This Article Abstract Full Text (PDF) All Versions of this Article: 40/5/687    most recent 01.HYP.0000036396.64769.C2v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ishibashi, M. Articles by Takeshita, A. Search for Related Content PubMed PubMed Citation Articles by Ishibashi, M. Articles by Takeshita, A. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Related Collections Growth factors/cytokines Endothelium/vascular type/nitric oxide Mechanism of atherosclerosis/growth factors Other Vascular biology

(Hypertension. 2002;40:687.)
© 2002 American Heart Association, Inc.

Scientific Contributions

Antiinflammatory and Antiarteriosclerotic Effects of Pioglitazone

Minako Ishibashi; Kensuke Egashira; Ken-ichi Hiasa; Shujiro Inoue; Weihua Ni; Qingwei Zhao; Makoto Usui; Shiro Kitamoto; Toshihiro Ichiki; Akira Takeshita

From the Department of Cardiovascular Medicine, Graduate School of Medical Science, Kyushu University, Fukuoka, Japan.

Correspondence to Kensuke Egashira, MD, PhD, Department of Cardiovascular Medicine, Graduate School of Medical Science, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. E-mail egashira{at}cardiol.med.kyushu-u.ac.jp

	Abstract

Top Abstract Introduction Materials and Methods Results Discussion References

 
Peroxisome proliferator-activated receptor- (PPAR) ligands are widely used in patients with insulin resistance and diabetes. Because coronary artery disease is a major complication for such patients, it is important to determine the effects of PPAR activation on arteriosclerosis. Long-term inhibition of endothelial NO synthesis by administration of N-nitro-L-arginine methyl ester (L-NAME) to rats induces coronary vascular inflammation (monocyte infiltration, monocyte chemoattractant protein-1 [MCP-1] expression) and subsequent arteriosclerosis. We examined the effects of pioglitazone (a PPAR ligand) in this rat model to determine whether PPAR activation with pioglitazone inhibits arteriosclerosis by its indirect effects on metabolic conditions or by direct effects on the cells participating to the pathogenesis of arteriosclerosis. We found that pioglitazone did not affect metabolic states, systolic blood pressure, or serum NO levels, but did prevent the L-NAME–induced coronary inflammation and arteriosclerosis. Pioglitazone did not reduce local expression of MCP-1 but markedly attenuated increased expression of the MCP-1 receptor C-C chemokine receptor 2 (CCR2) in lesional and circulating monocytes. PPAR activation with pioglitazone prevented coronary arteriosclerosis, possibly by its antiinflammatory effects (downregulation of CCR2 in circulating monocytes). Inhibition of the CCR2-mediated inflammation may represent novel antiinflammatory actions of pioglitazone beyond improvement of metabolic state.

Key Words: arteriosclerosis • leukocytes • nitric oxide • remodeling

	Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

 
Coronary arteriosclerosis and/or atherosclerosis is increasingly recognized to be a major complication of diabetes. It is the number 1 cause of death in subjects with diabetes.^1,2 Furthermore, prediabetic patients with insulin resistance have high risks for coronary atherosclerosis.³ Recently, peroxisome proliferator-activated receptor- (PPAR) agonists, oral insulin sensitizers, are used extensively in the treatment of insulin resistance and type 2 diabetes mellitus; therefore, the impact of PPAR agonists on atherosclerosis is important. PPAR, a nuclear receptor, is highly expressed in all major cell types participating in vascular injury and atherosclerotic plaque: endothelial cells, macrophages, and vascular smooth muscle cells.^4–6 PPAR ligands have been shown to inhibit inflammatory and proliferative process. PPAR ligands inhibited endothelial expression of adhesion molecules or chemokines, which induces monocyte infiltration into the vessel wall.^4,7–9

Treatment with PPAR ligand has been shown to reduce atherosclerosis in hypercholesterolemic mice^10–12 and neointimal hyperplasia after balloon injury.^13,14 PPAR ligands also inhibited neointimal formation of coronary arteries after stent placement in patients with diabetes.¹⁵ Because neointimal formation after vascular injury is usually caused by increased migration and proliferation of vascular smooth muscle cells, it is suggested that PPAR ligands might inhibit inflammatory and proliferative process that occurs after vascular injury. In addition, PPAR ligands improve endothelial dysfunction and activation.¹⁶ In contrast, PPAR activation might promote vascular disease because it activates the CD36 scavenger receptor on macrophages.^17,18 Furthermore, it remains unclear whether such beneficial vascular effects of PPAR ligands result from improvement of metabolic disorder or are attributable to direct vasculoprotective actions. Because arteriosclerotic and/or atherosclerotic coronary artery disease is the major cause of death in patients with type 2 diabetes, it is important to determine the effects of the PPAR agonist on the development of such vascular disease in vivo.

To determine whether PPAR activation exhibits pro- or antiarteriosclerotic actions in vivo, we examined the effect of the PPAR activation with pioglitazone in the rat model of long-term inhibition of NO synthesis. In this model, long-term administration of N-nitro-L-arginine methyl ester (L-NAME) induces early inflammation (monocyte infiltration into the coronary vessels) associated with induction of monocyte chemoattractant protein-1 (MCP-1) and causes subsequent arteriosclerosis.^19,20 We have previously demonstrated that (1) increased activity of angiotensin II (increased activity of ACE and angiotensin II type 1 [AT₁] receptor) is critical in the development of such inflammation and arteriosclerosis, and (2) such inflammation is mediated by increased activity of MCP-1 through nuclear factor-B.^21–24 Thus, some of these pathologic process are similar in human arteriosclerosis. We considered that this rat model is useful to investigate antiarteriosclerotic effects of PPAR activation, because administration of pioglitazone dose not change serum glucose or insulin levels.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

 
Experimental Animals
The study protocol was reviewed and approved by the Committee on the Ethics of Animal Experiments, Kyushu University Graduate School of Medical Sciences. A part of this study was performed at the Kyushu University Station for Collaborative Research and the Morphology Core, Kyushu University School of Medical Sciences.

