Donate Help Contact The AHA Sign In Home
American Heart Association
Arteriosclerosis, Thrombosis, and Vascular Biology
Search: search_blue_button Advanced Search
« Previous Article | Table of Contents | Next Article »
Arteriosclerosis, Thrombosis, and Vascular Biology. 2007;27:1276-1282
Published online before print April 5, 2007, doi: 10.1161/ATVBAHA.107.142638
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Data Supplement
Right arrow All Versions of this Article:
27/6/1276    most recent
ATVBAHA.107.142638v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Krönke, G.
Right arrow Articles by Leitinger, N.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Krönke, G.
Right arrow Articles by Leitinger, N.
Related Collections
Right arrow Other Vascular biology
Right arrow Pathophysiology
Right arrow Gene regulation
Right arrow Smooth muscle proliferation and differentiation
(Arteriosclerosis, Thrombosis, and Vascular Biology. 2007;27:1276.)
© 2007 American Heart Association, Inc.

Vascular Biology

Expression of Heme Oxygenase-1 in Human Vascular Cells Is Regulated by Peroxisome Proliferator-Activated Receptors

Gerhard Krönke; Alexandra Kadl; Elena Ikonomu; Stefan Blüml; Alexander Fürnkranz; Ian J. Sarembock; Valery N. Bochkov; Markus Exner; Bernd R. Binder; Norbert Leitinger

From Robert M. Berne Cardiovascular Research Center (G.K., A.K., I.J.S., N.L.) and Department of Pharmacology (N.L.), University of Virginia, Charlottesville, Va; Department of Vascular Biology and Thrombosis Research (G.K., E.I., A.F., V.N.B., B.R.B.), Institute of Immunology (S.B.), and Department of Medical and Chemical Laboratory Diagnostics (M.E.), Medical University of Vienna, Austria.

Correspondence to Norbert Leitinger, PhD, Robert M. Berne Cardiovascular Research Center, University of Virginia, Charlottesville, P.O. Box 801394 Charlottesville, VA. E-mail nl2q{at}virginia.edu


*    Abstract
up arrowTop
 *Abstract
down arrowIntroduction
 down arrowMaterials and Methods
 down arrowResults
 down arrowDiscussion
 down arrowReferences
 
Objective— Activation of peroxisome proliferator-activated receptors (PPARs) by lipid-lowering fibrates and insulin-sensitizing thiazolidinediones inhibits vascular inflammation, atherosclerosis, and restenosis. Here we investigate if the vasculoprotective and anti-inflammatory enzyme heme oxygenase-1 (HO-1) is regulated by PPAR ligands in vascular cells.

Methods and Results— We show that treatment of human vascular endothelial and smooth muscle cells with PPAR ligands leads to expression of HO-1. Analysis of the human HO-1 promoter in transient transfection experiments together with mutational analysis and gel shift assays revealed a direct transcriptional regulation of HO-1 by PPAR{alpha} and PPAR{gamma} via 2 PPAR responsive elements. We demonstrate that a clinically relevant polymorphism within the HO-1 promoter critically influences its transcriptional activation by both PPAR isoforms. Moreover, inhibition of HO-1 enzymatic activity reversed PPAR ligand-mediated inhibition of cell proliferation and expression of cyclooxygenase-2 in vascular smooth muscle cells.

Conclusion— We demonstrate that HO-1 expression is transcriptionally regulated by PPAR{alpha} and PPAR{gamma}, indicating a mechanism of anti-inflammatory and antiproliferative action of PPAR ligands via upregulation of HO-1. Identification of HO-1 as a target gene for PPARs provides new strategies for therapy of cardiovascular diseases and a rationale for the use of PPAR ligands in the treatment of other chronic inflammatory diseases.

We investigated if the vasculoprotective activity of PPARs could be mediated by expression of HO-1. We found that PPAR{alpha} and PPAR{gamma} ligands upregulate HO-1 expression in vascular cells via 2 PPAR responsive elements. In addition, PPAR-induced HO-1 mediated anti-inflammatory and antiproliferative action of PPAR{alpha}-ligands in smooth muscle cells.


Key Words: atherosclerosis • heme oxygenase • inflammation • PPAR


*    Introduction
up arrowTop
 up arrowAbstract
 *Introduction
down arrowMaterials and Methods
 down arrowResults
 down arrowDiscussion
 down arrowReferences
 
Hyperlipidemia and diabetes type II are major causes for vascular disorders including atherosclerosis. Among the drugs that are used to treat these metabolic diseases, both the lipid-lowering fibrates and the insulin-sensitizing thiazolidinediones exert their effects via activation of peroxisome proliferator-activated receptors (PPARs).1 PPARs are ligand-activated transcription factors that bind to specific PPAR-responsive elements (PPREs) as heterodimers together with the retinoid X receptor and govern the expression of genes involved in the regulation of lipid and glucose metabolism.1,2 The 3 PPAR isoforms PPAR{alpha}, PPARβ/{delta}, and PPAR{gamma} show unique tissue distribution and ligand specificity. Natural PPAR ligands include various fatty acids and fatty acid-derived compounds such as eicosanoids.1,2 Treatment with synthetic PPAR{alpha} and PPAR{gamma} ligands such as fibrates and thiazolidinediones,3 respectively, has been shown to potently inhibit the development of atherosclerosis3–6 and restenosis.7,8 Increasing evidence suggests that the beneficial effects of PPAR ligands on the vascular wall are not only caused by changes in systemic metabolic parameters9 but additionally involve local anti-atherogenic effects,6 such as inhibition of inflammation10–12 and vascular smooth muscle cell (VSMC) proliferation.8 The beneficial effects of synthetic PPAR agonists on cardiovascular disease outcome have been demonstrated in major clinical trials that have reported cardiovascular risk reduction in patients with dyslipidemia. However, the favorable alterations in plasma lipids can only partially explain the reduction in cardiovascular events in these studies. Therefore, many beneficial effects of PPARs have been attributed to their anti-inflammatory activity.

