PPARδ Agonist Promotes Type II Cartilage Formation in a Rabbit Osteochondral Defect Model.

Osteoarthritis (OA) is a chronic degenerative joint disease accompanied by an inflammatory milieu that results in painful joints. The pathogenesis of OA is multifactorial, with genetic predisposition, environmental factors, and traumatic injury resulting in the direct or indirect loss of cartilage. The articular cartilage can also be damaged by direct focal traumatic injury. Articular cartilage provides a smooth, deformable bearing surface with a low coefficient of friction, increased contact area, and reduced contact stress. Articular type II hyaline cartilage lines the synovial joints and, when injured, has a limited ability for repair, except for the most superficial layers via diffusion from the synovial fluid, secondary to no blood supply, a complex structure, and a low metabolic rate. Restoring the articular surface can relieve pain and restore function. Although many strategies have been developed to regenerate type II collagen based on the extent of the lesion, surgical treatments are still evolving. The peroxisome proliferator-activated receptor delta (PPARδ) agonist and collagen treatment of mesenchymal stem cells (MSCs) enhance the chondrogenic capacity in vitro. We present a novel technique for cartilage restoration in a rabbit cartilage osteochondral defect model using a PPARδ agonist (GW0742)-infused 3D collagen scaffold to induce type II cartilage from MSCs.

PPAR-δ activation reduces cisplatin-induced apoptosis via inhibiting p53/Bax/caspase-3 pathway without modulating autophagy in murine renal proximal tubular cells.

BACKGROUND: Cisplatin-induced injury of renal proximal tubular cells results basically from increased apoptosis via mitochondrial damage, and is mitigated by appropriate enhancement of autophagy. Peroxisome proliferator-activated receptor-delta (PPAR-δ) reportedly protects against not only mitochondrial damages but also enhances autophagy. Thus, PPAR-δ may protect against cisplatin-induced kidney injury. METHODS: We examined the protective effects of PPAR-δ activation on cisplatin-induced cellular injury and their detailed mechanisms in a murine renal proximal tubular (mProx) cell line using GW0742, an authentic PPAR-δ activator. Cisplatin-induced cell damages were evaluated by TUNEL assay and immunoblot analyses for p53, 14-3-3, Bax, Bcl2, cytochrome C, and activated caspases. Autophagy status was examined by immunoblot analyses for p62 and LC3. RESULTS: GW0742 suppressed cisplatin-induced apoptosis of mProx cells by reducing the activation of caspase-3 via attenuating the phosphorylation of p53 and 14-3-3, mitochondrial Bax accumulation, cytochrome C release from mitochondria to the cytosol and ensuing cytosolic caspase-9 activation. In contrast, GW0742 did not diminish cisplatin-enhanced activation of caspases-8 or -12 as extrinsic or endothelium reticulum apoptotic pathways, respectively. The inhibitory effect of GW0742 on cisplatin-induced caspase-3 activation was significantly diminished by silencing of the PPAR-δ gene expression. GW0742 itself had no influence on starvation-stimulated or cisplatin-induced autophagy in mProx cells, suggesting that the protective effects were not mediated by autophagy modification. CONCLUSION: Our results indicate that GW0742 may serve as a candidate agent to mitigate cisplatin nephrotoxicity via inhibiting the mitochondrial apoptotic pathway considerably depending on PPAR-δ, without modulating autophagy.

Endogenous hydrogen sulfide improves vascular remodeling through PPARδ/SOCS3 signaling.

