PPARdelta Research Articles

The PPARγ Agonist Efatutazone Increases the Spectrum of Well-Differentiated Mammary Cancer Subtypes Initiated by Loss of Full-Length BRCA1 in Association with TP53 Haploinsufficiency

Peroxisome proliferator-activated receptor gamma (PPARγ) agonists have anticancer activity and influence cell differentiation. We examined the impact of the selective PPARγ agonist efatutazone on mammary cancer pathogenesis in a mouse model of BRCA1 mutation. Mice with conditional loss of full-length BRCA1 targeted to mammary epithelial cells in association with germline TP53 insufficiency were treated with efatutazone through the diet starting at age 4 months and were euthanized at age 12months or when palpable tumor reached 1 cm(3). Although treatment did not reduce percentage of mice developing invasive cancer, it significantly reduced prevalence of noninvasive cancer and total number of cancers per mouse and increased prevalence of well-differentiated cancer subtypes not usually seen in this mouse model. Invasive cancers from controls were uniformly estrogen receptor α negative and undifferentiated, whereas well-differentiated estrogen receptor α-positive papillary invasive cancers appeared in efatutazone-treated mice. Expression levels of phosphorylated AKT and CDK6 were significantly reduced in the cancers developing in efatutazone-treated mice. Efatutazone treatment reduced rates of mammary epithelial cell proliferation and development of hyperplastic alveolar nodules and increased expression levels of the PPARγ target genes Adfp, Fabp4, and Pdhk4 in preneoplastic mammary tissue. Intervention efatutazone treatment in mice with BRCA1 deficiency altered mammary cancer development by promoting development of differentiated invasive cancer and reducing prevalence of noninvasive cancer and preneoplastic disease.

Metabolic Adaptations in the Liver of Born Long-Distance Running Mice

Long-distance runners have increased needs of energy supply. To unravel genetically based mechanisms required for efficient energy supply, we have analyzed hepatic metabolism of mice characterized by the inborn capacity to perform as long-distance runners. The mouse model had been established by phenotypic selection for high treadmill performance for 90 generations and was characterized by approximately 3.8-fold higher running capacities (Dummerstorf high Treadmill Performance mouse line [DUhTP]) compared with unselected and also untrained controls (Dummerstorf Control mouse line [DUC]). From 7-wk-old male mice, serum and liver samples were collected and analyzed for messenger RNA, protein, and metabolite levels, respectively. In livers from DUhTP mice, we identified significantly higher messenger RNA transcript levels of peroxisome proliferator-activated receptor delta and higher protein levels of sirtuin-1, acetyl-CoA-synthetase, acetyl-CoA-carboxylase, phosphoenolpyruvate carboxykinase, and glutamate-dehydrogenase, suggesting higher gluconeogenesis and lipogenesis in DUhTP mice. In fact, higher hepatic levels of glycogen and triglycerides as well as higher concentrations of carbohydrate, fatty acid, and cholesterol metabolites were found in DUhTP mice. In parallel, in DUhTP mice, which did not have access to running wheels, a marked hyperlipidemia (cholesterol = 160% ± 8%, triglycerides = 174% ± 14% of controls, respectively), and abdominal obesity (DUhTP = 0.396 ± 0.019 g, DUC = 0.291 ± 0.019 g) were found. From our data, we conclude that the physiological basis of genetically fixed higher endurance-running performance in DUhTP marathon mouse is related to increased hepatic gluconeogenesis and lipogenesis. Expression of sirtuin 1 as well as of gluconeogenic and lipogenic key enzymes may be related to peroxisome proliferator-activated receptor delta. Metabolic adaptations presented in our study represent inborn features of superior endurance-running performance.

Conjugated linoleic acid (CLA) activates PGC‐1alpha via AMPK and SIRT1 in C2C12 myotubes

