A growing body of evidence suggests that chronic kidney disease is a significant risk for cardiovascular events and stroke regardless of traditional risk factors. The aim of this study was to examine the effects of peroxisome proliferator-activated receptor (PPAR) agonists on the tissue damage affecting salt-loaded spontaneously hypertensive stroke-prone rats ( SHRSPs), an animal model that develops a complex pathology characterized by systemic inflammation, hypertension, and proteinuria and leads to end-organ injury (initially renal and subsequently cerebral). Compared with the PPARγ agonist rosiglitazone, the PPARα ligands fenofibrate and clofibrate significantly increased survival (p < 0.001) by delaying the occurrence of brain lesions monitored by magnetic resonance imaging (p < 0.001) and delaying increased proteinuria (p < 0.001). Fenofibrate completely prevented the renal disorder characterized by severe vascular lesions, tubular damage, and glomerular sclerosis, reduced the number of ED-1-positive cells and collagen accumulation, and decreased the renal expression of interleukin-1β, transforming growth factor β, and monocyte chemoattractant protein 1. It also prevented the plasma and urine accumulation of acute-phase and oxidized proteins, suggesting that the protection induced by PPARα agonists was at least partially caused by their anti-inflammatory and antioxidative properties. The results of this study demonstrate that PPAR agonism has beneficial effects on spontaneous brain and renal damage in SHRSPs by inhibiting systemic inflammation and oxidative stress, and they support carrying out future studies aimed at evaluating the effect of PPARα agonists on proteinuria and clinical outcomes in hypertensive patients with renal disease at increased risk of stroke.
High blood pressure and kidney disease have been associated with an increased incidence of stroke. Epidemiological studies report a high stroke risk in subjects with kidney disease even after adjusting for traditional risk factors (Townsend, 2008), suggesting that the pathophysiology of cerebrovascular diseases and renal dysfunction involve different phenomena arising from a common vascular origin (Ito et al., 2009).
Albuminuria and, even more, microalbuminuria may therefore be early signs of vascular damage, reflecting a generalized endothelial dysfunction that occurs in the kidney and cerebral vessels. Emerging data indicate that antiproteinuric pharmacological agents may not only assure a better renal prognosis, but also better cardiovascular and cerebrovascular outcomes (Park et al., 2003).
This view is supported by our own data concerning salt-loaded spontaneously hypertensive stroke-prone rats ( SHRSPs), a unique animal model of spontaneously developing progressive systemic inflammation, hypertension, proteinuria, histological lesions in the renal vasculature and parenchyma, and endothelial dysfunction contributing to the onset of brain injury (Blezer et al., 1998; Sironi et al., 2005), in which we have shown that renoprotective compounds also delay the appearance of brain damage (Sironi et al., 2004; Gianella et al., 2007; Gelosa et al., 2009).
In this context, one successful strategy for inducing neuroprotection in stroke could be to modulate multiple pathophysiological pathways simultaneously by using a combination of drugs or, even better, one pharmacological agent with pleiotropic effects. There is some evidence that drugs acting on peroxisome proliferator-activated receptors (PPARs), a family of nuclear transcriptional factor receptors, can induce such pleiotropic effects as they simultaneously regulate several genes. The three isoforms/subtypes of PPARs (α, β/δ, and γ) are involved in a number of physiological processes, including the regulation of lipoproteins, lipid metabolism, and glucose homeostasis.
In addition to their well known lipid-lowering activity, PPARα activators have pleiotropic properties that lead to anti-inflammatory and antioxidative effects in vivo (Besson et al., 2005). Emerging data suggest that PPARγ also down-regulates inflammatory processes at both peripheral and cerebral levels (Delerive et al., 2001; Bordet et al., 2006).
These findings prompted us to examine the effect of PPAR ligands in SHRSPs, an animal model of a complex pathology that is characterized by systemic inflammation, hypertension, and proteinuria and leads to end-organ damage at renal and, subsequently, cerebral levels.
