagby S.P. Obesity-initiated metabolic syndrome and the kidney: a recipe for chronic kidney disease?.J Am Soc Nephrol. 2004; 15: 2775-2791Crossref PubMed Scopus (214) Google Scholar Lipotoxicity in glomeruli is involved in the initiation of glomerular damage related to obesity and type 2 diabetes.4.Schaffer J.E. Lipotoxicity: when tissues overeat.Curr Opin Lipidol. 2003; 14: 281-287Crossref PubMed Scopus (650) Google Scholar, 5.Unger R.H. Orci L. Diseases of liporegulation: new perspective on obesity and related disorders.FASEB J. 2001; 15: 312-321Crossref PubMed Scopus (357) Google Scholar Glomerular lesion such as glomerulosclerosis and albuminuria are observed in the early stage of kidney disease associated with metabolic abnormalities. We and others have shown that intrarenal lipotoxicity resulting from enhanced renal lipogenesis and suppressed renal lipolysis contributes to the development of glomerular lesions in several animal experimental models, such as high-fat diet (HFD)-induced obese mice6.Kume S. Uzu T. Araki S. et al.Role of altered renal lipid metabolism in the development of renal injury induced by a high-fat diet.J Am Soc Nephrol. 2007; 18: 2715-2723Crossref PubMed Scopus (171) Google Scholar and type 2 diabetic db/db mice.7.Wang Z. Jiang T. Li J. et al.Regulation of renal lipid metabolism, lipid accumulation, and glomerulosclerosis in FVBdb/db mice with type 2 diabetes.Diabetes. 2005; 54: 2328-2335Crossref PubMed Scopus (213) Google Scholar Thus, therapeutic measures to combat lipolysis in glomerular cells may be beneficial in glomerular injury associated with metabolic syndrome or obesity. Another type of lipotoxicity is related to the development of tubulointerstitial lesion in proteinuric kidney disease regardless of metabolic syndrome and obesity.8.Thomas M.E. Schreiner G.F. Contribution of proteinuria to progressive renal injury: consequences of tubular uptake of fatty acid bearing albumin.Am J Nephrol. 1993; 13: 385-398Crossref PubMed Scopus (103) Google Scholar The reuptake of free fatty acid (FFA)-bound albumin, which is filtrated through the glomeruli, has been demonstrated to mediate tubulointerstitial damage in proteinuric kidney disease.9.Kamijo A. Kimura K. Sugaya T. et al.Urinary free fatty acids bound to albumin aggravate tubulointerstitial damage.Kidney Int. 2002; 62: 1628-1637Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar, 10.Thomas M.E. Harris K.P. Walls J. et al.Fatty acids exacerbate tubulointerstitial injury in protein-overload proteinuria.Am J Physiol Renal Physiol. 2002; 283: F640-F647Crossref PubMed Scopus (112) Google Scholar As the progressive nature of proteinuric kidney disease is dependent on the degree of tubulointerstitial damage,11.Risdon R.A. Sloper J.C. De Wardener H.E. Relationship between renal function and histological changes found in renal-biopsy specimens from patients with persistent glomerular nephritis.Lancet. 1968; 2: 363-366Abstract PubMed Google Scholar amelioration of tubulointerstitial lipotoxicity mediated by FFA could represent an additional therapeutic target to prevent the progression of tubulointerstitial lesion in proteinuric kidney disease. Peroxisome proliferator-activated receptor (PPAR)-α is a member of the nuclear hormone receptor superfamily of ligand-activated transcription factors, and positively regulates fatty acid β-oxidation, lipolysis, through the upregulation of expression levels of several enzymes involved in lipolysis.12.Dreyer C. Krey G. Keller H. et al.Control of the peroxisomal beta-oxidation pathway by a novel family of nuclear hormone receptors.Cell. 1992; 68: 879-887Abstract Full Text PDF PubMed Scopus (1168) Google Scholar, 13.Schoonjans K. Staels B. Auwerx J. Role of the peroxisome proliferator-activated receptor (PPAR) in mediating the effects of fibrates and fatty acids on gene expression.J Lipid Res. 1996; 37: 907-925Abstract Full Text PDF PubMed Google Scholar, 14.Aoyama T. Peters J.M. Iritani N. et al.Altered constitutive expression of fatty acid-metabolizing enzymes in mice lacking the peroxisome proliferator-activated receptor alpha (PPARalpha).J Biol Chem. 1998; 273: 5678-5684Crossref PubMed Scopus (717) Google Scholar PPARα exists in both glomerular and renal tubular cells, and regulates renal lipolysis.15.Braissant O. Foufelle F. Scotto C. et al.Differential expression of peroxisome proliferator-activated receptors (PPARs): tissue distribution of PPAR-alpha, -beta, and -gamma in the adult rat.Endocrinology. 1996; 137: 354-366Crossref PubMed Scopus (1736) Google Scholar Thus, the pharmacological activation of PPARα in the kidney may have a therapeutic potency through its lipolytic effect against lipotoxicity—related both to glomerular injury associated with metabolic abnormalities and to tubulointerstitial injury in proteinuric kidney disease. In the present study, we investigated whether fenofibrate, a PPARα agonist, attenuates glomerular injury in HFD-fed obese mice and tubulointerstitial injury in a mouse model of FFA-bound bovine serum albumin (BSA)-overload nephropathy. In addition, we performed in vitro studies using cultured mouse mesangial and proximal tubular cells to show the direct renoprotective role of PPARα agonist. The results showed that PPARα agonist attenuated lipotoxicity-mediated oxidative stress and renal injury with enhancement of renal lipolysis, suggesting its potential use in a new therapeutic strategy against the initiation and progression of CKD. At the end of the 12-week experimental period, treatment of HFD-fed obese mice with fenofibrate significantly reduced body weight, blood glucose, and plasma insulin levels, without affecting the food intake (Table 1). Furthermore, fenofibrate treatment significantly improved HFD-induced insulin resistance determined by intraperitoneal insulin tolerance test (Table 1). Fenofibrate treatment significantly decreased the plasma levels of triglyceride, although it did not affect the plasma levels of cholesterol and FFA (Table 1). No significant difference in systolic blood pressure was observed among all groups of mice (Table 1).Table 1Characteristics and results of IPITT at the end of 12-week experimental period in HFD modelLFDHFDControlFenofibrateControlFenofibrateCharacteristics Body weight (g)30.2±0.46bP<0.05 versus mice fed HFD.26.2±0.50bP<0.05 versus mice fed HFD.45.6±1.09aP<0.05 versus mice fed LFD and38.5±1.31a,b Fasting blood glucose (mg/dl)114.1±6.15bP<0.05 versus mice fed HFD.114.8±5.52bP<0.05 versus mice fed HFD.197.4±16.35aP<0.05 versus mice fed LFD and124.6±7.55bP<0.05 versus mice fed HFD. Plasma triglyceride (mg/dl)62.0±7.844.4±2.6bP<0.05 versus mice fed HFD.81.8±5.951.8±3.7bP<0.05 versus mice fed HFD. Plasma cholesterol (mg/dl)113.0±10.3bP<0.05 versus mice fed HFD.119.2±3.4bP<0.05 versus mice fed HFD.169.9±7.6aP<0.05 versus mice fed LFD and156.5±16.4aP<0.05 versus mice fed LFD and Plasma FFA (mEq/l)1.02±0.110.79±0.100.70±0.04aP<0.05 versus mice fed LFD and0.56±0.02aP<0.05 versus mice fed LFD and Hemoglobin A1c (%)3.1±0.12bP<0.05 versus mice fed HFD.3.3±0.07bP<0.05 versus mice fed HFD.3.9±0.08aP<0.05 versus mice fed LFD and3.7±0.10 Plasma insulin (ng/ml)0.32±0.16bP<0.05 versus mice fed HFD.0.18±0.06bP<0.05 versus mice fed HFD.1.47±0.38aP<0.05 versus mice fed LFD and0.48±0.12bP<0.05 versus mice fed HFD. Systolic blood pressure (mm Hg)106.4±4.994.6±7.5113.5±4.6109.1±2.3 Food intake (kcal/day)11.2±0.69bP<0.05 versus mice fed HFD.10.8±0.13bP<0.05 versus mice fed HFD.15.1±0.42aP<0.05 versus mice fed LFD and15.6±0.89aP<0.05 versus mice fed