Along with its effect on body fat reduction, dietary CLA has been reported to improve physical activity and endurance capacity in mice. It has been suggested these effects are due to up-regulation of skeletal muscle peroxisome proliferator-activated receptor δ (PPARδ), however, the mode of action is not completely understood. Thus, the purpose of the study was to determine the mechanisms of CLA on skeletal muscle PPA Rδ regulation to understand its effects on muscle metabolism. C2C12 myocytes were differentiated into myotubes and effects of CLA (cis-9, trans-11 and trans-10, cis-12) were investigated on major upstream regulators of PPARδ, AMP-activated protein kinase (AMPK), PPAR γ coactivator 1α (PGC-1α), and sirtuin 1 (SIRT1), including time- and dose-dependent effects. Both the cis-9, trans-11 and trans-10, cis-12 CLA isomers significantly increased expressions of AMPK (both total- and phosphorylated-AMPKs), most significantly at 50μM for 12 or 2 4 hour incubation compared to control. However, only the trans-10, cis-12 CLA isomer, but not the cis-9, trans-11 isomer, increased expressions of PGC-1α and SIRT1 (30% and 25%, respectively) compared to control. These results suggest that CLA isomers promote muscle metabolism via AMPK mediated mechanism, where the trans-10, cis-12 CLA isomer is more effective than the cis-9, trans-11 isomer by modulating both AMPK and SIRT1 mediated mechanisms. Grant Funding Source: None

Effect of Genetic Polymorphism +294T/C in Peroxisome Proliferator-Activated Receptor Delta on the Risk of Ischemic Stroke in a Tunisian Population

PPARδ +294T/C polymorphism was investigated in diabetics, in normolipidemic healthy controls, in dyslipidemic and nondyslipidemic coronary artery disease patients but never in ischemic stroke patients. The aim of this study was to explore, for the first time, the relationship between the genetic polymorphism of PPARδ and the risk of ischemic stroke among patients with diabetes. The study group consisted of 196 patients with ischemic stroke and 192 controls. Plasma concentrations of total cholesterol, triglycerides, low-, and high-density lipoprotein did not differ significantly between subjects carrying the TT genotype and those carrying the CC/TC genotype in both ischemic stroke patients (with or without diabetes) and control groups. The +294C allele (CC + CT genotypes) as compared with TT genotypes was found to be higher in total ischemic stroke patients than in controls. On the other hand, no interaction between diabetes and PPAR +294T/C polymorphism on the risk of ischemic stroke was found (p = 0.089). The PPARδ +294T/C polymorphism was associated with the risk of ischemic stroke in Tunisian subjects. This polymorphism has no influence on plasma lipoprotein concentrations and body mass index either in healthy subjects or in ischemic stroke patients with or without diabetes both in males and females.

Perfluorooctane sulfonate induces neuronal and oligodendrocytic differentiation in neural stem cells and alters the expression of PPARγ in vitro and in vivo

Perfluorinated compounds are ubiquitous chemicals of major concern for their potential adverse effects on the human population. We have used primary rat embryonic neural stem cells (NSCs) to study the effects of perfluorooctane sulfonate (PFOS) on the process of NSC spontaneous differentiation. Upon removal of basic fibroblast growth factor, NSCs were exposed to nanomolar concentrations of PFOS for 48h, and then allowed to differentiate for additional 5days. Exposure to 25 or 50nM concentration resulted in a lower number of proliferating cells and a higher number of neurite-bearing TuJ1-positive cells, indicating an increase in neuronal differentiation. Exposure to 50nM also significantly increased the number of CNPase-positive cells, pointing to facilitation of oligodendrocytic differentiation. PPAR genes have been shown to be involved in PFOS toxicity. By q-PCR we detected an upregulation of PPARγ with no changes in PPARα or PPARδ genes. One of the downstream targets of PPARs, the mitochondrial uncoupling protein 2 (UCP2) was also upregulated. The number of TuJ1- and CNPase-positive cells increased after exposure to PPARγ agonist rosiglitazone (RGZ, 3μM) and decreased after pre-incubation with the PPARγ antagonist GW9662 (5μM). RGZ also upregulated the expression of PPARγ and UCP2 genes. Meanwhile GW9662 abolished the UCP2 upregulation and decreased Ca2+ activity induced by PFOS. Interestingly, a significantly higher expression of PPARγ and UCP3 genes was also detected in mouse neonatal brain after prenatal exposure to PFOS. These data suggest that PPARγ plays a role in the alteration of spontaneous differentiation of NSCs induced by nanomolar concentrations of PFOS.

Peroxisome proliferator‐activated receptor delta protects against obesity‐related glomerulopathy through the P38 MAPK pathway

Obesity is a prominent component of metabolic syndrome and a major risk factor for renal disease. The aim of this study was to explore the effect of cross-talk between peroxisome proliferator-activated receptor (PPAR)δ and p38 mitogen-activated protein kinase (p38 MAPK) on obesity-related glomerulopathy. Male Wistar rats were randomly assigned to standard laboratory chow or a high-fat diet for 32 weeks. Glomerular mesangial cells HBZY-1 and mature differentiation 3T3-L1 cells were cocultured and were transfected with PPARδ-expressing vectors or treated with agonist or inhibitor of PPARδ or p38 MAPK. Rats on a high-fat diet showed typical characteristics of metabolic syndrome including obesity, dyslipidemia, insulin resistance, and hypertension. Rats on a high-fat diet also had significant glomerular hypertrophy and extracellular matrix accumulation, which were accompanied by increased p38 MAPK phosphorylation and decreased PPARδ expression in the kidney tissue. The roles of p38 MAPK and PPARδ in a coculture system of mesangial cells and mature differentiation 3T3-L1 cells were further explored. PPARδ suppression promoted laminin and type IV collagen secretion through p38 MAPK phosphorylation in mesangial cells, whereas PPARδ overexpression or PPARδ agonist attenuated phosphorylation of p38 MAPK and laminin and type IV collagen secretion. The characteristics of obesity-related glomerulopathy, which might be partly caused by PPARδ suppression-induced p38 MAPK activation and laminin and type IV collagen secretion was demonstrated.

A Comprehensive Nuclear Receptor Network for Breast Cancer Cells

In breast cancer, nuclear receptors (NRs) playa prominent role in governing gene expression, have prognostic utility, and are therapeutic targets. We built a regulatory map for 24 NRs, six chromatin state markers, and 14 breast-cancer-associated transcription factors (TFs) that are expressed in the breast cancer cell line MCF-7. The resulting network reveals a highly interconnected regulatory matrix where extensive crosstalk occurs among NRs and other breast -cancer-associated TFs. We show that large numbers of factors are coordinately bound to highly occupied target regions throughout the genome, and these regions are associated with active chromatin state and hormone-responsive gene expression. This network also provides a framework for stratifying and predicting patient outcomes, and we use it to show that the peroxisome proliferator-activated receptor delta binds to a set of genes also regulated by the retinoic acid receptors and whose expression is associated with poor prognosis in breast cancer.

Circadian clock gene expression is impaired in gestational diabetes mellitus

Dysfunction of the circadian clock genes is involved in the development of obesity and type 2 diabetes (T2D). Since gestational diabetes mellitus (GDM) and T2D share common genetic and phenotypic features, in the present study, we investigated the status of the circadian clock in a cohort of 40 Greek pregnant women with GDM, four with T2D and 20 normal controls. Peripheral blood mRNA transcript levels of 10 clock genes (CLOCK1, BMAL1, PER1, PER2, PER3, PPARΑ, PPARD, PPARG, CRY1 and CRY2) were determined by real-time quantitative PCR. GDM patients expressed significantly lower transcript levels of BMAL1, PER3, PPARD and CRY2 compared to control women (p < 0.05). No significant difference was documented between GDM women maintained either under insulin treatment or diet. A positive correlation was found between the expression of BMAL1 versus CRY2 (r = 0.45, p = 0.003) and BMAL1 versus PPARD (r = 0.43, p = 0.004). Further investigation on the functional relevance of these clock genes, disclosed that expression of PER3 correlated negatively with HbA1C levels (r = −0.36, p = 0.022). These data document for the first time that the expression of BMAL1, PER3, PPARD and CRY2 genes is altered in GDM compared to normal pregnant women and support the notion that deranged expression of clock genes may play a pathogenic role in GDM.

Olive oil polyphenols enhance the expression of cholesterol efflux related genes in vivo in humans. A randomized controlled trial

Both oleic acid and polyphenols have been shown to increase high-density lipoprotein (HDL) cholesterol and to protect HDL from oxidation, a phenomenon associated with a low cholesterol efflux from cells. Our goal was to determine whether polyphenols from olive oil could exert an in vivo nutrigenomic effect on genes related to cholesterol efflux in humans. In a randomized, controlled, cross-over trial, 13 pre/hypertensive patients were assigned 30 ml of two similar olive oils with high (961 mg/kg) and moderate (289 mg/kg) polyphenol content. We found an increase in ATP binding cassette transporter-A1, scavenger receptor class B type 1, peroxisome proliferator-activated receptor (PPAR)BP, PPARα, PPARγ, PPARδ and CD36 gene expression in white blood cells at postprandial after high polyphenol olive oil when compared with moderate polyphenol olive oil intervention (P<.017), with COX-1 reaching borderline significance (P=.024). Linear regression analyses showed that changes in gene expression were related to a decrease in oxidized low-density lipoproteins and with an increase in oxygen radical absorbance capacity and olive oil polyphenols (P<.05). Our results indicate a significant role of olive oil polyphenols in the up-regulation of genes involved in the cholesterol efflux from cells to HDL in vivo in humans. These results are in agreement with previous ones concerning the fact that benefits associated with polyphenol-rich olive oil consumption on cardiovascular risk could be mediated through an in vivo nutrigenomic effect in humans.

