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Objective(s): This study investigates the effect of naonoliposmal encapsulated hybrid extract (NEHE) obtained from three plants (green tea, garlic, and chicory) in treating diabetes through expression of PPARgamma, BGK, and GLUT2 genes involved in diabetes and their relationship with miR-27a. Materials and Methods: Herbal extracts were encapsulated in liposome. RIN-5F cells were exposed to streptozotocin (STZ) for 24 hr and divided into five groups to induce the diabetes model. Three groups received 0.4, 1, and 2 mg/ml of NEHE, a positive control group received 5 µg/ml of metformin for 72 hr, and a negative control group was only treated with FBS. Then, they were subjected to an MTT assay to check the toxicity of the extract. An immunohistochemical test was performed to check the level of insulin expression in different groups. Real Time-PCR test was performed to check the expression of desired genes. Results: MTT assay showed that the NEHE extract had no toxicity on the tested cells. Also, the immunohistochemistry results showed that insulin expression NEHE group was significantly higher than that of the metformin group (P

Objective(s): This study investigates the effect of naonoliposmal encapsulated hybrid extract (NEHE) obtained from three plants (green tea, garlic, and chicory) in treating diabetes through expression of PPARgamma, BGK, and GLUT2 genes involved in diabetes and their relationship with miR-27a. Materials and Methods: Herbal extracts were encapsulated in liposome. RIN-5F cells were exposed to streptozotocin (STZ) for 24 hr and divided into five groups to induce the diabetes model. Three groups received 0.4, 1, and 2 mg/ml of NEHE, a positive control group received 5 µg/ml of metformin for 72 hr, and a negative control group was only treated with FBS. Then, they were subjected to an MTT assay to check the toxicity of the extract. An immunohistochemical test was performed to check the level of insulin expression in different groups. Real Time-PCR test was performed to check the expression of desired genes. Results: MTT assay showed that the NEHE extract had no toxicity on the tested cells. Also, the immunohistochemistry results showed that insulin expression NEHE group was significantly higher than that of the metformin group (P<0.001), indicating the antidiabetic effect of the extract. In all groups treated with the extract, especially with the maximum extract concentration, the expression level of BGK, PPARgamma, miR-27a, and GLUT2 genes was significantly higher than that of the untreated control group. Furthermore, there was no significant difference in the expression of BGK and PPARgamma genes in groups trated with either metformin or plant extract at concentration of 2 mg/ml groups. NEHE showed an improved effect in treating diabetes in a dose-dependent manner by increasing insulin secretion from pancreatic cells. Conclusion: As a glucose transporter and sensor, GLUT2 controls the balance between intracellular and extracellular glucose concentrations. When glucose-induced insulin secretion is impaired, garlic can increase the half-life of insulin, while green tea and chicory reduce insulin resistance through the miR-27a pathway. [ABSTRACT FROM AUTHOR]

Exposure of pregnant F0 mouse dams to the obesogen tributyltin (TBT) predisposes unexposed male descendants to obesity and diverts mesenchymal stem cells (MSCs) toward the adipocytic lineage. TBT promotes adipogenic commitment and differentiation of MSCs in vitro. To identify TBT-induced factors predisposing MSCs toward the adipocytic fate, we exposed mouse MSCs to TBT, the peroxisome proliferator activated receptor gamma (PPARγ)-selective agonist rosiglitazone, or the retinoid X receptor (RXR)-selective agonist LG-100268. Then we determined their transcriptomal profiles to determine candidate microRNAs (miR) regulating adipogenic commitment and differentiation. Of the top 10 candidate microRNAs predicted by Ingenuity Pathway Analysis, miR-21, miR-33, and miR-223 were expressed consistent with an ability to differentially regulate target genes during adipogenesis. We found that 24-hour exposure to 50nM TBT caused miR-223 levels in MSCs to increase; expression of its target genes ZEB1, NFIB, and FOXP1 was decreased. Rosiglitazone and TBT increased miR-223 levels. This induction was inhibited by the PPARγ antagonist T0070907 but not by the RXR antagonists HX531 or UVI3003, placing miR-223 downstream of PPARγ. Chromatin immunoprecipitation confirmed TBT-induced binding of PPARγ to regulatory elements in the miR-223 promoter. miR-223 levels were elevated in white adipose tissue of F2 and F3 male descendants of pregnant F0 mouse dams exposed to 50nM TBT throughout gestation. miR-223 levels were potentiated in males fed an increased fat diet. We infer that TBT induced miR-223 expression and increased adipogenesis in MSCs through the PPARγ pathway and that transgenerationally increased expression of miR-223 plays an important role in the development of obesity caused by TBT exposure. [ABSTRACT FROM AUTHOR]

There is a growing prevalence of inflammatory bowel disease (IBD), a chronic inflammatory condition of the gastrointestinal tract, among the aging population. Ghrelin is a gut hormone that, in addition to controlling feeding and energy metabolism, has been shown to exert anti-inflammatory effects; however, the effect of ghrelin in protecting against colitis in old mice has not been assessed. Here, we subjected old female C57BL/6J mice to dextran sulfate sodium (DSS) in drinking water for six days, then switched back to normal drinking water, administered acyl-ghrelin or vehicle control from day 3 to 13, and monitored disease activities throughout the disease course. Our results showed that treatment of old mice with acyl-ghrelin attenuated DSS-induced colitis. Compared to the DSS group, ghrelin treatment decreased levels of the inflammation marker S100A9 in the colons collected on day 14 but not on day 8, suggesting that the anti-inflammatory effect was more prominent in the recovery phase. Ghrelin treatment also significantly reduced F4/80 and interleukin-17A on day 14. Moreover, acyl-ghrelin increased mitochondrial respiration and activated transcriptional activity of the peroxisome proliferator-activated receptor gamma (PPARγ) in Caco-2 cells. Together, our data show that ghrelin alleviated DSS-induced colitis, suggesting that ghrelin may promote tissue repair in part through regulating epithelial metabolism via PPARγ mediated signaling. [ABSTRACT FROM AUTHOR]

Abstract Background Well-differentiated and dedifferentiated liposarcomas are rare soft tissue tumors originating in adipose tissue that share genetic abnormalities but have significantly different metastatic potential. Dedifferentiated liposarcoma (DDLPS) is highly aggressive and has an overall 5-year survival rate of 30% as compared to 90% for well-differentiated liposarcoma (WDLPS). This discrepancy may be connected to their potential to form adipocytes, where WDLPS is adipogenic but DDLPS is adipogenic-impaired. Normal adipogenesis requires Zinc Finger Protein 423 (ZFP423), a transcriptional coregulator of Perixosome Proliferator Activated Receptor gamma (PPARG2) mRNA expression that defines committed preadipocytes. Expression of ZFP423 in preadipocytes is promoted by Seven-In-Absentia Homolog 2 (SIAH2)-mediated degradation of Zinc Finger Protein 521 (ZFP521). This study investigated the potential role of ZFP423, SIAH2 and ZFP521 in the adipogenic potential of WDLPS and DDLPS. Methods Human WDLPS and DDLPS fresh and paraffin-embedded tissues were used to assess the gene and protein expression of proadipogenic regulators. In parallel, normal adipose tissue stromal cells along with WDLPS and DDLPS cell lines were cultured, genetically modified, and induced to undergo adipogenesis in vitro. Results Impaired adipogenic potential in DDLPS was associated with reduced ZFP423 protein levels in parallel with reduced PPARG2 expression, potentially involving regulation of ZFP521. SIAH2 protein levels did not define a clear distinction related to adipogenesis in these liposarcomas. However, in primary tumor specimens, SIAH2 mRNA was consistently upregulated in DDLPS compared to WDLPS when assayed by fluorescence in situ hybridization or real-time PCR. Conclusions These data provide novel insights into ZFP423 expression in adipogenic regulation between WDLPS and DDLPS adipocytic tumor development. The data also introduces SIAH2 mRNA levels as a possible molecular marker to distinguish between WDLPS and DDLPS.