INTRODUCTION: Mounting evidences demonstrated the deficiency of hydrogen sulfide (H2S) facilitated the progression of cardiovascular diseases. However, the exact effects of H2S on vascular remodeling are not consistent. OBJECTIVES: This study aimed to investigate the beneficial role of endogenous H2S on vascular remodeling. METHODS: CSE inhibitor, DL-propargylglycine (PPG) was used to treat mice and vascular smooth muscle cells (VSMCs). Sodium hydrosulfide (NaHS) was given to provide hydrogen sulfide. Vascular tension, H&E staining, masson trichrome staining, western blot and CCK8 were used to determine the vascular remodeling, expressions of inflammatory molecules and proliferation of VSMCs. RESULTS: The deficiency of endogenous H2S generated vascular remodeling with aggravated active and passive contraction, thicken aortic walls, collagen deposition, increased phosphorylation of STAT3, decreased production of PPARδ and SOCS3 in aortas, which were reversed by NaHS. PPG inhibited expression of PPARδ and SOCS3, stimulated the phosphorylation of STAT3, increased inflammatory molecules production and proliferation rate of VSMCs which could all be corrected by NaHS supply. PPARδ agonist GW501516 offered protections similar to NaHS in PPG treated VSMCs. Aggravated active and passive contraction in PPG mice aortas, upregulated p-STAT3 and inflammatory molecules, downregulated SOCS3 and phenotype transformation in PPG treated VSMCs could be corrected by PPARδ agonist GW501516 treatment. On the contrary, PPARδ antagonist GSK0660 exhibited opposite effects on vascular contraction in aortas, expressions of p-STAT3 and SOCS3 in VSMCs compared with GW501516. CONCLUSION: In a word, endogenous H2S protected against vascular remodeling through preserving PPARδ/SOCS3 anti-inflammatory signaling pathway. Deficiency of endogenous H2S should be considered as a risk factor for VSMCs dysfunction.

CD137 signaling induces macrophage M2 polarization in atherosclerosis through STAT6/PPARδ pathway.

CD137 signaling plays an important role in the formation and development of atherosclerotic plaques. The purpose of the present study was to investigate the effects of CD137 signaling on macrophage polarization during atherosclerosis and to explore the underlying mechanisms. The effect of CD137 signaling on macrophage phenotype in atherosclerotic plaques was determined by intraperitoneal injection of agonist-CD137 recombinant protein in apolipoprotein E-deficient (ApoE-/-) mice, an established in vivo model of atherosclerosis. Murine peritoneal macrophages and RAW 264.7 cells were treated with AS1517499 and siPPARδ (peroxisome proliferator-activated receptor δ) to study the role of STAT6 (signal transducers and activators of transcription 6)/PPARδ signaling in CD137-induced M2 macrophage polarization in vitro. Results from both in vivo and in vitro experiments showed that CD137 signaling can transform macrophages into the M2 phenotype during the process of atherosclerotic plaque formation and regulate the angiogenic features of M2 macrophages. Furthermore, activation of the CD137 signaling pathway induces phosphorylation of STAT6 and enhances the expression of PPARδ. We further found that macrophage M2 polarization is reduced when the STAT6/PPARδ pathway is inhibited. Together, these data show a role for the STAT6/PPARδ signaling pathway in the CD137 signaling-induced M2 macrophage polarization pathway.

The association between peroxisome proliferator-activated receptor Δ rs3777744, rs3798343, and rs6922548 and coronary artery disease.

Objective: The aim of the present study is to investigate the association between the single nucleotide polymorphism (SNP) sites of peroxisome proliferator-activated receptor Δ (PPARD) and the risk of coronary artery disease (CAD). To this end, a prospective observational single-center study of the clinical data from 880 subjects in a Chinese population was conducted. Methods: A total of 880 subjects, including 609 CAD patients and 271 control subjects, were selected for the present study. All inpatients had 4 ml of venous blood drawn after 12 h of fasting, and then clinical tests were conducted to obtain the biochemical parameters. CAD patients and Controls were distinguished by coronary angiography. Statistical analysis was conducted with SPSS software (ver 16.0). Results: A significant association between the G-alleles of PPARD rs3777744 and rs3798343 and a decreased risk for CAD was found. Moreover, we found an interaction between high fasting high-density lipoprotein cholesterol (HDL-C) serum levels, low serum glucose levels and their genotypes, ultimately decreasing the risk of CAD. Haplotype analysis was conducted on the three SNP sites, rs3777744 and rs3798343 to form a block [r2 = 0.79, D' = 0.99). The A-C haplotypes were associated with an increased risk of CAD (odds ratio (OR), 95% confidence interval (CI): 1.321 (1.060-1.647), P=0.013], and the G-G haplotypes were associated with a decreased risk [OR, 95% CI: 0.714 (0.567-0.849), P=0.004]. Conclusions: Our study indicates a significant association between the G-alleles of PPARD rs3777744 and rs3798343 and a decreased CAD risk. In addition, genotypes interact with high serum HDL-C levels and low serum glucose levels, resulting in decreased prevalence of CAD.

