Expression Of PPARγ Target Genes Research Articles

High-fat diet induced obesity promotes inflammation, oxidative stress, and hepatotoxicity in female FVB/N mice.

Although obesity and subsequent liver injury are increasingly prevalent in women, female mouse models have generally shown resistance to high-fat diet (HFD)-induced obesity. We evaluated control and HFD-fed male and female FVB/N mice, a strain well-suited to transgenic analyses, for phenotypic, histological, and molecular markers related to control of glucose, lipids, and inflammation in serum, liver, and perigonadal white adipose tissues. Unlike many mouse models, HFD-fed FVB/N females gained more perigonadal and mesenteric fat mass and overall body weight than their male counterparts, with increased hepatic expression of lipogenic PPARγ target genes (Cd36, Fsp27, and Fsp27β), oxidative stress genes and protein (Nqo1 and CYP2E1), inflammatory gene (Mip-2), and the pro-fibrotic gene Pai-1, along with increases in malondialdehyde and serum ALT levels. Further, inherent to females (independently of HFD), hepatic antioxidant heme oxygenase-1 (HMOX1, HO-1) protein levels were reduced compared to their male counterparts. In contrast, males may have been relatively protected from HFD-induced oxidative stress and liver injury by elevated mRNA and protein levels of hepatic antioxidants BHMT and Gpx2, increased fatty acid oxidation genes in liver and adipocytes (Pparδ), despite disorganized and inflamed adipocytes. Thus, female FVB/N mice offer a valuable preclinical, genetically malleable model that recapitulates many of the features of diet-induced obesity and liver damage observed in human females.

Maternal obesity attenuates PPARγ nuclear migration impairing offspring adipogenesis.

Maternal obesity predisposes offspring to obesity in adulthood. Since the perinatal period is a critical window for adipose organogenesis, we evaluated if maternal obesity affects the perinatal offspring adipogenesis. Female mice were fed a standard diet (eutrophic dam, ED) or a high-fat diet supplemented with condensed milk (obese dam, OD) for 6 weeks before mating, and the diets were maintained until the end of the protocol. Inguinal adipose tissue of offspring at gestational day 16.5 (E16.5), postnatal day 0 (P0), and P2 was collected to analyze morphological and molecular features. In OD offspring, the number of preadipocytes increased at E16.5 and P0 compared to ED offspring. The cell cycle-related elements Ccnd1 and Ki67 were also upregulated in these groups. In parallel, lipid accumulation started at E16.5 in OD offspring, while ED offspring preadipocytes only accumulated lipids after P0. Peroxisome proliferator-activated receptor gamma (PPARγ) levels and activity were decreased in OD offspring due to impaired nuclear migration. Increased Hdac1 expression, which negatively regulates PPAR-responsive elements in the genome, was also detected. At P2, OD adipocytes presented abnormal features, including a clustered distribution and decreased expression of PPARγ target genes and Adbr3 and Slc2a4, which are highly expressed in mature functional adipocytes. The abnormal adipose tissue is one of the major factors promoting metabolic abnormalities in adulthood. This study demonstrates for the first time the morphological and molecular alterations induced by maternal obesity in vivo in the perinatal adipogenesis in murine inguinal adipose tissue.

234-LB: Cell-State Dependent Regulation of PPARg Signaling by Zbtb9 in Adipocytes