Mice lacking peroxisome-proliferator activated receptor-γ (PPARγ) in neurons do not become leptin resistant when placed on a high-fat diet (HFD). In male mice, this results in decreased food intake and increased energy expenditure, causing reduced body weight, but this difference in body weight is not observed in female mice. In addition, estrous cycles are disturbed and the ovaries present with hemorrhagic follicles. We observed that PPARγ was more highly expressed in astrocytes than neurons, so we created an inducible, conditional knockout of PPARγ in astrocytes (AKO). The AKO mice had impaired glucose tolerance and hepatic steatosis that did not worsen with HFD. Expression of gluconeogenic genes was elevated in the mouse livers, as was expression of several genes involved in lipogenesis, lipid transport, and storage. The AKO mice also had a reproductive phenotype with fewer estrous cycles, elevated plasma testosterone levels, reduced corpora lutea formation, and alterations in hypothalamic and ovarian gene expression. Thus, the phenotypes of the AKO mice were very different from those seen in the neuronal knockout mice, suggesting distinct roles for PPARγ in these two cell types.

Background: Well-differentiated and dedifferentiated liposarcomas are rare soft tissue tumors originating in adipose tissue that share genetic abnormalities but have significantly different metastatic potential. Dedifferentiated liposarcoma (DDLPS) is highly aggressive and has an overall 5-year survival rate of 30% as compared to 90% for well-differentiated liposarcoma (WDLPS). This discrepancy may be connected to their potential to form adipocytes, where WDLPS is adipogenic but DDLPS is adipogenic-impaired. Normal adipogenesis requires Zinc Finger Protein 423 (ZFP423), a transcriptional coregulator of Perixosome Proliferator Activated Receptor gamma (PPARG2) mRNA expression that defines committed preadipocytes. Expression of ZFP423 in preadipocytes is promoted by Seven-In-Absentia Homolog 2 (SIAH2)-mediated degradation of Zinc Finger Protein 521 (ZFP521). This study investigated the potential role of ZFP423, SIAH2 and ZFP521 in the adipogenic potential of WDLPS and DDLPS.Methods: Human WDLPS and DDLPS fresh and paraffin-embedded tissues were used to assess the gene and protein expression of proadipogenic regulators. In parallel, normal adipose tissue stromal cells along with WDLPS and DDLPS cell lines were cultured, genetically modified, and induced to undergo adipogenesis in vitro.Results: Impaired adipogenic potential in DDLPS was associated with reduced ZFP423 protein levels in parallel with reduced PPARG2 expression, potentially involving regulation of ZFP521. SIAH2 protein levels did not define a clear distinction related to adipogenesis in these liposarcomas. However, in primary tumor specimens, SIAH2 mRNA was consistently upregulated in DDLPS compared to WDLPS when assayed by fluorescence in situ hybridization or real-time PCR.Conclusions: These data provide novel insights into ZFP423 expression in adipogenic regulation between WDLPS and DDLPS adipocytic tumor development. The data also introduces SIAH2 mRNA levels as a possible molecular marker to distinguish between WDLPS and DDLPS. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) has a high incidence rate among women worldwide, and the mechanisms and etiology of this disease are not yet fully understood. The peroxisome proliferator-activated receptor gamma (PPARgamma), a nuclear hormone receptor that plays important roles in energy metabolism and cellular differentiation, is also suggested to be effective in cancer development. However, the results of studies investigating the cancer association with PPARgamma are inconsistent, creating a need for further investigation of the effects of this transcription factor on BC risk. We have examined the Pro12Ala-(rs1801282) and C161T-(rs3856806) polymorphisms of the PPARgamma gene in Turkish patients with BC in this case–control study. A total of 95 women diagnosed with BC as cases and 119 controls were genotyped for PPARgamma polymorphisms by polymerase chain reaction and restriction fragment length polymorphism techniques. The ProPro genotype and T161 allele were associated with an increased risk of BC comparing with the Ala12 allele and CC161 genotype, respectively (p < 0.001). The multivariate regression analysis confirmed that the ProPro genotype (p < 0.011), T161 allele (p < 0.001), smoking (p = 0.019), and advanced age (> 60 years) (p = 0.007) are risk factors for breast cancer. We also found that the PPARgamma Pro12Ala and C161T polymorphisms were in linkage disequilibrium (D':0.511, r2:0.099). It was determined that carrying ProPro-T161 risky PPARgamma haplotype was associated with a higher risk of BC compared to protective Ala12-CC161 haplotype (p < 0.01, OR:7.797, 95% CI:3.521–17.263). We concluded that PPARgamma Pro12Ala and C161T polymorphisms are associated with increased BC risk, and ProPro-T161 risky haplotype, which is in linkage disequilibrium, increases this effect. [ABSTRACT FROM AUTHOR]

Atherosclerotic cardiovascular disease is a metabolic and inflammatory disorder. In vitro studies have suggested that protein arginine methyltransferase 4 (PRMT4) may act as a transcriptional coactivator to modulate inflammatory and metabolic processes. Here we investigated the potential anti-atherogenic effect of PRMT4 inhibitor TP-064 in vivo. Male apolipoprotein E knockout mice fed a high cholesterol/high fat Western-type diet were intraperitoneally injected three times a week with 2.5 mg/kg (low dose) or 10 mg/kg (high dose) TP-064 or with DMSO control. TP-064 induced a dose-dependent decrease in lipopolysaccharide-induced ex vivo blood monocyte Tnfα secretion (p < 0.05 for trend) in the context of unchanged blood monocyte concentrations and neutrophilia induction (p < 0.01 for trend). A dose-dependent decrease in gonadal white adipose tissue expression levels of PPARγ target genes was detected, which translated into a reduced body weight gain after high dose TP-064 treatment (p < 0.05). TP-064 treatment also dose-dependently downregulated gene expression of the glycogen metabolism related protein G6pc in the liver (p < 0.001 for trend). In addition, a trend towards lower plasma insulin and higher blood glucose levels was observed, which was paralleled by a reduction in hepatic mRNA expression levels of the insulin-responsive genes Fasn (−55%; p < 0.001) and Gck (−47%; p < 0.001) in high dose-treated mice. Plasma triglyceride levels were reduced by high dose TP-064 treatment (−30%; p < 0.05). However, no change was observed in the size or composition of aortic root atherosclerotic lesions. The PRMT4 inhibitor TP-064 impacts both inflammatory and metabolic processes without changing atherosclerosis susceptibility of male apolipoprotein E knockout mice. [Display omitted] • Protein arginine methyltransferase 4 (PRMT4) is overexpressed in white blood cells of atherosclerotic patients. • TP-064 dose-dependently downregulates PRMT4-mediated transcription in vivo. • Chronic TP-064 treatment does not change atherosclerosis susceptibility. • PRMT4 can possibly serve as novel therapeutic target in obesity. [ABSTRACT FROM AUTHOR]

Abstract Background Obesity is associated with cardiovascular complications, including pulmonary hypertension (PH). Reports suggest that peroxisome proliferator-activated receptor-γ (PPARγ) has direct action in preventing vascular remodelling in PH. Here we dissected the specific role of high-fat-diet (HFD)-induced obesity and vascular smooth muscle cell (VSMC)-PPARγ for remodelling of small pulmonary arteries. Methods Wild-type (WT) and VSMC-specific PPARγ-knockout (SmPparγ −/−) mice were fed a low-fat-diet (LFD, 10% kcal from fat) or HFD (60% kcal from fat) for 24 weeks. Mice were metabolically phenotyped (e.g. weight development, insulin/glucose tolerance) at the beginning, and after 12 and 24 weeks, respectively. At 24 weeks additionally pulmonary pressure, heart structure, pulmonary vascular muscularization together with gene and protein expression in heart and lung tissues were determined. Results HFD increased right ventricular systolic pressure (RVSP) to a similar extent in WT and SmPparγ −/− mice. HFD decreased glucose tolerance and insulin sensitivity in both WT and SmPparγ −/− mice. Importantly, the increase in RVSP correlated with the degree of insulin resistance. However, VSMC-PPARγ deficiency increased pulmonary vascular muscularization independently of the diet-induced rise in RVSP. This increase was associated with elevated expression of early growth response protein 1 in heart and osteopontin in lung tissue. Conclusions Here we demonstrate a correlation of insulin resistance and pulmonary pressure. Further, deficiency of PPARγ in VSMCs diet-independently leads to increased pulmonary vascular muscularization.