Materials and Methods
Animals and Treatments.
Male SHRSPs aged 4 to 5 weeks were obtained from Charles River Italia (Calco, Italy). The procedures involving the animals and their care were carried out at the University of Milan's Department of Pharmacological Sciences in accordance with the institution's guidelines, which comply with Italian and international rules and policies. Baseline measurements were made in all of the rats when they were 6 weeks old, and all of the animals were then switched (day 0) to a specific permissive diet that was low in potassium and protein and high in sodium (Japanese permissive diet; Laboratorio Dr. Piccioni, Gessate, Italy; 18.7% protein, 0.63% potassium, 0.37% sodium) accompanied by 1% NaCl in drinking water. At the same time, they were randomly divided into the following groups for oral gavage treatment: group 1 (n = 14) received 0.5% sodium carboxymethylcellulose (vehicle); group 2 (n = 14) received 150 mg/kg/day fenofibrate; and group 3 (n = 6) received 30 mg/kg/day rosiglitazone. Two other groups received clofibrate dissolved in corn oil (300 mg/kg/day: n = 6) or the vehicle alone (n = 6), and one group continued a standard diet (normal diet group: n = 4), the experimental situation in which SHRSPs do not develop any histological or biochemical alterations.
To allow the amount of the administered drugs to be adjusted to changes in body weight, drug concentrations were recalculated weekly. Fenofibrate and clofibrate were purchased from Sigma-Aldrich (St. Louis, MO), and rosiglitazone was from Enzo Life Sciences, Inc. (Plymouth Meeting, PA). The drug doses were chosen on the basis of previous rat studies (Jiang et al., 2004; Morgan et al., 2006).
Once a week, all of the rats were weighed and then housed individually in metabolic cages for 24 h to measure their food and liquid intake, and their urine was collected. Systolic arterial blood pressure was measured weekly in conscious rats by means of tail-cuff plethysmography (PB Recorder 8006; Ugo Basile, Comerio, Italy). During each recording session, blood pressure was measured three times in each animal by at least two expert operators, who were blinded to the experimental groups. Twenty-four-hour urinary proteins were measured according to Bradford, 1976 , with bovine albumin being used as a standard. Proteinuria (protein levels of ≥40 mg/day) predicts the appearance of brain abnormalities in SHRSPs (Blezer et al., 1998; Guerrini et al., 2002) and was used to schedule the frequency of magnetic resonance imaging (MRI) every week until 24-h proteinuria exceeded 40 mg/day, and then every other day until brain damage was observed.
After 6 weeks of dietary treatment, and after collecting blood and 24-h urine samples, four animals from group 1 (vehicle-treated), group 2 (fenofibrate 150 mg/kg/day), and the normal diet group were randomly euthanized, and their brains and kidneys were harvested for immunohistochemical and molecular biology analyses. In the remaining animals, the treatments were continued for up to 98 days.
The behavior of all of the considered parameters (i.e., the development of proteinuria, body weight changes, the time to the development of brain abnormalities, and the biochemical parameters) was identical in the rats receiving 0.5% sodium carboxymethylcellulose or corn oil (the vehicles, respectively, used for fenofibrate/rosiglitazone or clofibrate), and so they will hereafter be considered a single group (also for the purposes of comparing measurements) and referred to as the “vehicle” group.
MRI Evaluation of Brain Damage.
The rats were anesthetized with 2% isofluorane in 70% N2/30% O2 and placed inside a Bruker Avance II 4.7T spectrometer (Bruker BioSpin, Ettlinger, Germany) with a micro-imaging accessory. After a sagittal scout image, 16 contiguous 1-mm-thick slices were analyzed caudally to the olfactory bulb by using a field of view of 4 × 3 cm2. A turbo spin echo T2-weighted sequence was used with 16 echoes per excitation and a 10-ms interecho time, 80-ms equivalent echo time, and 3-s repetition time. The images consisted of 128 × 96 points and had a spatial resolution of 0.312 mm/pixel. Sixteen images were averaged over 3 min, 30 s. The occurrence of lesions was identified as the presence of areas of high signal intensity in the T2-weighted images.