The discovery of new genes involved in adipocyte development and function can shed light on the pathophysiology of obesity and T2D and provide new therapeutic targets for these disorders. Towards this end, GWAS and rare variant association studies in the T2D Knowledge Portal revealed associations between ZBTB9 and T2D, BMI, and hip circumference. However, the function of ZBTB9 in organismal physiology was not known. We found that Zbtb9 plays a key role in adipocyte biology and differentiation via cell-state dependent regulation of PPARγ signaling. Zbtb9 directly interacts with the PPARγ/RXRα complex as shown by co-IP, and acts as a positive regulator of PPARγ transcriptional activity as shown by a PPARγ reporter assay, together establishing Zbtb9 as a new PPARγ cofactor. In line with these findings, endogenous PPARγ target gene expression was decreased in the context of Zbtb9 deficiency in adipocytes. Given the central role of PPARγ in adipogenesis, we also sought to investigate the role of Zbtb9 in this process. Surprisingly, loss of Zbtb9 in preadipocytes increased adipogenesis and the expression of adipogenic genes, including PPARγ. Transcriptome analysis prior to and during adipogenesis revealed enhanced expression of E2F target genes prior to the initiation of differentiation and later induction of adipogenic gene expression. Treatment of Zbtb9 deficient preadipocytes with an E2F1 inhibitor (HLM006474) blocked the effects of Zbtb9 deficiency on adipogenic gene expression and adipogenesis. Collectively, these results demonstrate that Zbtb9 inhibits adipogenesis as a negative regulator of PPARγ expression via altered E2F1 signaling. We further demonstrated that E2F1 activation by Zbtb9 is associated with increased phosphorylation of Rb, which is known to cause dissociation of the Rb/E2F complex, and activation of E2F. Our findings revealed cell-state dependent roles of Zbtb9 in adipocytes, identifying a new molecule that may be important in the pathogenesis and treatment of obesity and T2D. Disclosure X. Xu: None. D. Buchner: None. Funding National Institutes of Health (DK119305)

The involvement of peroxisome proliferator-activated receptor gamma (PPARγ) in anti-inflammatory activity of N-stearoylethanolamine

BackgroundN-stearoylethanolamine (NSE) is a bioactive lipid amine with a wide range of biological activities. Anti-inflammatory properties of NSE were previously confirmed on multiple animal models. However, the molecular mechanisms of anti-inflammatory action of NSE remain unclear. In the current study, we examined the involvement of nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) in the NF-kB –dependent pathway of anti-inflammatory action of NSE using different methodological approaches. MethodsMolecular modeling calculated the possibility of NSE binding PPAR. Ex vivo experiment, using selective agonist of PPARα/γ - LY-171883 and antagonist of PPARγ - GW9662, examined the role of PPARα/γ in the NSE’s effect on nuclear NF-kB translocation in LPS-activated rat peritoneal macrophages. Finally, the NSE’s action on mRNA level of PPARγ-dependent genes was studied in the liver of insulin-resistant rats. ResultsThe results of molecular docking showed that NSE could bind to PPARγ and compete for the binding site with antagonist GW9662 and agonist LY-171883. These data was supported by in vitro study where pre-treatment with NSE prevented further LPS-induced NF-kB translocation into the nuclei of rat peritoneal macrophages. NSE treatment before GW9662 and LPS addition normalized the level of NF-kB translocation and IL-1β content. This finding confirmed a competitive binding of NSE with GW9662 for the ligand-binding domain of PPARγ. Additional in vivo study showed that NSE administration changed the mRNA expression of several PPARγ target genes, including SLC27A1 encoding fatty acid transport protein-1 and IL1RN - interleukin-1 receptor antagonist in insulin resistant rats. ConclusionNSE suppressed nuclear translocation of NF-κB in LPS-stimulated peritoneal macrophages via PPARγ and changed hepatic mRNA expression of PPARγ target genes (SLC27A1, IL1RN) in insulin resistant rats.

Aerobic exercise improves adipogenesis in diet-induced obese mice via the lncSRA/p38/JNK/PPARγ pathway

Adipogenesis is one of the triggers of obesity, which is a risk factor for various metabolic diseases. Long noncoding RNA steroid receptor RNA activator (lncRNA-SRA) is closely related to adipogenesis and p38/JNK mitogen-activated protein kinase mediates lipid production by regulating peroxisome proliferator-activated receptor gamma (PPARγ). Aerobic exercise can be efficient in improving adiposity and losing weight. Hence, we hypothesize that aerobic exercise ameliorates obesity by affecting the SRA/p38/JNK/PPARγ pathway and downstream target genes. The broad approaches used to test hypotheses are as follows. Spectrophotometer detected C57BL/6J mice blood lipid level; hematoxylin and eosin-stained fat tissue to check the grade of epididymis fat; quantitative polymerase chain reaction and Western blot detected messenger RNA expression and protein levels. Injected lncRNA-SRA virus vector to overexpress SRA. After 8 weeks of aerobic exercise intervention, obese mice showed significant improvements in body weight, white fat weight, lipid levels, and the Lee index. Aerobic exercise significantly inhibited the expression of SRA, activated the p38/JNK signaling pathway, further inhibited the expression of PPARγ and downstream target genes, and improved obesity. Aerobic exercise intervention improved lipid metabolism in obese mice, and the mechanism may be related to the regulation of the LncSRA/p38/JNK/PPARγ signaling pathway.