Background: Marrow adipose tissue (MAT) has been shown to be vital for regulating metabolism and maintaining skeletal homeostasis in the bone marrow (BM) niche. As a reflection of BM remodeling, MAT is highly responsive to nutrient fluctuations, hormonal changes, and metabolic disturbances such as obesity and diabetes mellitus. Expansion of MAT has also been strongly associated with bone loss in mice and humans. However, the regulation of BM plasticity remains poorly understood, as does the mechanism that links changes in marrow adiposity with bone remodeling. Methods: We studied deletion of Adipsin, and its downstream effector, C3, in C57BL/6 mice as well as the bone-protected PPARγ constitutive deacetylation 2KR mice to assess BM plasticity. The mice were challenged with thiazolidinedione treatment, calorie restriction, or aging to induce bone loss and MAT expansion. Analysis of bone mineral density and marrow adiposity was performed using a μCT scanner and by RNA analysis to assess adipocyte and osteoblast markers. For in vitro studies, primary bone marrow stromal cells were isolated and subjected to osteoblastogenic or adipogenic differentiation or chemical treatment followed by morphological and molecular analyses. Clinical data was obtained from samples of a previous clinical trial of fasting and high-calorie diet in healthy human volunteers. Results: We show that Adipsin is the most upregulated adipokine during MAT expansion in mice and humans in a PPARγ acetylation-dependent manner. Genetic ablation of Adipsin in mice specifically inhibited MAT expansion but not peripheral adipose depots, and improved bone mass during calorie restriction, thiazolidinedione treatment, and aging. These effects were mediated through its downstream effector, complement component C3, to prime common progenitor cells toward adipogenesis rather than osteoblastogenesis through inhibiting Wnt/β-catenin signaling. Conclusions: Adipsin promotes new adipocyte formation and affects skeletal remodeling in the BM niche. Our study reveals a novel mechanism whereby the BM sustains its own plasticity through paracrine and endocrine actions of a unique adipokine. Funding: This work was supported by the National Institutes of Health T32DK007328 (NA), F31DK124926 (NA), R01DK121140 (JCL), R01AR068970 (BZ), R01AR071463 (BZ), R01DK112943 (LQ), R24DK092759 (CJR), and P01HL087123 (LQ).

Preterm birth is a risk factor for growth failure and development of respiratory disease in children and young adults. Their early exposure to oxygen may contribute to lung disease because adult mice exposed to hyperoxia as neonates display reduced lung function, changes in the host response to respiratory viral infections, and develop pulmonary hypertension and heart failure that shortens their lifespan. Here, we provide new evidence that neonatal hyperoxia also impairs growth by inhibiting fat accumulation. Failure to accumulate fat may reflect a systemic defect in adipogenic potential of stem cells because bone marrow-derived mesenchymal cells (BMSCs) isolated from the mice grew slower and were more oxidized compared to controls. They also displayed reduced capacity to accumulate lipid and differentiate into adipocytes. BMSCs from adult mice exposed to neonatal hyperoxia express lower levels of peroxisome proliferator-activated receptor gamma (PPARγ), a transcription factor that drives adipocyte differentiation. The defect in adipogenesis was rescued by expressing PPARγ in these cells. These findings reveal early life exposure to high levels of oxygen may suppresses fat accumulation and impair adipogenic differentiation upstream of PPARγ signaling, thus potentially contributing to growth failure seen in people born preterm. [Display omitted] • Mice exposed to neonatal hyperoxia accumulate less fat and weight over their lifespan. • Bone marrow derived stem cells (BMSCs) from adult mice exposed to neonatal hyperoxia show elevated reactive oxygen species. • These BMSCs show a defect in proliferation and fat cell formation (adipogenesis). • Adipogenesis can be restored in these BMSCs by expressing PPARγ. [ABSTRACT FROM AUTHOR]

Knowledge of biological responses to proton therapy (PT) in comparison to X-ray remains in its infancy. Identification of PT specific molecular signals is an important opportunity for the discovery of biomarkers and synergistic drugs to advance clinical application. Since PT is used for the treatment of lymphoma, we report here transcriptomic responses of lymphoma cell lines to PT vs X-ray and identify potential therapeutic targets. Two lymphoma cell lines of human (BL41) and murine (J3D) origin were irradiated by X-ray and PT. Differential transcriptome regulation was quantified by RNA sequencing for each radiation type at 12 hours post irradiation. Gene-set enrichment analysis revealed deregulated molecular pathways and putative targets for lymphoma cell sensitization to PT. Transcriptomic gene set enrichment analyses uncovered pathways that contribute to the unfolded protein response (UPR) and mitochondrial transport. Functional validation at multiple time points demonstrated increased UPR activation and decreased protein translation, perhaps due to increased oxidative stress and oxidative protein damage after PT. PPARgamma was identified as a potential regulator of the PT transcriptomic response. Inhibition of PPARgamma by two compounds, T0070907 and SR2595, sensitized lymphoma cells to PT. Proton vs X-ray radiation leads to the transcriptional regulation of a specific subset of genes in line with diminished protein translation and UPR activation that may be due to oxidative stress. This study demonstrates that different radiation qualities trigger distinct cellular responses in lymphoma cells, and identifies PPARgamma inhibition as a potential strategy for the sensitization of lymphoma to PT. [ABSTRACT FROM AUTHOR]

The nuclear receptor PPARγ is essential to maintain whole-body glucose homeostasis and insulin sensitivity, acting as a master regulator of adipogenesis, lipid, and glucose metabolism. Its activation through natural or synthetic ligands induces the recruitment of coactivators, leading to transcription of target genes such as cytokines and hormones. More recently, post translational modifications, such as PPARγ phosphorylation at Ser273 by CDK5 in adipose tissue, have been linked to insulin resistance trough the dysregulation of expression of a specific subset of genes. Here, we investigate how this phosphorylation may disturb the interaction between PPARγ and some coregulator proteins as a new mechanism that may leads to insulin resistance. Through cellular and in vitro assays, we show that PPARγ phosphorylation inhibition increased the activation of the receptor, therefore the increased recruitment of PGC1-α and TIF2 coactivators, whilst decreases the interaction with SMRT and NCoR corepressors. Moreover, our results show a shift in the coregulators interaction domains preferences, suggesting additional interaction interfaces formed between the phosphorylated PPARγ and some coregulator proteins. Also, we observed that the CDK5 presence disturb the PPARγ-coregulator’s synergy, decreasing interaction with PGC1-α, TIF2, and NCoR, but increasing coupling of SMRT. Finally, we conclude that the insulin resistance provoked by PPARγ phosphorylation is linked to a differential coregulators recruitment, which may promote dysregulation in gene expression.