Urine Analysis by Means of One-Dimensional Gel Electrophoresis.
One-dimensional electrophoresis (1-DE) analyses of the weekly urine samples were run in the presence of SDS without sample reduction in a discontinuous buffer system on polyacrylamide gradients of 4 to 12% T; the sample load was 50 μg of urine proteins.
The gels were stained with Colloidal blue before densitometric acquisition (GS800; Bio-Rad Laboratories, Milan, Italy). In brief, the gels were fixed with a fixing solution containing 40% methanol and 10% acetic acid and stained overnight with a solution containing 0.12% Coomassie brilliant blue G250, 8% (NH4)2SO4, 1.6% phosphoric acid, and 20% methanol. They were then destained by using a solution containing 25% methanol. Densitometry was performed by using Quantity One version 4.5.2 (Bio-Rad Laboratories) after background subtraction by evaluating the percentages of low molecular weight and high molecular weight protein density in relation to total protein density.
Analysis of mRNA for MCP-1, TGF-β1, and IL-1β.
Total RNA was prepared by guanidium thiocyanate denaturation from the frozen kidneys collected from the vehicle- and fenofibrate (150 mg/kg/day)-treated rats sacrificed after 6 weeks of dietary treatment (n = 4 in each group) and from four animals in the normal diet group fed a standard diet during the same time period. The expression of monocyte chemoattractant protein-1 (MCP-1), transforming growth factor-β1 (TGF-β1), and interleukin-1β (IL-1β) was measured by semiquantitative RT-PCR (Sironi et al., 2004), with GAPDH being amplified as a standard. The RT-PCR products were separated on 1.5% agarose gel and visualized by ethidium bromide staining. The intensity of each band was quantified by using National Institutes of Health (Bethesda, MD) Image software and expressed in arbitrary units (A.U.). The densities of the MCP-1, TGF-β1, and IL-1β bands were normalized by using the corresponding GAPDH signal.
Histology and Immunohistochemistry.
As described above, after 6 weeks of dietary treatment, four animals treated with vehicle and four treated with fenofibrate (150 mg/kg/day) were euthanized. After being fixed in Carnoy reagent (Merck, Darmstadt, Germany) and embedded in paraplast (Sigma-Aldrich), renal sections of 5 μm were cut and used for hematoxylin/eosin staining and immunohistochemical studies as described previously (Gianella et al., 2007; Gelosa et al., 2009). The presence of inflammatory infiltrates was visualized by using an anti-ED-1 primary antibody (1:100; Serotec, Oxford, UK), and the presence of collagen was visualized by staining with Sirius red F3BA (0.5% in saturated aqueous picric acid) (Sigma-Aldrich).
Advanced Oxidation Protein Product Assay.
The determination of urinary AOPP after 6 weeks of dietary treatment was based on spectrophotometric detection according to Witko-Sarsat et al. (1996); 200 μl of plasma diluted 1:5 in phosphate-buffered saline was placed in each well of a 96-well microtiter plate and, immediately after the addition of 10 μl of 1.16 M potassium iodide and 20 μl of acetic acid, absorbance was measured at 340 nm with a Mithras LP940 fluorescence spectrometer (Berthold Technologies Italia, Milan, Italy).
Quantification of Protein Carbonyl Groups.