Twenty-week-old male Wistar-Kyoto rats were obtained from an established colony at the Animal Research Institution of Kyushu University, Faculty of Medicine (Fukuoka, Japan). Several groups of 20-week-old male Wistar-Kyoto rats were studied. The control group received untreated chow and drinking water. The L-NAME group received L-NAME in drinking water (1.0 mg/mL). The L+Pio group received L-NAME in drinking water (1.0 mg/mL) and pioglitazone (3 or 20 mg/kg per day; Takeda Chemical Industries Ltd) in chow. The L+CS866 group received L-NAME in drinking water and CS866 (75 µg/g; Sankyo Pharmaceitocal Co),²⁵ an AT₁ receptor blocker, in chow. Treatment with pioglitazone or CS866 was started 4 days before L-NAME administration was begun.

Serum pioglitazone concentration was measured with the use of high-performance liquid chromatography/mass spectrometry.²⁶ Serum pioglitazone concentrations in the L+Pio3 and L+Pio20 groups were 0.46±0.13 and 2.16±0.31 µg/mL, respectively. The concentrations of pioglitazone achieved by 3 and 20 mg doses in rats were equivalent to the concentration reported in humans who were administered 15 and 30 mg doses of pioglitazone, respectively. Effectiveness of pioglitazone as a PPAR ligand was determined by assessing gene expression of lipoprotein lipase (LPL) in adipose tissues by Northern blot analysis; LPL mRNA levels (ratio of LPL to GAPDH relative to control group, which was assigned to be 1.0) in the L-NAME, L+Pio3, and L+Pio20 groups were 1.0±0.1, 2.8±0.2 (P<0.01), and 3.8±0.3 (P<0.01 versus control and L-NAME group), respectively.

Systolic blood pressure (by the tail-cuff method) was measured on days 3, 7, 14, and 28 of L-NAME treatment. On days 3 or 28, arterial blood was collected after anesthesia, and the rats were killed for morphometric, immunohistochemical, or biochemical analysis.

Biochemical Analysis
Rats were fasted overnight before the collection of the blood samples. Plasma glucose levels were immediately measured on Glutest Pro (Sanwa Kagaku Co). Serum total cholesterol, triglycerides, HDL cholesterol, and insulin levels were detected with commercially available kits (Wako). Serum NO_x (NO₂ plus NO₃) concentration was measured with use of fluorometric assay kit (Wako).²⁷ The ACE activity was measured in cardiac tissue by fluorometric assay as described.¹⁹

Histopathology and Immunohistochemistry
Histopathology and immunohistochemistry were performed as described.¹⁹ Some sections were subjected to immunostaining using antibodies against rat macrophage/monocytes (ED-1, Serotec Inc), proliferating cell nuclear antigen (PCNA, DAKO), MCP-1, transforming growth factor-ß₁ (TGF-ß1;Biosource International), and C-C chemokine receptor 2 (CCR2) (Santa Cruz Biotechnology Inc). Cell enumeration was performed as described.¹⁹ Evaluation of the medial thickness and perivascular fibrosis of coronary arteries on day 28 was performed as previously described.¹⁹

Northern Blot Analysis
Northern blot hybridization was performed as described.²⁰ A rat MCP-1 cDNA probe, rat TGF-ß1 probe, and a mouse GAPDH cDNA probe were used.

TaqMan Real-Time Reverse Transcription–Polymerase Chain Reaction Analysis
Total RNA was extracted from unfixed hearts or 10 to 20 million peripheral blood mononuclear cells (PBMCs) by guanidinium thiocyanate-phenol-chloroform extraction (Isogen, Nippon Gene Co). Transcripts from 1 µg of total RNA were reverse-transcribed by ReverTra Ace using oligo(dT)10 primer, and the resultant cDNA was amplified by TaqMan real-time reverse transcription–polymerase chain reaction (RT-PCR). The PCR reaction is directly monitored by the ABI Prism 7000 sequence detection system.²⁸ The PCR primers and TaqMan probes were designed with a software program from Primer Express, version 2.0. Real-time RT-PCR for CCR2 was amplified in the same way with the following primers: sense primer, 5'-ACCTGTTCAACCTGGCCATCT-3'; antisense primer, 5'-AGACCCACTCATTTGCAGCAT-3'; and probe oligonucleotides, 5'-ACCTGCTCTTCCTGCTCACACTCCCA-3'. The GAPDH probe was obtained from Applied Biosystems.

Flow Cytometry Analysis
Rat PBMCs were purified by centrifugation on Lympholyte-M and were washed with ice-cold PBS supplemented with 1% bovine serum albumin and 0.1% sodium azide. The isolated PBMCs (1x10⁶) were preincubated with 4 µg of goat IgG for 15 minutes at room temperature and then were incubated with 4 µg of biotin anti-rat Mononuclear Phagocyte (Becton Dickinson Biosciences) and 4 µg of goat anti-rat CCR2 antibody (Santa Cruz Biotechnology Inc) for 30 minutes at 4°C. After washing, cells were stained with 4 µg of phycoerythrin-conjugated streptavidin (Becton Dickinson Biosciences) and 4 µg of fluorescein-labeled mouse anti-goat IgG (Santa Cruz Biotechnology Inc) for 30 minutes at 4°C. Stained cells were analyzed by FACSCalibur instrument using CELL QUEST software (Becton Dickinson Biosciences). In control experiments, fluorescein-conjugated nonspecific goat IgG was used to measure nonspecific binding.