Effects of short- and long-chain fatty acids on the expression of stearoyl-CoA desaturase and other lipogenic genes in bovine mammary epithelial cells

Stearoyl-CoA desaturase (SCD) in the bovine mammary gland introduces a cis-double bond at the Δ9 position in a wide range of fatty acids (FA). Several long-chain polyunsaturated fatty acids (PUFA) inhibit expression of SCD, but information on the effect of short-chain fatty acids on mammary SCD expression is scarce. We used a bovine mammary cell line (MAC-T) to assess the effect of acetic acid (Ac) and β-hydroxybutyric acid (BHBA) in comparison with the effect of various long-chain fatty acids on the mRNA expression of the lipogenic enzymes SCD, acetyl-CoA carboxylase (ACACA), fatty acid synthase (FASN) and their associated gene regulatory proteins sterol regulatory element binding transcription factor 1 (SREBF1), insulin-induced gene 1 protein (INSIG1) and peroxisome proliferator-activated receptor alpha (PPARA)and peroxisome proliferator-activated receptor delta (PPARD) by quantitative real-time PCR. MAC-T cells were treated for 12 h without FA additions (CON) or with either 5 mM Ac, 5 mM BHBA, a combination of 5 mM Ac + 5 mM BHBA, 100 μM C16:0, 100 μM C18:0, 100 μM C18:1 cis-9, 100 μM C18:1 trans-11, 100 μM C18:2 cis-9,12 or 100 μM C18:3 cis-9,12,15. Compared with control, mRNA expression of SCD1 was increased by Ac (+61%) and reduced by C18:1 cis-9 (−61%), C18:2 cis-9,12 (−84%) and C18:3 cis-9,12,15 (−88%). In contrast to native bovine mammary gland tissue, MAC-T cells did not express SCD5. Expression of ACACA was increased by Ac (+44%) and reduced by C18:2 cis-9,12 (−48%) and C18:3 cis-9,12,15 (−49%). Compared with control, FASN expression was not significantly affected by the treatments. The mRNA level of SREBF1 was not affected by Ac or BHBA, but was reduced by C18:1 cis-9 (−44%), C18:1 trans-11 (−42%), C18:2 cis-9,12 (−62%) and C18:3 cis-9,12,15 (−68%) compared with control. Expression of INSIG1 was downregulated by C18:0 (−37%), C18:1 cis-9 (−63%), C18:1 trans-11 (−53%), C18:2 cis-9,12 (−81%) and C18:3 cis-9,12,15 (−91%). Both PPARA and PPARD expression were not significantly affected by the treatments. Our results show that Ac upregulated mRNA expression of SCD1 and ACACA in MAC-T cells. The opposite effect of the PUFA C18:2 cis-9,12 and C18:3 cis-9,12,15 on the these genes and the failure of Ac to mimic the PUFA-inhibited SREBF1 and INSIG1 mRNA expression, suggest that Ac can stimulate mammary lipogenesis via a transcriptional regulatory mechanism different from PUFA.

Pleiotropic Effects of Peroxisome Proliferator-Activated Receptor γ and δ in Vascular Diseases