The PPARδ agonist GW0742 restores neuroimmune function by regulating Tim-3 and Th17/Treg-related signaling in the BTBR autistic mouse model.

Autism spectrum disorders (ASD) are neurodevelopmental disorders that are characterized by repetitive behaviors, and impairments in communication and social interaction. Studies have shown that activation of peroxisome proliferator-activated receptor-delta (PPARδ) causes anti-inflammatory effects in animal models of neuroinflammatory diseases. We investigated the possible anti-inflammatory effect of a PPARδ agonist, GW0742 in the BTBR T+ Itpr3tf/J (BTBR) mouse model of autism. BTBR and C57BL/6 (B6) mice were treated orally with GW0742 (30 mg/kg, p.o., once daily) for 7 days. Effect of GW0742 treatment on repetitive behavior, marble burying, and thermal sensitivity response was assessed on day 8. We further examined the effect of GW0742 treatment on immunological parameters in splenocytes using flow cytometry (CD4+TIM-3+, IL-17A+TIM-3+, IL-17A+CD4+, RORγT+TIM-3+, RORγT+CD4+, Stat3+TIM-3+, Foxp3+TIM-3+, Foxp3+CD4+, and IFN-γ+CD4+). We also explored the effects of GW0742 on mRNA and protein expression of TIM-3, IL-17A, RORγT, Stat3, IFN-γ, Foxp3, and IL-10 in the brain tissue using RT-PCR and western blot analyses. GW0742 treatment substantially decreased repetitive behaviors, and lowered thermal sensitivity response in BTBR mice. GW0742 attenuated the expression of inflammatory markers such as IL-17A, RORγT, Stat3, TIM-3, and IFN-γ, while upregulating anti-inflammatory markers such as IL-10/Foxp3 both in the brain and periphery of BTBR mice. In conclusion, this study suggests that GW0742 corrects neurobehavioral dysfunction in BTBR mice which is concurrent with modulation of multiple signaling pathways.

In Vivo Optical Imaging of Myelination Events in a Myelin Basic Protein Promoter-Driven Luciferase Transgenic Mouse Model.

The compact myelin sheath is important for axonal function, and its loss can lead to neuronal cell death and irreversible functional deficits. Myelin is vulnerable to a variety of metabolic, toxic, and autoimmune insults. In diseases like multiple sclerosis, there is currently no therapy to stop myelin loss, underscoring the need for neuroprotective and remyelinating therapies. Noninvasive, robust techniques are also needed to confirm the effect of such therapies in animal models. This article describes the generation, characterization, and potential uses for a myelin basic protein-luciferase (MBP-luci) transgenic mouse model, in which the firefly luciferase reporter gene is selectively controlled by the MBP promoter. In vivo bioluminescence imaging can be used to visualize and quantify demyelination and remyelination at the transcriptional level, noninvasively, and in real time. Transgenic mice were assessed in the cuprizone-induced model of demyelination, and luciferase activity highly correlated with demyelination and remyelination events as confirmed by both magnetic resonance imaging and postmortem histological analysis. Furthermore, MBP-luci mice demonstrated enhanced luciferase signal and remyelination in the cuprizone model after treatment with a peroxisome proliferator activated receptor-delta selective agonist and quetiapine. Imaging sensitivity was further enhanced by using CycLuc 1, a luciferase substrate, which has greater blood-brain barrier penetration. We demonstrated the utility of MBP-luci model in tracking myelin changes in real time and supporting target and therapeutic validation efforts.

Chitinase-3-like protein 1 ameliorates atherosclerotic responses via PPARδ-mediated suppression of inflammation and ER stress.