LFD and Urinary albumin excretion (μg/day)18.8±4.8bP<0.05 versus mice fed HFD.6.0±1.0bP<0.05 versus mice fed HFD.50.4±10.0aP<0.05 versus mice fed LFD and20.2±4.6bP<0.05 versus mice fed HFD.IPITT Glucose level at basal (mg/dl)142.5±9.5bP<0.05 versus mice fed HFD.127.3±6.3bP<0.05 versus mice fed HFD.261.5±31.4aP<0.05 versus mice fed LFD and118.0±10.5bP<0.05 versus mice fed HFD. Glucose level at 15 min (mg/dl)80.5±2.6bP<0.05 versus mice fed HFD.75.8±7.4bP<0.05 versus mice fed HFD.260.0±40.3aP<0.05 versus mice fed LFD and71.0±6.6bP<0.05 versus mice fed HFD. Glucose level at 30 min (mg/dl)63.8±5.0bP<0.05 versus mice fed HFD.63.5±5.7bP<0.05 versus mice fed HFD.221.3±37.3aP<0.05 versus mice fed LFD and55.8±5.4bP<0.05 versus mice fed HFD. Glucose level at 60 min (mg/dl)55.8±11.1bP<0.05 versus mice fed HFD.69.8±7.0bP<0.05 versus mice fed HFD.206.8±30.3aP<0.05 versus mice fed LFD and60.8±7.2bP<0.05 versus mice fed HFD.Abbreviations: FFA, free fatty acid; HFD, high-fat diet; IPITT, intraperitoneal insulin tolerance test; LFD, low-fat diet.Data are mean±s.e.m.; n=8 (except for food intake and IPITT) in each group and n=4 (food intake and IPITT) in each group.a P<0.05 versus mice fed LFD andb P<0.05 versus mice fed HFD. Open table in a new tab Abbreviations: FFA, free fatty acid; HFD, high-fat diet; IPITT, intraperitoneal insulin tolerance test; LFD, low-fat diet. Data are mean±s.e.m.; n=8 (except for food intake and IPITT) in each group and n=4 (food intake and IPITT) in each group. Next, we examined the effect of fenofibrate on HFD-induced renal injury. Fenofibrate treatment significantly attenuated the development of albuminuria in HFD-fed mice (Table 1). Histologically, fenofibrate treatment significantly attenuated HFD-induced increases of mesangial matrix area (Figure 1a and b) and glomerular volume (Figure 1a and c). Fenofibrate treatment also ameliorated glomerular fibrosis determined by immunostaining for fibronectin (Figure 1d and e), type I collagen (Figure 1f and g), and type IV collagen (Figure 1h and i). Furthermore, fenofibrate treatment significantly inhibited HFD-induced increases of mRNA expression levels of fibronectin and plasminogen activator inhibitor-1 (PAI-1) in the renal cortex, both of which are indices of renal fibrosis (Table 2). Neither HFD feeding nor fenofibrate treatment influenced PPARα gene expression in the renal cortex (Table 2).Table 2The mRNA expression levels in the renal cortex at the end of 12-week experimental period in HFD modelLFDHFDControlFenofibrateControlFenofibrateFibrosis Fibronectin0.95±0.12b1.36±0.131.86±0.33a0.98±0.14b PAI-11.43±0.19b0.88±0.11b2.89±0.39a1.00±0.16bLipolysis PPARα0.96±0.111.82±0.341.12±0.081.74±0.15 ACO1.11±0.181.41±0.120.58±0.051.47±0.32b MCAD0.88±0.101.54±0.12a,b0.58±0.031.39±0.20b CPT-11.02±0.111.24±0.070.87±0.071.29±0.10bLipogenesis ACCα1.29±0.141.71±0.231.79±0.211.65±0.10 PPARγ0.72±0.090.84±0.080.88±0.060.70±0.11Anti-oxidant Catalase0.91±0.080.92±0.050.84±0.061.26±0.09a,b Cu/Zn-SOD0.92±0.061.06±0.080.80±0.111.89±0.20a,b Mn-SOD0.86±0.041.19±0.131.04±0.061.54±0.12a,bAbbreviations: ACC, acetyl-CoA carboxylase; ACO, acyl-CoA oxidase; CPT-1, carnitine palmitoyltransferase-1; HFD, high-fat diet; LFD, low-fat diet; MCAD, medium-chain acyl-CoA dehydrogenase; PAI-1, plasminogen activator inhibitor-1; PPAR, peroxisome proliferator-activated receptor; SOD, superoxide dismutase.Data are mean±s.e.m.; n=6 in each group.aP<0.05 versus mice fed LFD and bP<0.05 versus mice fed HFD. Open table in a new tab Abbreviations: ACC, acetyl-CoA carboxylase; ACO, acyl-CoA oxidase; CPT-1, carnitine palmitoyltransferase-1; HFD, high-fat diet; LFD, low-fat diet; MCAD, medium-chain acyl-CoA dehydrogenase; PAI-1, plasminogen activator inhibitor-1; PPAR, peroxisome proliferator-activated