Peroxisome proliferator-activated receptors gamma (PPARγ) and delta (PPARδ) are nuclear receptors that have significant physiological effects on glucose and lipid metabolism. Experimental studies in animal models of metabolic disease have demonstrated their effects on improving lipid profile, insulin sensitivity, and reducing inflammatory responses. PPARγ and -δ are also expressed in the vasculature and their beneficial effects have been examined in various cardiovascular disease models such as atherosclerosis, hypertension, diabetic vascular complications, etc. using pharmacological ligands or genetic tools including viral vectors and transgenic mice. These studies suggest that PPARγ and δ are antiinflammatory, antiatherogenic, antioxidant, and antifibrotic against vascular diseases. Several signaling pathways, effector molecules, as well as coactivators/repressors have been identified as responsible for the protective effects of PPARγ and -δ in the vasculature. We discuss the pleiotropic effect of PPARγ and δ in vascular dysfunction, including atherosclerosis, hypertension, vascular remodeling, vascular injury, and diabetic vasculopathy, in various animal models, and the major underlying mechanisms. We also compare the phenotypes of several endothelial cell/vascular smooth muscle-specific PPARγ and -δ knockout and overexpressing transgenic mice in various disease models, and the implications underlying the functional importance of vascular PPARγ and δ in regulating whole-body homeostasis.

Beneficial effects of omega-3 fatty acids on the consequences of a fructose diet are not mediated by PPAR delta or PGC1 alpha

To study, in high-fructose-fed rats, the effect of a dietary enrichment in omega-3 polyunsaturated fatty acids (n-3 PUFA) on the expression of genes involved in lipid metabolism and cardiovascular function. Twenty-eight male "Wistar Han" rats received for 8 weeks, either a standard chow food or an isocaloric 67% fructose diet enriched or not in alpha-linolenic acid (ALA) or in docosahexaenoic (DHA) and eicosapentaenoic acids (EPA) mix (DHA/EPA). After sacrifice, blood was withdrawn for biochemical analyses; heart, periepididymal adipose tissue and liver were collected and analyzed for the expression of 22 genes by real-time PCR. Fructose intake resulted in an increase in liver weight and triglyceride content, plasma triglyceride and cholesterol concentrations, although no difference in glucose and insulin. In the liver, lipogenesis was promoted as illustrated by an increase in stearoyl-CoA desaturase and fatty acid synthase (Fasn) together with a decrease in PPAR gamma, delta and PPAR gamma coactivator 1 alpha (PGC1 alpha) expression. In the heart, Fasn and PPAR delta expression were increased. The addition of ALA or DHA/EPA into the diet resulted in a protection against fructose effects except for the decreased expression of PPARs in the liver that was not counterbalanced by n-3 PUFA suggesting that n-3 PUFA and fructose act independently on the expression of PPARs and PGC1 alpha. In liver, but not in heart, the fructose-enriched diet induces an early tissue-specific reduction in PPAR gamma and delta expression, which is insensitive to n-3 PUFA intake and dissociated from lipogenesis.

Phosphoiodyns A and B, Unique Phosphorus-Containing Iodinated Polyacetylenes from a Korean Sponge Placospongia sp.

Two unprecedented phosphorus-containing iodinated polyacetylenes, phosphoiodyns A and B (1-2), were isolated from a Korean marine sponge Placospongia sp. Their structures were elucidated by spectroscopic data analysis. Phosphoiodyn A exhibited potent agonistic activity on human peroxisome proliferator-activated receptor delta (hPPARδ) with an EC(50) of 23.7 nM.

Increase of Peroxisome Proliferator-activated Receptor δ (PPARδ) by Digoxin to Improve Lipid Metabolism in the Heart of Diabetic Rats

Increase of peroxisome proliferator-activated receptor δ (PPARδ) expression by digoxin in the heart of diabetic rats has been documented. The present study investigated the mediation of PPARδ in lipid metabolism improved by digoxin in the heart of diabetic rats and in the hyperglycemia-treated cardiomyocytes using the primary cultured cardiomyocytes from neonatal rat. The lipid deposition within the heart section was assessed in diabetic rats by oil red O staining. The fatty acid oxidation genes in cardiomyocytes were also examined. Inhibitor of calcium ions and siRNA-PPARδ were employed to investigate the potential mechanisms. After a 20-day digoxin treatment, the PPARδ expression was elevated in hearts of diabetic rats while the cardiac lipid deposition was reduced. In neonatal cardiomyocytes, digoxin also caused an increase in expressions of PPARδ and fatty acid oxidation genes. But both actions of digoxin were blocked by BAPTA-AM to chelate calcium ions and by siRNA-PPARδ in cardiomyocytes. The obtained results show that increase of PPARδ by digoxin is related to regulation of fatty acid oxidation genes in cardiac cells mediated by calcium-triggered signals.

