Receptor PPARα Research Articles

Differential effects on lung cancer cell proliferation by agonists of glucocorticoid and PPARα receptors

Glucocorticoids (GCs) are well-known anti-inflammatory compounds, but they also inhibit cell proliferation depending on cell type. Similarly, peroxisome proliferator-activated receptors (PPARα, PPARδ, and PPARγ) also possess anti-proliferation properties beyond their canonical roles as metabolic mediators. In the present study, we investigated the potential additive or synergistic inhibitory effects on cancer cell proliferation by simultaneous application of fenofibrate and budesonide, agonists for PPARα and glucocorticoid receptor, respectively. We observed differential effects on cell proliferation in A549 and SK-MES-1 lung cancer cells by budesonide and fenofibrate. Fenofibrate inhibited cell proliferation in both TP53 wild type and deficient lung cancer cells. The anti-proliferation effect of budesonide in TP53 wild type A549 cells was abolished in SK-MES-1 cells that do not have wild type TP53 protein. An additive effect against cell proliferation by budesonide and fenofibrate combination was observed only in TP53 wild type A549 cancer cells. Analysis of cell cycle distribution and cyclin profile indicated that the inhibition of cell proliferation was associated with G1 cell cycle arrest. The suppression of NF-κB activity and ERK signaling may contribute to the inhibition of cell proliferation by budesonide and or fenofibrate. The additive inhibitory effect on cell proliferation by budesonide and fenofibrate combination suggests that the same or greater therapeutic effect could be achieved with reduced dosage and side effects when the two compounds are applied simultaneously.

Nutrigenomics – bioactive dietary components

Nutrigenomics analyzes relations between diet and genes, and identifies mechanisms in which food and nutrition affect health and lifestyles and noncommunicable diseases (R. Chadwick, 2004). Bioactive dietary components are signal molecules that carry information from the external environment and affect in terms of quantity and quality in the process of gene expression. The biological effect of bioactive dietary components depends on various of physiological processes that can occur within a few genes. Polymorphism of genes can change their function and physiological response of the body for nutrients. Bioactive dietary components work on at least two levels of the expression of genes as factors regulating chromatin structure and as factors directly regulate the activity of nuclear receptors. The processes of synthesis and DNA repair are regulated by some of vitamins, macro-and micro-elements. They provide, among others, cofactors of enzymes that catalyze the replication of DNA methylation and its repair. DNA methylation profile may change under the influence of diet, single nucleotide polymorphisms and environmental factors. Bioactive dietary components may directly affect the process of gene expression by acting as ligands for nuclear receptors. Sensitive to dietary group of nuclear receptors are sensory receptors. This group includes, among others receptor PPAR (peroxisome proliferator activated), responsible for energy metabolism and receptors LXR (liver X receptor), FXR (farnesoid X receptor) and RXR, which is responsible for the metabolism of cholesterol.