receptor; SOD, superoxide dismutase. Data are mean±s.e.m.; n=6 in each group. aP<0.05 versus mice fed LFD and bP<0.05 versus mice fed HFD. Oil-red O staining showed that fenofibrate treatment reduced HFD-induced neutral lipid accumulation mainly in the glomeruli (Figure 2a). Consistent with this result, fenofibrate significantly attenuated HFD-induced increase in renal triglyceride content (Figure 2b). Immunohistochemical analysis of 4-hydroxynonenal-modified proteins, a marker of lipid peroxidation, showed that fenofibrate treatment reduced 4-hydroxynonenal accumulation in glomeruli (Figure 2c). Similarly, in the renal cortex of HFD-fed mice, fenofibrate treatment significantly reduced the content of malondialdehyde, which reflects the degree of lipid peroxidation (Figure 2d). In addition, fenofibrate treatment significantly increased the expression levels of lipolytic genes, such as acyl-CoA oxidase (ACO), medium-chain acyl-CoA dehydrogenase, and carnitine palmitoyltransferase-1 (CPT-1), in the kidney of HFD-fed mice (Table 2). These changes in the expression levels of ACO and CPT-1 were confirmed by western blot analysis (Figure 2e, f and g). Sterol regulatory element-binding transcription factor-1c (SREBP-1c) is a transcriptional factor regulating gene expression of lipogenic enzymes such as acetyl-CoA carboxylase. Protein expression of the active form of SREBP-1c tended to increase in the kidney of HFD-fed mice, which was not affected by fenofibrate treatment (Figure 2e and h). Accordingly, renal protein and mRNA expression of acetyl-CoA carboxylase-α in four groups of mice showed a similar tendency to the expression level of SREBP-1c (Figure 2, Table 2, Table 2, Table 2). No significant difference in renal expression of another transcriptional regulator of lipogenesis, PPARγ, was observed in all groups (). Furthermore, fenofibrate significantly increased the mRNA expression levels of antioxidant genes, such as catalase, Cu/Zn-superoxide dismutase, and Mn-superoxide dismutase, in the renal cortex of HFD-fed mice (). Taken together, fenofibrate seems to attenuate albuminuria and glomerular fibrosis in HFD-fed obese mice, accompanied by enhancement of renal lipolysis and antioxidant defenses. We next investigated the mRNA expression of PAI-1 in cultured mesangial cells stimulated by palmitate, a saturated fatty acid, to determine the direct protective effect of fenofibrate on lipotoxicity-mediated glomerular injury. In mesangial cells, palmitate significantly increased the mRNA expression level of PAI-1 (Figure 3a). Pretreatment with fenofibrate significantly attenuated palmitate-induced overexpression of PAI-1 in a dose-dependent manner (Figure 3a). In addition, pretreatment with fenofibrate significantly increased the mRNA expression levels of ACO and CPT-1 in a dose-dependent manner, and tended to increase the mRNA expression level of medium-chain acyl-CoA dehydrogenase in palmitate-stimulated mesangial cells (Figure 3b–d). Furthermore, pretreatment with fenofibrate significantly reduced the production of reactive oxygen species (ROS) in palmitate-stimulated mesangial cells (Figure 3e). To determine whether PPARα agonist has a protective role against tubulointerstitial lesion in proteinuric kidney disease, we examined the effect of fenofibrate on FFA-bound BSA-overload nephropathy model. Neither BSA overload nor fenofibrate treatment influenced the systemic parameters at the end of the experiment (Table 3). Fenofibrate treatment tended to reduce proteinuria in FFA-bound BSA-overloaded mice, although the effect was not significant (Table 3). Morphological analysis of the kidneys of FFA-bound BSA-overloaded mice showed tubular dilatation, tubular vacuolation, and detachment of proximal tubular cells from the