Angiopoietin-Like 4 Mediates PPAR Delta Effect on Lipoprotein Lipase-Dependent Fatty Acid Uptake but Not on Beta-Oxidation in Myotubes

Peroxisome proliferator-activated receptor (PPAR) delta is an important regulator of fatty acid (FA) metabolism. Angiopoietin-like 4 (Angptl4), a multifunctional protein, is one of the major targets of PPAR delta in skeletal muscle cells. Here we investigated the regulation of Angptl4 and its role in mediating PPAR delta functions using human, rat and mouse myotubes. Expression of Angptl4 was upregulated during myotubes differentiation and by oleic acid, insulin and PPAR delta agonist GW501516. Treatment with GW501516 or Angptl4 overexpression inhibited both lipoprotein lipase (LPL) activity and LPL-dependent uptake of FAs whereas uptake of BSA-bound FAs was not affected by either treatment. Activation of retinoic X receptor (RXR), PPAR delta functional partner, using bexarotene upregulated Angptl4 expression and inhibited LPL activity in a PPAR delta dependent fashion. Silencing of Angptl4 blocked the effect of GW501516 and bexarotene on LPL activity. Treatment with GW501516 but not Angptl4 overexpression significantly increased palmitate oxidation. Furthermore, Angptl4 overexpression did not affect the capacity of GW501516 to increase palmitate oxidation. Basal and insulin stimulated glucose uptake, glycogen synthesis and glucose oxidation were not significantly modulated by Angptl4 overexpression. Our findings suggest that FAs-PPARdelta/RXR-Angptl4 axis controls the LPL-dependent uptake of FAs in myotubes, whereas the effect of PPAR delta activation on beta-oxidation is independent of Angptl4.

INTERACTIONS OF SEVERAL GENETIC POLYMORPHISMS AND CIGARETTE SMOKING ON BLOOD PRESSURE LEVELS

ObjectivesAlthough the relationship between cigarette smoking and cardiovascular diseases is well established, the interactions of some single nucleotide polymorphisms (SNPs) and cigarette smoking on blood pressure levels are still limited....

Several Lipid-Related Gene Polymorphisms Interact with Overweight/Obesity to Modulate Blood Pressure Levels

Little is known about the interactions of single nucleotide polymorphisms (SNPs) and overweight/obesity on blood pressure levels. The present study was undertaken to detect 10 lipid-related gene SNPs and their interactions with overweight/obesity on blood pressure levels. Genotyping of ATP-binding cassette transporter A1 (ABCA-1) V825I, acyl-CoA:cholesterol acyltransferase-1 (ACAT-1) rs1044925, low density lipoprotein receptor (LDL-R) AvaII hepatic lipase gene (LIPC) −250G > A, endothelial lipase gene (LIPG) 584C > T, methylenetetrahydrofolate reductase (MTHFR) 677C > T, the E3 ubiquitin ligase myosin regulatory light chain-interacting protein (MYLIP) rs3757354, proprotein convertase subtilisin-like kexin type 9 (PCSK9) E670G, peroxisome proliferator-activated receptor delta (PPARD) +294T > C, and Scavenger receptor class B type 1 (SCARB1) rs5888 was performed in 978 normal weight and 751 overweight/obese subjects. The interactions were detected by factorial regression analysis. The genotypes of ACAT-1 AC, LIPC GA and AA, and SCARB1 TT; LDL-R A-A- and LIPC GA; and SCARB1 TT were interacted with overweight/obesity to increase systolic, diastolic blood pressure (SBP, DBP) and pulse pressure (PP) levels; respectively. The genotypes of ACAT-1 CC; ACAT-1 AA and CC were interacted with overweight/obesity to decrease SBP, PP levels (p < 0.01–0.001); respectively. The differences in blood pressure levels between normal weight and overweight/obese subjects might partly result from different interactions of several SNPs and overweight/obesity.