Despite the large number of known target genes for PPARs, little is known about induction of expression of anti-inflammatory and antiproliferative genes by PPARs. We hypothesized that the local upregulation of anti-atherogenic genes by PPAR ligands contributes to their beneficial effects on the vascular wall. Recently, the presence of PPREs in the antioxidant enzymes catalase13 and manganese superoxide dismutase14 have been reported.

Numerous studies demonstrated a protective role for heme oxygenase-1 (HO-1) during atherogenesis, restenosis, and other inflammatory vascular disorders.15–20 HO-1 is the rate-limiting enzyme of heme catabolism, catalyzing the breakdown of heme into iron, biliverdin, and carbon monoxide. Both biliverdin and carbon monoxide have been shown to act as anti-inflammatory agents and to inhibit VSMC growth.17,21–23 It has been shown that the length of a GT-repeat within the proximal region of the human HO-1 promoter is highly polymorphic, which seems to influence the transcription of the HO-1 gene, rendering people possessing longer (≥29 repeats) GT fragments more susceptible to inflammatory disorders such as coronary atherosclerosis.24

Here we demonstrate induction of HO-1 expression by PPAR{alpha} and PPAR{gamma} ligands in cultured vascular cells and we show that HO-1 enzymatic activity mediates anti-inflammatory and antiproliferative effects exerted by PPAR ligands. Moreover, we show that HO-1 is a direct PPAR target gene and that a clinically relevant (GT)n dinucleotide length polymorphism within the human HO-1 promoter significantly influences the transcriptional regulation of HO-1 by both PPAR{alpha} and PPAR{gamma}.


*    Materials and Methods
up arrowTop
 up arrowAbstract
 up arrowIntroduction
 *Materials and Methods
down arrowResults
 down arrowDiscussion
 down arrowReferences
 
Expression of HO-1 and PPAR isotypes was studied using real-time polymerase chain reaction or Western blotting. The constructs (4.9 kb to 0.3 kb) used for human HO-1 promoter analysis were described previously.25 Site-directed mutagenesis of the HO-1 promoter was performed by a polymerase chain reaction-based technique using the QuickChange XL Site-Directed Mutagenesis Kit (Stratagene, La Jolla, Calif). Electromobility shift assays were performed using the TNT reticulocyte lysate kit (Promega). For more details, please see the supplemental Materials available online at http://atvb.ahajournals.org.


*    Results
up arrowTop
 up arrowAbstract
 up arrowIntroduction
 up arrowMaterials and Methods
 *Results
down arrowDiscussion
 down arrowReferences
 
Ligands for PPAR{gamma} and PPAR{alpha} Induce HO-1 Expression in Human Vascular Cells
Activation of both PPAR{gamma} and PPAR{alpha} was shown to inhibit the inflammatory response in endothelial cells, VSMCs, and macrophages,3,26,27 and ligands for both receptors have been shown to reduce atherosclerosis in mice3–5 and humans.28,29 Therefore we determined the influence of ligands specific for either PPAR{alpha} (Wy-14,643 and fenofibrate) or PPAR{gamma} (rosiglitazone and troglitazone) on HO-1 expression in vascular cells. Western blot analysis (Figure 1A) and immunofluorescence (Figure 1B) demonstrated that both PPAR{alpha} and PPAR{gamma} ligands induced HO-1 expression in human umbilical venous endothelial cells (human umbilical venous endothelial cells) as well as in human VSMCs.


Figure 1
View larger version (45K):
[in this window]
[in a new window]

  		Figure 1. Induction of HO-1 protein expression by ligands for PPAR{alpha} and PPAR{gamma}. Human umbilical venous endothelial cells and human VSMCs were treated with Wy-14,643 (250 µmol/L), fenofibrate (200 µmol/L), rosiglitazone (5 µmol/L), or troglitazone (5 µmol/L) for 20 hours. A, Western blot analysis of HO-1 expression in extracts from human umbilical venous endothelial cells and human VSMCs. B, Immunofluorescence in human umbilical venous endothelial cells.

We found a maximum expression of HO-1 protein at 18 to 20 hours after incubation with PPAR ligands, and dose-response experiments revealed a concentration-dependent increase of HO-1 protein (Figure 2A) and mRNA (Figure 2B and 2C) after stimulation with the PPAR{alpha} ligand Wy-14,643 or the PPAR{gamma} ligand rosiglitazone in both human umbilical venous endothelial cells and VSMC.


Figure 2
View larger version (27K):
[in this window]
[in a new window]

  		Figure 2. HO-1 expression is induced in human umbilical venous endothelial cells and SMC by ligands for PPAR{alpha} and PPAR{gamma} in a dose-dependent manner. A, Western blot analysis in human umbilical venous endothelial cells and human VSMC that had been treated with Wy-14,643 or rosiglitazone at indicated concentrations for 20 hours. B, Measurement of HO-1 mRNA levels in human umbilical venous endothelial cells by quantitative polymerase chain reaction. C, Measurement of HO-1 mRNA levels in human VSMC by quantitative polymerase chain reaction. D, Measurement of PPAR{alpha} and PPAR{gamma} mRNA expression levels. *P<0.05 compared with the control value.

Quantification of PPAR{alpha} and PPAR{gamma} mRNA levels showed that PPAR{alpha} was the predominant PPAR isotype in both human umbilical venous endothelial cells and VSMC (Figure 2D). Accordingly, measurement of HO-1 mRNA levels in human umbilical venous endothelial cells and VSMC by quantitative polymerase chain reaction showed a marked increase in HO-1 mRNA after activation of PPAR{alpha} by Wy-14,643, whereas treatment with the PPAR{gamma}-specific ligand rosiglitazone induced HO-1 mRNA to a lesser extent (Figure 2B and 2C). Together, these data suggest that depending on the expression levels of the respective PPAR isotypes, PPAR{alpha} and PPAR{gamma} ligands induce HO-1 in vascular cells.