Protein carbonyls were measured in the plasma samples and brain homogenates of animals sacrificed after 6 weeks of dietary treatment (Banfi et al., 2008) by using a Zentech PC test ELISA in accordance with the manufacturer's instructions (Biocell, Papatoetoe, Auckland, New Zealand). The plasma was collected in a citrate solution (0.129 M) and prepared by centrifugation at 1500g for 10 min at 4°C. The tissues were washed in cold phosphate-buffered saline and homogenized in ice-cold buffer containing 10 mM Tris, 10 mM EDTA, 1 mM EGTA, and protease inhibitors (0.1 mM phenylmethylsulfonyl fluoride, 1 mM benzamidine, 2 μg/ml leupeptin, and 1 mM sodium orthovanadate; all from Sigma-Aldrich), and the samples were briefly sonicated and centrifuged at 18,000g for 5 min at 4°C. In brief, the plasma samples and brain homogenates were reacted with 2,4-dinitrophenylhydrazine (DNPH) before probing with an antibody against protein-conjugated DNPH, and the absorbances were read by using a 450-nm filter on a Mithras LP940 fluorescence spectrometer (Berthold Technologies Italia). The standard curve of oxidized albumin and the samples were assayed in triplicate; the intraassay and interassay coefficients of variation were less than 2.1 and 3.2%, respectively.
Western Blotting of Carbonylated Proteins.
Protein carbonyl immunoblotting was carried out on cerebral cortex, and urine was collected after 6 weeks of treatment from non–salt-loaded rats (normal diet group; n = 4), vehicle-treated salt-loaded rats (n = 4), and fenofibrate (150 mg/kg/day)-treated salt-loaded rats (n = 4). The brain homogenate supernatant (50 μg) and urine proteins (50 μg) were then dissolved in 24 μl of 6% SDS and mixed with 8 μl of 20 mM DNPH in 20% trifluoroacetic acid. Derivatization was allowed for 15 to 30 min at room temperature, after which 32 μl of sample buffer (2 M Tris buffer containing 30% glycerol and 1 M β-mercaptoethanol) was added. The proteins were then separated on 12% SDS polyacrylamide gel and transferred to nitrocellulose membranes (Banfi et al., 2003), which were incubated with primary biotinylated antibodies directed against DNP adducts (1:5000; Promega, Milan, Italy). The blots were subsequently incubated with horseradish peroxidase-conjugated avidin (1:1000, Bio-Rad Laboratories), and the bands were visualized by enhanced chemiluminescence (ECL System; GE Healthcare, Little Chalfont, Buckinghamshire, UK). The densitometric analysis were performed with Quantity One version 4.5.2 (Bio-Rad Laboratories).
Between-group differences were computed by analysis of variance followed by an appropriate post hoc test; the between-group differences in proteinuria and low molecular weight and high molecular weight protein density were computed by analysis of variance for repeated measurements over time followed by Tukey's post hoc test. An unpaired t test was used to compare the normal diet and vehicle-treated group data. The results are expressed as mean values ± S.D.; p < 0.05 was considered statistically significant.
The salt-loaded SHRSPs treated with vehicle showed a progressive increase in 24-h proteinuria, which rose sharply after 4 to 5 weeks of permissive diet and reached 300 mg/day after 7 weeks (Fig. 1 A). Chronic treatment with rosiglitazone (30 mg/kg/day) had no effect, whereas fenofibrate (150 mg/kg/day) and clofibrate (300 mg/kg/day) completely prevented any increase in proteinuria (Fig. 1A).
The SHRSPs exposed to salt loading developed severe hypertension, which was not affected by any of the drugs used in the study (data not shown). Body weight increased similarly in all groups, but decreased 2 weeks before the appearance of brain abnormalities in the vehicle- and rosiglitazone-treated rats (Fig. 1B).
Appearance of Brain Damage and Survival.
The vehicle-treated SHRSPs developed brain abnormalities detected by MRI during 7 weeks of the permissive diet (Fig. 1C). Treatment with rosiglitazone failed to delay the occurrence of brain damage, whereas the animals treated with clofibrate or fenofibrate remained free of MRI-detectable lesions for 14 weeks of dietary treatment, after which the animals were euthanized (Fig. 1C).
The difference in survival between the groups treated with vehicle or rosiglitazone and those treated with fenofibrate or clofibrate was statistically significant (Fig. 1D): the vehicle- and rosiglitazone-treated rats died, respectively, 43 ± 8 and 50 ± 2 days after the start of dietary treatment, whereas clofibrate and fenofibrate reduced mortality to zero throughout the 14 weeks of observation (Fig. 1D).