Statistical Analysis
Data are expressed as mean±SE. Statistical analysis of differences was compared by ANOVA and Bonferroni’s multiple comparison test. A level of P<0.05 was considered statistically significant.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
Systolic Arterial Pressure and Serum Metabolic State
The L-NAME group showed a progressive rise in systolic arterial pressure throughout the study (Table 1). Treatment with pioglitazone did not affect the L-NAME–induced increase in systolic blood pressure. Pioglitazone also did not affect blood glucose and serum concentrations of insulin, total cholesterol (TC), triglycerides (TG), and HDL cholesterol (HDL-C) (Table 2).

View this table: [in this window] [in a new window]   TABLE 1. Systolic Blood Pressure and Tissue ACE Activitiy

View this table: [in this window] [in a new window]   TABLE 2. Blood Glucose Levels and Serum Insulin, Lipid, and NOx Concentrations

Effects of Pioglitazone on Local ACE Activity and Serum NO_x Concentration
Cardiac ACE activity was increased in the L-NAME group on day 3 (Table 1). This increased ACE activity was decreased by treatment with pioglitazone. Serum NO_x concentration was significantly decreased in the L-NAME group (Table 2). This decreased serum NO_x was not influenced by treatment of pioglitazone.

Effects of Pioglitazone on Inflammatory and Proliferative Changes
As we have reported,^22–24 on day 3, attachment of mononuclear leukocytes to endothelium of coronary vessels and their infiltration into the vessel wall were noted in the L-NAME group. The majority of these cells were found to be ED-1–positive monocytes. Nuclear staining with PCNA antibody was observed in some endothelial cells, vascular smooth muscle cells in the media, monocytes, and myofibroblast-like cells. We found no evidence of such inflammation in the control rats.

The number of immunopositive cells per section was significantly greater in L-NAME group than in the control group (Figure 1A). The increases in ED-1–positive cells and PCNA-positive cells were significantly reduced by treatment with low and high doses of pioglitazone (Figure 1A).

View larger version (36K): [in this window] [in a new window]   Figure 1. Effect of pioglitazone on early inflammatory and proliferative changes and late arteriosclerosis. A, Coronary artery sections were stained immunohistochemically for ED-1 and PCNA on day 3 of L-NAME treatment. Number of ED-1–positive monocytes infiltrated into the coronary vessels and PCNA-positive cells in the control, L-NAME, L+Pio3, L+Pio20 groups are shown. *P<0.05 vs L-NAME group; P<0.05 vs control group; n=7. B, Coronary artery sections stained with Masson’s trichrome on day 28. C, Wall-to-lumen ratio and perivascular fibrosis of coronary arteries. *P<0.05 vs L-NAME group; P<0.05 vs control group; n=6.

Effects of Pioglitazone on Coronary Arteriosclerosis
On day 28, the increases in medial thickening (wall-to-lumen ratio) and perivascular fibrosis of coronary arteries were seen in the L-NAME group. These changes were inhibited by treatment with low and high doses of pioglitazone (Figure 1B and 1C).

Effects of Pioglitazone on MCP-1 and TGF-ß1 Gene Expression
On day 3, both MCP-1 and TGF-ß1 mRNA levels were significantly increased in L-NAME group (Figure 2A and 2B). The increased expression of MCP-1 and TGF-ß1 gene was not reduced by treatment with low and high doses of pioglitazone. As we previously reported,^22–24 immunohistochemical study showed the L-NAME–induced increases of MCP-1 and TGF-ß1 immunoreactivity in coronary arteries, which were not reduced in rats from the L+Pio group (Figure 2C).

View larger version (51K): [in this window] [in a new window]   Figure 2. Effect of pioglitazone on cardiac MCP-1 and TGF-ß1 gene expression. A, Representative autogram of Northern blot analysis of MCP-1, TGF-ß1, and GAPDH mRNA. B, Summary of densitometric analysis of data. Data are expressed as ratio of MCP-1 and TGF-ß1 mRNA to GAPDH mRNA relative to control, which was assigned an arbitrary value of 1. C, Coronary artery sections were stained with MCP-1 and TGF-ß1 on day 3 of L-NAME treatment. *P<0.05 vs L-NAME group; P<0.05 vs control group; n=8.

Effects of Pioglitazone on the MCP-1 Receptor Expression in Lesional and Circulating Monocytes
On day 3, CCR2 mRNA levels in PBMC and left ventricle were significantly higher in the L-NAME group (Figure 3A). This upregulation of CCR2 was prevented by treatment with the low and high doses of pioglitazone. We determined the cell type expressing CCR2 in hearts by fluorescence immunohistochemistry (Figure 3B) and found that immunoreactive CCR2 was present exclusively in the monocytes/macrophages that had infiltrated into the intima and adventitia of coronary arteries in the L-NAME group. No such CCR2-positive cells were noted in the control group. The appearance of CCR2-positive cells was significantly reduced by treatment with low and high doses of pioglitazone. The CCR2 mRNA levels in PBMCs and left ventricle, and the number of CCR2-positive cells, were significantly reduced by treatment with CS-866, an AT₁ receptor blocker. The CCR2 immunoreactivity was also observed in endothelial cell layer of coronary arteries, the degrees of which did not differ among the 4 groups.