The nuclear receptor PPARγ is essential to maintain whole-body glucose homeostasis and insulin sensitivity, acting as a master regulator of adipogenesis, lipid, and glucose metabolism. Its activation through natural or synthetic ligands induces the recruitment of coactivators, leading to transcription of target genes such as cytokines and hormones. More recently, post translational modifications, such as PPARγ phosphorylation at Ser273 by CDK5 in adipose tissue, have been linked to insulin resistance trough the dysregulation of expression of a specific subset of genes. Here, we investigate how this phosphorylation may disturb the interaction between PPARγ and some coregulator proteins as a new mechanism that may leads to insulin resistance. Through cellular and in vitro assays, we show that PPARγ phosphorylation inhibition increased the activation of the receptor, therefore the increased recruitment of PGC1-α and TIF2 coactivators, whilst decreases the interaction with SMRT and NCoR corepressors. Moreover, our results show a shift in the coregulators interaction domains preferences, suggesting additional interaction interfaces formed between the phosphorylated PPARγ and some coregulator proteins. Also, we observed that the CDK5 presence disturb the PPARγ-coregulator's synergy, decreasing interaction with PGC1-α, TIF2, and NCoR, but increasing coupling of SMRT. Finally, we conclude that the insulin resistance provoked by PPARγ phosphorylation is linked to a differential coregulators recruitment, which may promote dysregulation in gene expression. [ABSTRACT FROM AUTHOR]

Despite the use of antiretroviral therapy for the treatment of HIV‐1 infection, cognitive impairments, that is, HIV‐1‐associated neurocognitive disorders remain prevalent potentially due to persistent viral replication, production of viral proteins, associated brain inflammation or in certain instances, antiretroviral neurotoxicity. Cellular targets in the brain include microglia which in response to infection release inflammatory markers and viral proteins. Evidence suggests that PPARγ agonists exert anti‐inflammatory properties in neurological disorders. However, these agonists namely, thiazolidinediones have limited use in the clinic due to reported adverse side effects. INT131 is a novel non‐thiazolidinedione compound that belongs to a new class of drugs known as selective PPARγ modulators. INT131 is considered to have a safer profile; however, its neuroprotective role in vivo is not known.The goal of this study was to examine the effect of INT131 in the context of EcoHIV‐induced inflammation in vitro, in primary cultures of mouse glial cells and in vivo, in a mouse model of EcoHIV‐associated brain inflammation, as well as characterize its pharmacokinetic properties and brain penetration. In primary cultures of glial cells and in the in vivo mouse model, EcoHIV exposure resulted in a significant elevation of inflammatory markers such as TNFα, IL‐1β, CCL3, and C3 which were attenuated with INT131 treatment. Pharmacokinetic analyses revealed that INT131 penetrates into the brain with a brain to blood partition ratio Kp value of 8.5%. Overall, this is the first report to demonstrate that INT131 could be a potential candidate for the treatment of HIV‐1‐associated brain inflammation. [ABSTRACT FROM AUTHOR]

Adipose tissue is an organ that shows great plasticity and is able to adapt to the conditions to which the body is subdued. It participates in the regulation of energetic homeostasis and has endocrine functions. Recent studies have shown how the parathyroid hormone-related protein (PTHrP)/Parathyroid Hormone Receptor 1 (PTH1R) axis participates in the regulation of adipogenesis, opposing the action of Peroxisome proliferator-activated receptor gamma (PPARγ). In addition to this, PTHrP is overexpressed in adipose tissue in situations of wear and tear of the body, favoring browning and lipolysis in this tissue. It is also overexpressed in adipose tissue in stressful situations but in the opposite direction, in obesity, metabolic syndrome, type 2 diabetes mellitus (T2DM) and gestational diabetes mellitus (GDM). In conclusion, the PTHrP/PTH1R axis has a main role in adipose tissue, participating in its differentiation and remodeling. PTHrP might be used in obesity treatment and its complications for its ability to reprogram adipogenesis and adipose tissue expansion, WAT browning and for the improvement of the insulin sensitivity. In addition, PTHrP could even be used as a marker of placental status and maternal adaptations to prevent future metabolic problems in mothers and children, as well as in the treatment of bone-related diseases such as osteoporosis.

Abstract Lactase (LCT) deficiency affects approximately 75% of the world's adult population and may lead to lactose malabsorption and intolerance. Currently, the regulation of LCT gene expression remains poorly known. Peroxisome proliferator activator receptorγ (PPARγ) is a key player in carbohydrate metabolism. While the intestine is essential for carbohydrate digestion and absorption, the role of PPARγ in enterocyte metabolic functions has been poorly investigated. This study aims at characterizing PPARγ target genes involved in intestinal metabolic functions. In microarray analysis, the LCT gene was the most upregulated by PPARγ agonists in Caco‐2 cells. We confirmed that PPARγ agonists were able to increase the expression and activity of LCT both in vitro and in vivo in the proximal small bowel of rodents. The functional response element activated by PPARγ was identified in the promoter of the human LCT gene. PPARγ modulation was able to improve symptoms induced by lactose‐enriched diet in weaned rats. Our results demonstrate that PPARγ regulates LCT expression, and suggest that modulating intestinal PPARγ activity might constitute a new therapeutic strategy for lactose malabsorption.

With thymic senescence the epithelial network shrinks to be replaced by adipose tissue. Transcription factor TBX-1 controls thymus organogenesis, however, the same TBX-1 has also been reported to orchestrate beige adipose tissue development. Given these different roles of TBX-1, we have assessed if thymic TBX-1 expression persists and demonstrates this dualism during adulthood. We have also checked whether thymic adipose involution could yield beige adipose tissue. We have used adult mouse and human thymus tissue from various ages to evaluate the kinetics of TBX-1 expression, as well as mouse (TEP1) and human (1889c) thymic epithelial cells (TECs) for our studies. Electron micrographs show multi-locular lipid deposits typical of beige adipose cells. Histology staining shows the accumulation of neutral lipid deposits. qPCR measurements show persistent and/or elevating levels of beige-specific and beige-indicative markers (TBX-1, EAR-2, UCP-1, PPAR-gamma). We have performed miRNome profiling using qPCR-based QuantStudio platform and amplification-free NanoString platform. We have observed characteristic alterations, including increased miR21 level (promoting adipose tissue development) and decreased miR34a level (bias toward beige adipose tissue differentiation). Finally, using the Seahorse metabolic platform we have recorded a metabolic profile (OCR/ECAR ratio) indicative of beige adipose tissue. In summary, our results support that thymic adipose tissue emerging with senescence is bona fide beige adipose tissue. Our data show how the borders blur between a key immune tissue (the thymus) and a key metabolic tissue (beige adipose tissue) with senescence. Our work contributes to the understanding of cross talk between the immune system and metabolism.

With thymic senescence the epithelial network shrinks to be replaced by adipose tissue. Transcription factor TBX-1 controls thymus organogenesis, however, the same TBX-1 has also been reported to orchestrate beige adipose tissue development. Given these different roles of TBX-1, we have assessed if thymic TBX-1 expression persists and demonstrates this dualism during adulthood. We have also checked whether thymic adipose involution could yield beige adipose tissue. We have used adult mouse and human thymus tissue from various ages to evaluate the kinetics of TBX-1 expression, as well as mouse (TEP1) and human (1889c) thymic epithelial cells (TECs) for our studies. Electron micrographs show multi-locular lipid deposits typical of beige adipose cells. Histology staining shows the accumulation of neutral lipid deposits. qPCR measurements show persistent and/or elevating levels of beige-specific and beige-indicative markers (TBX-1, EAR-2, UCP-1, PPAR-gamma). We have performed miRNome profiling using qPCR-based QuantStudio platform and amplification-free NanoString platform. We have observed characteristic alterations, including increased miR21 level (promoting adipose tissue development) and decreased miR34a level (bias toward beige adipose tissue differentiation). Finally, using the Seahorse metabolic platform we have recorded a metabolic profile (OCR/ECAR ratio) indicative of beige adipose tissue. In summary, our results support that thymic adipose tissue emerging with senescence is bona fide beige adipose tissue. Our data show how the borders blur between a key immune tissue (the thymus) and a key metabolic tissue (beige adipose tissue) with senescence. Our work contributes to the understanding of cross talk between the immune system and metabolism. [ABSTRACT FROM AUTHOR]

In this Annual Review Issue of The Journal of Pathology, we present 15 invited reviews on topical aspects of pathology, ranging from the impacts of the microbiome in human disease through mechanisms of cell death and autophagy to recent advances in immunity and the uses of genomics for understanding, classifying and treating human cancers. Each of the reviews is authored by experts in their fields and our intention is to provide comprehensive updates in specific areas of pathology in which there has been considerable recent progress. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Peroxisome proliferator-activated receptor γ (PPARγ) is a member of the nuclear receptor family and plays an important role in adipocyte differentiation, glucose homeostasis, and insulin sensitivity. Thiazolidinediones (TZDs), synthetic ligands of PPARγ, have been used for the treatment of diabetes mellitus for two decades. TZDs were expected to be amazing drugs not only for type 2 diabetes but also for metabolic syndrome and atherosclerotic vascular disease because they can reduce both insulin resistance and inflammation in experimental studies. However, serious unwanted effects pushed TZDs back to an optional second-tier drug for type 2 diabetes. Nevertheless, PPARγ is still one of the most important targets for the treatment of insulin resistance and diabetes mellitus, and novel strategies to modulate PPARγ activity to enhance its beneficial effects and reduce unwanted adverse effects are anticipated. Recent studies showed that post-translational modification (PTM) of PPARγ regulates PPARγ activity or stability and may be a novel way to optimize PPARγ activity with reduced adverse effects. In this review, we will focus on recent advances in PTM of PPARγ and the mechanisms regulating PPARγ function as well as in the development of PPARγ modulators or agonists.

This article describes qPCR analysis for the Adig/Smaf1 gene in multiple in vitro adipocyte differentiation models including white and brown adipogenesis, cell lines and primary cultures. The article also contains qPCR data for transcript levels of Adig/Smaf1 in a wide panel of murine tissues. Expression of Adig/Smaf1 transcript in white and brown adipose tissue in fasted and refed mice is reported and also data for Adig/Smaf1 transcript expression in genetically obese ob/ob mice. Data on the effects of siRNA-mediated knockdown of Srebp1c on Adig/Smaf1 transcript levels in 3T3-L1 adipocytes are shown. Luciferase reporter assays provide data for regulation of an ~ 2 kb fragment of the 5′ flanking region of Adig/Smaf1 gene by PPARγ/RXRα. This data is related to a research article describing Adig/Smaf1 protein expression, “Expression, regulation and functional assessment of the 80 amino acid Small Adipocyte Factor 1 (Smaf1) protein in adipocytes” (G. Ren, P. Eskandari, S. Wang, C.M. Smas, 2016) [1].

Although glucocorticoids (GC) represent the most frequently used immunosuppressive drugs, their effects are still not well understood. In our previous studies, we have shown that treatment of monocytes with GC does not cause a global suppression of monocytic effector functions, but rather induces differentiation of a specific anti-inflammatory phenotype. The anti-inflammatory role of peroxisome proliferator-activated receptor (PPAR)-γ has been extensively studied during recent years. However, a relationship between GC treatment and PPAR-γ expression in macrophages has not been investigated so far. Studies using PPAR-γ-deficient mice have frequently provided controversial results. A potential reason is the use of primary cells, which commonly represent inhomogeneous populations burdened with side effects and influenced by bystander cells. To overcome this constraint, we established ER-Hoxb8-immortalized bone marrow-derived macrophages from Ppargfl/fl and LysM-Cre Ppargfl/fl mice in this study. In contrast to primary macrophages, the ER-Hoxb8 system allows the generation of a homogeneous and well-defined population of resting macrophages. We could show that the loss of PPAR-γ resulted in delayed kinetic of differentiation of monocytes into macrophages as assessed by reduced F4/80, but increased Ly6C expression in early phases of differentiation. As expected, PPAR-γ-deficient macrophages displayed an increased pro-inflammatory phenotype upon long-term LPS stimulation characterized by an elevated production of pro-inflammatory cytokines TNF-α, IL1-β, IL-6, IL-12 and a reduced production of anti-inflammatory cytokine IL-10 compared to PPAR-γ WT cells. Moreover, PPAR-γ-deficient macrophages showed impaired phagocytosis. GC treatment of macrophages led to the upregulation of PPAR-γ expression. However, there were no differences in GC-induced suppression of cytokines between both cell types, implicating a PPAR-γ-independent mechanism. Intriguingly, GC treatment resulted in an increased in vitro migration only in PPAR-γ-deficient macrophages. Performing a newly developed in vivo cell-tracking experiment, we could confirm that GC induces an increased recruitment of PPAR-γ KO, but not PPAR-γ WT macrophages to the site of inflammation. Our findings suggest a specific effect of PPAR-γ on GC-induced migration in macrophages. In conclusion, we could demonstrate that PPAR-γ exerts anti-inflammatory activities and shapes macrophage functions. Moreover, we identified a molecular link between GC and PPAR-γ and could show for the first time that PPAR-γ modulates GC-induced migration in macrophages.

Highlights • The ketogenic diet increases hippocampal catalase mRNA and protein. • The ketogenic diet does not increase catalase mRNA or protein in the hippocampus of mice lacking PPARγ2. • Inhibiting catalase occluded the KD antiseizure effects. Abstract We have previously found that the transcription factor PPARγ2 contributes to the mechanism of action of the ketogenic diet (KD), an established treatment for pediatric refractory epilepsy. Among the wide-array of genes regulated by PPARγ, previous studies have suggested that antioxidants such as catalase may have prominent roles in KD neuroprotective and antiseizure effects. Here, we tested the hypothesis that the KD increases catalase through activation of PPARγ2, and that this action is part of the mechanism of antiseizure efficacy of the KD. We determined catalase mRNA and protein expression in hippocampal tissue from epileptic Kcna1 −/− mice, Ppar γ2+/+ mice and Ppar γ2−/− mice. We found that a KD increases hippocampal catalase expression in Kcna1 −/− and Ppar γ2+/+ mice, but not Ppar γ2−/− mice. Next, we determined whether catalase contributes to KD seizure protection. We found that the KD reduces pentylenetetrazole (PTZ)-induced seizures; however, pretreatment with a catalase inhibitor occluded KD effects on PTZ seizures. These results suggest that the KD regulates catalase expression through PPARγ2 activation, and that catalase may contribute to the KD antiseizure efficacy. [ABSTRACT FROM AUTHOR]

Context: The metabolic function of peroxisome proliferator-activated receptor gamma (PPARγ) in lung cancer remains unclear. Objectives: To determine the relationship of PPARγ on ALDH1A3-induced lipid peroxidation to inhibit lung cancer cell growth. Materials and methods: In silico analysis using microarray dataset was performed to screen the positive correlation between PPARγ and all ALDH isoforms. NUBIscan software and ChIP assay were used to identify the binding sites (BSs) of PPARγ on ALDH1A3 promoter. The expression of ALDH1A3 under thiazolidinedione (TZD) treatment was evaluated by QPCR and Western Blot in HBEC and H1993 cell lines. Upon treatment of TZD, colony formation assay was used to check cell growth inhibition and 4-hydroxy-2-nonenal (4HNE) production as lipid peroxidation marker was determined by Western Blot in PPARγ positive cell H1993 and PPARγ negative cell H1299. Results: Compared to other ALDH isoforms, ALDH1A3 showed the highest positive correlation to PPARγ expression. ALDH1A3 upregulated PPARγ expression while PPARγ activation suppressed ALDH1A3. Among 2 potential screened PPARγ response elements, BS 1 and 2 in the promoter of ALDH1A3 gene, PPARγ bound directly to BS2. Ligand activation of PPARγ suppressed mRNA and protein expression of ALDH1A3. Growth inhibition was observed in H1993 (PPARγ positive cell) treated with PPARγ activator and ALDH inhibitor compared to H1299 (PPARγ negative cell). PPARγ activation increased 4HNE which is known to be suppressed by ALDH1A3. Conclusions: ALDH1A3 suppression could be one of PPARγ tumor suppressive function. This study provides a better understanding of the role of PPARγ in lung cancer. [ABSTRACT FROM AUTHOR]

Although glucocorticoids (GC) represent the most frequently used immunosuppressive drugs, their effects are still not well understood. In our previous studies, we have shown that treatment of monocytes with GC does not cause a global suppression of monocytic effector functions, but rather induces differentiation of a specific anti-inflammatory phenotype. The anti-inflammatory role of peroxisome proliferator-activated receptor (PPAR)-γ has been extensively studied during recent years. However, a relationship between GC treatment and PPAR-γ expression in macrophages has not been investigated so far. Studies using PPAR-γ-deficient mice have frequently provided controversial results. A potential reason is the use of primary cells, which commonly represent inhomogeneous populations burdened with side effects and influenced by bystander cells. To overcome this constraint, we established ER-Hoxb8-immortalized bone marrow-derived macrophages from Ppargfl/fl and LysM-Cre Ppargfl/fl mice in this study. In contrast to primary macrophages, the ER-Hoxb8 system allows the generation of a homogeneous and well-defined population of resting macrophages. We could show that the loss of PPAR-γ resulted in delayed kinetic of differentiation of monocytes into macrophages as assessed by reduced F4/80, but increased Ly6C expression in early phases of differentiation. As expected, PPAR-γ-deficient macrophages displayed an increased pro-inflammatory phenotype upon long-term LPS stimulation characterized by an elevated production of pro-inflammatory cytokines TNF-α, IL1-β, IL-6, IL-12 and a reduced production of anti-inflammatory cytokine IL-10 compared to PPAR-γ WT cells. Moreover, PPAR-γ- deficient macrophages showed impaired phagocytosis. GC treatment of macrophages led to the upregulation of PPAR-γ expression. However, there were no differences in GC-induced suppression of cytokines between both cell types, implicating a PPAR- γ-independent mechanism. Intriguingly, GC treatment resulted in an increased in vitro migration only in PPAR-γ-deficient macrophages. Performing a newly developed in vivo cell-tracking experiment, we could confirm that GC induces an increased recruitment of PPAR-γ KO, but not PPAR-γ WT macrophages to the site of inflammation. Our findings suggest a specific effect of PPAR-γ on GC-induced migration in macrophages. In conclusion, we could demonstrate that PPAR-γ exerts anti-inflammatory activities and shapes macrophage functions. Moreover, we identified a molecular link between GC and PPAR-γ and could show for the first time that PPAR-γ modulates GC-induced migration in macrophages. [ABSTRACT FROM AUTHOR]

The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) is an essential regulator of placental development. To gain deeper insights into placental PPARγ signaling, we dissected its regulation of the Mud promoter. We find that, unlike prototypic target activation by heterodimeric receptors, which is either stimulated by or refractory to retinoid X receptor (RXR) ligands (rexinoids), the induction of Mud by liganded PPARγ requires RXRα but is inhibited by rexinoids. We demonstrate that this inhibition is mediated by the activation function 2 (AF2) domain of RXRα and that Mud activation entails altered AF2 structures of both PPARγ and RXRα. This unique regulation of Mud reflects specific coactivation of PPARγ-RXRα heterodimers by the transcription cofactor ligand-dependent corepressor (LCoR), corroborated by significant downregulation of Mud in Lcor-null placentas. LCoR interacts with PPARγ and RXRα in a synergistic fashion via adjacent noncanonical protein motifs, and the AF2 domain of ligand-bound RXRα inhibits this interaction. We further identify the transcription factor Krüppel-like factor 6 (KLF6) as a critical regulator of placental development and a component of Muc1 regulation in cooperation with PPARγ, RXRα, and LCoR. Combined, these studies reveal new principles and players in nuclear receptor function in general and placental PPARγ signaling in particular. [ABSTRACT FROM AUTHOR]

Obesity and dyslipidemia is the two facet of metabolic syndrome, which needs further attention. Recent studies indicate triazole and indole derivatives have remarkable anti-obesity/antidyslipidemic activity. To harness the above-mentioned potential, a series of novel triazole clubbed indole derivatives were prepared using click chemistry and evaluated for anti-adipogenic activity. Based on the structure-activity relationship, essential functional groups which potentiate anti-adipogenic activity were identified. The lead compound 13m exhibited potent anti-adipogenic activity compared to its parent compounds with the IC-50 value of 1.67 μ M. Further evaluation of anti-adipogenic activity was conducted in different cell lines such as C3H10T1/2 and hMSC with positive result. The anti-adipogenic effect of compound 13m was most prominent in the early phase of adipogenesis, which is driven by the G1 to S phase cell cycle arrest during mitotic clonal expansion. The mechanistic study suggests that compound 13m exhibit anti-adipogenic property by activating Wnt3a/β-catenin pathway, a known suppressor of key adipogenic genes PPARγ and C/EBPα. It is noteworthy that the compound 13m also reduced serum triglyceride, LDL and total cholesterol in Syrian Golden hamster model of dyslipidemia. The anti-adipogenic activity of compound 13m can also be correlated with decreased expression of PPARγ and increased expression of β-catenin in epididymal white adipose tissue (eWAT) in vivo . The compound 13m also increased the expression of genes involved in reverse cholesterol transport (RCT) such as PPARα and LXR1α indicating another mechanism by which compound 13m ameliorates dyslipidemia in Syrian Golden hamster model. Overall this study provides a unique perspective into the anti-adipogenic/antidyslipidemic property of triazole and indole hybrids molecules with further scope to increase the anti-adipogenic potency for therapeutic intervention of obesity and metabolic syndrome. [ABSTRACT FROM AUTHOR]

Podocytes are specialized epithelial cells that maintain the glomerular filtration barrier. These cells are susceptible to lipotoxicity in the obese state and irreversibly lost during kidney disease leading to proteinuria and renal injury. PPARγ is a nuclear receptor whose activation can be renoprotective. This study examined the role of PPARγ in the lipotoxic podocyte using a PPARγ knockout (PPARγKO) cell line and since the activation of PPARγ by Thiazolidinediones (TZD) is limited by their side effects, it explored other alternative therapies to prevent podocyte lipotoxic damage. Wild-type and PPARγKO podocytes were exposed to the fatty acid palmitic acid (PA) and treated with the TZD (Pioglitazone) and/or the Retinoid X receptor (RXR) agonist Bexarotene (BX). It revealed that podocyte PPARγ is essential for podocyte function. PPARγ deletion reduced key podocyte proteins including podocin and nephrin while increasing basal levels of oxidative and ER stress causing apoptosis and cell death. A combination therapy of low-dose TZD and BX activated both the PPARγ and RXR receptors reducing PA-induced podocyte damage. This study confirms the crucial role of PPARγ in podocyte biology and that their activation in combination therapy of TZD and BX may be beneficial in the treatment of obesity-related kidney disease. [Display omitted] • PPARγ is crucial for the proper functioning and for cell survival of the podocytes • PPARγ is involved in the number and size of the lipid droplets • A combination treatment of TZD+BX exhibits synergistic effect and protective role in the podocyte [ABSTRACT FROM AUTHOR]

Thymic senescence contributes to increased incidence of infection, cancer and autoimmunity at senior ages. This process manifests as adipose involution. As with other adipose tissues, thymic adipose involution is also controlled by PPARgamma. This is supported by observations reporting that systemic PPARgamma activation accelerates thymic adipose involution. Therefore, we hypothesized that decreased PPARgamma activity could prevent thymic adipose involution, although it may trigger metabolic adverse effects. We have confirmed that both human and murine thymic sections show marked staining for PPARgamma at senior ages. We have also tested the thymic lobes of PPARgamma haplo-insufficient and null mice. Supporting our working hypothesis both adult PPARgamma haplo-insufficient and null mice show delayed thymic senescence by thymus histology, thymocyte mouse T-cell recombination excision circle qPCR and peripheral blood naive T-cell ratio by flow-cytometry. Delayed senescence showed dose–response with respect to PPARgamma deficiency. Functional immune parameters were also evaluated at senior ages in PPARgamma haplo-insufficient mice (null mice do not reach senior ages due to metabolic adverse affects). As expected, sustained and elevated T-cell production conferred oral tolerance and enhanced vaccination efficiency in senior PPARgamma haplo-insufficient, but not in senior wild-type littermates according to ELISA IgG measurements. Of note, humans also show increased oral intolerance issues and decreased protection by vaccines at senior ages. Moreover, PPARgamma haplo-insufficiency also exists in human known as a rare disease (FPLD3) causing metabolic adverse effects, similar to the mouse. When compared to age- and metabolic disorder-matched other patient samples (FPLD2 not affecting PPARgamma activity), FPLD3 patients showed increased human Trec (hTrec) values by qPCR (within healthy human range) suggesting delayed thymic senescence, in accordance with mouse results and supporting our working hypothesis. In summary, our experiments prove that systemic decrease of PPARgamma activity prevents thymic senescence, albeit with metabolic drawbacks. However, thymic tissue-specific PPARgamma antagonism would likely solve the issue.