Urinary Accumulation of Inflammatory Proteins.
At the beginning of the experiment (week 1; Fig. 2 A), one of the most abundantly excreted proteins in 24-h urine was the major urinary protein (or α-2-globulin, the main protein excreted in the urine of healthy male rats) together with other low molecular weight proteins that accounted for approximately 70% of total protein content. The protein composition of urine changed over time in the vehicle-treated SHRSPs (n = 4); in particular, high molecular weight proteins (previously identified as markers of inflammation) (Sironi et al., 2001) started accumulating within 5 weeks of treatment at the same time as the percentage of low molecular weight proteins decreased (Fig. 2, A and B). The representative 1-DE image of the urine samples collected from a rat treated with fenofibrate at 150 mg/kg/day (Fig. 2C) and the densitometric analysis of the excreted proteins in the fenofibrate-treated rats (Fig. 2D) show that high molecular weight proteins did not increase to more than 50% of total protein content even after 14 weeks of treatment.
Effects of Fenofibrate on Renal Disorder.
The kidneys taken from the vehicle-treated rats after 6 weeks of dietary treatment showed severe vascular lesions, tubular damage, and glomerular sclerosis (Fig. 3 A), massive ED-1-positive inflammatory cell infiltration (Fig. 3B), and a massive accumulation of collagen (data not shown). Fenofibrate at 150 mg/kg/day completely prevented the development of renal lesions and abolished the tissue infiltration of ED-1-positive cells (Fig. 3, A and B) and collagen accumulation.
Moreover, in comparison with the animals fed a normal diet, the vehicle-treated rats fed a permissive diet showed a substantial increase in the expression of mRNA for MCP-1, IL-1β, and TGF-1β in renal tissue detected by RT-PCR, whereas treatment with fenofibrate at 150 mg/kg/day significantly attenuated the increased transcription of these cytokines (Fig. 3C).
In comparison with the nonsalt-loaded rats (normal diet group), plasma levels of the markers of protein oxidation (AOPPs) increased after 6 weeks of dietary treatment in the vehicle-treated salt-loaded rats and were significantly reduced by treatment with fenofibrate at 150 mg/kg/day (Fig. 4 A). The plasma levels of carbonylated proteins were also higher in the vehicle-treated salt-loaded rats than in the rats treated with the normal diet, but treatment with the PPARα agonist fenofibrate significantly reduced the plasma levels of protein carbonyls in comparison with the vehicle-treated rats (Fig. 4B).
The markers of oxidative stress were also assessed in the brains of salt-loaded SHRSPs treated with vehicle or fenofibrate. Quantitative ELISA determinations of the carbonylated proteins in the brain did not show any significant difference between the groups (not shown), a finding that was confirmed by immunoblotting analysis (Fig. 4, C and D).
Estimates of carbonylation in urine revealed that the high molecular weight proteins (the major component of excreted proteins 4 to 5 weeks after the start of dietary treatment) underwent strong oxidation in the vehicle-treated animals. Figure 5 A shows the results of a representative immunoblotting analysis of carbonylated proteins in urine samples from one vehicle-treated rat during 7 weeks of dietary treatment; the bar graph in Fig. 5C shows the results of the densitometric analysis of three immunoblotting experiments involving three vehicle-treated animals. Fenofibrate at 150 mg/kg/day significantly prevented the accumulation of carbonylated proteins at the seventh week (Fig. 5, B and D).
The main finding of this study is that fenofibrate and clofibrate, two activators of PPARα, provided end-organ protection in SHRSP rats, a well established model of spontaneous hypertension and systemic inflammation, whereas rosiglitazone, a PPARγ activator, failed to exert any beneficial effect. In particular, fenofibrate prevented the body fluid accumulation of acute-phase and carbonylated proteins, suggesting that the protection induced by the PPARα agonists was at least partially caused by their anti-inflammatory and antioxidative properties.