Inhibitory effects of Rocaglamide-A on PPARγ-driven adipogenesis through regulation of mitotic clonal expansion involving the JAK2/STAT3 pathway

Inhibition of adipogenesis is an important strategy for obesity treatment. Rocaglamide-A (Roc-A) is a natural herbal medicine isolated from the genus Aglaia (family Meliaceae), which has a cyclopenta[b]benzofuran core structure. Roc-A exhibits various pharmacological effects against diverse human cancer cells. However, the exact role of Roc-A during adipogenesis in adipocytes has not been studied at all. In this study, we demonstrate that Roc-A is crucial for reducing adipogenesis via downregulating PPARγ transcriptional activity. Consistently, Western-blot and RT-PCR analyses clearly showed that Roc-A inhibits the expression of PPARγ target genes and lipogenic markers in a dose-dependent manner along with suppression of lipid accumulation, in both 3T3-L1 cells and mouse adipose-derived stem cells. Mechanistically, Roc-A significantly decreased STAT3 phosphorylation in a dose-dependent manner in 3T3-L1 adipocytes. In particular, we confirmed that Roc-A effectively suppressed the expression of genes involved in cell-cycle regulation, such as cyclin A, B, D1, and E1, early during mitotic clonal expansion in 3T3-L1 adipocytes, and this effect was abolished by the JAK2/STAT3 activator FGF2. Taken together, our results demonstrated that Roc-A reduces adipogenesis by inhibiting PPARγ transactivation and STAT3 phosphorylation and thus may serve as a therapeutic target in obesity.

Rab2A regulates the progression of nonalcoholic fatty liver disease downstream of AMPK-TBC1D1 axis by stabilizing PPARγ.

Nonalcoholic fatty liver disease (NAFLD) affects approximately a quarter of the population worldwide, and persistent overnutrition is one of the major causes. However, the underlying molecular basis has not been fully elucidated, and no specific drug has been approved for this disease. Here, we identify a regulatory mechanism that reveals a novel function of Rab2A in the progression of NAFLD based on energy status and PPARγ. The mechanistic analysis shows that nutrition repletion suppresses the phosphorylation of AMPK-TBC1D1 signaling, augments the level of GTP-bound Rab2A, and then increases the protein stability of PPARγ, which ultimately promotes the hepatic accumulation of lipids in vitro and in vivo. Furthermore, we found that blocking the AMPK-TBC1D1 pathway in TBC1D1S231A-knock-in (KI) mice led to a markedly increased GTP-bound Rab2A and subsequent fatty liver in aged mice. Our studies also showed that inhibition of Rab2A expression alleviated hepatic lipid deposition in western diet-induced obesity (DIO) mice by reducing the protein level of PPARγ and the expression of PPARγ target genes. Our findings not only reveal a new molecular mechanism regulating the progression of NAFLD during persistent overnutrition but also have potential implications for drug discovery to combat this disease.

Structural Basis for PPARs Activation by The Dual PPARα/γ Agonist Sanguinarine: A Unique Mode of Ligand Recognition.

Peroxisome proliferator-activated receptors (PPARs) play crucial roles in glucose and lipid metabolism and inflammation. Sanguinarine is a natural product that is isolated from Sanguinaria Canadensis, a potential therapeutic agent for intervention in chronic diseases. In this study, biochemical and cell-based promoter-reporter gene assays revealed that sanguinarine activated both PPARα and PPARγ, and enhanced their transcriptional activity; thus, sanguinarine was identified as a dual agonist of PPARα/γ. Similar to fenofibrate, sanguinarine upregulates the expression of PPARα-target genes in hepatocytes. Sanguinarine also modulates the expression of key PPARγ-target genes and promotes adipocyte differentiation, but with a lower adipogenic activity compared with rosiglitazone. We report the crystal structure of sanguinarine bound to PPARα, which reveals a unique ligand-binding mode of sanguinarine, dissimilar to the classic Y-shaped binding pocket, which may represent a new pharmacophore that can be optimized for selectively targeting PPARα. Further structural and functional studies uncover the molecular basis for the selectivity of sanguinarine toward PPARα/γ among all three PPARs. In summary, our study identifies a PPARα/γ dual agonist with a unique ligand-binding mode, and provides a promising and viable novel template for the design of dual-targeting PPARs ligands.