HO-1 Is a Direct PPAR Target Gene
To determine if HO-1 was a direct PPAR target gene, we analyzed the activity of a full-length (4.9 kb) human HO-1 promoter construct that had been previously cloned in our laboratory.25 HO-1 promoter activity was induced by cotransfection of expression vectors for PPAR{alpha} or PPAR{gamma} together with a plasmid encoding for retinoid X receptor-{alpha} in HEK293 cells followed by treatment with the respective PPAR ligand. Analysis of deletion mutants demonstrated that PPAR{alpha} or PPAR{gamma}-induced HO-1 promoter activity was dependent on a fragment located between –1.4 and –2.2 kb upstream of the transcription start site (Figure 3A). To narrow down the responsive promoter elements we created a second series of deletion mutants, which enabled us to locate the PPAR-responsive promoter region between –1740 kb and –1826 kb (Figure 3B). Bioinformatic analysis identified 2 putative PPAR-responsive elements (PPRE1 and PPRE2 GGGACAAAGGTTG and AGGTGAAAGGCCG) in DR1 configuration located within the 86-bp region.


Figure 3
View larger version (20K):
[in this window]
[in a new window]

  		Figure 3. Regulation of the human HO-1 promoter by PPAR{alpha} and PPAR{gamma}. A and B, HEK293 cells were transfected with HO-1 promoter luciferase reporter constructs containing promotor fragments with the indicated length or the full-length promoter and cotransfected with PPAR{alpha} or PPAR{gamma} and retinoid X receptor-{alpha} expression plasmids.

To confirm the functional relevance of the 2 identified PPREs, we introduced mutations in each of these elements, which abolished the PPAR{alpha}- or PPAR{gamma}-induced activity of the full-length HO-1 promoter construct (Figure 4A). Electrophoretic mobility shift assays showed that both PPRE1 and PPRE2 bound to in vitro-translated PPAR{alpha}/retinoid X receptor-{alpha} and PPAR{gamma}/retinoid X receptor-{alpha} heterodimers (Figure 4B), whereas we did not observe binding to retinoid X receptor-{alpha} homodimers (data not shown). The specificity of the bands was confirmed by competition with excess cold wild-type probes or cold probes containing point mutations. Together, these data demonstrate that HO-1 is a direct PPAR{alpha} or PPAR{gamma} target gene regulated via 2 PPREs.


Figure 4
View larger version (39K):
[in this window]
[in a new window]

  		Figure 4. Functional properties of identified PPREs. A, HEK293 cells were transfected with either the wild-type (wt) HO-1 promoter constructs or constructs with the indicated mutations of PPRE1 or PPRE2 and cotransfected with PPAR{alpha} or PPAR{gamma} and retinoid X receptor-{alpha} expression plasmids. HO-1 promotor activity was analyzed measuring firefly luciferase activity normalized to renilla luciferase activity. Activity in the absence of PPAR expression vectors was set to 1. B, Binding of in vitro-translated PPAR/retinoid X receptor heterodimers to the identified PPREs of the human HO-1 promoter was determined by electrophoretic mobility shift assay. Competition was performed using a 100-fold excess of nonlabeled oligonucleotides.

A Polymorphism Within the HO-1 Promoter Critically Affects Its Regulation by PPAR{alpha} or PPAR{gamma}
The HO-1 promoter is highly polymorphic within the human population in that the length of a proximal (GT)n dinucleotide repeat varies24 (Figure 5A). Longer repeats have been reported to negatively influence HO-1 expression and thus are associated with a higher risk for coronary artery disease and restenosis.30,31 To determine if the length of the GT-repeat affects the transcriptional regulation of the human HO-1 promoter by PPARs, we generated 3 full-length (4.9 kb) HO-1 promoter constructs differing in the length of their proximal GT-repeats (11, 24, and 29) by amplification of the respective DNA fragments in samples isolated from genotyped individuals. Subsequent analysis revealed a strong negative correlation between the PPAR{alpha}- or PPAR{gamma}-induced promoter activity and the increasing length of the GT-repeat within the HO-1 promoter (Figure 5B).


Figure 5
View larger version (20K):
[in this window]
[in a new window]

  		Figure 5. Influence of an HO-1 promoter polymorphism on PPAR-induced HO-1 promoter activity. A, Structure of the human HO-1 promoter. B, Promoter activity of full-length (4.9 kb) HO-1 promoter constructs carrying 11, 24, or 29 GT-repeats after cotransfection with PPAR{alpha}, PPAR{gamma}, and retinoid X receptor-{alpha} expression plasmids and PPAR ligand treatment (24 hours). HO-1 promoter activity was analyzed measuring firefly luciferase activity normalized to renilla luciferase activity. Activity in the absence of PPAR expression vectors was set to 1.

HO-1 Enzymatic Activity Contributes to the Anti-Inflammatory and Antiproliferative Effects of PPAR{alpha} in VSMCs
Ligands for PPAR{alpha} have been shown to inhibit the cytokine-induced expression of inflammatory genes in vascular cells.26,27 To examine the contribution of HO-1 to anti-inflammatory effects exerted by PPAR ligands, we analyzed tumor necrosis factor-induced cyclooxygenase-2 (COX-2) expression in VSMCs, which was inhibited by the PPAR{alpha} ligand Wy14,643 in a concentration-dependent manner (Figure 6A). Addition of the specific HO-1 inhibitor tin protoporphyrin (SnPP) abrogated the inhibition of COX-2 expression by Wy-14,643 (Figure 6A), demonstrating the involvement of HO-1 as a mediator of anti-inflammatory effects of PPAR ligands in VSMCs. In endothelial cells, in contrast, expression of COX-2 was not inhibited by Wy-14,643 (Figure 6B).


Figure 6
View larger version (15K):
[in this window]
[in a new window]

  		Figure 6. Inhibition of COX-2 expression by PPAR ligands is HO-1–dependent in SMCs. VSMCs (A) or human umbilical venous endothelial cells (B) were incubated with 150 µmol/L, 200 µmol/L, or 250 µmol/L Wy-14,643 in presence or absence of SnPP (5 µmol/L) for 18 hours. Cells were then stimulated with tumor necrosis factor-{alpha} (100 U/mL) for 5 hours. COX-2 protein expression was measured in cellular extracts by Western blot analysis.