Previous in vivo studies have shown that treatment with PPARα agonists has neuroprotective effects in models of focal ischemia (Deplanque et al., 2003; Collino et al., 2006), and observations that these effects are absent in PPARα- deficient mice (Zhao et al., 2009) reinforce the hypothesis that they are receptor dependent. Furthermore, fenofibrate treatment decreases the neurological deficit induced by traumatic brain injury caused by lateral fluid percussion in rats (Besson et al., 2005), and it has been shown that 14 days of preventive treatment with fenofibrate reduces susceptibility to stroke in apolipoprotein E-deficient mice and decreases cerebral infarct volume in wild-type mice (Deplanque et al., 2003). Another study has shown that two different PPARα agonists [fenofibrate and [[4-chloro-6-[(2,3-dimethylphenyl)amino]-2-pyrimidinyl]thio]-acetic acid (WY-14643)] provide similar brain protection when administered respectively 3 and 7 days before the induction of cerebral ischemia (Inoue et al., 2003). It has been demonstrated that PPARα agonists can also exert an acute neuroprotective effect when administered just before cerebral ischemia or during the reperfusion period (Ouk et al., 2005; Collino et al., 2006).
It is noteworthy that we found that the PPARγ agonist rosiglitazone failed to protect the brain against the damage that spontaneously develops in SHRSP rats. This finding is in contrast with the growing body of evidence suggesting that PPARγ is a regulator of cell inflammatory and ischemic responses. It has been found that the administration of the PPARγ agonists troglitazone or pioglitazone (24 or 72 h before and at the time of cerebral infarction) dramatically reduces infarct volume and improves neurological function after transient middle cerebral artery occlusion in rats by negatively regulating the expression of proinflammatory genes (Sundararajan et al., 2005). However, the fact that pioglitazone reduces infarct size in the case of transient but not permanent middle cerebral artery occlusion suggests that the neuroprotective role of PPARγ is specific to events occurring during reperfusion. It has also been demonstrated that rosiglitazone reduces inflammation but does not provide neuroprotection or have any beneficial effect on brain edema after surgical brain injury (Hyong et al., 2008). These data seem to suggest that rosiglitazone does not specifically target edema, particularly the vasogenic edema occurring in our experimental model (Guerrini et al., 2002).
The neuroprotection observed after treatment with PPARα agonists has been related to a number of mechanisms, including oxidative stress modulation and anti-inflammatory effects. It has been shown that the PPARα agonist-induced neuroprotective effect is associated with a decrease in cerebral oxidative stress, which may depend on the increased activity of numerous antioxidant enzymes such as Cu/Zn superoxide dismutase and glutathione peroxidase (Deplanque et al., 2003). This modulation of antioxidant enzymes is responsible for a decrease in the ischemia-induced production of reactive oxygen species and lipid peroxidation (Collino et al., 2006). Moreover, fenofibrate decreases susceptibility to stroke in apolipoprotein E-deficient mice partly by inhibiting chronically high oxidative stress levels in their brains by reducing the level of thiobarbituric acid-reactive substances, an index of lipid peroxidation (Deplanque et al., 2003).
We now add evidence that fenofibrate reduces systemic protein oxidation in both plasma and urine. Protein carbonyls are considered a marker of oxidative stress and used to quantify the degree of oxidative damage in polypeptide chains (Dalle-Donne et al., 2003). This parameter may have some advantages over the analysis of lipid peroxidation products, because the formation of protein-bound carbonyl groups seems to be commonly associated with protein oxidation, and oxidized proteins are formed relatively early and relative stably. It is also known that cells degrade oxidized proteins over hours or days, whereas lipid peroxidation products are detoxified within minutes (Siems et al., 1997). It is noteworthy that protein carbonyl groups are formed early and circulate in the blood for longer than the other parameters of oxidative stress (Pantke et al., 1999). In addition, their chemical stability makes them suitable for laboratory measurement (Griffiths, 2000). The detection of high levels of protein carbonyls is a sign of oxidative stress and protein dysfunction (Dalle-Donne et al., 2003; Banfi et al., 2008), but we cannot exclude the possibility that other types of oxidative modification occur in salt-loaded and vehicle-treated SHRSPs.