1944 UPREGULATION OF ANGIOTENSIN II TYPE 1 AND TYPE 2 RECEPTORS IN AGED RATS WITH BLADDER DYSFUNCTION

INTRODUCTION AND OBJECTIVES: Angiotensin II is reportedly involved in the bladder remodeling underlying bladder dysfunction induced by bladder outlet obstruction, and exerts its effects through angiotensin II type 1 (AT1) and type 2 receptors (AT2). AT1 activation induces organ fibrosis, MAP kinase (MAPK) activation and junctional protein Cx43 expression while AT2 activation increases PPARactivation and adiponectin (Adipo), which protects against AT1-induced fibrosis. This study examined changes in AT1 or AT2 mechanisms in the development of bladder dysfunction during aging. Therefore, we investigated alternations in bladder function and expression of AT1, AT2 and related molecules of the bladder and urethra in aged rats with bladder dysfunction. METHODS: Female SD rats of three different ages at 8 weeks old (8W) (n 5), 9 months old (9M) (n 5) and 15 months old (15M) (n 5) were used. Cystometry was performed under urethane anesthesia, and urodynamic parameters were evaluated, including non-voiding contraction (NVC), pressure threshold (PT), maximum voiding pressure (MVP) and post-void residual volume (RV). After cystometry, the bladder and urethra were excised. The bladder was separated into mucosa and detrusor muscle layers under a microscope. The protein and mRNA expression levels of AT1, Cx43, MAPK, AT2 PPAR, Adipo and adiponectin receptor (Adipo-R) were investigated by Western blot and RT-PCR, respectively. RESULTS: PT, RV and the number of NVCs were significantly increased in 15M vs. 8W rats (P 0.05). MVP was significantly decreased in 15M rats, compared to 8W rats (P 0.05). However, there was no significant difference in any parameters between 8W and 9M rats. In bladder and urethral tissues, the protein and mRNA expression of AT1, Cx43, and MAPK1/2 were significantly increased in 15M than in 8W rats (P 0.05). AT2, PPAR, Adipo, and AdipoR1 were also significantly increased in 15M vs. 8W rats. CONCLUSIONS: These results suggest that, during the aging process, both bladder overactivity and impaired voiding, which are often seen in aged patients, are induced by upregulation of AT1, which can increase Cx43 expression via activation of MAPK signaling pathways. Furthermore, increased adiponectin through PPARactivation, which is regulated by AT2, may protect against AT1-induced bladder fibrosis and dysfunction. Therefore, suppression of AT1 and/or stimulation of PPARvia AT2 could be effective for aged patients with bladder dysfunction.

Novel Acylethanolamide Derivatives That Modulate Body Weight through Enhancement of Hypothalamic Pro-Opiomelanocortin (POMC) and/or Decreased Neuropeptide Y (NPY)

Newly synthesized acylethanolamide derivatives oleoyl-L-valinolamide (1), oleoyl-D-valinolamide (2), elaidoyl-L-valinolamide (3), elaidoyl-D-valinolamide (4) stearoyl-L-valinolamide (5), and palmitoyl-L-valinolamide (6) were investigated in mice as antiobesity compounds. Compounds 1, 2, 5, 6 significantly decreased body weight by 6.57% following eight injections of 1 mg/kg i.p. during 39 days, while 3 and 4 showed no such activity. Receptor binding indicated that no compound activated CB1, CB2, PPARα, or TRPV1 receptors. Hypothalamic RT-PCR showed that mRNA expression of the anorexigenic genes POMC and CART was up-regulated by 1, 2, 5 and 1, 2, respectively, while that of the orexigenic genes NPY and CaMKK2 was down-regulated by the respective compounds 1, 5, 6 and 1, 2, 5. Oleoyl-L-valinolamide enhances anorectic pathways and lead to decreased glucose levels, enhanced locomotor activity, and improved cognition. Effects of oleoyl-L-valinolamide on weight were dose-dependent, and it could be given orally. 1, 2, 4, 5 down-regulated FAAH mRNA expression.

Open tubular columns containing the immobilized ligand binding domain of peroxisome proliferator-activated receptors α and γ for dual agonists characterization by frontal affinity chromatography with mass spectrometry detection

The peroxisome proliferator-activated receptors (PPARs) belong to the nuclear receptor superfamily. In the last years novel PPARs ligands have been identified and these include PPARα/γ dual agonists. To rapidly identify novel PPARs dual ligands, a robust binding assay amenable to high-throughput screening toward PPAR isoforms would be desirable. In this work we describe a parallel assay based on the principles of frontal affinity chromatography coupled to mass spectrometry (FAC-MS) that can be used to characterize dual agonists. For this purpose the ligand binding domain of PPARα receptor was immobilized onto the surface of open tubular capillaries to create new PPAR-alpha-OT columns to be used in parallel with PPAR-gamma-OT columns. The two biochromatographic systems were used in both ranking and Kd experiments toward new ureidofibrate-like dual agonists for subtype selectivity ratio determination. In order to validate the system, the Kd values determined by frontal analysis chromatography were compared to the affinity constants obtained by ITC experiments. The results of this study strongly demonstrate the specific nature of the interaction of the ligands with the two immobilized receptor subtypes.