View larger version (24K): [in this window] [in a new window]   Figure 3. Effect of pioglitazone on CCR2 gene expression. A, CCR2 gene expression by real-time RT-PCR in left ventricular tissues and PBMCs. Data are expressed as ratio of CCR2 mRNA to GAPDH mRNA relative to control, which was assigned an arbitrary value of 1. *P<0.05 vs L-NAME group; P<0.05 vs control group; n=7 to 8. B, Immunofluorescence images of coronary arteries. Indirect immunofluorescence was performed on paraffin-fixed tissue section, using anti-rat CCR2 antibody or anti-rat ED-1. The bound antibody is visualized with a fluorescein-conjugated secondary antibody (green fluorescence) or rhodamine-conjugated secondary antibody (red fluorescence): CCR2 immunofluorescence (left; labeled green with fluorescein), ED-1 immunofluorescence (middle; red with rhodamine), and their overlap (right; yellow). CCR2 located exclusively at infiltrated monocyte/macrophages, resulting in overlap with ED-1 (yellow).

We further analyzed the appearance of CCR2 on circulating monocytes by flow cytometry (Figure 4). The CCR2 expression in monocytes was significantly higher in L-NAME group than in the control group. Treatment with pioglitazone or CS-866 prevented the L-NAME–induced increase in CCR2 expression (Figure 4).

View larger version (30K): [in this window] [in a new window]   Figure 4. Effect of pioglitazone on CCR2 surface expression of circulating monocytes, which was determined by flow cytometry. A, CCR2 surface expression on monocytes shown by 2-color immunofluorescence using anti-rat mononuclear phagocyte and anti-rat CCR2 antibody. B, Overlays of 4 histograms that illustrate control group (black line), L-NAME group (red line), L+Pio3 group (yellow line), and L+Pio20 group (blue line). C, Summary of mean log fluorescence intensity, which was calculated as an expression index of CCR2 on the surface of the circulating monocytes. *P<0.05 vs L-NAME group; P<0.05 vs control group; n=6.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

 
We have demonstrated herein that PPAR activation with pioglitazone prevented coronary arteriosclerosis, possibly by its antiinflammatory action on the MCP-1 receptor (CCR2) in this rat model. Our present data suggest that the antiinflammatory effects of pioglitazone may not be explained by its indirect effects on metabolic state, by antihypertensive effect, or by restoration of NO production.

An important feature that emerged in the present study is that pioglitazone attenuated inflammatory (monocyte infiltration) and proliferative (appearance of PCNA-positive cells) disorders, suggesting that pioglitazone prevented coronary arteriosclerosis by its direct antiinflammatory actions in this model. Because we have previously demonstrated that (1) MCP-1 mediates inflammation and arteriosclerosis, and (2) TGF-ß1 mediates fibrosis in this rat model,^22–24 we expected that pioglitazone might suppress increased expression of MCP-1 and TGF-ß1. In contrast, pioglitazone did not affect expression of such local factors, indicating that other factors must be involved in antiinflammatory and antiarteriosclerotic actions of pioglitazone. Most previous studies that investigated inflammatory aspects of vascular disease focused exclusively on lipid- or stress-induced changes in inflammation-driving factors, such as MCP-1 in cells of the arterial wall,^29,30 whereas pathobiological changes in CCR2 have not been attracted attention. The monocyte/macrophage infiltration and atherosclerotic lesion formation were less in CCR2^-/- and apoE^-/- mice than in apoE^-/- mice.³¹ We found here that pioglitazone prevented the L-NAME–induced increase in CCR2 expression in lesional and circulating monocytes.

Our present data are in accordance with recent data by Han et al, ¹⁰ who demonstrated inhibition of monocyte CCR2 expression by rosiglitazone in LDL receptor mice. The increase in CCR2 intensity by 40% in L-NAME group and its prevention by pioglitazone are implicated to be pathophysiologically significant findings, because prior reports have shown that small changes (ie, 20%) in CCR2 expression in monocytes result in biologically meaningful (2x) enhancement in chemotaxis activity.^10,32 These findings suggest that in addition to MCP-1, the enhanced expression and activity of CCR2 in lesional and circulating monocytes contributed not only to monocyte recruitment into the arterial wall by MCP-1 but also to monocyte activation (production of growth factors and/or cytokines).

We have previously shown the critical role of local activity of angiotensin II (increased activity of tissue ACE and AT₁ receptor) in the development of vascular inflammation and arteriosclerosis induced by long-term inhibition of NO synthesis.^19,22 Therefore, our present data on prevention of local ACE activation by pioglitazone may be an interesting observation. Because previous reports by us²² and others^33,34 showed that ACE and AT₁ receptor activities are increased in lesional monocytes in experimental and human arteriosclerotic lesions, pioglitazone might have decreased local ACE activity secondary to its antiinflammatory effects. Further studies are needed to elucidate the molecular mechanism involved in decreased ACE activity by pioglitazone.