Ligands for PPAR{alpha} have also been shown to inhibit smooth muscle cell (SMC) proliferation.32 Therefore, we investigated the effect of PPAR-induced HO-1 on SMC proliferation. Inhibition of HO-1 by zinc protoporphyrin dose-dependently increased SMC proliferation, induced by either platelet-derived growth factor or 15% fetal bovine serum. Moreover, treatment of SMCs with the PPAR{alpha}-ligand Wy-14,643 inhibited fetal bovine serum-induced proliferation, while concomitant inhibition of HO-1 with zinc protoprhyrin reversed this effect (supplemental Figure I, available online at http://atvb.ahajournals.com). Together these data demonstrate a HO-1–dependent and cell type-specific modulation of COX-2 expression and VSMC proliferation by a PPAR{alpha} ligand.


*    Discussion
up arrowTop
 up arrowAbstract
 up arrowIntroduction
 up arrowMaterials and Methods
 up arrowResults
 *Discussion
down arrowReferences
 
In addition to their beneficial influence on glucose and lipid metabolism, pharmacological PPAR ligands have been shown to exert direct anti-inflammatory and antiproliferative effects on cells of the vascular wall and hence inhibit the development of atherosclerosis and reduce intimal growth.3–9,11 Nevertheless, the exact mechanisms behind these versatile qualities of PPARs are poorly understood. One important anti-inflammatory mechanism of PPARs is transcriptional transrepression of inflammatory genes, which involves both a direct interaction with other transcription factors and competition for cofactors.12 Recently, it was shown that PPAR{gamma} inhibits transcription of proinflammatory genes by SUMOylation.33 In addition, upregulation of protective genes by PPARs may significantly contribute to their anti-inflammatory properties. With HO-1 we have identified a novel PPAR target gene that has been shown to exert multiple beneficial effects on cells of the vascular wall and to inhibit the development of atherosclerosis and restenosis in vivo.16–18,20

We show that ligands for PPAR{alpha} or PPAR{gamma} upregulate HO-1 expression in cultured human endothelial cells and VSMCs. Although it has been shown that the vast majority of Wy-14,643–induced genes are regulated in a PPAR{alpha}-dependent manner,35 Wy-14,643 can exert pan-PPAR agonistic activity.34 Therefore, we confirmed PPAR{alpha}-dependent induction of HO-1 expression in endothelial cells and VSMCs after treatment with another PPAR{alpha} activator, fenofibrate. PPAR{alpha} was the predominant PPAR isotype expressed in both human umbilical venous endothelial cells and SMCs, and PPAR{alpha} ligands seemed to be more effective than PPAR{gamma} ligands in inducing HO-1 expression.

PPAR ligands can also display receptor-independent effects,35,36 especially at high concentrations. Wy-14,643 is a fibric acid derivative with an EC50 of 5 µmol/L for human PPAR{alpha}.37 Although we demonstrate that Wy-14,643 at pharmacological doses significantly increases HO-1 expression, we also used higher doses (150 to 250 µmol/L), which likely have additional unspecific effects. However, the induction of HO-1 expression by receptor-specific concentrations of ligands as well as our promoter studies indicated a transcriptional regulation of HO-1 expression by PPAR{alpha} or PPAR{gamma}. Detailed analysis of the human HO-1 promoter showed that HO-1 is indeed a direct PPAR-target gene, whose transcription is regulated by both PPAR{alpha} and PPAR{gamma} via 2 PPREs.

Previous studies have highlighted the importance of stress-responsive elements (StREs) in the transcriptional regulation of HO-1.38 The repressor protein Bach1 constitutively binds to StREs and is displaced on activation of transcription factors such as Nrf2.39 15-deoxy-delta12,14 prostaglandin J2, which exerts multiple PPAR-dependent and PPAR-independent effects,36 has been shown to activate the Nrf2 regulatory pathway via Keap140 and to induce HO-1 expression via StREs in an PPAR-independent manner.41–44 However, prostaglandin J2-induced transcription of glutathione S-transferase involves synergistic activation of Nrf2 and PPAR{gamma}.45 In our present study the StRE-containing enhancer region of the human HO-1 promoter25,38 was dispensable for the PPAR-induced transcriptional regulation of the HO-1 promoter, because the 3.8-kb and 2.2-kb HO-1 promoter constructs, which lack this enhancer, but include the PPREs, were still PPAR-responsive. Moreover, mutation of the StREs did not affect PPAR-induced HO-1 promoter activity (data not shown). With the 2 identified PPREs located between the StREs and the basic HO-1 promoter, it will be nevertheless interesting to determine a possible influence of the Bach1/Nrf2 system on the PPAR-mediated transcriptional regulation of HO-1 and to analyze the accessibility of the 2 PPREs under various conditions of cellular stress.

We demonstrate that PPAR-induced HO-1 promoter activity inversely correlates with the length of a polymorphic GT-repeat in the human HO-1 promoter. This polymorphism has been reported to affect hydrogen peroxide-induced HO-1 promoter activity as well as HO-1 mRNA expression and enzymatic activity in lymphoblastoid cells.46,47 It has been suggested that longer GT-repeats promote changes in DNA conformation, which in turn negatively affect transcriptional activity.24 Moreover, the length of the respective GT-repeat is associated with the susceptibility for various inflammatory diseases including coronary heart disease and restenosis.30,31 It will be important to determine whether the vasculoprotective effects of PPAR ligands correlate with this HO-1 promoter polymorphism, which would provide a possibility to better assess the individual benefit of the treatment with fibrates or thiazolidinediones.