Triterpenoid-Rich Fraction from Ilex hainanensis Merr. Attenuates Non-Alcoholic Fatty Liver Disease Induced by High Fat Diet in Rats

Non-alcoholic fatty liver disease (NAFLD) has become a major challenge to the healthcare system. This study was designed to evaluate the effect of the triterpenoid-rich fraction (TF) from Ilex hainanensis Merr. on NAFLD. Male Sprague-Dawley (SD) rats were fed a normal diet (control) or high fat diet (NAFLD model). After four weeks, the high fat diet group was orally administrated TF (250 mg/kg) for another two weeks. High fat diet fed rats displayed hyperlipidemia and a decline in liver function compared with control. However, administration with TF could effectively improve these symptoms, as demonstrated by decreasing the plasma levels of triglyceride (p <0.05), total cholesterol (p < 0.01), low-density lipoprotein cholesterol (p < 0.05), alanine transaminase (p < 0.05), aspartate aminotransferase (p < 0.01), liver index (p < 0.05) and insulin resistance index (p < 0.05) while increasing the high-density lipoprotein cholesterol (p < 0.05). Meanwhile, histopathological examination of livers also showed that TF could reduce the incidence of liver lesions induced by high fat diet. Furthermore, TF could alleviate oxidative stress and inflammation status indicated by the decline malondialdehyde and superoxide dismutase levels (p < 0.01, both) and levels of interleukin 6 and tumor necrosis factor-α (p < 0.05). In addition, immunohistochemistry showed TF evidently elevated the peroxisome proliferator-activated receptor (PPARα) expression (p < 0.01), while it diminished the Cytochrome P450 2E1 (CYP2E1) expression (p < 0.01) in liver. These results demonstrate that TF has potential ability to protect liver against NAFLD by regulating lipids metabolism and alleviating insulin resistance, inflammation and oxidative stress. This effect might be associated with regulating PPARα and CYP2E1 expression.

Targeting peroxisome proliferator-activated receptors (ppar) for the treatment l-dopa-induced dyskinesias

L administration of L-DOPA to Parkinson’s disease (PD) patients is accompanied by fluctuations in its duration of action and chorea-like motor complications known as L-DOPA-induced dyskinesias (LID). As L-DOPA is the most effective treatment for PD, there is a strong need to identify new pharmacological targets to control LID while maintaining L-DOPA beneficial effects.LID can be modeled in rats with unilateral 6-OHDA lesions via chronic administration of L-DOPA, which induces increasingly severe axial, limb and oro-facial abnormal involuntary movements (AIMs). Previous studies in our laboratory showed that systemic administration of the PPAR gamma agonist rosiglitazone (5 & 10 mg/kg, i.p.) ameliorated axial, oral and limb AIMs without reducing L-DOPA’s anti-parkinsonian activity. Rosiglitazone also attenuated the L-DOPA-induced elevation of dyskinesia (Zif-268 and prodynorphin) and inflammation (COX-2) markers within the denervated striatum. Interestingly, chronic L-DOPA significantly increased the PPAR beta/delta isoform in dyskinetic rats and this elevation was reversed by rosiglitazone treatment. These data suggest that targeting PPAR gamma/delta receptors can ameliorate dyskinetic symptoms, possibly by reducing the underlying neuroinflammatory response observed in PD. Supported by RO1 -NS050401-06 (A.G).

Oleoylethanolamide enhances β-adrenergic-mediated thermogenesis and white-to-brown adipocyte phenotype in epididymal white adipose tissue in rat