Chitinase 3-like protein 1 (CHI3L1) is a novel biomarker of systemic inflammation. However, the effects of CHI3L1 on the progression of atherosclerosis remain to be explored. In the current study, we found that CHI3L1 induces peroxisome proliferator-activated receptor delta (PPARδ) expression, leading to a dose-dependent increase in oxygen-regulated protein 150 (ORP150) expression. We demonstrated that CHI3L1 suppresses atherosclerotic reactions caused by LPS treatment via a PPARδ-dependent pathway. Treatment of HUVECs and THP-1 cells with CHI3L1 suppressed LPS-induced phosphorylation of nuclear factor kappa B (NFκB) and secretion of proinflammatory cytokines such as TNFα and MCP-1. In HUVECs, expression of adhesion molecules and LPS-stimulated adhesion of THP-1 cells to the endothelium were significantly reduced after CHI3L1 treatment. Furthermore, LPS-induced endoplasmic reticulum (ER) stress and cell apoptosis were significantly ameliorated after treatment of HUVECs with CHI3L1. Particularly, all of the pro-atherosclerotic effects were significantly mitigated by treatment with small interfering (si) RNA for PPARδ. In conclusion, CHI3L1 ameliorates LPS-induced atherosclerotic reactions via PPARδ-mediated suppression of inflammation and ER stress.

Sirtuins 1-7 expression in human adipose-derived stem cells from subcutaneous and visceral fat depots: influence of obesity and hypoxia.

The sirtuin family comprises seven NAD+-dependent deacetylases which control the overall health of organisms through the regulation of pleiotropic metabolic pathways. Sirtuins are important modulators of adipose tissue metabolism and their expression is higher in lean than obese subjects. At present, the role of sirtuins in adipose-derived stem cells has not been investigated yet. Therefore, in this study, we evaluated the expression of the complete panel of sirtuins in adipose-derived stem cells isolated from both subcutaneous and visceral fat of non-obese and obese subjects. We aimed at investigating the influence of obesity on sirtuins' levels, their role in obesity-associated inflammation, and the relationship with the peroxisome proliferator-activated receptor delta, which also plays functions in adipose tissue metabolism. The mRNA levels in the four types of adipose-derived stem cells were evaluated by quantitative polymerase chain reaction, in untreated cells and also after 8 h of hypoxia exposure. Correlations among sirtuins' expression and clinical and molecular parameters were also analyzed. We found that sirtuin1-6 exhibited significant higher mRNA expression in visceral adipose-derived stem cells compared to subcutaneous adipose-derived stem cells of non-obese subjects. Sirtuin1-6 levels were markedly reduced in visceral adipose-derived stem cells of obese patients. Sirtuins' expression in visceral adipose-derived stem cells correlated negatively with body mass index and C-reactive protein and positively with peroxisome proliferator-activated receptor delta. Finally, only in the visceral adipose-derived stem cells of obese patients hypoxia-induced mRNA expression of all of the sirtuins. Our results highlight that sirtuins' levels in adipose-derived stem cells are consistent with protective effects against visceral obesity and inflammation, and suggest a transcriptional mechanism through which acute hypoxia up-regulates sirtuins in the visceral adipose-derived stem cells of obese patients.

Association of functional genetic variants in PPARδ encoding peroxisome proliferator-activated receptor delta with ischemic stroke in Chinese Uyghur population.

PPARδ belongs to a receptor family of ligand-activated transcription factors involved in the regulation of inflammation, cellular glucose uptake, protection against atherosclerosis and endothelial cell function. Through these effects, they might be involved with the ischemic stroke (IS). We recruited 200 subjects (100 IS patients diagnosed by CTs or/and magnetic resonance imaging (MRI) and 100 normal healthy controls from Chinese Uyghur population) to assess the nature of the functional polymorphisms of PPARδ +294T/C and any links with IS in this unique population. We found that the C allele of the PPARδ +294T/C polymorphism was more common in controls than IS subjects in the Uyghur population. C allele carriage may be associated with an increased risk of IS in Uyghurs with a strong trend (OR 1.79, 95% CI: 1.11-2.89). Additionally, the PPARδ CC and TC genotypes were less frequent in Uyghur population than in Han population. Our population and ethnic-based study demonstrates that the PPARδ +294C allele maybe an independent risk of IS in Chinese Uyghurs especially in the male (OR 1.99, 95%CI:1.06-3.72) and obesity populations (OR 2.36, 95%CI: 1.19-4.67), which were consistent with Tunisian Population. Moreover, total cholesterol, fasting blood glucose, waist-to-hip ratio, hypertension, history of heart diseases, and negative events may increase the risk of IS, with a trend for HDL to be a protective factor for IS in the Uyghur population. However, larger populations are warranted to validate our findings.