Our present observation of inhibition of enhanced expression of CCR2 by AT₁ receptor blocker may imply that an AT₁ receptor might be involved in the pathogenesis of upregulation of CCR2 in the present study. Compared with the regulation of MCP-1 expression by angiotensin II, a relationship between angiotensin II and CCR2 expression has not extensively been investigated. A recent in vitro study demonstrated increased CCR2 expression by oxidative stress.^32,35 Because oxidative stress has been shown to mediate various proinflammatory and proarteriosclerotic actions of angiotensin II,³⁶ and because oxidative stress is increased through increased activity of angiotensin II in this rat model of long-term inhibition of NO synthesis,³⁷ it is possible that oxidative stress driven by angiotensin II might mediate the upregulation of CCR2 in the present study. Further studies are needed to elucidate the mechanism of downregulation of CCR2 by angiotensin II receptor blockade. A caveat to interpret our present data is that the observed antiinflammatory effect of pioglitazone was dependent or independent of PPAR activity. We showed an in vivo dose-related response of pioglitazone in increasing LPL mRNA, whereas pioglitazone at the low and high doses exhibited equivalent antiinflammatory effect. This discrepant action of pioglitazone suggests that the observed antiinflammatory actions may occur independent of PPAR activity as reported by Chawla et al³⁸ Further studies are needed to determine the molecular mechanisms by which pioglitazone downregulated CCR2 in monocytes in vivo.

Perspectives
Pioglitazone prevented coronary arteriosclerosis, possibly by its antiinflammatory effects. The antiinflammatory and antiarteriosclerotic effects of pioglitazone may be mediated by downregulation of CCR2 in circulating and lesional monocytes. Inhibition of the CCR2-mediated inflammation may represent novel antiinflammatory actions of pioglitazone beyond metabolic effects.

	Acknowledgments

 
This study was supported by Grants-in-Aid for Scientific Research (11470164, 11158216, 14657172, and 14207036) from the Ministry of Education, Science and Culture, Tokyo, Japan.

Received May 15, 2002; first decision June 6, 2002; accepted August 23, 2002.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 
1. Haffner SM, Lehto S, Ronnemaa T, Pyorala K, Laakso M. Mortality from coronary heart disease in subjects with type 2 diabetes and in nondiabetic subjects with and without prior myocardial infarction. N Engl J Med. 1998; 339: 229–234.[Abstract/Free Full Text]

2. Mukamal KJ, Nesto RW, Cohen MC, Muller JE, Maclure M, Sherwood JB, Mittleman MA. Impact of diabetes on long-term survival after acute myocardial infarction: comparability of risk with prior myocardial infarction. Diabetes Care. 2001; 24: 1422–1427.[Abstract/Free Full Text]

3. Howard G, O’Leary DH, Zaccaro D, Haffner S, Rewers M, Hamman R, Selby JV, Saad MF, Savage P, Bergman R. Insulin sensitivity and atherosclerosis: the Insulin Resistance Atherosclerosis Study (IRAS) Investigators. Circulation. 1996; 93: 1809–1817.[Abstract/Free Full Text]

4. Ricote M, Huang J, Fajas L, Li A, Welch J, Najib J, Witztum JL, Auwerx J, Palinski W, Glass CK. Expression of the peroxisome proliferator-activated receptor- (PPAR) in human atherosclerosis and regulation in macrophages by colony stimulating factors and oxidized low density lipoprotein. Proc Natl Acad Sci U S A. 1998; 95: 7614–7619.[Abstract/Free Full Text]

5. Xin X, Yang S, Kowalski J, Gerritsen ME. Peroxisome proliferator-activated receptor- ligands are potent inhibitors of angiogenesis in vitro and in vivo. J Biol Chem. 1999; 274: 9116–9121.[Abstract/Free Full Text]

6. Law RE, Goetze S, Xi XP, Jackson S, Kawano Y, Demer L, Fishbein MC, Meehan WP, Hsueh WA. Expression and function of PPAR in rat and human vascular smooth muscle cells. Circulation. 2000; 101: 1311–1318.[Abstract/Free Full Text]

7. Jiang C, Ting AT, Seed B. PPAR- agonists inhibit production of monocyte inflammatory cytokines. Nature. 1998; 391: 82–86.[CrossRef][Medline] [Order article via Infotrieve]

8. Marx N, Schonbeck U, Lazar MA, Libby P, Plutzky J. Peroxisome proliferator-activated receptor- activators inhibit gene expression and migration in human vascular smooth muscle cells. Circ Res. 1998; 83: 1097–1103.[Abstract/Free Full Text]

9. Pasceri V, Wu HD, Willerson JT, Yeh ET. Modulation of vascular inflammation in vitro and in vivo by peroxisome proliferator-activated receptor- activators. Circulation. 2000; 101: 235–238.[Abstract/Free Full Text]

10. Han KH, Chang MK, Boullier A, Green SR, Li A, Glass CK, Quehenberger O. Oxidized LDL reduces monocyte CCR2 expression through pathways involving peroxisome proliferator-activated receptor-. J Clin Invest. 2000; 106: 793–802.[Medline] [Order article via Infotrieve]

11. Collins AR, Meehan WP, Kintscher U, Jackson S, Wakino S, Noh G, Palinski W, Hsueh WA, Law RE. Troglitazone inhibits formation of early atherosclerotic lesions in diabetic and nondiabetic low-density lipoprotein receptor–deficient mice. Arterioscler Thromb Vasc Biol. 2001; 21: 365–371.[Abstract/Free Full Text]

12. Chen Z, Ishibashi S, Perrey S, Osuga J, Gotoda T, Kitamine T, Tamura Y, Okazaki H, Yahagi N, Iizuka Y, Shionoiri F, Ohashi K, Harada K, Shimano H, Nagai R, Yamada N. Troglitazone inhibits atherosclerosis in apolipoprotein E–knockout mice: pleiotropic effects on CD36 expression and HDL. Arterioscler Thromb Vasc Biol. 2001; 21: 372–377.[Abstract/Free Full Text]

13. Law RE, Meehan WP, Xi XP, Graf K, Wuthrich DA, Coats W, Faxon D, Hsueh WA. Troglitazone inhibits vascular smooth muscle cell growth and intimal hyperplasia. J Clin Invest. 1996; 98: 1897–1905.[Medline] [Order article via Infotrieve]