β-adrenergic receptor activation promotes brown adipose tissue (BAT) β-oxidation and thermogenesis by burning fatty acids during uncoupling respiration. Oleoylethanolamide (OEA) can inhibit feeding and stimulate lipolysis by activating peroxisome proliferator-activating receptor-α (PPARα) in white adipose tissue (WAT). Here we explore whether PPARα activation potentiates the effect of β3-adrenergic stimulation on energy balance mediated by the respective agonists OEA and CL316243. The effect of this pharmacological association on feeding, thermogenesis, β-oxidation, and lipid and cholesterol metabolism in epididymal (e)WAT was monitored. CL316243 (1 mg/kg) and OEA (5 mg/kg) co-administration over 6 days enhanced the reduction of both food intake and body weight gain, increased the energy expenditure and reduced the respiratory quotient (VCO2/VO2). This negative energy balance agreed with decreased fat mass and increased BAT weight and temperature, as well as with lowered plasma levels of triglycerides, cholesterol, nonessential fatty acids (NEFAs), and the adipokines leptin and TNF-α. Regarding eWAT, CL316243 and OEA treatment elevated levels of the thermogenic factors PPARα and UCP1, reduced p38-MAPK phosphorylation, and promoted brown-like features in the white adipocytes: the mitochondrial (Cox4i1, Cox4i2) and BAT (Fgf21, Prdm16) genes were overexpressed in eWAT. The enhancement of the fatty-acid β-oxidation factors Cpt1b and Acox1 in eWAT was accompanied by an upregulation of de novo lipogenesis and reduced expression of the unsaturated-fatty-acid-synthesis enzyme gene, Scd1. We propose that the combination of β-adrenergic and PPARα receptor agonists promotes therapeutic adipocyte remodelling in eWAT, and therefore has a potential clinical utility in the treatment of obesity.

Expression of adiponectin receptors 1 (AdipoR1) and 2 (AdipoR2) gene in the porcine pituitary during the oestrous cycle

Aquaporins (AQPs) are trans-membrane proteins which allow the movement of water and glycerol required by hepatic stellate cells (HSC) for triglyceride formation and lipid storage. Adiponectin (ADPQ) is a hormone produced by the adipose tissue, which is known to increase AQP3 expression. Since ADPQ receptor signals via the nuclear receptor PPAR we aimed to explore the role of this pathway in AQP3 regulation by ADPQ in HSC. AQP3 and CPT1α expression increased only after ADPQ but not rosiglitazone stimulation. In LX2 cells co-transfected with plasmids expressing PPARα or PPARγ coupled to a luciferase reporter gene, only PPARα increased luciferase activity after ADPQ stimulation. Collectively, our findings demonstrate that ADPQ increases AQP3 expression through PPARα-mediated signaling in HSC.

Antiepileptic action of N-palmitoylethanolamine through CB1 and PPAR-α receptor activation in a genetic model of absence epilepsy

N-palmitoylethanolamine (PEA), an endogenous fatty acid ethanolamide, plays a key role in the regulation of the inflammatory response and pain through, among others, activation of nuclear peroxisome proliferator-activated receptors (PPAR-α). Endogenous cannabinoids play a protective role in several central nervous system (CNS) disorders, particularly those associated with neuronal hyperexcitability. We investigated the effects of PEA and the role of PPAR-α in absence epilepsy using the WAG/Rij rat model. PEA, anandamide (AEA), a PPAR-α antagonist (GW6471) and a synthetic CB1 receptor antagonist/inverse agonist (SR141716) were administered to WAG/Rij rats in order to evaluate the effects on epileptic spike-wave discharges (SWDs) on EEG recordings. We studied also the effects of PEA co-administration with SR141716 and GW6471 and compared these effects with those of AEA to evaluate PEA mechanism of action and focusing on CB1 receptors and PPAR-α. Both PEA and AEA administration significantly decreased SWDs parameters (absence seizures). In contrast, GW6471 was devoid of effects while SR141716 had pro-absence effects. The co-administration of SR141716 with PEA or AEA completely blocked the anti-absence effects of these compounds. GW6471 antagonized PEA's effects whereas it did not modify AEA's effects.Furthermore, we have also measured PEA, AEA and 2-AG (2-arachidonoylglycerol) brain levels identifying significant differences between epileptic and control rats such as decreased PEA levels in both thalamus and cortex that might contribute to absence epilepsy. Our data demonstrate that PEA has anti-absence properties in the WAG/Rij rat model and that such properties depend on PPAR-α and indirect activation of CB1 receptors.This article is part of the Special Issue entitled ‘New Targets and Approaches to the Treatment of Epilepsy’.

Design and synthesis of alkoxyindolyl-3-acetic acid analogs as peroxisome proliferator-activated receptor-γ/δ agonists

A series of carbazole or phenoxazine containing alkoxyindole-3-acetic acid analogs were prepared as PPARγ/δ agonists and their transactivation activities for PPAR receptor subtypes (α, γ and δ) were investigated. Structure–activity relationship studies disclosed the effect of the lipophilic tail, attaching position of the alkoxy group and N-benzyl substitution at indole. Compound 1b was the most potent for PPARδ and 3b for PPARγ. Molecular modeling suggested two different binding modes of our alkoxyindole-3-acetic acid analogs providing the insight into their PPAR activity.

Oleoylethanolamide dose‐dependently attenuates cocaine‐induced behaviours through a PPARα receptor‐independent mechanism

Oleoylethanolamide (OEA) is an acylethanolamide that acts as an agonist of nuclear peroxisome proliferator-activated receptor alpha (PPARα) to exert their biological functions, which include the regulation of appetite and metabolism. Increasing evidence also suggests that OEA may participate in the control of reward-related behaviours. However, direct experimental evidence for the role of the OEA-PPARα receptor interaction in drug-mediated behaviours, such as cocaine-induced behavioural phenotypes, is lacking. The present study explored the role of OEA and its receptor PPARα on the psychomotor and rewarding responsiveness to cocaine using behavioural tests indicative of core components of addiction. We found that acute administration of OEA (1, 5 or 20 mg/kg, i.p.) reduced spontaneous locomotor activity and attenuated psychomotor activation induced by cocaine (20 mg/kg) in C57Bl/6 mice. However, PPARα receptor knockout mice showed normal sensitization, although OEA was capable of reducing behavioural sensitization with fewer efficacies. Furthermore, conditioned place preference and reinstatement to cocaine were intact in these mice. Our results indicate that PPARα receptor does not play a critical, if any, role in mediating short- and long-term psychomotor and rewarding responsiveness to cocaine. However, further research is needed for the identification of the targets of OEA for its inhibitory action on cocaine-mediated responses.

Scaffold-Based Pan-Agonist Design for the PPARα, PPARβ and PPARγ Receptors

As important members of nuclear receptor superfamily, Peroxisome proliferator-activated receptors (PPAR) play essential roles in regulating cellular differentiation, development, metabolism, and tumorigenesis of higher organisms. The PPAR receptors have 3 identified subtypes: PPARα, PPARβ and PPARγ, all of which have been treated as attractive targets for developing drugs to treat type 2 diabetes. Due to the undesirable side-effects, many PPAR agonists including PPARα/γ and PPARβ/γ dual agonists are stopped by US FDA in the clinical trials. An alternative strategy is to design novel pan-agonist that can simultaneously activate PPARα, PPARβ and PPARγ. Under such an idea, in the current study we adopted the core hopping algorithm and glide docking procedure to generate 7 novel compounds based on a typical PPAR pan-agonist LY465608. It was observed by the docking procedures and molecular dynamics simulations that the compounds generated by the core hopping and glide docking not only possessed the similar functions as the original LY465608 compound to activate PPARα, PPARβ and PPARγ receptors, but also had more favorable conformation for binding to the PPAR receptors. The additional absorption, distribution, metabolism and excretion (ADME) predictions showed that the 7 compounds (especially Cpd#1) hold high potential to be novel lead compounds for the PPAR pan-agonist. Our findings can provide a new strategy or useful insights for designing the effective pan-agonists against the type 2 diabetes.