Molecular Characterization and Expression Analysis of the Peroxisome Proliferator Activated Receptor Delta (PPARδ) Gene before and after Exercise in Horse.

While athletic abilities such as speed, endurance and recovery are important in the horse, genes related to these abilities have not been extensively investigated. Here, we characterized the horse peroxisome proliferator-activated receptor delta (PPARδ) gene and analyzed the expression of PPARδ during exercise. PPARδ is a known regulator of ß-oxidation, muscle fiber transformation, and running endurance. Through evolutionary analysis using the synonymous and non-synonymous mutation ratio, it was revealed that positive selection occurred in the horse PPARδ gene. Two important domains related to nuclear hormone receptors, C4 zinc finger and ligand binding domain, were also found to be conserved well in horse PPARδ. Horse PPARδ was expressed ubiquitously in many tissues, but the expression level was various depending on the tissues. In the skeletal muscle, PPARδ increased about 2.5 folds after 30 min of exercise. Unlike in muscle, the increase of PPARδ expression was observed at 60 min but not 30 min of exercise in leukocytes. This finding might be useful for testing the endurance of horse using blood samples. Conclusively, the horse PPARδ gene is evolutionarily conserved well and can be used as a biomarker of endurance in horse.

PPARδ deficiency disrupts hypoxia-mediated tumorigenic potential of colon cancer cells.

Peroxisome proliferator-activated receptor (PPAR) δ is highly expressed in colon epithelial cells and closely linked to colon carcinogenesis. However, the role of PPARδ in colon cancer cells in a hypoxic tumor microenvironment is not fully understood. We found that expression of the tumor-promoting cytokines, IL-8 and VEGF, induced by hypoxia (<1% O2) and deferoxamine (a hypoxia mimetic) was significantly attenuated in PPARδ-deficient HCT116 colon cancer cells. Consequently, PPARδ-knockout colon cancer cells exposed to hypoxia and deferoxamine failed to stimulate endothelial cell vascularization and macrophage migration/proliferation, whereas wild-type cells were able to induce angiogenesis and macrophage activation in response to hypoxic stress. Hypoxic stress induced transcriptional activation of PPARδ, but not its protein expression, in HCT116 cells. Exogenous expression of p300 potentiated deferoxamine-induced PPARδ transactivation, while siRNA knockdown of p300 abolished hypoxia- and deferoxamine-induced PPARδ transactivation. PPARδ associated with p300 upon hypoxic stress as demonstrated by coimmunoprecipitation studies. PI3K inhibitors or siRNA knockdown of Akt suppressed the PPARδ transactivation induced by hypoxia and deferoxamine in HCT116 cells, leading to decreased expression of IL-8 and VEGF. Collectively, these results reveal that PPARδ is required for hypoxic stress-mediated cytokine expression in colon cancer cells, resulting in promotion of angiogenesis, macrophage recruitment, and macrophage proliferation in the tumor microenvironment. p300 and the PI3K/Akt pathway play a role in the regulation of PPARδ transactivation induced by hypoxic stress. Our results demonstrate the positive crosstalk between PPARδ in tumor cells and the hypoxic tumor microenvironment and provide potential therapeutic targets for colon cancer.

Peroxisome proliferator-activated receptor delta and cardiovascular disease.

Recent reports have shown that peroxisome proliferator-activated receptor delta (PPARD) plays an important role in different vascular processes suggesting that PPARD is a significant modulator of cardiovascular disease. This review will focus on PPARD in relation to cardiovascular risk factors based on cell, animal and human data. Mouse studies suggest that Ppard is an important metabolic modulator that may have implications for cardiovascular disease (CVD). Specific human PPARD gene variants show no clear association with CVD but interactions between variants and lifestyle factors might influence disease risk. During recent years, development of specific and potent PPARD agonists has also made it possible to study the effects of PPARD activation in humans. PPARD agonists seem to exert beneficial effects on dyslipidemia and insulin-resistant syndromes but safety issues have been raised due to the role that PPARD plays in cell proliferation. Thus, large long term outcome as well as detailed safety and tolerability studies are needed to evaluate whether PPARD agonists could be used to treat CVD in humans.