tubular basement membrane and interstitial edema (Figure 4a and b). Furthermore, the deposition of fibronectin (Figure 4c and d), type I collagen (Figure 4e and f) and IV collagen (Figure 4g and h), and infiltration of macrophages in the interstitium (Figure 4i and j) were significantly increased in the FFA-bound BSA-overloaded mice. Fenofibrate treatment significantly attenuated these histological abnormalities in the FFA-bound BSA-overloaded mice (Figure 4a–j). Compatible with these morphological changes, FFA-bound BSA-overload-induced increases in the mRNA expression levels of fibronectin, PAI-1, and monocyte chemoattractant protein-1 (MCP-1), a marker of inflammation, in the kidney were significantly inhibited by fenofibrate treatment (Table 4). The renal mRNA expression level of PPARα was significantly decreased by FFA-bound BSA-overload, which was not affected by fenofibrate treatment (Table 4).Table 3Characteristics at the end of experimental period in mice with FFA-bound BSA-overload modelPBSFFA-bound BSAControlFenofibrateControlFenofibrateBody weight (g)22.9±0.5222.5±0.4223.7±0.2523.4±0.58Blood glucose (mg/dl)159.2±9.0142.0±11.3141.0±8.1137.5±11.3Plasma insulin (ng/ml)0.96±0.380.92±0.391.11±0.281.03±0.34Urinary protein excretion (mg/day)3.86±0.672.95±0.326.92±1.513.79±0.40Abbreviations: BSA, bovine serum albumin; FFA, free fatty acid; PBS, phosphate-buffered saline.Data are mean±s.e.m.; n=5 in each group. Open table in a new tab Table 4The mRNA expression levels in the kidney at the end of experimental period in mice with FFA-bound BSA-overload modelPBSFFA-bound BSAControlFenofibrateControlFenofibrateFibrosis and inflammation Fibronectin1.04±0.18b0.71±0.03b1.62±0.13a0.88±0.07b PAI-10.61±0.11b0.72±0.20b1.53±0.13a0.71±0.07b MCP-10.55±0.13b0.31±0.03b1.50±0.24a0.62±0.12bLipolysis PPARα1.61±0.261.85±0.10b0.87±0.10a1.14±0.171 ACO1.20±0.161.71±0.211.06±0.153.06±0.57a,b MCAD1.16±0.161.25±0.060.85±0.091.73±0.16a,b CPT-11.35±0.141.32±0.130.96±0.182.01±0.30bLipogenesis ACCα0.75±0.130.66±0.071.27±0.240.87±0.14 PPARγ0.72±0.090.84±0.080.88±0.060.70±0.11Anti-oxidant Catalase1.22±0.130.97±0.030.82±0.04a0.76±0.06a Cu/Zn-SOD1.00±0.021.27±0.241.35±0.311.01±0.10 Mn-SOD0.99±0.110.99±0.081.01±0.161.00±0.13Abbreviations: ACC, acetyl-CoA carboxylase; ACO, acyl-CoA oxidase; BSA, bovine serum albumin; CPT-1, carnitine palmitoyltransferase-1; FFA, free fatty acid; MCP-1, monocyte chemoattractant protein-1; MCAD, medium-chain acyl-CoA dehydrogenase; PAI-1, plasminogen activator inhibitor-1; PBS, phosphate-buffered saline; PPAR, peroxisome proliferator-activated receptor; SOD, superoxide dismutase.Data are mean±s.e.m.; n=6 in each group.aP<0.05 versus PBS and bP<0.05 versus FFA-bound BSA. Open table in a new tab Abbreviations: BSA, bovine serum albumin; FFA, free fatty acid; PBS, phosphate-buffered saline. Data are mean±s.e.m.; n=5 in each group. Abbreviations: ACC, acetyl-CoA carboxylase; ACO, acyl-CoA oxidase; BSA, bovine serum albumin; CPT-1, carnitine palmitoyltransferase-1; FFA, free fatty acid; MCP-1, monocyte chemoattractant protein-1; MCAD, medium-chain acyl-CoA dehydrogenase; PAI-1, plasminogen activator inhibitor-1; PBS, phosphate-buffered saline; PPAR, peroxisome proliferator-activated receptor; SOD, superoxide dismutase. Data are mean±s.e.m.; n=6 in each group. aP<0.05 versus PBS and bP<0.05 versus FFA-bound BSA. Immunohistochemical analysis showed that fenofibrate treatment attenuated the FFA-bound BSA-overload-induced increase in accumulation of 4-hydroxynonenal-modified proteins in the renal interstitium (Figure 5a). Similarly, increased renal malondialdehyde contents in FFA-bound BSA-overloaded mice were significantly reduced by fenofibrate treatment (Figure 5b). Fenofibrate treatment increased the mRNA expression levels