Gene-gene interactions among the peroxisome proliferator-activated receptor polymorphisms for hypertriglyceridemia

To investigate the association of ten SNP at peroxisome proliferator-activated receptors (PPARα, δ, γ) with hypertriglyceridemia and the gene-gene interaction. Participants were recruited from the Prevention of MetS and Multi-metabolic Disorders in Jiangsu province of China Study (PMMJS). A total of 820 subjects were selected from the 4083 participants who had received follow-up examination, by using simple random sampling. Participants in baseline and follow-up study surveys were both collected blood samples 11 ml in the morning after at least 8 hours of fasting. Blood samples which collected at the baseline were subjected to PPARα, PPARδ and PPARγ genotype analyses. Blood samples which collected at the follow-up were used to measure serum triglyceride levels. The logistic regression model was used to analyze the association between different SNP and hypertriglyceridemia, and the generalized multifactor dimensionality reduction (GMDR) was applied to explore the gene-gene interaction. The samples included 474 in the non-hypertriglyceridemia group and 346 in the hypertriglyceridemia group. The genotype frequencies of rs1800206 in the hypertriglyceridemia group were 211 (61.0%) for LL, 132 (38.2%) for LV and 3 (0.9%) for VV, and in the non-hypertriglyceridemia group were 411 (86.7%) for LL, 59 (12.4%) for LV and 4(0.8%) for VV (χ(2) = 74.18, P < 0.01). V allele frequencies of rs1800206 in the hypertriglyceridemia group was 138(19.9%), and in the non-hypertriglyceridemia group was 67 (7.1%) (χ(2) = 60.62, P < 0.01). The genotype frequencies of rs2016520 in the hypertriglyceridemia group were 177 (51.2%) for TT, 154 (44.5%) for TC and 15 (4.3%) for CC, and in the non-hypertriglyceridemia group were 211 (44.5%) for TT, 212 (44.7%) for TC and 51 (10.8%) for CC(χ(2) = 15.93, P < 0.01). C allele frequencies of rs2016520 in the hypertriglyceridemia group was 184(26.6%), and in the non-hypertriglyceridemia group was 314 (33.1%) (χ(2) = 8.07, P < 0.01). The genotype frequencies of rs3856806 in the hypertriglyceridemia group were 149 (43.1%) for CC, 156 (45.1%) for CT and 41 (11.8%) for TT, and in the non-hypertriglyceridemia group were 269 (56.8%) for CC, 170 (35.9%) for CT and 35 (7.4%) for TT (χ(2) = 15.93, P < 0.01). T allele frequencies of rs3856806 in the hypertriglyceridemia group was 238(34.4%), and was 240 (25.3%) in the non-hypertriglyceridemia group (χ(2) = 15.96, P < 0.01). The genotype frequencies of rs1805192 in the hypertriglyceridemia group were 145 (41.9%) for PP, 158(45.7%) for PA and 43(12.4%) for AA, and in the non-hypertriglyceridemia group were 314 (66.2%) for PP, 137(28.9%) for PA and 23(4.9%) for AA (χ(2) = 50.92, P < 0.01). A allele frequencies of rs1805192 in the hypertriglyceridemia group was 244(35.2%), and was 183 (19.3%) in the non-hypertriglyceridemia group(χ(2) = 52.89, P < 0.01). After adjusting age, gender, smoking, alcohol consumption, high-fat diet, low -fiber diet and occupational physical activity factors, rs1800206, rs2016520, rs3856806 and rs1805192 were significantly associated with hypertriglyceride, while the OR (95%CI) was 3.88 (2.69 - 5.60), 0.71 (0.52 - 0.96), 1.40 (1.03 - 1.90) and 2.56 (1.88 - 3.49), respectively (P < 0.05). GMDR model analysis showed that the second-order model (rs1800206 and rs1805192) was the best model when quality traits of triglyceride was chosen as outcome (P < 0.01); while third-order model (rs1800206, rs1805192 and rs2016520) was the best model when quantitative traits of triglyceride was chosen as outcome (P < 0.01). The rs1800206, rs2016520, rs3856806 and rs1805192 were significantly associated with hypertriglyceridemia. There was a gene-gene interaction between multiple SNP.