Both PPAR ligands and HO-1 have been described to block the inflammatory response in vascular cells and to potently inhibit VSMC proliferation.6,8,17,21,26,27,32,48–50 Furthermore, inhibition of COX-2 expression and prostaglandin synthesis by HO-1 was reported.51–56 We determined the contribution of HO-1 to the described anti-inflammatory and antiproliferative effects of PPAR{alpha} ligands in VSMCs27 and show that inhibition of HO-1 enzymatic activity by zinc protoprhyrin abrogated the inhibitory effect of PPAR{alpha} ligands on cytokine-induced expression of COX-2 as well as on proliferation in VSMCs. Interestingly, Wy-14,643–induced HO-1 did not affect COX-2 expression in endothelial cells, and while the expression of vascular cell adhesion molecule-1 in endothelial cells was strongly inhibited by the PPAR{alpha} ligand, inhibition of HO-1 did not reverse this effect (data not shown). Moreover, we show that treatment of SMCs with the PPAR{alpha}-ligand Wy-14,643 inhibited platelet-derived growth factor and serum-induced proliferation, while concomitant inhibition of HO-1 reversed this effect. Interestingly, it has been shown that upregulation of HO-1 in endothelial cells increases proliferation.57 These data strongly indicate cell type specific mechanistic differences in the anti-inflammatory action of PPAR ligand-induced HO-1.

Since PPARs have been recognized as key transcription factors regulating the expression of genes involved in lipid and glucose metabolism, HO-1 represents a novel type of PPAR target gene. The finding that HO-1 expression is transcriptionally regulated by PPAR{alpha} or PPAR{gamma} provides an important additional link between these ligand-activated transcription factors and their described anti-inflammatory and antiproliferative properties and thus contributes to our understanding of the pleiotropic beneficial effects exerted by PPAR ligands in inflammatory vascular disorders.


*    Acknowledgments
 
Sources of Funding

This work was supported in part by National Institutes of Health Grant RO1 HL084422-01 (to N.L.), by the Partner’s Fund at the University of Virginia (to N.L.), the Sixth Framework Programme of the European Community LSHM-CT-2004-005206, and the European Network of Excellence EVGN LSHM-CT-2003-503254 (to V.B. and B.R.B.). G.K. and A.K. were supported by Max Kade Fellowships.

Disclosures

None.


*    Footnotes
 
Original received August 15, 2006; final version accepted March 15, 2007.


*    References
up arrowTop
 up arrowAbstract
 up arrowIntroduction
 up arrowMaterials and Methods
 up arrowResults
 up arrowDiscussion
 *References
 