14. Goetze S, Kintscher U, Kim S, Meehan WP, Kaneshiro K, Collins AR, Fleck E, Hsueh WA, Law RE. Peroxisome proliferator-activated receptor- ligands inhibit nuclear but not cytosolic extracellular signal–regulated kinase/mitogen-activated protein kinase–regulated steps in vascular smooth muscle cell migration. J Cardiovasc Pharmacol. 2001; 38: 909–921.[CrossRef][Medline] [Order article via Infotrieve]

15. Takagi T, Akasaka T, Yamamuro A, Honda Y, Hozumi T, Morioka S, Yoshida K. Troglitazone reduces neointimal tissue proliferation after coronary stent implantation in patients with non-insulin dependent diabetes mellitus: a serial intravascular ultrasound study. J Am Coll Cardiol. 2000; 36: 1529–1535.[Abstract/Free Full Text]

16. Kato K, Satoh H, Endo Y, Yamada D, Midorikawa S, Sato W, Mizuno K, Fujita T, Tsukamoto K, Watanabe T. Thiazolidinediones down-regulate plasminogen activator inhibitor type 1 expression in human vascular endothelial cells: a possible role for PPAR in endothelial function. Biochem Biophys Res Commun. 1999; 258: 431–435.[CrossRef][Medline] [Order article via Infotrieve]

17. Nagy L, Tontonoz P, Alvarez JG, Chen H, Evans RM. Oxidized LDL regulates macrophage gene expression through ligand activation of PPAR. Cell. 1998; 93: 229–240.[CrossRef][Medline] [Order article via Infotrieve]

18. Tontonoz P, Nagy L, Alvarez JG, Thomazy VA, Evans RM. PPAR promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell. 1998; 93: 241–252.[CrossRef][Medline] [Order article via Infotrieve]

19. Takemoto M, Egashira K, Usui M, Numaguchi K, Tomita H, Tsutsui H, Shimokawa H, Sueishi K, Takeshita A. Important role of tissue angiotensin-converting enzyme activity in the pathogenesis of coronary vascular and myocardial structural changes induced by long-term blockade of nitric oxide synthesis in rats. J Clin Invest. 1997; 99: 278–287.[Medline] [Order article via Infotrieve]

20. Tomita H, Egashira K, Kubo-Inoue M, Usui M, Koyanagi M, Shimokawa H, Takeya M, Yoshimura T, Takeshita A. Inhibition of NO synthesis induces inflammatory changes and monocyte chemoattractant protein-1 expression in rat hearts and vessels. Arterioscler Thromb Vasc Biol. 1998; 18: 1456–1464.[Abstract/Free Full Text]

21. Katoh M, Egashira K, Usui M, Ichiki T, Tomita H, Shimokawa H, Rakugi H, Takeshita A. Cardiac angiotensin II receptors are upregulated by long-term inhibition of nitric oxide synthesis in rats. Circ Res. 1998; 83: 743–751.[Abstract/Free Full Text]

22. Usui M, Egashira K, Tomita H, Koyanagi M, Katoh M, Shimokawa H, Takeya M, Yoshimura T, Matsushima K, Takeshita A. Important role of local angiotensin II activity mediated via type 1 receptor in the pathogenesis of cardiovascular inflammatory changes induced by chronic blockade of nitric oxide synthesis in rats. Circulation. 2000; 101: 305–310.[Abstract/Free Full Text]

23. Kitamoto S, Egashira K, Kataoka C, Koyanagi M, Katoh M, Shimokawa H, Morishita R, Kaneda Y, Sueishi K, Takeshita A. Increased activity of nuclear factor-B participates in cardiovascular remodeling induced by chronic inhibition of nitric oxide synthesis in rats. Circulation. 2000; 102: 806–812.[Abstract/Free Full Text]

24. Koyanagi M, Egashira K, Kitamoto S, Ni W, Shimokawa H, Takeya M, Yoshimura T, Takeshita A. Role of monocyte chemoattractant protein-1 in cardiovascular remodeling induced by chronic blockade of nitric oxide synthesis. Circulation. 2000; 102: 2243–2248.[Abstract/Free Full Text]

25. Takemoto M, Egashira K, Tomita H, Usui M, Okamoto H, Kitabatake A, Shimokawa H, Sueishi K, Takeshita A. Chronic angiotensin-converting enzyme inhibition and angiotensin II type 1 receptor blockade: effects on cardiovascular remodeling in rats induced by the long-term blockade of nitric oxide synthesis. Hypertension. 1997; 30: 1621–1627.[Abstract/Free Full Text]

26. Zhong WZ, Williams MG. Simultaneous quantitation of pioglitazone and its metabolites in human serum by liquid chromatography and solid phase extraction. J Pharm Biomed Anal. 1996; 14: 465–473.[CrossRef][Medline] [Order article via Infotrieve]

27. Ni W, Egashira K, Kataoka C, Kitamoto S, Koyanagi M, Inoue S, Takeshita A. Antiinflammatory and antiarteriosclerotic actions of HMG-CoA reductase inhibitors in a rat model of chronic inhibition of nitric oxide synthesis. Circ Res. 2001; 89: 415–421.[Abstract/Free Full Text]

28. Heid CA, Stevens J, Livak KJ, Williams PM. Real-time quantitative PCR. Genome Res. 1996; 6: 986–994.[Abstract/Free Full Text]

29. Glass CK, Witztum JL. Atherosclerosis: the road ahead. Cell. 2001; 104: 503–516.[CrossRef][Medline] [Order article via Infotrieve]