Adipocyte-specific deletion of zinc finger protein 407 results in lipodystrophy and insulin resistance in mice

PPARγ deficiency in humans and model organisms impairs the transcriptional control of adipogenesis and mature adipocyte function resulting in lipodystrophy and insulin resistance. Zinc finger protein 407 (ZFP407) positively regulates PPARγ target gene expression and insulin-stimulated glucose uptake in cultured adipocytes. The in vivo physiological role of ZFP407 in mature adipocytes, however, remains to be elucidated. Here we generated adipocyte-specific ZFP407 knockout (AZKO) mice and discovered a partial lipodystrophic phenotype with reduced fat mass, hypertrophic adipocytes in inguinal and brown adipose tissue, and reduced adipogenic gene expression. The lipodystrophy was further exacerbated in AZKO mice fed a high-fat diet. Glucose and insulin tolerance tests revealed decreased insulin sensitivity in AZKO mice compared to control littermates. Cell-based assays demonstrated that ZFP407 is also required for adipogenesis, which may also contribute to the lipodystrophic phenotype. These results demonstrate an essential in vivo role of ZFP407 in brown and white adipose tissue formation and organismal insulin sensitivity.

FSH promotes fat accumulation by activating PPARγ signaling in surgically castrated, but not immunocastrated, male pigs

Follicle-stimulating hormone (FSH) was recently implicated as a novel regulator of fat accumulation. Surgical castration causes high FSH concentrations and increases fat accumulation, whereas immunocastration results in low FSH concentrations and less fat in immunocastrated boars versus barrows. However, detailed information regarding the role of FSH in regulation of fat accumulation in male pigs is unclear. First, expression of FSH receptor was confirmed (real-time quantitative PCR) in subcutaneous and visceral adipose tissues (SAT and VAT, respectively) of boars. Then, surgical castration (high FSH model) was compared to immunocastration (low FSH model) to investigate potential roles of FSH in adipogenesis and fat accumulation. High FSH concentrations after surgical castration activated PPARγ signaling by upregulating expression of CREB (P < 0.05), and then recruited an array of PPARγ target adipogenic genes, including transcription factor (C/EBPα), long-chain fatty acid uptake (LPL), fatty acid de novo synthesis (FASN, ACACA) and lipid droplet formation (PLIN1) in both SAT and VAT, promoting fat accumulation in barrows. In contrast, much lower serum FSH concentrations in immunocastrates attenuated (P < 0.05) expressions of PPARγ and PPARγ target genes in both SAT and VAT, resulting in less fat accumulation in immunocastrated boars versus barrows. We concluded that the substantially elevated FSH concentrations in barrows promoted fat accumulation by activating the PPARγ signaling pathway in adipose tissues, whereas immunocastrates accumulated less fat due to low FSH.

PHACTR1, a pro-atherosclerotic mechanosensitive PPARgamma corepressor in endothelial cells