of lipolytic enzymes such as ACO, medium-chain acyl-CoA dehydrogenase, and CPT-1 in the kidneys of both phosphate-buffered saline (PBS)-injected and FFA-bound BSA-overloaded mice (Table 4). These changes in the expression levels of ACO and CPT-1 were confirmed by western blot analysis (Figure 5c–e). As for the expression of renal lipogenic enzymes, significant changes in protein and mRNA expression of lipogenic genes, such as SREBP-1c, acetyl-CoA carboxylase-α and PPARγ, were not observed in the kidneys of all groups (Figure 5, Table 4, Table 4). Furthermore, fenofibrate treatment did not affect the mRNA expression levels of antioxidant genes in FFA-bound BSA-overloaded mice (). These results indicate that fenofibrate attenuates tubulointerstitial fibrosis and inflammation, with enhancement of renal lipolysis in the FFA-bound BSA-overloaded mice. To examine the direct renoprotective effect of fenofibrate on tubulointerstitial inflammation and fibrosis, we examined the mRNA expression levels of MCP-1 and PAI-1 in palmitate-stimulated cultured murine proximal tubular cells. Palmitate stimulation significantly increased the mRNA expression levels of MCP-1 and PAI-1 (Figure 6a and b). Pretreatment of fenofibrate significantly attenuated the increased mRNA expression levels of MCP-1 and PAI-1 (Figure 6a and b). Similar to the results for mesangial cells, pretreatment with fenofibrate significantly increased the mRNA expression levels of ACO, medium-chain acyl-CoA dehydrogenase, and CPT-1 in palmitate-stimulated proximal tubular cells (Figure 6c–e). Furthermore, pretreatment with fenofibrate significantly reduced ROS production in palmitate-stimulated proximal tubular cells (Figure 6f). The present study showed that fenofibrate, a PPARα agonist, attenuated glomerular injury in HFD-fed obese mice and tubulointerstitial injury in FFA-bound BSA-overload nephropathy mice. These renoprotective effects of fenofibrate were accompanied by enhancement of renal lipolysis, which consequently prevented lipid accumulation, oxidative stress, fibrosis, and inflammation. These results highlight the importance of PPARα-mediated enhancement of renal lipolysis as novel therapeutic strategy for prevention of both initiation and progression of CKD. We have previously reported that mice with HFD-induced obesity exhibit renal lesions similar to CKD in patients with metabolic syndrome.6.Kume S. Uzu T. Araki S. et al.Role of altered renal lipid metabolism in the development of renal injury induced by a high-fat diet.J Am Soc Nephrol. 2007; 18: 2715-2723Crossref PubMed Scopus (171) Google Scholar, 16.Deji N. Kume S. Araki S. et al.Structural and functional changes in the kidneys of high-fat diet-induced obese mice.Am J Physiol Renal Physiol. 2009; 296: F118-F126Crossref PubMed Scopus (172) Google Scholar The present study provided the first evidence for the renobeneficial effects of fenofibrate against albuminuria and glomerular lesions in HFD-fed mice. PPARα is expressed in several organs, such as liver and skeletal muscle. PPARα agonists induce lipolysis in these metabolic organs and subsequently attenuate insulin resistance in diet-induced obese model17.Nagai Y. Nishio Y. Nakamura T. et al.Amelioration of high fructose-induced metabolic derangements by activation of PPARalpha.Am J Physiol Endocrinol Metab. 2002; 282: E1180-E1190Crossref PubMed Scopus (164) and type 2 diabetic db/db mice,18.Koh E. Kim M. Park J. et al.Peroxisome proliferator-activated receptor (PPAR)-alpha activation prevents diabetes in OLETF rats: comparison with PPAR-gamma activation.Diabetes. 