30. Lusis AJ. Atherosclerosis. Nature. 2000; 407: 233–241.[CrossRef][Medline] [Order article via Infotrieve]

31. Boring L, Gosling J, Cleary M, Charo IF. Decreased lesion formation in CCR2^-/- mice reveals a role for chemokines in the initiation of atherosclerosis. Nature. 1998; 394: 894–897.[CrossRef][Medline] [Order article via Infotrieve]

32. Wang G, O K. Homocysteine stimulates the expression of monocyte chemoattractant protein-1 receptor (CCR2) in human monocytes: possible involvement of oxygen free radicals. Biochem J. 2001; 357: 233–240.[CrossRef][Medline] [Order article via Infotrieve]

33. Diet F, Pratt RE, Berry GJ, Momose N, Gibbons GH, Dzau VJ. Increased accumulation of tissue ACE in human atherosclerotic coronary artery disease. Circulation. 1996; 94: 2756–2767.[Abstract/Free Full Text]

34. Potter DD, Sobey CG, Tompkins PK, Rossen JD, Heistad DD. Evidence that macrophages in atherosclerotic lesions contain angiotensin II. Circulation. 1998; 98: 800–807.[Abstract/Free Full Text]

35. Saccani A, Saccani S, Orlando S, Sironi M, Bernasconi S, Ghezzi P, Mantovani A, Sica A. Redox regulation of chemokine receptor expression. Proc Natl Acad Sci U S A. 2000; 97: 2761–2766.[Abstract/Free Full Text]

36. Dzau VJ. Tissue angiotensin and pathobiology of vascular disease: a unifying hypothesis. Hypertension. 2001; 37: 1047–1052.[Abstract/Free Full Text]

37. Usui M, Egashira K, Kitamoto S, Koyanagi M, Katoh M, Kataoka C, Shimokawa H, Takeshita A. Pathogenic role of oxidative stress in vascular angiotensin-converting enzyme activation in long-term blockade of nitric oxide synthesis in rats. Hypertension. 1999; 34: 546–551.[Abstract/Free Full Text]

38. Chawla A, Barak Y, Nagy L, Liao D, Tontonoz P, Evans RM. PPAR-– dependent and independent effects on macrophage-gene expression in lipid metabolism and inflammation. Nat Med. 2001; 7: 48–52.[CrossRef][Medline] [Order article via Infotrieve]

This article has been cited by other articles:

				M. R. Dasu, S. Park, S. Devaraj, and I. Jialal Pioglitazone Inhibits Toll-Like Receptor Expression and Activity in Human Monocytes and db/db Mice Endocrinology, August 1, 2009; 150(8): 3457 - 3464. [Abstract] [Full Text] [PDF]

				T. Takagi, H. Okura, Y. Kobayashi, T. Kataoka, H. Taguchi, I. Toda, K. Tamita, A. Yamamuro, Y. Sakanoue, A. Ito, et al. A Prospective, Multicenter, Randomized Trial to Assess Efficacy of Pioglitazone on In-Stent Neointimal Suppression in Type 2 Diabetes: POPPS (Prevention of In-Stent Neointimal Proliferation by Pioglitazone Study) J. Am. Coll. Cardiol. Intv., June 1, 2009; 2(6): 524 - 531. [Abstract] [Full Text] [PDF]

				H. Nakaya, B. D. Summers, A. C. Nicholson, A. M. Gotto Jr., D. P. Hajjar, and J. Han Atherosclerosis in LDLR-Knockout Mice Is Inhibited, but Not Reversed, by the PPAR{gamma} Ligand Pioglitazone Am. J. Pathol., June 1, 2009; 174(6): 2007 - 2014. [Abstract] [Full Text] [PDF]

				K. K. Koh, P. C. Oh, and M. J. Quon Does reversal of oxidative stress and inflammation provide vascular protection? Cardiovasc Res, March 1, 2009; 81(4): 649 - 659. [Abstract] [Full Text] [PDF]

				T. Nakamura, E. Yamamoto, K. Kataoka, T. Yamashita, Y. Tokutomi, Y.-F. Dong, S. Matsuba, H. Ogawa, and S. Kim-Mitsuyama Beneficial Effects of Pioglitazone on Hypertensive Cardiovascular Injury Are Enhanced by Combination With Candesartan Hypertension, February 1, 2008; 51(2): 296 - 301. [Abstract] [Full Text] [PDF]

				G. Ceolotto, A. Gallo, I. Papparella, L. Franco, E. Murphy, E. Iori, E. Pagnin, G. P. Fadini, M. Albiero, A. Semplicini, et al. Rosiglitazone Reduces Glucose-Induced Oxidative Stress Mediated by NAD(P)H Oxidase via AMPK-Dependent Mechanism Arterioscler Thromb Vasc Biol, December 1, 2007; 27(12): 2627 - 2633. [Abstract] [Full Text] [PDF]

				G Rogler Significance of anti-inflammatory effects of PPAR{gamma} agonists? Gut, August 1, 2006; 55(8): 1067 - 1069. [Full Text] [PDF]

				R. Agarwal Anti-inflammatory effects of short-term pioglitazone therapy in men with advanced diabetic nephropathy Am J Physiol Renal Physiol, March 1, 2006; 290(3): F600 - F605. [Abstract] [Full Text] [PDF]