The etiology of inflammatory bowel diseases remains largely unknown. We previously demonstrated that the expression of the peroxisome proliferator activated receptor-gamma (PPARγ) is downregulated in colonic epithelial cells of patients with ulcerative colitis (UC). PPARγ is a nuclear receptor that modulates inflammation. We hypothesized that its deficiency may play a role in the loss of intestinal homeostasis through the control of immunomodulatory factors. We found that thymic stromal lymphopoietin (TSLP), an epithelial cytokine with pleiotropic functions, is regulated by PPARγ. While this cytokine possesses two isoforms, only the short form (sfTSLP) was regulated by PPARγ. sfTSLP mRNA expression was decreased both in PPARγ knock-down Caco2 cells and cells treated with PPARγ antagonist, whereas PPARγ agonists induced the expression of sfTSLP in Caco2 and T-84 cells. The response element activated by PPARγ was identified in the promoter of the sfTSLP gene by chromatin immunoprecipitation and gene reporter assays. The expression of sfTSLP was significantly decreased in the colonic mucosa of UC patients compared to controls and was correlated with PPARγ expression. Our results identified sfTSLP as a new PPARγ-target gene and support the hypothesis that, in UC, PPARγ deficiency in colonic mucosa could play a role in the loss of intestinal tolerance through an impaired sfTSLP expression.

Thymic senescence contributes to increased incidence of infection, cancer and autoimmunity at senior ages. This process manifests as adipose involution. As with other adipose tissues, thymic adipose involution is also controlled by PPARgamma. This is supported by observations reporting that systemic PPARgamma activation accelerates thymic adipose involution. Therefore, we hypothesized that decreased PPARgamma activity could prevent thymic adipose involution, although it may trigger metabolic adverse effects. We have confirmed that both human and murine thymic sections show marked staining for PPARgamma at senior ages. We have also tested the thymic lobes of PPARgamma haplo-insufficient and null mice. Supporting our working hypothesis both adult PPARgamma haplo-insufficient and null mice show delayed thymic senescence by thymus histology, thymocyte mouse T-cell recombination excision circle qPCR and peripheral blood naive T-cell ratio by flow-cytometry. Delayed senescence showed dose-response with respect to PPARgamma deficiency. Functional immune parameters were also evaluated at senior ages in PPARgamma haplo-insufficient mice (null mice do not reach senior ages due to metabolic adverse affects). As expected, sustained and elevated T-cell production conferred oral tolerance and enhanced vaccination efficiency in senior PPARgamma haplo-insufficient, but not in senior wild-type littermates according to ELISA IgG measurements. Of note, humans also show increased oral intolerance issues and decreased protection by vaccines at senior ages. Moreover, PPARgamma haplo-insufficiency also exists in human known as a rare disease (FPLD3) causing metabolic adverse effects, similar to the mouse. When compared to age- and metabolic disorder-matched other patient samples (FPLD2 not affecting PPARgamma activity), FPLD3 patients showed increased human Trec (hTrec) values by qPCR (within healthy human range) suggesting delayed thymic senescence, in accordance with mouse results and supporting our working hypothesis. In summary, our experiments prove that systemic decrease of PPARgamma activity prevents thymic senescence, albeit with metabolic drawbacks. However, thymic tissue-specific PPARgamma antagonism would likely solve the issue. [ABSTRACT FROM AUTHOR]

Lactase (LCT) deficiency affects approximately 75% of the world's adult population and may lead to lactose malabsorption and intolerance. Currently, the regulation of LCT gene expression remains poorly known. Peroxisome proliferator activator receptorγ (PPARγ) is a key player in carbohydrate metabolism. While the intestine is essential for carbohydrate digestion and absorption, the role of PPARγ in enterocyte metabolic functions has been poorly investigated. This study aims at characterizing PPARγ target genes involved in intestinal metabolic functions. In microarray analysis, the LCT gene was the most upregulated by PPARγ agonists in Caco-2 cells. We confirmed that PPARγ agonists were able to increase the expression and activity of LCT both in vitro and in vivo in the proximal small bowel of rodents. The functional response element activated by PPARγ was identified in the promoter of the human LCT gene. PPARγ modulation was able to improve symptoms induced by lactose-enriched diet in weaned rats. Our results demonstrate that PPARγ regulates LCT expression, and suggest that modulating intestinal PPARγ activity might constitute a new therapeutic strategy for lactose malabsorption. [ABSTRACT FROM AUTHOR]

The etiology of inflammatory bowel diseases remains largely unknown. We previously demonstrated that the expression of the peroxisome proliferator activated receptor-gamma (PPAR&3947;) is downregulated in colonic epithelial cells of patients with ulcerative colitis (UC). PPAR&3947; is a nuclear receptor that modulates inflammation. We hypothesized that its deficiency may play a role in the loss of intestinal homeostasis through the control of immunomodulatory factors. We found that thymic stromal lymphopoietin (TSLP), an epithelial cytokine with pleiotropic functions, is regulated by PPAR&3947;. While this cytokine possesses two isoforms, only the short form (sfTSLP) was regulated by PPAR&3947;. sfTSLP mRNA expression was decreased both in PPAR&3947; knock-down Caco2 cells and cells treated with PPAR&3947; antagonist, whereas PPAR&3947; agonists induced the expression of sfTSLP in Caco2 and T-84 cells. The response element activated by PPAR&3947; was identified in the promoter of the sfTSLP gene by chromatin immunoprecipitation and gene reporter assays. The expression of sfTSLP was significantly decreased in the colonic mucosa of UC patients compared to controls and was correlated with PPAR&3947; expression. Our results identified sfTSLP as a new PPAR&3947;-target gene and support the hypothesis that, in UC, PPAR&3947; deficiency in colonic mucosa could play a role in the loss of intestinal tolerance through an impaired sfTSLP expression. [ABSTRACT FROM AUTHOR]

Background Cognitive impairment in humans with Alzheimer's disease (AD) and in animal models of Aβ-pathology can be ameliorated by treatments with the nuclear receptor peroxisome proliferator-activated receptor-gamma (PPARγ) agonists, such as rosiglitazone (RSG). Previously, we demonstrated that in the Tg2576 animal model of AD, RSG treatment rescued cognitive deficits and reduced aberrant activity of granule neurons in the dentate gyrus (DG), an area critical for memory formation. Methods We used a combination of mass spectrometry, confocal imaging, electrophysiology and split-luciferase assay and in vitro phosphorylation and Ingenuity Pathway Analysis. Results Using an unbiased, quantitative nano-LC-MS/MS screening, we searched for potential molecular targets of the RSG-dependent rescue of DG granule neurons. We found that S226 phosphorylation of fibroblast growth factor 14 (FGF14), an accessory protein of the voltage-gated Na + (Nav) channels required for neuronal firing, was reduced in Tg2576 mice upon treatment with RSG. Using confocal microscopy, we confirmed that the Tg2576 condition decreased PanNav channels at the AIS of the DG, and that RSG treatment of Tg2576 mice reversed the reduction in PanNav channels. Analysis from previously published data sets identified correlative changes in action potential kinetics in RSG-treated T2576 compared to untreated and wildtype controls. In vitro phosphorylation and mass spectrometry confirmed that the multifunctional kinase GSK–3β, a downstream target of insulin signaling highly implicated in AD, phosphorylated FGF14 at S226. Assembly of the FGF14:Nav1.6 channel complex and functional regulation of Nav1.6-mediated currents by FGF14 was impaired by a phosphosilent S226A mutation. Bioinformatics pathway analysis of mass spectrometry and biochemistry data revealed a highly interconnected network encompassing PPARγ, FGF14, SCN8A (Nav 1.6), and the kinases GSK–3 β, casein kinase 2β, and ERK1/2. Conclusions These results identify FGF14 as a potential PPARγ-sensitive target controlling Aβ-induced dysfunctions of neuronal activity in the DG underlying memory loss in early AD. [ABSTRACT FROM AUTHOR]