Ligand activation of peroxisome proliferator‐activated receptor delta suppresses cathepsin B expression in human endothelial cells in a posttranslational manner

Peroxisome proliferator-activated receptor (PPAR) delta agonists are known to have distinct anti-inflammatory and antitumor effects; though, the knowledge regarding their mode of action has thus far been limited. Different cathepsins have been shown to be upregulated in a broad range of pathological events, such as rheumatoid arthritis, psoriasis, atherosclerosis and diverse tumor entities, for example melanoma. Recent work demonstrated that cathepsin B in particular is an important pro-angiogenic protease in various pathological conditions. We therefore analysed whether cathepsins are a valid target for PPARδ agonists. This study reveals an inhibitory effect of two commonly used PPARδ agonists, GW501516 and L-165,041, on the protein expression and enzyme activity of cathepsin B in human endothelial cells. In contrast, no inhibitory effects were observed on cathepsin L and cathepsin D protein expression after treatment with PPARδ agonists. Furthermore, the results substantiate that PPARδ activators mediate their inhibitory action in a PPARδ-dependent manner and that the underlying regulatory mechanism is not based on a transcriptional but rather on a posttranslational mode of action, via the reduction in the cathepsin B protein half-life. Mechanisms conveying the suppressive effect by 5'-alternative splicing, a 3'-UTR-dependent way or by miRNA could be excluded. The data of this study explore cathepsin B as a new valid target for PPARδ agonists in endothelial cells. The results bolster other studies demonstrating PPARδ agonists as anti-inflammatory and anticarcinogenic agents and thus might have the potential to help to develop new pharmaceutical drugs.

PPARδ agonist GW501516 prevents uncoupling of endothelial nitric oxide synthase in cerebral microvessels of hph-1 mice

Peroxisome proliferator-activated receptor delta (PPARδ) is ubiquitously expressed in the vasculature, including cerebral circulation. The role of PPARδ in metabolism of tetrahydrobiopterin (BH₄) has not been studied in the cerebral microvasculature. In the present study, the effects of PPARδ agonist GW501516 on uncoupling of endothelial nitric oxide synthase (eNOS) were determined in cerebral microvessels of BH₄-deficient hph-1 mice. Wild-type (B6CBA) and hph-1 mice were orally gavaged with a selective PPARδ activator, GW501516 (2 mg/kg/day) for 14 days, and thereafter, cerebral microvessels were isolated and studied. Treatment of hph-1 mice with GW501516 significantly reduced oxidation of BH₄ and increased the ratio of BH₄ to 7,8-BH₂ (P<0.05, n=6-9). Attenuation of L-NAME-inhibitable superoxide anion levels by GW501516 demonstrated that activation of PPARδ might prevent uncoupling of endothelial nitric oxide synthase (eNOS, P<0.05, n=6-9). Western blotting studies demonstrated that GW501516 selectively increased the endothelial expressions of CuZn superoxide dismutase (P<0.05, n=6-9) and catalase (P<0.05, n=6-8). PPARδ activation increased the total nitrite and nitrate (NO₂+NO₃) content in cerebral microvessels (P<0.05, n=6). Obtained results suggest that in vivo activation of PPARδ prevents eNOS uncoupling, restores bioavailability of NO and may help preserve endothelial function in the BH₄-deficient cerebral circulation.

Peroxisome proliferator-activated receptor delta agonist attenuates nicotine suppression effect on human mesenchymal stem cell-derived osteogenesis and involves increased expression of heme oxygenase-1

Smoking has long been associated with osteoporosis, decreased bone mineral density, increased risk of bone fracture, and increased health costs. Nicotine, the main component of cigarette smoke, has major negative effects on bone metabolism and skeletal remodeling in vivo. Although osteoblasts and osteoblast-like cells have been used extensively to study the impact of nicotine, few studies have been performed on human mesenchymal stem cells (hMSCs). In this context, we examined the impact of nicotine on (a) hMSCs proliferation, (b) osteoblastic differentiation, (c) alkaline phosphatase (ALP) activity, and (d) expression of canonical genes during differentiation of hMSCs. MSCs isolated from human bone marrow were treated with different concentrations (0, 0.1, 1 and 10μM) of nicotine for 7days. Nicotine caused a dose-dependent decrease in cell proliferation, decreased heme oxygenase-1 (HO-1) expression (p<0.05) and attenuated osteogenesis (p<0.05) in hMSCs (45% reduction at day 14). In addition, nicotine caused a dose-dependent decrease in alizarin red staining for calcium and staining for ALP. Induction of HO-1 by peroxisome proliferator-activated receptor delta agonist (GW0742) prevented the effect of nicotine. Nicotine caused a dose-dependent reduction in the expression of BMP-2, a well-known marker for bone formation; however, this was prevented by GW0742 treatment. Therefore, induction of HO-1 prevents the deleterious effects of nicotine on osteogenesis in hMSC. This offers insight into both how nicotine affects bone remodeling and a therapeutic approach to prevent fracture and osteoporosis in smokers.