The relationships between protein oxidation, protein dysfunction, and diseases remain largely undefined, but it is known that oxidative changes in enzymes and structural proteins play a significant role in the pathophysiology of diseases such as stroke (Lakhan et al., 2009) and Alzheimer's disease (Butterfield et al., 1997). The finding that high levels of oxidized proteins in the plasma and urine of vehicle-treated SHRSPs correlates with the development of proteinuria supports the hypothesis that, together with inflammation, reactive oxygen species are considerably involved in the progression of renal diseases.
It is noteworthy that there is growing evidence that chronic kidney disease is a significant risk factor for cardiovascular events and stroke regardless of traditional risk factors such as hypertension, dyslipidemia, and diabetes (Khella and Bleicher, 2007). In addition to its antioxidative effects and brain protection, we found that fenofibrate delayed the onset of proteinuria and prevented morphological renal alterations. This finding is relevant because it corroborates the results of the FIELD study of patients with type 2 diabetes mellitus (Keech et al., 2005), which showed that fenofibrate treatment had an important protective effect on the secondary outcome of total cardiovascular disease events such as stroke, the progression of albuminuria, and microvasculature damage, thus confirming the results of the Diabetes Atherosclerosis Intervention Study (Ansquer et al., 2005).
The beneficial effects of fenofibrate observed in this study were also at least partially attributable to its anti-inflammatory action: the decrease in low molecular weight urinary proteins and reduction in local tissue inflammation showed that it prevented systemic inflammation and also significantly decreased renal mRNA expression of the inflammatory cytokines MCP-1, IL-1β, and TGF-1β. These findings are in line with those of previous in vitro and in vivo experimental studies showing that PPARα ligands decrease the expression of cytokines and adhesion molecules (Marx et al., 1999) and repress the expression of a number of acute-phase proteins in rodent liver (Kockx et al., 1999). Similar findings have been described in clinical trials in which fibrate-treated patients with atherosclerosis and cardiovascular diseases had reduced plasma levels of cytokines, C-reactive protein, and fibrinogen (Madej et al., 1998; Staels et al., 1998).
Although studies of the molecular mechanisms of action of PPAR-α have shown that their anti-inflammatory effects are caused by their antagonizing the activity of nuclear factor-κB and the inducible cyclooxygenase-2 signaling pathway (Poynter and Daynes, 1998; Staels et al., 1998; Marx et al., 1999), the molecular determinants underlying the effects of PPARα agonists are not entirely clear.
In conclusion, we demonstrate that PPARα activators have beneficial effects on spontaneous brain and renal damage in SHRSPs, and that these effects are at least partially caused by the inhibition of systemic inflammation and oxidative stress. These properties of PPARα activators may be useful in patients undergoing cardiovascular procedures or in hypertensive patients with renal diseases, who are at high risk of stroke.
We thank Loredana Bonacina and Andrea Mangolini for looking after the animals.
This work was supported by Fondo per gli Investimenti della Ricerca di Base (Progetto Reti FIRB Ricerca e Sviluppo de Farmaco, CHEM-PROFARMA-NET) (to A.P.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
- spontaneously hypertensive stroke-prone rats
- peroxisome proliferator-activated receptor
- magnetic resonance imaging
- transforming growth factor |gb
- monocyte chemoattractant protein-1
- glyceraldehyde-3-phosphate dehydrogenase
- reverse transcription-polymerase chain reaction
- one-dimensional electrophoresis
- arbitrary units
- enzyme-linked immunosorbent assay
- advanced oxidation protein product
- [[4-chloro-6-[(2,3-dimethylphenyl)amino]-2-pyrimidinyl]thio]-acetic acid.
- Received June 8, 2010.
- Accepted July 28, 2010.
- Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics