A2E-induced inflammation and angiogenesis in RPE cells in vitro are modulated by PPAR-α, -ß/δ, -γ, and RXR antagonists and by norbixin.

N-retinylidene-N-retinylethanolamine (A2E) plays a central role in age-related macular degeneration (AMD) by inducing angiogenesis and inflammation. A2E effects are mediated at least partly via the retinoic acid receptor (RAR)-α. Here we show that A2E binds and transactivates also peroxisome proliferator-activated receptors (PPAR) and retinoid X receptors (RXR). 9'-cis-norbixin, a di-apocarotenoid is also a ligand of these nuclear receptors (NR). Norbixin inhibits PPAR and RXR transactivation induced by A2E. Moreover, norbixin reduces protein kinase B (AKT) phosphorylation, NF-κB and AP-1 transactivation and mRNA expression of the inflammatory interleukins (IL) -6 and -8 and of vascular endothelial growth factor (VEGF) enhanced by A2E. By contrast, norbixin increases matrix metalloproteinase 9 (MMP9) and C-C motif chemokine ligand 2 (CCL2) mRNA expression in response to A2E. Selective PPAR-α, -ß/δ and -γ antagonists inhibit the expression of IL-6 and IL-8 while only the antagonist of PPAR-γ inhibits the transactivation of NF-κB following A2E exposure. In addition, a cocktail of all three PPARs antagonists and also HX531, an antagonist of RXR reproduce norbixin effects on inflammation. Altogether, A2E's deleterious biological effects could be inhibited through PPAR and RXR regulation. Moreover, the modulation of these NR by norbixin may open new avenues for the treatment of AMD.

Nuclear factor kappa B/p65 plays a positive role in peroxisome proliferator-activated receptor δ expression in orange-spotted grouper Epinephelus coioides.

The transcription factor nuclear factor kappa B (NF-κB) is a critical regulator of immune and inflammatory responses with crucial roles in various pathophysiologic conditions involving cell survival and death. Recent studies in mammals showed that NF-κB was also involved in peroxisome proliferator-activated receptors (PPARs)-mediated immune responses However, the mechanism by which NF-κB regulates PPARδ in teleosts remains unclear. In the present study, we analyzed the potential role of NF-κB/p65 (Ecp65) in the immune response stimulated by various pathogens in the grouper Epinephelus coioides. Ecp65 expression was significantly induced soon after infection with lipopolysaccharide, nervous necrosis virus, poly(I:C), and zymosan A. We also analyzed the promoter to determine the regulatory effect of Ecp65 on PPARδ expression, using progressive EcPPARδ promoter deletion mutations. Among the five truncated mutants, the luciferase reporter activity of the PPARδ-5 promoter region was highest in response to Ecp65, indicating that the core p65-binding region was located in the PPARδ-5 promoter region (+122 bp to +383 bp). Mutation analyses indicated that the luciferase reporter activity of the EcPPARδ promoter was dramatically decreased by mutation of the M3 (+305 bp to +324 bp) and M4 (+346 bp to +365 bp) binding sites, respectively. We further confirmed that Ecp65 bound to the M3 and M4 binding sites in the 5'-untranslated region of EcPPARδ by electrophoretic mobility shift assay. Finally, overexpression of Ecp65 in vitro notably promoted the transcription of EcPPARδ, interferon-related genes, and several inflammatory cytokines. This study demonstrated that Ecp65 plays an important role in modulating the innate immune responses in groupers. These results also further our understanding of the mechanisms involved in the transcriptional regulation of PPARs by p65 in bony fish.

SIRT4 silencing in tumor-associated macrophages promotes HCC development via PPARδ signalling-mediated alternative activation of macrophages.

BACKGROUND: The activation of tumour-associated macrophages (TAMs) contributes to the progression of hepatocellular carcinoma (HCC). SIRT4 acts as a tumour suppressor of tumour growth by regulating cell metabolism, inflammation, and anti-tumourigenesis. However, the involvement of SIRT4 in the activation of TAMs is unknown. Based on previous findings, the expression of SIRT4 in distinct groups of TAMs as well as the effect of SIRT4 silencing on macrophage polarization was investigated. METHODS: The expression of SIRT4 in HCC tissues and peritumour tissues was tested by qRT-PCR, western blotting and histological analysis. A Kaplan-Meier survival curve was generated based on the expression of SIRT4 in the HCC samples. Next, immunofluorescence staining was used to evaluate distinct groups of TAMs in human HCC samples, and the expression of SIRT4 in M1 and M2 TAMs was examined by flow cytometry. A homograft mouse model was used to assess the effect of SIRT4 silencing in TAMs on the development of HCC cells. RESULTS: SIRT4 was significantly downregulated in HCC tumour tissues, and the expression of SIRT4 in peritumour tissues was positively associated with survival in patients. We further found that downregulation of SIRT4 was associated with increased macrophage infiltration and a high ratio of M2/M1 macrophages in HCC peritumour tissues. Using gene interference, we found that SIRT4 silencing in TAMs significantly modulated the alternative activation of macrophages and promoted in vitro and in vivo HCC cell growth. Mechanistically, we revealed that HCM restricted the expression of SIRT4 in macrophages and promoted alternative activation of macrophages via the FAO-PPARδ-STAT3 axis. Furthermore, we also revealed that elevated MCP-1 expression induced by SIRT4 downregulation was responsible for increased TAM infiltration in peritumour tissues. CONCLUSIONS: Overall, our results demonstrate that downregulation of SIRT4 in TAMs modulates the alternative activation of macrophages and promotes HCC development via the FAO-PPARδ-STAT3 axis. These results could provide a new therapeutic target for the treatment of HCC.

Gymnemic acid alleviates inflammation and insulin resistance via PPARδ- and NFκB-mediated pathways in db/db mice.

Gymnemic acid (GA) is a naturally occurring herbal ingredient that improves glucose metabolism in patients with diabetes mellitus. In this study, we evaluated the ameliorative effects of GA on obesity-induced inflammation and insulin resistance (IR), and identified the mechanisms for these effects in db/db mice. In these mice, GA effectively lowered fasting blood glucose concentrations from 26.3 ± 4.09 to 17.4 ± 3.38 mmol L-1, and improved oral glucose and insulin tolerance. Furthermore, GA treatment accelerated lipid transport and promoted fatty acid oxidation, which reduced lipid accumulation and inhibited expression of inflammatory cytokines, including those involved in the proliferator-activated receptor δ (PPARδ)- and nuclear factor κB (NFκB)-mediated signaling pathways. In addition, the anti-inflammatory effects increased the ratio of insulin to glucagon. It also regulated the insulin signal transduction with reduced phosphorylation of IRS-1 (Ser) and increased phosphorylation of IRS (Tyr) in liver, skeletal muscle and adipose tissue. In summary, we demonstrated in db/db mice that GA induces fatty acid oxidation, and alleviates inflammation and IR in liver, skeletal muscle and adipose tissue through PPARδ- and NFκB-mediated signaling pathways.

PPAR-δ of orange-spotted grouper exerts antiviral activity against fish virus and regulates interferon signaling and inflammatory factors.

Peroxisome proliferator-activated receptor δ (PPAR-δ), also called PPAR-ß or PPAR-ß/δ, is a member of the peroxisome proliferator-activated receptor (PPAR) family, which belongs to the nuclear steroid receptor superfamily. Activated PPARs participate in the regulation of lipid and glucose metabolism and also affect cellular proliferation, differentiation, and apoptosis, and the immune responses. To investigate the roles of PPAR-δ in Singapore grouper iridovirus (SGIV) infection, we cloned and characterized the gene encoding a PPAR-δ homologue from the orange-spotted grouper, Epinephelus coioides (EcPPAR-δ). EcPPAR-δ encodes a 514-amino-acid polypeptide, with 95.29% and 74.76% homologue to the Seriola dumerili and human proteins, respectively. EcPPAR-δ contains a typical DNA-binding domain and a ligand-binding domain. Its expression was induced by SGIV infection in vitro. A subcellular localization analysis showed that EcPPAR-δ localizes throughout the cytoplasm and nucleus, with a diffuse intracellular expression pattern. SGIV replication was reduced by EcPPAR-δ overexpression, which was evident in the reduced severity of the cytopathic effect, reduced viral gene transcription, and the reduced expression of the viral capsid protein. The replication of SGIV increased with the knockdown of EcPPAR-δ. The overexpression and silencing of EcPPAR-δ in grouper spleen cells showed that EcPPAR-δ plays a positive role in the regulation of the interferon signaling pathway, but has an anti-inflammatory effect on the inflammatory response. The anti-inflammatory effect of EcPPAR-δ may be related to its function in maintaining cell homeostasis. Because the interferon signaling pathway plays an important role in antiviral immune responses, we speculate that the activation of the interferon signaling pathway by EcPPAR-δ overexpression underlies its inhibitory effect on SGIV replication. Together, our data greatly extend our understanding of the roles of the EcPPAR-δ family members in the pathogenesis of fish viruses.

The Role of PPAR-δ in Metabolism, Inflammation, and Cancer: Many Characters of a Critical Transcription Factor.

Peroxisome proliferator-activated receptor-delta (PPAR-δ), one of three members of the PPAR group in the nuclear receptor superfamily, is a ligand-activated transcription factor. PPAR-δ regulates important cellular metabolic functions that contribute to maintaining energy balance. PPAR-δ is especially important in regulating fatty acid uptake, transport, and ß-oxidation as well as insulin secretion and sensitivity. These salutary PPAR-δ functions in normal cells are thought to protect against metabolic-syndrome-related diseases, such as obesity, dyslipidemia, insulin resistance/type 2 diabetes, hepatosteatosis, and atherosclerosis. Given the high clinical burden these diseases pose, highly selective synthetic activating ligands of PPAR-δ were developed as potential preventive/therapeutic agents. Some of these compounds showed some efficacy in clinical trials focused on metabolic-syndrome-related conditions. However, the clinical development of PPAR-δ agonists was halted because various lines of evidence demonstrated that cancer cells upregulated PPAR-δ expression/activity as a defense mechanism against nutritional deprivation and energy stresses, improving their survival and promoting cancer progression. This review discusses the complex relationship between PPAR-δ in health and disease and highlights our current knowledge regarding the different roles that PPAR-δ plays in metabolism, inflammation, and cancer.

Dose-dependent effects of peroxisome proliferator-activated receptors ß/δ agonist on systemic inflammation after haemorrhagic shock.

INTRODUCTION: PPARß/δ agonists are known to modulate the systemic inflammatory response after sepsis. In this study, inflammation modulation effects of PPARß/δ are investigated using the selective PPARß/δ agonist (GW0742) in a model of haemorrhagic shock (HS)-induced sterile systemic inflammation. METHODS: Blood pressure-controlled (35±5mmHg) HS was performed in C57/BL6 mice for 90min. Low-dose GW0742 (0.03mg/kg/BW) and high-dose GW0742 (0.3mg/kg/BW) were then administered at the beginning of resuscitation. Mice were sacrificed 6h after induction of HS. Plasma levels of IL-6, IL-1ß, IL-10, TNFα, KC, MCP-1, and GM-CSF were determined by ELISA. Myeloperoxidase (MPO) activity in pulmonary and liver tissues was analysed with standardised MPO kits. RESULTS: In mice treated with high-dose GW0742, plasma levels of IL-6, IL-1ß, and MCP-1 were significantly increased compared to the control group mice. When compared to mice treated with low-dose GW0742 plasma levels of IL-6, IL-1ß, GM-CSF, KC, and MCP-1 were significantly elevated in high-dose-treated mice. Low-dose GW0742 treatment was associated with a non-significant downtrend of inflammatory factors in mice with HS. No significant changes of MPO activity in lung and liver were observed between the control group and the GW0742 treatment groups. CONCLUSION: This study identified dose-dependent effects of GW0742 on systemic inflammation after HS. While high-dose GW0742 substantially enhanced the systemic inflammatory response, low-dose GW0742 led to a downtrend of pro-inflammation cytokine expression. The exact mechanisms are yet unknown and need to be assessed in further studies.

Endothelial, but not smooth muscle, peroxisome proliferator-activated receptor ß/δ regulates vascular permeability and anaphylaxis.

BACKGROUND: Remodeling of quiescent vessels with increases in permeability, vasodilatation, and edema are hallmarks of inflammatory disorders. Factors involved in this type of remodeling represent potential therapeutic targets. OBJECTIVES: We investigated whether the nuclear hormone receptor peroxisome proliferator-activated receptor (PPAR) ß/δ, a regulator of metabolism, fibrosis, and skin homeostasis, is involved in regulation of this type of remodeling. METHODS: Wild-type and various Pparb/d mutant mice were used to monitor dermal acute vascular hyperpermeability (AVH) and passive systemic anaphylaxis-induced hypothermia and edema. PPARß/δ-dependent kinase activation and remodeling of endothelial cell-cell junctions were addressed by using human endothelial cells. RESULTS: AVH and dilatation of dermal microvessels stimulated by vascular endothelial growth factor A, histamine, and thrombin are severely compromised in PPARß/δ-deficient mice. Selective deletion of the Pparb/d-encoding gene in endothelial cells in vivo similarly limits dermal AVH and vasodilatation, providing evidence that endothelial PPARß/δ is the major player in regulating acute dermal microvessel remodeling. Furthermore, endothelial PPARß/δ regulatory functions are not restricted to the skin vasculature because its deletion in the endothelium, but not in smooth muscle cells, also leads to reduced systemic anaphylaxis, the most severe form of allergic reaction, in which an acute vascular response plays a key role. PPARß/δ-dependent AVH activation likely involves the activation of mitogen-activated protein kinase and Akt pathways and leads to downstream destabilization of endothelial cell-cell junctions. CONCLUSION: These results unveil not only a novel function of PPARß/δ as a direct regulator of acute vessel permeability and dilatation but also provide evidence that antagonizing PPARß/δ represents an important strategy to consider for moderating diseases with altered endothelial integrity, such as acute inflammatory and allergic disorders.

Telmisartan, a possible PPAR-δ agonist, reduces TNF-α-stimulated VEGF-C production by inhibiting the p38MAPK/HSP27 pathway in human proximal renal tubular cells.

Vascular endothelial growth factor-C (VEGF-C) is a main inducer of inflammation-associated lymphangiogenesis in various inflammatory disorders including chronic progressive kidney diseases, for which angiotensin II receptor type 1 blockers (ARBs) are widely used as the main treatment. Although proximal renal tubular cells may affect the formation of lymphatic vessels in the interstitial area by producing VEGF-C, the molecular mechanisms of VEGF-C production and its manipulation by ARB have not yet been examined in human proximal renal tubular epithelial cells (HPTECs). In the present study, TNF-α dose-dependently induced the production of VEGF-C in HPTECs. The TNF-α-induced production of VEGF-C was mediated by the phosphorylation of p38MAPK and HSP27, but not by that of ERK or NFkB. Telmisartan, an ARB that can activate the peroxisome proliferator-activated receptor (PPAR), served as a PPAR-δ activator and reduced the TNF-α-stimulated production of VEGF-C. This reduction was partially attributed to a PPAR-δ-dependent decrease in p38MAPK phosphorylation. Our results indicate that TNF-α induced the production of VEGF-C in HPTECs by activating p38MAPK/HSP27, and this was partially inhibited by telmisartan in a PPAR-δ dependent manner. These results provide a novel insight into inflammation-associated lymphangiogenesis.

PPARß/δ activation of CD300a controls intestinal immunity.

Macrophages are important for maintaining intestinal immune homeostasis. Here, we show that PPARß/δ (peroxisome proliferator-activated receptor ß/δ) directly regulates CD300a in macrophages that express the immunoreceptor tyrosine based-inhibitory motif (ITIM)-containing receptor. In mice lacking CD300a, high-fat diet (HFD) causes chronic intestinal inflammation with low numbers of intestinal lymph capillaries and dramatically expanded mesenteric lymph nodes. As a result, these mice exhibit triglyceride malabsorption and reduced body weight gain on HFD. Peritoneal macrophages from Cd300a-/- mice on HFD are classically M1 activated. Activation of toll-like receptor 4 (TLR4)/MyD88 signaling by lipopolysaccharide (LPS) results in prolonged IL-6 secretion in Cd300a-/- macrophages. Bone marrow transplantation confirmed that the phenotype originates from CD300a deficiency in leucocytes. These results identify CD300a-mediated inhibitory signaling in macrophages as a critical regulator of intestinal immune homeostasis.

Allergen-induced dermatitis causes alterations in cutaneous retinoid-mediated signaling in mice.

Nuclear receptor-mediated signaling via RARs and PPARδ is involved in the regulation of skin homeostasis. Moreover, activation of both RAR and PPARδ was shown to alter skin inflammation. Endogenous all-trans retinoic acid (ATRA) can activate both receptors depending on specific transport proteins: Fabp5 initiates PPARδ signaling whereas Crabp2 promotes RAR signaling. Repetitive topical applications of ovalbumin (OVA) in combination with intraperitoneal injections of OVA or only intraperitoneal OVA applications were used to induce allergic dermatitis. In our mouse model, expression of IL-4, and Hbegf increased whereas expression of involucrin, Abca12 and Spink5 decreased in inflamed skin, demonstrating altered immune response and epidermal barrier homeostasis. Comprehensive gene expression analysis showed alterations of the cutaneous retinoid metabolism and retinoid-mediated signaling in allergic skin immune response. Notably, ATRA synthesis was increased as indicated by the elevated expression of retinaldehyde dehydrogenases and increased levels of ATRA. Consequently, the expression pattern of genes downstream to RAR was altered. Furthermore, the increased ratio of Fabp5 vs. Crabp2 may indicate an up-regulation of the PPARδ pathway in allergen-induced dermatitis in addition to the altered RAR signaling. Thus, our findings suggest that ATRA levels, RAR-mediated signaling and signaling involved in PPARδ pathways are mainly increased in allergen-induced dermatitis and may contribute to the development and/or maintenance of allergic skin diseases.

Obesity impairs apoptotic cell clearance in asthma.

BACKGROUND: Asthma in obese adults is typically more severe and less responsive to glucocorticoids than asthma in nonobese adults. OBJECTIVE: We sought to determine whether the clearance of apoptotic inflammatory cells (efferocytosis) by airway macrophages was associated with altered inflammation and reduced glucocorticoid sensitivity in obese asthmatic patients. METHODS: We investigated the relationship of efferocytosis by airway (induced sputum) macrophages and blood monocytes to markers of monocyte programming, in vitro glucocorticoid response, and systemic oxidative stress in a cohort of adults with persistent asthma. RESULTS: Efferocytosis by airway macrophages was assessed in obese (n=14) and nonobese (n=19) asthmatic patients. Efferocytosis by macrophages was 40% lower in obese than nonobese subjects, with a mean efferocytic index of 1.77 (SD, 1.07) versus 3.00 (SD, 1.25; P<.01). A similar reduction of efferocytic function was observed in blood monocytes of obese participants. In these monocytes there was also a relative decrease in expression of markers of alternative (M2) programming associated with efferocytosis, including peroxisome proliferator-activated receptor δ and CX3 chemokine receptor 1. Macrophage efferocytic index was significantly correlated with dexamethasone-induced mitogen-activated protein kinase phosphatase 1 expression (ρ=0.46, P<.02) and baseline glucocorticoid receptor α expression (ρ=0.44, P<.02) in PBMCs. Plasma 4-hydroxynonenal levels were increased in obese asthmatic patients at 0.33 ng/mL (SD, 0.15 ng/mL) versus 0.16 ng/mL (SD, 0.08 ng/mL) in nonobese patients (P=.006) and was inversely correlated with macrophage efferocytic index (ρ=-0.67, P=.02). CONCLUSIONS: Asthma in obese adults is associated with impaired macrophage/monocyte efferocytosis. Impairment of this anti-inflammatory process is associated with altered monocyte/macrophage programming, reduced glucocorticoid responsiveness, and systemic oxidative stress.

Immunomodulatory action of dietary fish oil and targeted deletion of intestinal epithelial cell PPARδ in inflammation-induced colon carcinogenesis.

The ligand-activated transcription factor peroxisome proliferator-activated receptor (PPAR)-δ is highly expressed in colonic epithelial cells; however, the role of PPARδ ligands, such as fatty acids, in mucosal inflammation and malignant transformation has not been clarified. Recent evidence suggests that the anti-inflammatory/chemoprotective properties of fish oil (FO)-derived n-3 polyunsaturated fatty acids (PUFAs) may be partly mediated by PPARδ. Therefore, we assessed the role of PPARδ in modulating the effects of dietary n-3 PUFAs by targeted deletion of intestinal epithelial cell PPARδ (PPARδ(ΔIEpC)). Subsequently, we documented changes in colon tumorigenesis and the inflammatory microenvironment, i.e., local [mesenteric lymph node (MLN)] and systemic (spleen) T cell activation. Animals were fed chemopromotive [corn oil (CO)] or chemoprotective (FO) diets during the induction of chronic inflammation/carcinogenesis. Tumor incidence was similar in control and PPARδ(ΔIEpC) mice. FO reduced mucosal injury, tumor incidence, colonic STAT3 activation, and inflammatory cytokine gene expression, independent of PPARδ genotype. CD8(+) T cell recruitment into MLNs was suppressed in PPARδ(ΔIEpC) mice. Similarly, FO reduced CD8(+) T cell numbers in the MLN. Dietary FO independently modulated MLN CD4(+) T cell activation status by decreasing CD44 expression. CD11a expression by MLN CD4(+) T cells was downregulated in PPARδ(ΔIEpC) mice. Lastly, splenic CD62L expression was downregulated in PPARδ(ΔIEpC) CD4(+) and CD8(+) T cells. These data demonstrate that expression of intestinal epithelial cell PPARδ does not influence azoxymethane/dextran sodium sulfate-induced colon tumor incidence. Moreover, we provide new evidence that dietary n-3 PUFAs attenuate intestinal inflammation in an intestinal epithelial cell PPARδ-independent manner.

Biological function and prognostic significance of peroxisome proliferator-activated receptor δ in rectal cancer.

PURPOSE: To investigate the expression significance of PPAR ß/δ in relation to radiotherapy (RT), clinicopathologic, and prognostic variables of rectal cancer patients. EXPERIMENTAL DESIGN: We included 141 primary rectal cancer patients who participated in a Swedish clinical trial of preoperative RT. Tissue microarray samples from the excised rectal cancers and the adjacent or distant normal mucosa and lymph node metastases were stained with PPAR δ antibody. Survival probability was computed by the Kaplan-Meier method and Cox regression model. The proliferation of colon cancer cell lines KM12C, KM12SM, and KM12L4a was assayed after PPAR δ knockdown. RESULTS: PPAR δ was increased from adjacent or distant normal mucosa to primary cancers, whereas it decreased from primary cancers to lymph node metastases. After RT, PPAR δ was increased in normal mucosa, whereas it decreased in primary cancers and lymph node metastases. In primary cancers, the high expression of PPAR δ was related to higher frequency of stage I cases, lower lymph node metastasis rate, and low expression of Ki-67 in the unirradiated cases, and related to favorable survival in the cases either with or without RT. The proliferation of the KM12C, KM12SM, or KM12L4a cells was significantly accelerated after PPAR δ knockdown. CONCLUSIONS: RT decreases the PPAR δ expression in primary rectal cancers and lymph node metastases. PPAR δ is related to the early development of rectal cancer and inhibits the proliferation of colorectal cancer cells. Increase of PPAR δ predicts favorable survival in the rectal cancer patients either with or without preoperative RT.

PPARδ deficient mice develop elevated Th1/Th17 responses and prolonged experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS) is a neurological disorder that affects more than a million people worldwide. The etiology of MS is not known and there is no medical treatment that can cure MS. Earlier studies have shown that peroxisome proliferator-activated receptor (PPARs) agonists ameliorate MS-like disease in experimental allergic encephalomyelitis (EAE). In this study we have used PPARδ deficient mice to determine its physiological role in the regulation of CNS EAE and MS. We found that PPARδ(-/-) mice develop EAE with similar day of onset and disease incidence compared to C57BL/6 wild type mice. Interestingly, both male and female PPARδ(-/-) mice showed prolonged EAE with resistance to remission and recovery. PPARδ(-/-) mice with EAE expressed elevated levels of IFNγ and IL-17 along with IL-12p35 and IL-12p40 in the brain and spleen. PPARδ(-/-) mice also developed augmented neural antigen-specific Th1/Th17 responses and impaired Th2/Treg responses compared to wild type mice. These findings indicate that PPARδ(-/-) mice develop prolonged EAE in association with augmented Th1/Th17 responses, suggesting a critical physiological role for PPARδ in the remission and recovery of EAE.

Peroxisome proliferator-activated receptor delta agonists inhibit T helper type 1 (Th1) and Th17 responses in experimental allergic encephalomyelitis.

Multiple sclerosis (MS) is a neurological disorder that affects more than a million people world-wide. The aetiology of MS is not known and there is no medical treatment available that can cure MS. Experimental autoimmune encephalomyelitis (EAE) is a T-cell-mediated autoimmune disease model of MS. The pathogenesis of EAE/MS is a complex process involving activation of immune cells, secretion of inflammatory cytokines and destruction of myelin sheath in the central nervous system (CNS). Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptor transcription factors that regulate cell growth, differentiation and homeostasis. PPAR agonists have been used in the treatment of obesity, diabetes, cancer and inflammation. We and others have shown that PPARgamma, alpha and delta agonists inhibit CNS inflammation and demyelination in the EAE model of MS. In this study we show that the PPARdelta agonists GW501516 and L165041 ameliorate MOGp35-55-induced EAE in C57BL/6 mice by blocking interferon (IFN)-gamma and interleukin (IL)-17 production by T helper type 1 (Th1) and Th17 cells. The inhibition of EAE by PPARdelta agonists was also associated with a decrease in IL-12 and IL-23 and an increase in IL-4 and IL-10 expression in the CNS and lymphoid organs. These findings indicate that PPARdelta agonists modulate Th1 and Th17 responses in EAE and suggest their use in the treatment of MS and other autoimmune diseases.

Comparison of PPARδ and PPARγ in inhibiting the pro-inflammatory effects of C-reactive protein in endothelial cells.

BACKGROUND: Inflammation associated with endothelial cell dysfunction is a key step of atherogenesis. C-reactive protein (CRP), used to serve as a nonspecific clinical inflammation marker, has now emerged as a new marker for cardiovascular diseases. Recently, PPARδ has revealed benefits for dealing with inflammation. The relationship between CRP-induced inflammation and PPARδ agonist remains unclear. METHODS: Human umbilical vein endothelial cells (HUVECs) were separated into the following groups: 25 µg CRP alone for 15 hours; CRP-treated with 1 µM PPARδ(L-165041) or 10 µM PPARγ(troglitazone) agonists, and untreated HUVECs. This research focused on the CRP underlying signaling pathways and the effects of PPAR agonists on monocyte attachment to endothelial cells. RESULTS: Levels of interleukin-6 (IL-6) and IL-8 increased by CRP were both significantly attenuated by pretreatment with PPARδ or PPARγ agonists, but the needed dose of PPARδ to reach the same effect was less than PPARγ agonist. After incubation with CRP, immunoblotting showed a significant increase in NF-κB activation and CD32 receptor. These changes were associated with a significant increase of MCP-1 and VCAM-1 expression. PPARδ treatment not only decreased these pro-inflammatory effects in HUVECs but also significantly attenuated monocyte adhesion to endothelial cells in less dosage than PPARγ. CONCLUSIONS: The results suggest that PPARδ attenuated CRP-induced pro-inflammatory effects may through CD32 and NF-κB pathway. PPARδ may serve as a more potent therapeutic target than PPARγ in atherosclerosis or inflammatory therapy.

Suppression of pro-inflammatory adhesion molecules by PPAR-delta in human vascular endothelial cells.

OBJECTIVE: Endothelial activation is implicated in atherogenesis and diabetes. The role of peroxisome proliferator-activated receptor-delta (PPAR-delta) in endothelial activation remains poorly understood. In this study, we investigated the anti-inflammatory effect of PPAR-delta and the mechanism involved. METHODS AND RESULTS: In human umbilical vein endothelial cells (HUVECs), the synthetic PPAR-delta ligands GW0742 and GW501516 significantly inhibited tumor necrosis factor (TNF)-alpha-induced expression of vascular cell adhesion molecule-1 and E-selectin (assayed by real-time RT-PCR and Northern blotting), as well as the ensuing endothelial-leukocyte adhesion. Activation of PPAR-delta upregulated the expression of antioxidant genes superoxide dismutase 1, catalase, and thioredoxin and decreased reactive oxygen species production in ECs. Chromatin immunoprecipitation assays showed that GW0742 switched the association of BCL-6, a transcription repressor, from PPAR-delta to the vascular cell adhesion molecule (VCAM)-1 promoter. Small interfering RNA reduced endogenous PPAR-delta expression but potentiated the suppressive effect of GW0742 on EC activation, which suggests that the nonliganded PPAR-delta may have an opposite effect. CONCLUSIONS: We have demonstrated that ligand activation of PPAR-delta in ECs has a potent antiinflammatory effect, probably via a binary mechanism involving the induction of antioxidative genes and the release of nuclear corepressors. PPAR-delta agonists may have a potential for treating inflammatory diseases such as atherosclerosis and diabetes.

Expression profiling of peroxisome proliferator-activated receptor-delta (PPAR-delta) in mouse tissues using tissue microarray.

Peroxisome proliferator-activated receptor-delta (PPAR-delta) is known as a transcription factor involved in the regulation of fatty acid oxidation and mitochondrial biogenesis in several tissues, such as skeletal muscle, liver and adipose tissues. In this study, to elucidate systemic physiological functions of PPAR-delta, we examined the tissue distribution and localization of PPAR-delta in adult mouse tissues using tissue microarray (TMA)-based immunohistochemistry. PPAR-delta positive signals were observed on variety of tissues/cells in multiple systems including cardiovascular, urinary, respiratory, digestive, endocrine, nervous, hematopoietic, immune, musculoskeletal, sensory and reproductive organ systems. In these organs, PPAR-delta immunoreactivity was generally localized on the nucleus, although cytoplasmic localization was observed on several cell types including neurons in the nervous system and cells of the islet of Langerhans. These expression profiling data implicate various physiological roles of PPAR-delta in multiple organ systems. TMA-based immunohistochemistry enables to profile comprehensive protein localization and distribution in a high-throughput manner.

Essential role of Smad3 in the inhibition of inflammation-induced PPARbeta/delta expression.

Wound healing proceeds by the concerted action of a variety of signals that have been well identified. However, the mechanisms integrating them and coordinating their effects are poorly known. Herein, we reveal how PPARbeta/delta (PPAR: peroxisome proliferator-activated receptor) follows a balanced pattern of expression controlled by a crosstalk between inflammatory cytokines and TGF-beta1. Whereas conditions that mimic the initial inflammatory events stimulate PPARbeta/delta expression, TGF-beta1/Smad3 suppresses this inflammation-induced PPARbeta/delta transcription, as seen in the late re-epithelialization/remodeling events. This TGF-beta1/Smad3 action involves an inhibitory effect on AP-1 activity and DNA binding that results in an inhibition of the AP-1-driven induction of the PPARbeta/delta promoter. As expected from these observations, wound biopsies from Smad3-null mice showed sustained PPARbeta expression as compared to those of their wild-type littermates. Together, these findings suggest a mechanism for setting the necessary balance between inflammatory signals, which trigger PPARbeta/delta expression, and TGF-beta1/Smad3 that governs the timely decrease of this expression as wound healing proceeds to completion.