Abstract Background Numerous genome-wide association studies revealed that SNPs at phosphatase and actin regulator 1 (PHACTR1) locus are strongly correlated with coronary artery disease (CAD). However, the mechanism linking these variants to CAD remains uncertain. Purpose We studied the biological functions and molecular mechanisms of PHACTR1 in atherosclerosis. Methods and results Analysis of GTEx database showed that CAD-related SNPs in PHACTR1 are cis-eQTLs for PHACTR1 in arteries. Therefore, we generated Phactr1 knockout mice and crossed them with apolipoprotein E-deficient (ApoE−/−) mice to induce atherosclerosis by high-fat/high-cholesterol (HF-HC) diet. Phactr1 deficiency significantly inhibited atherosclerosis with decreased inflammatory cell infiltration. Western blot showed that PHACTR1 was restricted to endothelial cells (ECs) in mice. Mechanistically, RNAseq of aortic ECs revealed that the major molecular function of PHACTR1 was transcriptional regulation. PPARγ/RXRα was the top transcription factor, and PPARγ target gene expression substantially increased in Phactr1−/− mice. Moreover, we generated endothelial cell specific Phactr1−/−, ApoE−/− mice and found decreased atherosclerotic plaque area in aortic sinus. In vitro, PHACTR1 associated with PPARγ and inhibited PPARγ transcriptional activity. The inhibitory effect of PHACTR1 on PPARγ required its shuttling from cytosol to nucleus triggered by disturbed flow, a well-established pro-atherosclerotic stimulus. Conclusion Our results identified PHACTR1 as a mechanosensitive corepressor of PPARγ in ECs to promote atherosclerosis. Endothelial PHACTR1 is a potential therapeutic target for atherosclerosis treatment. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): National Natural Science Foundation of China (NSFC), China Postdoctoral Science Foundation (CPSF)

LJ-529, a partial peroxisome proliferator-activated receptor gamma (PPARγ) agonist and adenosine A3 receptor agonist, ameliorates elastase-induced pulmonary emphysema in mice.

Chronic obstructive pulmonary disease (COPD) is the leading cause of human death worldwide. Currently available therapies for COPD mainly relieve symptoms and preserve lung function, suggesting the need to develop novel therapeutic or preventive regimens. Because chronic inflammation is a mechanism of emphysematous lesion formation and because adenosine A3 receptor signaling and peroxisome proliferator-activated receptor gamma (PPARγ) regulate inflammation, we investigated the effect of LJ-529, a selective adenosine A3 receptor agonist and partial PPARγ agonist, on inflammation in vitro and elastase-induced pulmonary emphysema in vivo. LJ-529 markedly ameliorated elastase-induced emphysematous lesion formation in the lungs in vivo, as indicated by the restoration of pulmonary function, suppression of airspace enlargement, and downregulation of elastase-induced matrix metalloproteinase activity and apoptotic cell death in the lungs. LJ-529 induced the expression of PPARγ target genes, the activity of PPARγ and several cytokines involved in inhibiting inflammation and inducing anti-inflammatory M2-like phenotypes. Moreover, LJ-529 did not exhibit significant cytotoxicity in normal cell lines derived from various organs in vitro and induced minimal changes in body weight in vivo, suggesting no overt toxicity of LJ-529 in vitro or in vivo. These results indicate the potential of LJ-529 as a novel therapeutic/preventive agent for emphysema with limited toxicity.

Comprehensive analysis of PPARγ agonist activities of stereo-, regio-, and enantio-isomers of hydroxyoctadecadienoic acids.

Hydroxyoctadecadienoic acids (HODEs) are produced by oxidation and reduction of linoleates. There are several regio- and stereo-isomers of HODE, and their concentrations in vivo are higher than those of other lipids. Although conformational isomers may have different biological activities, comparative analysis of intracellular function of HODE isomers has not yet been performed. We evaluated the transcriptional activity of peroxisome proliferator-activated receptor γ (PPARγ), a therapeutic target for diabetes, and analyzed PPARγ agonist activity of HODE isomers. The lowest scores for docking poses of 12 types of HODE isomers (9-, 10-, 12-, and 13-HODEs) were almost similar in docking simulation of HODEs into PPARγ ligand-binding domain (LBD). Direct binding of HODE isomers to PPARγ LBD was determined by water-ligand observed via gradient spectroscopy (WaterLOGSY) NMR experiments. In contrast, there were differences in PPARγ agonist activities among 9- and 13-HODE stereo-isomers and 12- and 13-HODE enantio-isomers in a dual-luciferase reporter assay. Interestingly, the activity of 9-HODEs was less than that of other regio-isomers, and 9-(E,E)-HODE tended to decrease PPARγ-target gene expression during the maturation of 3T3-L1 cells. In addition, 10- and 12-(Z,E)-HODEs, which we previously proposed as biomarkers for early-stage diabetes, exerted PPARγ agonist activity. These results indicate that all HODE isomers have PPARγ-binding affinity; however, they have different PPARγ agonist activity. Our findings may help to understand the biological function of lipid peroxidation products.