				K. Benkirane, E. C. Viel, F. Amiri, and E. L. Schiffrin Peroxisome Proliferator-Activated Receptor {gamma} Regulates Angiotensin II-Stimulated Phosphatidylinositol 3-Kinase and Mitogen-Activated Protein Kinase in Blood Vessels In Vivo Hypertension, January 1, 2006; 47(1): 102 - 108. [Abstract] [Full Text] [PDF]

				V. R. Babaev, P. G. Yancey, S. V. Ryzhov, V. Kon, M. D. Breyer, M. A. Magnuson, S. Fazio, and M. F. Linton Conditional Knockout of Macrophage PPAR{gamma}Increases Atherosclerosis in C57BL/6 and Low-Density Lipoprotein Receptor-Deficient Mice Arterioscler Thromb Vasc Biol, August 1, 2005; 25(8): 1647 - 1653. [Abstract] [Full Text] [PDF]

				E. L. Schiffrin More Evidence of Cardiorenal Protective Effects of Peroxisome Proliferator-Activated Receptor Activation Hypertension, August 1, 2005; 46(2): 267 - 268. [Full Text] [PDF]

				V. Gaillard, D. Casellas, C. Seguin-Devaux, H. Schohn, M. Dauca, J. Atkinson, and I. Lartaud Pioglitazone Improves Aortic Wall Elasticity in a Rat Model of Elastocalcinotic Arteriosclerosis Hypertension, August 1, 2005; 46(2): 372 - 379. [Abstract] [Full Text] [PDF]

				A. Pfutzner, N. Marx, G. Lubben, M. Langenfeld, D. Walcher, T. Konrad, and T. Forst Improvement of Cardiovascular Risk Markers by Pioglitazone Is Independent From Glycemic Control: Results From the Pioneer Study J. Am. Coll. Cardiol., June 21, 2005; 45(12): 1925 - 1931. [Abstract] [Full Text] [PDF]

				M. Abdelrahman, A. Sivarajah, and C. Thiemermann Beneficial effects of PPAR-{gamma} ligands in ischemia-reperfusion injury, inflammation and shock Cardiovasc Res, March 1, 2005; 65(4): 772 - 781. [Abstract] [Full Text] [PDF]

				D. Li, K. Chen, N. Sinha, X. Zhang, Y. Wang, A. K. Sinha, F. Romeo, and J. L. Mehta The effects of PPAR-{gamma} ligand pioglitazone on platelet aggregation and arterial thrombus formation Cardiovasc Res, March 1, 2005; 65(4): 907 - 912. [Abstract] [Full Text] [PDF]

				E. L. Schiffrin Peroxisome proliferator-activated receptors and cardiovascular remodeling Am J Physiol Heart Circ Physiol, March 1, 2005; 288(3): H1037 - H1043. [Abstract] [Full Text] [PDF]

				D. Choi, S.-K. Kim, S.-H. Choi, Y.-G. Ko, C.-W. Ahn, Y. Jang, S.-K. Lim, H.-C. Lee, and B.-S. Cha Preventative Effects of Rosiglitazone on Restenosis After Coronary Stent Implantation in Patients With Type 2 Diabetes Diabetes Care, November 1, 2004; 27(11): 2654 - 2660. [Abstract] [Full Text] [PDF]

				N. Brettenthaler, C. De Geyter, P. R. Huber, and U. Keller Effect of the Insulin Sensitizer Pioglitazone on Insulin Resistance, Hyperandrogenism, and Ovulatory Dysfunction in Women with Polycystic Ovary Syndrome J. Clin. Endocrinol. Metab., August 1, 2004; 89(8): 3835 - 3840. [Abstract] [Full Text] [PDF]

				S. E. Epstein, E. Stabile, T. Kinnaird, C. W. Lee, L. Clavijo, and M. S. Burnett Janus Phenomenon: The Interrelated Tradeoffs Inherent in Therapies Designed to Enhance Collateral Formation and Those Designed to Inhibit Atherogenesis Circulation, June 15, 2004; 109(23): 2826 - 2831. [Full Text] [PDF]

				M. Ishibashi, K.-i. Hiasa, Q. Zhao, S. Inoue, K. Ohtani, S. Kitamoto, M. Tsuchihashi, T. Sugaya, I. F. Charo, S. Kura, et al. Critical Role of Monocyte Chemoattractant Protein-1 Receptor CCR2 on Monocytes in Hypertension-Induced Vascular Inflammation and Remodeling Circ. Res., May 14, 2004; 94(9): 1203 - 1210. [Abstract] [Full Text] [PDF]

				K. T. Weber From Inflammation to Fibrosis: A Stiff Stretch of Highway Hypertension, April 1, 2004; 43(4): 716 - 719. [Full Text] [PDF]

				C. Baylis, E.-A. Atzpodien, G. Freshour, and K. Engels Peroxisome Proliferator-Activated Receptor {gamma} Agonist Provides Superior Renal Protection versus Angiotensin-Converting Enzyme Inhibition in a Rat Model of Type 2 Diabetes with Obesity J. Pharmacol. Exp. Ther., December 1, 2003; 307(3): 854 - 860. [Abstract] [Full Text] [PDF]

				J. M. Fernandez-Real and W. Ricart Insulin Resistance and Chronic Cardiovascular Inflammatory Syndrome Endocr. Rev., June 1, 2003; 24(3): 278 - 301. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) All Versions of this Article: 40/5/687    most recent 01.HYP.0000036396.64769.C2v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ishibashi, M. Articles by Takeshita, A. Search for Related Content PubMed PubMed Citation Articles by Ishibashi, M. Articles by Takeshita, A. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Related Collections Growth factors/cytokines Endothelium/vascular type/nitric oxide Mechanism of atherosclerosis/growth factors Other Vascular biology