Selective regulation of peroxisome proliferator-activated receptors on fatty acid binding protein-4 in human syncytiotrophoblast cells

To observe the selective regulation of peroxisome proliferator-activated receptors (PPAR) on fatty acid binding protein-4 (FABP4) in human syncytiotrophoblasts. Cultivate normal human syncytiotrophoblast cells, and put in the specific antagonists and agonists of PPAR each subtypes receptors, then observe the different expression of FABP4 mRNA and protein. Pretreated the human syncytiotrophoblast cells with the agonists (GW7647, GW0742) and antagonists (GW6471, GSK0660) of PPARα and PPARβ receptors, the expression of the FABP4 was not significantly change (P > 0.05). However pretreated with PPAR γ agonists (rosiglitazone, 1×10(-9), 1×10(-8), 1×10(-7) and 1×10(-6) mol/L), the expression of FABP4 mRNA and protein could be dose dependent-promoted significantly (mRNA: 1.27 ± 0.12, 1.45 ± 0.14, 1.57 ± 0.14, 1.72 ± 0.12, protein:1.10 ± 0.08, 1.37 ± 0.09, 1.60 ± 0.13, 1.79 ± 0.14; P < 0.05), furthermore, the promotion can be dose dependent-reversed by specific antagonists GW9662 (mRNA:0.92 ± 0.06, 0.77 ± 0.06, 0.64 ± 0.05, 0.55 ± 0.05, protein:0.91 ± 0.03, 0.78 ± 0.06, 0.70 ± 0.07, 0.55 ± 0.06; P < 0.05). In normal human syncytiotrophoblast cells, FABP4 is a target factor of PPARγ. PPARγ regulated the expression of FABP4 mRNA and protein selectively. And the regulation will not be influenced by the other two PPAR subtypes.

Peroxisome Proliferator-Activated Receptor α Mediates Acute Effects of Palmitoylethanolamide on Sensory Neurons

The amplitude of the depolarization-evoked Ca2+ transient is larger in dorsal root ganglion (DRG) neurons from tumor-bearing mice compared with that of neurons from naive mice, and the change is mimicked by coculturing DRG neurons with the fibrosarcoma cells used to generate the tumors (Khasabova et al., 2007). The effect of palmitoylethanolamide (PEA), a ligand for the peroxisome proliferator-activated receptor α (PPARα), was determined on the evoked-Ca2+ transient in the coculture condition. The level of PEA was reduced in DRG cells from tumor-bearing mice as well as those cocultured with fibrosarcoma cells. Pretreatment with PEA, a synthetic PPARα agonist (GW7647), or ARN077, an inhibitor of the enzyme that hydrolyzes PEA, acutely decreased the amplitude of the evoked Ca2+ transient in small DRG neurons cocultured with fibrosarcoma cells. The PPARα antagonist GW6471 blocked the effect of each. In contrast, the PPARα agonist was without effect in the control condition, but the antagonist increased the amplitude of the Ca2+ transient, suggesting that PPARα receptors are saturated by endogenous ligand under basal conditions. Effects of drugs on mechanical sensitivity in vivo paralleled their effects on DRG neurons in vitro. Local injection of ARN077 decreased mechanical hyperalgesia in tumor-bearing mice, and the effect was blocked by GW6471. These data support the conclusion that the activity of DRG neurons is rapidly modulated by PEA through a PPARα-dependent mechanism. Moreover, agents that increase the activity of PPARα may provide a therapeutic strategy to reduce tumor-evoked pain.

Human-specific SNP in obesity genes, adrenergic receptor beta2 (ADRB2), Beta3 (ADRB3), and PPAR γ2 (PPARG), during primate evolution.

Adrenergic-receptor beta2 (ADRB2) and beta3 (ADRB3) are obesity genes that play a key role in the regulation of energy balance by increasing lipolysis and thermogenesis. The Glu27 allele in ADRB2 and the Arg64 allele in ADRB3 are associated with abdominal obesity and early onset of non-insulin-dependent diabetes mellitus (NIDDM) in many ethnic groups. Peroxisome proliferator-activated receptor γ (PPARG) is required for adipocyte differentiation. Pro12Ala mutation decreases PPARG activity and resistance to NIDDM. In humans, energy-expense alleles, Gln27 in ADRB2 and Trp64 in ADRB3, are at higher frequencies than Glu27 and Arg64, respectively, but Ala12 in PPARG is at lower frequency than Pro12. Adaptation of humans for lipolysis, thermogenesis, and reduction of fat accumulation could be considered by examining which alleles in these genes are dominant in non-human primates (NHP). All NHP (P. troglodytes, G. gorilla, P. pygmaeus, H. agilis and macaques) had energy-thrifty alleles, Gly16 and Glu27 in ADRB2, and Arg64 in ADRB3, but did not have energy-expense alleles, Arg16, Gln27 and Trp64 alleles. In PPARG gene, all NHP had large adipocyte accumulating type, the Pro12 allele.ConclusionsThese results indicate that a tendency to produce much more heat through the energy-expense alleles developed only in humans, who left tropical rainforests for savanna and developed new features in their heat-regulation systems, such as reduction of body hair and increased evaporation of water, and might have helped the protection of entrails from cold at night, especially in glacial periods.

Synthesis, radiolabeling and initial in vivo evaluation of [11C]KSM-01 for imaging PPAR-α receptors

Peroxisome proliferator-activated receptor alpha (PPAR-α) is a ligand-activated nuclear receptor transcription factor that regulates the fatty acid β-oxidation. An in vitro assay identified the p-methoxy phenyl ureido thiobutyric acid derivative KSM-01 (IC50=0.28±0.09nM) having a higher affinity to activate PPAR-α than the PPAR-α agonist GW7647 (IC50=0.46±0.19nM). In this study, we report the synthesis and initial in vivo evaluation of [11C]KSM-01. The radiosynthesis was carried out by first alkylating the corresponding p-phenol precursor with [11C]MeI in DMF using NaOH, followed by deprotection of the t-butyl ester group by TFA, yielding [11C]KSM-01. SUV analysis of dynamic micro PET/CT imaging data showed that [11C]KSM-01 accumulation was ∼2.0-fold greater in cardiac-specific PPAR-α overexpressing transgenic mice compared to wild-type littermates. The post-PET biodistribution studies were consistent with these results and demonstrated 2.5-fold greater radiotracer uptake in the heart of transgenic mice compared to the wild-type littermates. These results demonstrate the potential utility of PPAR-α agonists as PET radiopharmaceuticals.

Effects of Palmitoylethanolamide on Aqueous Humor Outflow

To study the effects of palmitoylethanolamide (PEA), a fatty acid ethanolamide, on aqueous humor outflow facility. The effects of PEA on outflow facility were measured using a porcine anterior segment-perfused organ culture model. The involvements of different receptors in PEA-induced changes were investigated using receptor antagonists and adenovirus delivered small hairpin RNAs (shRNAs). PEA-induced activation of p42/44 mitogen-activated protein kinase (MAPK) was determined by Western blot analysis using an antiphospho p42/44 MAPK antibody. PEA caused a concentration-dependent enhancement of outflow facility, with the maximum effect (151.08 ± 11.12% of basal outflow facility) achieved at 30 nM of PEA. Pretreatment of anterior segments with 1 μM cannabinoid receptor 2 antagonist SR144528 and 1 μM PPARα antagonist GW6471, but not 1 μM cannabinoid receptor 1 antagonist SR141716A, produced a partial antagonism on the PEA-induced increase of outflow facility. Treatment of TM cells with PEA for 10 minutes activated phosphorylation of p42/44 MAPK, which was blocked by pretreatment with SR1444528 and GW6471, but not SR141716A. Knocking down the expression of either GPR55 or PPARα receptors with specific shRNAs for these receptors partially blocked PEA-induced increase in outflow facility and abolished PEA-induced phosphorylation of p42/44 MAPK. PD98059, an inhibitor of the p42/44 MAPK pathway, blocked both PEA-induced enhancement of aqueous humor outflow facility and PEA-induced phosphorylation of p42/44 MAPK. Our results demonstrate that PEA increases aqueous humor outflow through the TM pathway and these effects are mediated by GPR55 and PPARα receptors through activation of p42/44 MAPK.

Constituents from Bupleurum chinense and their effects on PPARα and related nuclear receptors

Constituents from Bupleurum chinense and their effects on PPARα and related nuclear receptors