Identification and Characterization of Cannabimovone, a Cannabinoid from Cannabis sativa, as a Novel PPARγ Agonist via a Combined Computational and Functional Study.

Phytocannabinoids (pCBs) are a large family of meroterpenoids isolated from the plant Cannabis sativa. Δ9-Tetrahydrocannabinol (THC) and cannabidiol (CBD) are the best investigated phytocannabinoids due to their relative abundance and interesting bioactivity profiles. In addition to various targets, THC and CBD are also well-known agonists of peroxisome proliferator-activated receptor gamma (PPARγ), a nuclear receptor involved in energy homeostasis and lipid metabolism. In the search of new pCBs potentially acting as PPARγ agonists, we identified cannabimovone (CBM), a structurally unique abeo-menthane pCB, as a novel PPARγ modulator via a combined computational and experimental approach. The ability of CBM to act as dual PPARγ/α agonist was also evaluated. Computational studies suggested a different binding mode toward the two isoforms, with the compound able to recapitulate the pattern of H-bonds of a canonical agonist only in the case of PPARγ. Luciferase assays confirmed the computational results, showing a selective activation of PPARγ by CBM in the low micromolar range. CBM promoted the expression of PPARγ target genes regulating the adipocyte differentiation and prevented palmitate-induced insulin signaling impairment. Altogether, these results candidate CBM as a novel bioactive compound potentially useful for the treatment of insulin resistance-related disorders.

DEC1 regulates the rhythmic expression of PPARγ target genes involved in lipid metabolism in white adipose tissue.

Previously, we found that the basic helix-loop-helix transcriptional repressor DEC1 interacts with the PPARγ:RXRα heterodimer, a master transcription factor for adipogenesis and lipogenesis, to suppress transcription from PPARγ target genes (Noshiro et al., Genes to Cells, 2018, 23:658-669). Because the expression of PPARγ and several of its target genes exhibits circadian rhythmicity in white adipose tissue (WAT), we examined the expression profiles of PPARγ target genes in wild-type and Dec1-/- mice. We found that the expression of PPARγ target genes responsible for lipid metabolism, including the synthesis of triacylglycerol from free fatty acids (FFAs), lipid storage and the lipolysis of triacylglycerol to FFAs, oscillates in a circadian manner in WAT. Moreover, DEC1 deficiency led to a marked increase in the expression of these genes at night (Zeitgeber times 16 and 22), resulting in disruption of circadian rhythms. Serum FFA levels in wild-type mice also showed circadian oscillations, but these were disrupted by DEC1 deficiency, leading to reduced FFA levels. These results suggest that PPARγ:RXRα and DEC1 cooperatively generate the circadian expression of PPARγ target genes through PPAR-responsive elements in WAT.

Anti-cancer activity of сuraxin CBL0137 on the models of acute leukemia in vitro

Background. Curaxin CBL0137 is a novel non-genotoxic compound with anti-cancer activity based on CBL0137 ability of non-covalent interaction with DNA causing histone chaperone FACT relocation. Anti-cancer activity of this drug was demonstrated previously on the wide panel of solid cancer models in vitro and in vivo.Objectives. Estimation of anticancer effects of CBL0137 on the acute myeloblastic leukemia cells (THP-1) and acute lymphoblastic leukemia (CCRF-CEM).Materials and methods. CBL0137 cytotoxicity was analyzed using the MTT test, the effects on the cell cycle and the induction of apoptosis was assessed by flow cytometry, the activity of signaling pathways in cells treated with CBL0137 was determined by real-time polymerase chain reaction.Results. Cell treatment with CBL0137 led to cell cycle arrest and apoptosis induction. In the study of CBL0137 effect on target gene clusters of 10 signal transduction pathways involved in the pathogenesis of acute leukemia we have showed that CBL0137 inhibited the expression of down-stream genes of WNT and Hedgehog signaling in both cell lines. In THP-1 cells we also observed the inhibition of the expression of PPARγ target and hypoxia-activated genes. In CCRF-CEM cells CBL0137 also induced the expression of Notch signaling target genes.Conclusion. The antitumor activity of CBL0137 was demonstrated on acute leukemia cell cultures, the drug possesses cytotoxicity, causes cell cycle arrest and activation of apoptosis. Significant changes in the expression of efferent gene clusters of several signaling pathways were observed in the cells treated with CBL0137.