2003; 52: 2331-2337Crossref PubMed Scopus (131) Google Scholar which was confirmed in our study. The present study showed for the first time that fenofibrate could enhance renal lipolysis as well as these metabolic organs. Furthermore, recent reports showed that fenofibrate attenuated renal injuries in spontaneously hypertensive rats fed a HFD19.Shin S. Lim J. Chung S. et al.Peroxisome proliferator-activated receptor-alpha activator fenofibrate prevents high-fat diet-induced renal lipotoxicity in spontaneously hypertensive rats.Hypertens Res. 2009; 32: 835-845Crossref PubMed Scopus (52) Google Scholar and type 2 diabetic db/db mice,20.Park C. Zhang Y. Zhang X. et al.PPARalpha agonist fenofibrate improves diabetic nephropathy in db/db mice.Kidney Int. 2006; 69: 1511-1517Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar although the mechanisms of the renoprotective effects of fenofibrate in these models were not determined. As renal lipotoxicity is involved in the pathogenesis of renal injury in these models,7.Wang Z. Jiang T. Li J. et al.Regulation of renal lipid metabolism, lipid accumulation, and glomerulosclerosis in FVBdb/db mice with type 2 diabetes.Diabetes. 2005; 54: 2328-2335Crossref PubMed Scopus (213) Google Scholar, 19.Shin S. Lim J. Chung S. et al.Peroxisome proliferator-activated receptor-alpha activator fenofibrate prevents high-fat diet-induced renal lipotoxicity in spontaneously hypertensive rats.Hypertens Res. 2009; 32: 835-845Crossref PubMed Scopus (52) Google Scholar and based on our results, amelioration of intrarenal lipotoxicity with enhancement of renal lipolysis could explain the mechanism underlying fenofibrate-mediated renoprotective effects in these models. In proteinuric kidney disease regardless of the metabolic disorder, tubulointerstitial damage, including fibrosis, proximal tubular cell damage, and inflammation, is closely associated with renal prognosis.21.Perico N. Codreanu I. Schieppati A. et al.Pathophysiology of disease progression in proteinuric nephropathies.Kidney Int Suppl. 2005; 67: S79-S82Abstract Full Text Full Text PDF Google Scholar The increased reabsorption of FFA-bound albumin by proximal tubular cells has been confirmed as the pathogenic process responsible for tubulointerstitial damage in a FFA-bound BSA-overload nephropathy mouse model.9.Kamijo A. Kimura K. Sugaya T. et al.Urinary free fatty acids bound to albumin aggravate tubulointerstitial damage.Kidney Int. 2002; 62: 1628-1637Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar, 10.Thomas M.E. Harris K.P. Walls J. et al.Fatty acids exacerbate tubulointerstitial injury in protein-overload proteinuria.Am J Physiol Renal Physiol. 2002; 283: F640-F647Crossref PubMed Scopus (112) Google Scholar FFA-bound BSA overload resulted in severer tubular damage in PPARα-knockout22.Kamijo Y. Hora K. Kono K. et al.PPARalpha protects proximal tubular cells from acute fatty acid toxicity.J Am Soc Nephrol. 2007; 18: 3089-3100Crossref PubMed Scopus (57) Google Scholar and interleukin-18 receptor-knockout mice.23.Sugiyama M. Kinoshita K. Kishimoto K. et al.Deletion of IL-18 receptor ameliorates renal injury in bovine serum albumin-induced glomerulonephritis.Clin Immunol. 2008; 128: 103-108Crossref PubMed Scopus (13) Google Scholar However, no study has so far reported an effective therapeutic approach against this type of renal injury. We showed here that treatment with PPARα agonist, fenofibrate, improved tubulointerstitial injury, with enhancement of lipolysis and reduction of fibrosis, inflammation, and oxidative stress, suggesting that activation of endogenous level of PPARα seems to have a protective role against tubulointerstitial injury in proteinuric kidney disease by mediating renal lipid homeostasis. Overexpression of lipogenic enzymes24.Ishigaki N. Yamamoto T. Shimizu Y. et al.Involvement of glomerular SREBP-1c in diabetic nephropathy.
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