  1. Evans RM, Barish GD, Wang YX. PPARs and the complex journey to obesity. Nat Med. 2004; 10: 355–361.[CrossRef][Medline] [Order article via Infotrieve]
  2. Lee CH, Olson P, Evans RM. Minireview: lipid metabolism, metabolic diseases, and peroxisome proliferator-activated receptors. Endocrinology. 2003; 144: 2201–2207.[Abstract/Free Full Text]
  3. Li AC, Binder CJ, Gutierrez A, Brown KK, Plotkin CR, Pattison JW, Valledor AF, Davis RA, Willson TM, Witztum JL, Palinski W, Glass CK. Differential inhibition of macrophage foam-cell formation and atherosclerosis in mice by PPARalpha, beta/delta, and gamma. J Clin Invest. 2004; 114: 1564–1576.[CrossRef][Medline] [Order article via Infotrieve]
  4. Li AC, Brown KK, Silvestre MJ, Willson TM, Palinski W, Glass CK. Peroxisome proliferator-activated receptor gamma ligands inhibit development of atherosclerosis in LDL receptor-deficient mice. J Clin Invest. 2000; 106: 523–531.[Medline] [Order article via Infotrieve]
  5. Duez H, Chao YS, Hernandez M, Torpier G, Poulain P, Mundt S, Mallat Z, Teissier E, Burton CA, Tedgui A, Fruchart JC, Fievet C, Wright SD, Staels B. Reduction of atherosclerosis by the peroxisome proliferator- activated receptor alpha agonist fenofibrate in mice. J Biol Chem. 2002; 277: 48051–48057.[Abstract/Free Full Text]
  6. Marx N, Duez H, Fruchart JC, Staels B. Peroxisome proliferator-activated receptors and atherogenesis: regulators of gene expression in vascular cells. Circ Res. 2004; 94: 1168–1178.[Abstract/Free Full Text]
  7. Phillips JW, Barringhaus KG, Sanders JM, Yang Z, Chen M, Hesselbacher S, Czarnik AC, Ley K, Nadler J, Sarembock IJ. Rosiglitazone reduces the accelerated neointima formation after arterial injury in a mouse injury model of type 2 diabetes. Circulation. 2003; 108: 1994–1999.[Abstract/Free Full Text]
  8. 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]
  9. Kooistra T, Verschuren L, van der WJ, Koenig W, Toet K, Princen HM, Kleemann R. Fenofibrate reduces atherogenesis in apoE*3 Leiden mice. Evidence for multiple antiatherogenic effects besides lowering plasma cholesterol. Arterioscler Thromb Vasc Biol. 2006; 26: 2322–2330.[Abstract/Free Full Text]
  10. Kleemann R, Verschuren L, de Rooij BJ, Lindeman J, de Maat MM, Szalai AJ, Princen HM, Kooistra T. Evidence for anti-inflammatory activity of statins and PPARalpha activators in human C-reactive protein transgenic mice in vivo and in cultured human hepatocytes in vitro. Blood. 2004; 103: 4188–4194.[Abstract/Free Full Text]
  11. Kleemann R, Princen HM, Emeis JJ, Jukema JW, Fontijn RD, Horrevoets AJ, Kooistra T, Havekes LM. Rosuvastatin reduces atherosclerosis development beyond and independent of its plasma cholesterol-lowering effect in APOE*3-Leiden transgenic mice: evidence for antiinflammatory effects of rosuvastatin. Circulation. 2003; 108: 1368–1374.[Abstract/Free Full Text]
  12. Daynes RA, Jones DC. Emerging roles of PPARs in inflammation and immunity. Nat Rev Immunol. 2002; 2: 748–759.[CrossRef][Medline] [Order article via Infotrieve]
  13. Girnun GD, Domann FE, Moore SA, Robbins ME. Identification of a functional peroxisome proliferator-activated receptor response element in the rat catalase promoter. Mol Endocrinol. 2002; 16: 2793–2801.[Abstract/Free Full Text]
  14. Yoo HY, Chang MS, Rho HM. Induction of the rat Cu/Zn superoxide dismutase gene through the peroxisome proliferator-responsive element by arachidonic acid. Gene. 1999; 234: 87–91.[CrossRef][Medline] [Order article via Infotrieve]
  15. Ishikawa K, Navab M, Leitinger N, Fogelman AM, Lusis AJ. Induction of heme oxygenase-1 inhibits the monocyte transmigration induced by mildly oxidized LDL. J Clin Invest. 1997; 100: 1209–1216.[Medline] [Order article via Infotrieve]
  16. Ishikawa K, Sugawara D, Wang X, Suzuki K, Itabe H, Maruyama Y, Lusis AJ. Heme oxygenase-1 inhibits atherosclerotic lesion formation in ldl-receptor knockout mice. Circ Res. 2001; 88: 506–512.[Abstract/Free Full Text]
  17. Otterbein LE, Zuckerbraun BS, Haga M, Liu F, Song R, Usheva A, Stachulak C, Bodyak N, Smith RN, Csizmadia E, Tyagi S, Akamatsu Y, Flavell RJ, Billiar TR, Tzeng E, Bach FH, Choi AM, Soares MP. Carbon monoxide suppresses arteriosclerotic lesions associated with chronic graft rejection and with balloon injury. Nat Med. 2003; 9: 183–190.[CrossRef][Medline] [Order article via Infotrieve]
  18. Juan SH, Lee TS, Tseng KW, Liou JY, Shyue SK, Wu KK, Chau LY. Adenovirus-mediated heme oxygenase-1 gene transfer inhibits the development of atherosclerosis in apolipoprotein E-deficient mice. Circulation. 2001; 104: 1519–1525.[Abstract/Free Full Text]
  19. Togane Y, Morita T, Suematsu M, Ishimura Y, Yamazaki JI, Katayama S. Protective roles of endogenous carbon monoxide in neointimal development elicited by arterial injury. Am J Physiol Heart Circ Physiol. 2000; 278: H623–H632.[Abstract/Free Full Text]
  20. Chen S, Kapturczak MH, Wasserfall C, Glushakova OY, Campbell-Thompson M, Deshane JS, Joseph R, Cruz PE, Hauswirth WW, Madsen KM, Croker BP, Berns KI, Atkinson MA, Flotte TR, Tisher CC, Agarwal A. Interleukin 10 attenuates neointimal proliferation and inflammation in aortic allografts by a heme oxygenase-dependent pathway. Proc Natl Acad Sci U S A. 2005; 102: 7251–7256.[Abstract/Free Full Text]
  21. Otterbein LE, Soares MP, Yamashita K, Bach FH. Heme oxygenase-1: unleashing the protective properties of heme. Trends Immunol. 2003; 24: 449–455.[CrossRef][Medline] [Order article via Infotrieve]
  22. Nakao A, Murase N, Ho C, Toyokawa H, Billiar TR, Kanno S. Biliverdin administration prevents the formation of intimal hyperplasia induced by vascular injury. Circulation. 2005; 112: 587–591.[Abstract/Free Full Text]
  23. Ollinger R, Bilban M, Erat A, Froio A, McDaid J, Tyagi S, Csizmadia E, Graca-Souza AV, Liloia A, Soares MP, Otterbein LE, Usheva A, Yamashita K, Bach FH. Bilirubin: a natural inhibitor of vascular smooth muscle cell proliferation. Circulation. 2005; 112: 1030–1039.[Abstract/Free Full Text]
  24. Exner M, Minar E, Wagner O, Schillinger M. The role of heme oxygenase-1 promoter polymorphisms in human disease. Free Radic Biol Med. 2004; 37: 1097–1104.[CrossRef][Medline] [Order article via Infotrieve]
  25. Kronke G, Bochkov VN, Huber J, Gruber F, Bluml S, Furnkranz A, Kadl A, Binder BR, Leitinger N. Oxidized phospholipids induce expression of human heme oxygenase-1 involving activation of cAMP-responsive element-binding protein. J Biol Chem. 2003; 278: 51006–51014.[Abstract/Free Full Text]
  26. Marx N, Sukhova GK, Collins T, Libby P, Plutzky J. PPARalpha activators inhibit cytokine-induced vascular cell adhesion molecule-1 expression in human endothelial cells. Circulation. 1999; 99: 3125–3131.[Abstract/Free Full Text]
  27. Staels B, Koenig W, Habib A, Merval R, Lebret M, Torra IP, Delerive P, Fadel A, Chinetti G, Fruchart JC, Najib J, Maclouf J, Tedgui A. Activation of human aortic smooth-muscle cells is inhibited by PPARalpha but not by PPARgamma activators. Nature. 1998; 393: 790–793.[CrossRef][Medline] [Order article via Infotrieve]
  28. Ericsson CG, Nilsson J, Grip L, Svane B, Hamsten A. Effect of bezafibrate treatment over five years on coronary plaques causing 20% to 50% diameter narrowing (The Bezafibrate Coronary Atherosclerosis Intervention Trial [Bendothelial cellsAIT]). Am J Cardiol. 1997; 80: 1125–1129.[CrossRef][Medline] [Order article via Infotrieve]
  29. Minamikawa J, Tanaka S, Yamauchi M, Inoue D, Koshiyama H. Potent inhibitory effect of troglitazone on carotid arterial wall thickness in type 2 diabetes. J Clin Endocrinol Metab. 1998; 83: 1818–1820.[Abstract/Free Full Text]
  30. Chen YH, Chau LY, Lin MW, Chen LC, Yo MH, Chen JW, Lin SJ. Heme oxygenase-1 gene promotor microsatellite polymorphism is associated with angiographic restenosis after coronary stenting. Eur Heart J. 2004; 25: 39–47.[Abstract/Free Full Text]
  31. Chen YH, Lin SJ, Lin MW, Tsai HL, Kuo SS, Chen JW, Charng MJ, Wu TC, Chen LC, Ding YA, Pan WH, Jou YS, Chau LY. Microsatellite polymorphism in promoter of heme oxygenase-1 gene is associated with susceptibility to coronary artery disease in type 2 diabetic patients. Hum Genet. 2002; 111: 1–8.[CrossRef][Medline] [Order article via Infotrieve]
  32. Gizard F, Amant C, Barbier O, Bellosta S, Robillard R, Percevault F, Sevestre H, Krimpenfort P, Corsini A, Rochette J, Glineur C, Fruchart JC, Torpier G, Staels B. PPAR alpha inhibits vascular smooth muscle cell proliferation underlying intimal hyperplasia by inducing the tumor suppressor p16INK4a. J Clin Invest. 2005; 115: 3228–3238.[CrossRef][Medline] [Order article via Infotrieve]
  33. Pascual G, Fong AL, Ogawa S, Gamliel A, Li AC, Perissi V, Rose DW, Willson TM, Rosenfeld MG, Glass CK. A SUMOylation-dependent pathway mediates transrepression of inflammatory response genes by PPAR-gamma. Nature. 2005; 437: 759–763.[CrossRef][Medline] [Order article via Infotrieve]
  34. Anderson SP, Howroyd P, Liu J, Qian X, Bahnemann R, Swanson C, Kwak MK, Kensler TW, Corton JC. The transcriptional response to a peroxisome proliferator-activated receptor alpha agonist includes increased expression of proteome maintenance genes. J Biol Chem. 2004; 279: 52390–52398.[Abstract/Free Full Text]
  35. Chawla A, Barak Y, Nagy L, Liao D, Tontonoz P, Evans RM. PPAR-gamma 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]
  36. Straus DS, Pascual G, Li M, Welch JS, Ricote M, Hsiang CH, Sengchanthalangsy LL, Ghosh G, Glass CK. 15-deoxy-delta 12,14-prostaglandin J2 inhibits multiple steps in the NF-kappa B signaling pathway. Proc Natl Acad Sci U S A. 2000; 97: 4844–4849.[Abstract/Free Full Text]
  37. Willson TM, Brown PJ, Sternbach DD, Henke BR. The PPARs: from orphan receptors to drug discovery. J Med Chem. 2000; 43: 527–550.[CrossRef][Medline] [Order article via Infotrieve]
  38. Alam J, Cook JL. Transcriptional regulation of the heme oxygenase-1 gene via the stress response element pathway. Curr Pharm Des. 2003; 9: 2499–2511.[CrossRef][Medline] [Order article via Infotrieve]
  39. Sun J, Brand M, Zenke Y, Tashiro S, Groudine M, Igarashi K. Heme regulates the dynamic exchange of Bach1 and NF-E2-related factors in the Maf transcription factor network. Proc Natl Acad Sci U S A. 2004; 101: 1461–1466.[Abstract/Free Full Text]
  40. Hosoya T, Maruyama A, Kang MI, Kawatani Y, Shibata T, Uchida K, Warabi E, Noguchi N, Itoh K, Yamamoto M. Differential responses of the Nrf2-Keap1 system to laminar and oscillatory shear stresses in endothelial cells. J Biol Chem. 2005; 280: 27244–27250.[Abstract/Free Full Text]
  41. Colville-Nash PR, Qureshi SS, Willis D, Willoughby DA. Inhibition of inducible nitric oxide synthase by peroxisome proliferator-activated receptor agonists: correlation with induction of heme oxygenase 1. J Immunol. 1998; 161: 978–984.[Abstract/Free Full Text]
  42. Gong P, Stewart D, Hu B, Li N, Cook J, Nel A, Alam J. Activation of the mouse heme oxygenase-1 gene by 15-deoxy-Delta(12,14)-prostaglandin J(2) is mediated by the stress response elements and transcription factor Nrf2. Antioxid Redox Signal. 2002; 4: 249–257.[CrossRef][Medline] [Order article via Infotrieve]
  43. varez-Maqueda M, El BR, Alba G, Monteseirin J, Chacon P, Vega A, Martin-Nieto J, Bedoya FJ, Pintado E, Sobrino F. 15-deoxy-delta 12,14-prostaglandin J2 induces heme oxygenase-1 gene expression in a reactive oxygen species-dependent manner in human lymphocytes. J Biol Chem. 2004; 279: 21929–21937.[Abstract/Free Full Text]
  44. Liu JD, Tsai SH, Lin SY, Ho YS, Hung LF, Pan S, Ho FM, Lin CM, Liang YC. Thiol antioxidant and thiol-reducing agents attenuate 15-deoxy-delta 12,14-prostaglandin J2-induced heme oxygenase-1 expression. Life Sci. 2004; 74: 2451–2463.[CrossRef][Medline] [Order article via Infotrieve]
  45. Park EY, Cho IJ, Kim SG. Transactivation of the PPAR-responsive enhancer module in chemopreventive glutathione S-transferase gene by the peroxisome proliferator-activated receptor-gamma and retinoid X receptor heterodimer. Cancer Res. 2004; 64: 3701–3713.[Abstract/Free Full Text]
  46. Yamada N, Yamaya M, Okinaga S, Nakayama K, Sekizawa K, Shibahara S, Sasaki H. Microsatellite polymorphism in the heme oxygenase-1 gene promoter is associated with susceptibility to emphysema. Am J Hum Genet. 2000; 66: 187–195.[CrossRef][Medline] [Order article via Infotrieve]
  47. Hirai H, Kubo H, Yamaya M, Nakayama K, Numasaki M, Kobayashi S, Suzuki S, Shibahara S, Sasaki H. Microsatellite polymorphism in heme oxygenase-1 gene promoter is associated with susceptibility to oxidant-induced apoptosis in lymphoblastoid cell lines. Blood. 2003; 102: 1619–1621.[Abstract/Free Full Text]
  48. Ollinger R, Bilban M, Erat A, Froio A, McDaid J, Tyagi S, Csizmadia E, Graca-Souza AV, Liloia A, Soares MP, Otterbein LE, Usheva A, Yamashita K, Bach FH Bilirubin. A natural inhibitor of vascular smooth muscle cell proliferation. Circulation. 2005; 112: 1030–1039.[Abstract/Free Full Text]
  49. Soares MP, Seldon MP, Gregoire IP, Vassilevskaia T, Berberat PO, Yu J, Tsui TY, Bach FH. Heme oxygenase-1 modulates the expression of adhesion molecules associated with endothelial cell activation. J Immunol. 2004; 172: 3553–3563.[Abstract/Free Full Text]
  50. Lee TS, Chang CC, Zhu Y, Shyy JY. Simvastatin induces heme oxygenase-1: a novel mechanism of vessel protection. Circulation. 2004; 110: 1296–1302.[Abstract/Free Full Text]
  51. Kobayashi H, Takeno M, Saito T, Takeda Y, Kirino Y, Noyori K, Hayashi T, Ueda A, Ishigatsubo Y. Regulatory role of heme oxygenase 1 in inflammation of rheumatoid arthritis. Arthritis Rheum. 2006; 54: 1132–1142.[CrossRef][Medline] [Order article via Infotrieve]
  52. Abraham NG. Heme oxygenase attenuated angiotensin II-mediated increase in cyclooxygenase activity and decreased isoprostane F2alpha in endothelial cells. Thromb Res. 2003; 110: 305–309.[CrossRef][Medline] [Order article via Infotrieve]
  53. Li VG, Seta F, Schwartzman ML, Nasjletti A, Abraham NG. Heme oxygenase attenuates angiotensin II-mediated increase in cyclooxygenase-2 activity in human femoral endothelial cells. Hypertension. 2003; 41: 715–719.[Abstract/Free Full Text]
  54. Botros FT, Laniado-Schwartzman M, Abraham NG. Regulation of cyclooxygenase- and cytochrome p450-derived eicosanoids by heme oxygenase in the rat kidney. Hypertension. 2002; 39: 639–644.[Abstract/Free Full Text]
  55. Haider A, Olszanecki R, Gryglewski R, Schwartzman ML, Lianos E, Kappas A, Nasjletti A, Abraham NG. Regulation of cyclooxygenase by the heme-heme oxygenase system in microvessel endothelial cells. J Pharmacol Exp Ther. 2002; 300: 188–194.[Abstract/Free Full Text]
  56. Botros FT, Schwartzman ML, Stier CT, Jr., Goodman AI, Abraham NG. Increase in heme oxygenase-1 levels ameliorates renovascular hypertension. Kidney Int. 2005; 68: 2745–2755.[CrossRef][Medline] [Order article via Infotrieve]
  57. Bussolati B, Ahmed A, Pemberton H, Landis RC, Di CF, Haskard DO, Mason JC. Bifunctional role for VEGF-induced heme oxygenase-1 in vivo: induction of angiogenesis and inhibition of leukocytic infiltration. Blood. 2004; 103: 761–766.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 J. Lipid Res.Home page
S. J. Peterson, G. Drummond, D. H. Kim, M. Li, A. L. Kruger, S. Ikehara, and N. G. Abraham
L-4F treatment reduces adiposity, increases adiponectin levels, and improves insulin sensitivity in obese mice
J. Lipid Res., August 1, 2008; 49(8): 1658 - 1669.
[Abstract] [Full Text] [PDF]

Home page
 Circ. Res.Home page
H.-K. Jyrkkanen, E. Kansanen, M. Inkala, A. M. Kivela, H. Hurttila, S. E. Heinonen, G. Goldsteins, S. Jauhiainen, S. Tiainen, H. Makkonen, et al.
Nrf2 Regulates Antioxidant Gene Expression Evoked by Oxidized Phospholipids in Endothelial Cells and Murine Arteries In Vivo
Circ. Res., July 3, 2008; 103(1): e1 - e9.
[Abstract] [Full Text] [PDF]

Home page
 J. Pharmacol. Exp. Ther.Home page
D. H. Kim, A. P. Burgess, M. Li, P. L. Tsenovoy, F. Addabbo, J. A. McClung, N. Puri, and N. G. Abraham
Heme Oxygenase-Mediated Increases in Adiponectin Decrease Fat Content and Inflammatory Cytokines Tumor Necrosis Factor-{alpha} and Interleukin-6 in Zucker Rats and Reduce Adipogenesis in Human Mesenchymal Stem Cells
J. Pharmacol. Exp. Ther., June 1, 2008; 325(3): 833 - 840.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
M. Li, D. H. Kim, P. L. Tsenovoy, S. J. Peterson, R. Rezzani, L. F. Rodella, W. S. Aronow, S. Ikehara, and N. G. Abraham
Treatment of Obese Diabetic Mice With a Heme Oxygenase Inducer Reduces Visceral and Subcutaneous Adiposity, Increases Adiponectin Levels, and Improves Insulin Sensitivity and Glucose Tolerance
Diabetes, June 1, 2008; 57(6): 1526 - 1535.
[Abstract] [Full Text] [PDF]

Home page
 Cardiovasc ResHome page
P. J.H. Smeets, B. E.J. Teunissen, P. H.M. Willemsen, F. A. van Nieuwenhoven, A. E. Brouns, B. J.A. Janssen, J. P.M. Cleutjens, B. Staels, G. J. van der Vusse, and M. van Bilsen
Cardiac hypertrophy is enhanced in PPAR{alpha}-/- mice in response to chronic pressure overload
Cardiovasc Res, April 1, 2008; 78(1): 79 - 89.
[Abstract] [Full Text] [PDF]

Home page
 Pharmacol. Rev.Home page
N. G. Abraham and A. Kappas
Pharmacological and Clinical Aspects of Heme Oxygenase
Pharmacol. Rev., March 1, 2008; 60(1): 79 - 127.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Data Supplement
Right arrow All Versions of this Article:
27/6/1276    most recent
ATVBAHA.107.142638v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Krönke, G.
Right arrow Articles by Leitinger, N.