CDK5 Participates in Amyloid-β Production by Regulating PPARγ Phosphorylation in Primary Rat Hippocampal Neurons

Cyclin-dependent kinase 5 (CDK5) in adipose tissue mediates peroxisome proliferator-activated receptor γ (PPARγ) phosphorylation at Ser273 to inhibit its activity, causing PPARγ target gene expression changes. Among these, insulin-degrading enzyme (IDE) degrades amyloid-β peptide (Aβ), the core pathological product of Alzheimer's disease (AD), whereas β-amyloid cleavage enzyme 1 (BACE1) hydrolyzes amyloid-β protein precursor (AβPP). Therefore, we speculated that CDK5 activity in the brain might participate in Aβ production, thereby functioning as a key molecule in AD pathogenesis. To confirm this hypothesis, we transduced primary rat hippocampal neurons using CDK5-expressing lentiviral vectors. CDK5 overexpression increased PPARγ Ser273 phosphorylation, decreased IDE expression, increased BACE1 and AβPP expression, increased Aβ levels, and induced neuronal apoptosis. The CDK5 inhibitor roscovitine effectively reversed these CDK5 overexpression-mediated effects. Moreover, silencing of the Cdk5 gene via CDK5 shRNA-expressing lentiviral vectors in primary hippocampal neurons did not exert any protective effect against normal neuronal apoptosis, nor were significant effects observed on Aβ levels, PPARγ phosphorylation, or PPARγ target gene expression in the cells. However, Cdk5 gene silencing exhibited a neuroprotective effect in the Aβ-induced AD neuron model by effectively inhibiting the Aβ-induced neuronal apoptosis, PPARγ phosphorylation, PPARγ expression downregulation, and PPARγ target gene expression changes, and reducing Aβ levels. In conclusion, this study demonstrated that CDK5 played an important role in the pathogenesis of AD. Specifically, CDK5 participated in Aβ production by regulating PPARγ phosphorylation. Targeted therapy against CDK5 could effectively reduce and reverse the neurotoxic effects of Aβ and may represent a novel approach for AD treatment.

A novel specific peroxisome proliferator-activated receptor γ (PPARγ) modulator YR4-42 ameliorates hyperglycaemia and dyslipidaemia and hepatic steatosis in diet-induced obese mice.

AimsTo evaluate a novel tetrahydroisoquinoline derivative YR4‐42 as a selective peroxisome proliferator‐activated receptor γ (PPARγ) modulator (SPPARM) and explore its anti‐diabetic effects in vitro and in vivo.Materials and methodsUsing two standard full PPARγ agonists rosiglitazone and pioglitazone as controls, the PPARγ binding affinity and transactivation action of YR4‐42 were evaluated using biochemical and cell‐based reporter gene assays. The capacity of YR4‐42 to recruit coactivators of PPARγ was also assessed. The effects of YR4‐42 on adipogenesis and glucose consumption and PPARγ Ser273 phosphorylation were investigated in 3T3‐L1 adipocytes. The effects of YR4‐42 and pioglitazone, serving as positive control, on glucose and lipids metabolism were investigated in high‐fat diet‐induced obese (DIO) C57BL/6J mice. The expression of PPARγ target genes involved in glucose and lipid metabolism was also assessed in vitro and in vivo.Results In vitro biochemical and cell‐based functional assays showed that YR4‐42 has much weaker binding affinity, transactivation, and recruitment to PPARγ of the coactivators thyroid hormone receptor‐associated protein complex 220 kDa component (TRAP220) and PPARγ coactivator 1‐α (PGC1α) compared to full agonists. In 3 T3‐L1 adipocytes, YR4‐42 significantly improved glucose consumption without a lipogenesis effect, while blocking tumour necrosis factor α‐mediated phosphorylation of PPARγ at Ser273, thereby upregulating the expression of the PPARγ Ser273 phosphorylation‐dependent genes. Furthermore, in DIO mice, oral administration of YR4‐42 ameliorated the hyperglycaemia, with a similar insulin sensitization effect to that of pioglitazone. Importantly, YR4‐42 also improved hyperlipidaemia‐associated hepatic steatosis without weight gain, which avoids a major side effect of pioglitazone. Thus, YR4‐42 appeared to selectively modulate PPARγ responses. This finding was supported by the gene expression analysis, which showed that YR4‐42 selectively targets PPARγ‐regulated genes mapped to glucose and lipid metabolism in DIO mice.ConclusionsWe conclude that YR4‐42 is a novel anti‐diabetic drug candidate with significant advantages compared to standard PPARγ agonists. YR4‐42 should be further investigated in preclinical and clinical studies.

Procyanidin B2 Activates PPARγ to Induce M2 Polarization in Mouse Macrophages.

Procyanidins, a subclass of flavonoids found in commonly consumed foods, possess potential anti-inflammatory activity. Manipulation of M1/M2 macrophage homeostasis is an effective strategy for the treatment of metabolic inflammatory diseases. The objective of this study was to determine the effect of procyanidins on macrophage polarization. Procyanidin B2 (PCB2), the most widely distributed natural procyanidins, enhanced the expressions of M2 macrophage markers (Arg1, Ym1, and Fizz1). PCB2 activated peroxisome proliferator-activated receptor γ (PPARγ) activity and increased the expressions of PPARγ target genes (CD36 and ABCG1) in macrophages. Inhibition of PPARγ using siRNA or antagonist GW9662 attenuated the PCB2-induced expressions of M2 macrophage markers. In addition, we identified cognate PPAR-responsive elements (PPREs) within the 5'-flanking regions of the mouse Arg1, Ym1, and Fizz1 genes. Furthermore, macrophages isolated from db/db diabetic mice showed lower expressions of M2 markers. PCB2 effectively restored the Arg1, Ym1, and Fizz1 expressions in a PPARγ-dependent manner. These findings support the notion that PCB2 regulated macrophage M2 polarization via the activation of PPARγ. Our results provide a new mechanism by which procyanidins exert their beneficial anti-inflammatory effects.

Pioglitazone Reduces β Amyloid Levels via Inhibition of PPARγ Phosphorylation in a Neuronal Model of Alzheimer's Disease.

It has been demonstrated that peroxisome proliferator-activated receptor γ (PPARγ) can regulate the transcription of its target gene, insulin-degrading enzyme (IDE), and thus enhance the expression of the IDE protein. The protein can degrade β amyloid (Aβ), a core pathological product of Alzheimer’s disease (AD). PPARγ can also regulate the transcription of other target gene, β-amyloid cleavage enzyme 1 (BACE1), and thus inhibit the expression of the BACE1 protein. BACE1 can hydrolyze amyloid precursor protein (APP), the precursor of Aβ. In adipose tissue, PPARγ agonists can inhibit the phosphorylation of PPARγ by inhibiting cyclin-dependent kinase 5 (CDK5), which in turn affects the expression of target genes regulated by PPARγ. PPARγ agonists may also exert inhibitory effects on the phosphorylation of PPARγ in the brain, thereby affecting the expression of the aforementioned PPARγ target genes and reducing Aβ levels. The present study confirmed this hypothesis by showing that PPARγ agonist pioglitazone attenuated the neuronal apoptosis of primary rat hippocampal neurons induced by Aβ1–42, downregulated CDK5 expression, weakened the binding of CDK5 to PPARγ, reduced PPARγ phosphorylation, increased the expression of PPARγ and IDE, decreased the expression of BACE1, reduced APP production, and downregulated intraneuronal Aβ1–42 levels. These effects were inhibited by the PPARγ antagonist GW9662. After CDK5 silencing with CDK5 shRNA, the above effect of pioglitazone was not observed, except when upregulating the expression of PPARγ in Aβ1–42 treated neurons. In conclusion, this study demonstrated that pioglitazone could inhibit the phosphorylation of PPARγ in vitro by inhibiting CDK5 expression, which in turn affected the expression of PPARγ target genes Ide and Bace1, thereby promoting Aβ degradation and reducing Aβ production. This reduced Aβ levels in the brain, thereby exerting neuroprotective effects in an AD model.