Inhibition of tissue non-specific alkaline phosphatase (TNALP) decreases intracellular lipid accumulation in human preadipocytes and the murine preadipocyte cell line, 3T3-L1. Therefore, the current study was performed to determine if TNALP is required for intracellular lipid deposition in the human hepatocyte cell line, HepG2. Intracellular lipid accumulation, TNALP activity and peroxisome proliferator activated receptor (PPAR) γ gene expression were measured in HepG2 and 3T3-L1 cells in the presence and absence of the TNALP inhibitors levamisole and histidine. Sub-cellular TNALP activity was localized using cytochemical analysis. Both PPARγ gene expression and TNALP activity increased during intracellular lipid accumulation in HepG2 and 3T3-L1 cells. Inhibition of TNALP blocked intracellular lipid accumulation but did not alter expression of the PPARγ gene. In HepG2 cells, TNALP co-localized with adipophilin on the lipid droplet membrane. These data suggest a role for TNALP in lipid droplet formation, possibly downstream from PPARγ, within HepG2 and 3T3-L1 cells. [ABSTRACT FROM AUTHOR]

Mansonic schistosomiasis is a disease caused by the trematode Schistosoma mansoni , endemic to tropical countries. S. mansoni infection induces the formation of granulomas and potent polarization of Th2-type immune response. There is great interest in understanding the mechanisms used by this parasite that causes a modulation of the immune system. Recent studies from our group demonstrated that lipids of S. mansoni , including lysophosphatidylcholine (LPC) have immunomodulatory activity. In the present study, our aim was to investigate the role of lipids derived from S. mansoni in the activation and polarization of macrophages and to characterize the mechanisms involved in this process. Peritoneal macrophages obtained from wild type C57BL/6mice or bone marrow derived macrophages were stimulated in vitro with lipids extracted from adult worms of S. mansoni . We demonstrated that total schistosomal-derived lipids as well as purified LPC induced alternatively activated macrophages/M2 profile observed by increased expression of arginase-1, mannose receptor, Chi3l3, TGFβ and production of IL-10 and PGE 2 24 h after stimulation. The involvement of the nuclear receptor PPARγ in macrophage response against LPC was investigated. Through Western blot and immunofluorescence confocal microscopy we demonstrated that schistosomal-derived LPC induces increased expression of PPARγ in macrophages. The LPC-induced increased expression of arginase-1 were significantly inhibited by the PPAR-γ antagonist GW9662. Together, these results demonstrate an immunomodulatory role of schistosomal-derived LPC in activating macrophages to a profile of the type M2 through PPARγ-dependent mechanisms, indicating a novel pathway for macrophage polarization triggered by parasite-derived LPC with potential implications to disease pathogenesis. [ABSTRACT FROM AUTHOR]

Tumor-associated macrophage (TAM) significantly contributes to cancer progression. Human cancer is enhanced by PPARγ loss-of-function mutations, but inhibited by PPARγ agonists such as TZD diabetes drugs including rosiglitazone. However, it remains enigmatic whether and how macrophage contributes to PPARγ tumor-suppressive functions. Here we report that macrophage PPARγ deletion in mice not only exacerbates mammary tumor development but also impairs the anti-tumor effects of rosiglitazone. Mechanistically, we identify Gpr132 as a novel direct PPARγ target in macrophage whose expression is enhanced by PPARγ loss but repressed by PPARγ activation. Functionally, macrophage Gpr132 is pro-inflammatory and pro-tumor. Genetic Gpr132 deletion not only retards inflammation and cancer growth but also abrogates the anti-tumor effects of PPARγ and rosiglitazone. Pharmacological Gpr132 inhibition significantly impedes mammary tumor malignancy. These findings uncover macrophage PPARγ and Gpr132 as critical TAM modulators, new cancer therapeutic targets, and essential mediators of TZD anti-cancer effects.

Background: Angiogenesis is important both in normal tissue function and disease and represents a key target in lung cancer (LC) therapy. Unfortunately, the two main subtypes of non-small-cell lung cancers (NSCLC) namely, adenocarcinoma (AC) and squamous cell carcinoma (SCC) respond differently to anti-angiogenic e.g. anti-vascular endothelial growth factor (VEGF)-A treatment with life-threatening side effects, often pulmonary hemorrhage in SCC. The mechanisms behind such adverse reactions are still largely unknown, although peroxisome proliferator activator receptor (PPAR) gamma as well as Wnt-s have been named as molecular regulators of the process. As the Wnt microenvironments in NSCLC subtypes are drastically different, we hypothesized that the particularly high levels of non-canonical Wnt5a in SCC might be responsible for alterations in blood vessel growth and result in serious adverse reactions. Methods: PPARgamma, VEGF-A, Wnt5a, miR-27b and miR-200b levels were determined in resected adenocarcinoma and squamous cell carcinoma samples by qRT-PCR and TaqMan microRNA assay. The role of PPARgamma in VEGF-A expression, and the role of Wnts in overall regulation was investigated using PPARgamma knock-out mice, cancer cell lines and fully human, in vitro 3 dimensional (3D), distal lung tissue aggregates. PPARgamma mRNA and protein levels were tested by qRT-PCR and immunohistochemistry, respectively. PPARgamma activity was measured by a PPRE reporter system. The tissue engineered lung tissues expressing basal level and lentivirally delivered VEGF-A were treated with recombinant Wnts, chemical Wnt pathway modifiers, and were subjected to PPARgamma agonist and antagonist treatment. Results: PPARgamma down-regulation and VEGF-A up-regulation are characteristic to both AC and SCC. Increased VEGF-A levels are under direct control of PPARgamma. PPARgamma levels and activity, however, are under Wnt control. Imbalance of both canonical (in AC) and non-canonical (in SCC) Wnts leads to PPARgamma downregulation. While canonical Wnts down-regulate PPARgamma directly, non-canonical Wnt5a increases miR27b that is known regulator of PPARgamma. Conclusion: During carcinogenesis the Wnt microenvironment alters, which can downregulate PPARgamma leading to increased VEGF-A expression. Differences in the Wnt microenvironment in AC and SCC of NSCLC lead to PPARgamma decrease via mechanisms that differentially alter endothelial cell motility and branching which in turn can influence therapeutic response. [ABSTRACT FROM AUTHOR]

Two consecutive multi-center phase II trials were designed to prove the hypothesis, whether therapeutic modeling of tumor-associated infl ammatory processes could result in improved tumor response. Therapy in both trials consisted of low-dose capecitabine 1g/m2 twice daily p.o. for 14 days, every 3 weeks, day 1+, and rofecoxib 25 mg daily p.o., day 1+ (from 11/04 etoricoxib 60 mg daily instead) plus pioglitazone 60 mg daily p.o., day 1+. In study II low-dose IFN-a 4.5 MU sc. three times a week, week 1+, was added until disease progression. Eighteen, and 33 patients, respectively, with clear cell renal carcinoma and progressive disease were enrolled. Objective response (48%) was exclusively observed in study II (PR 35%, CR 13%), and paralleled by a strong CRP response after 4 weeks on treatment, p = 0.0005, in all 29 pts (100%) with elevated CRP levels. Median progression-free survival could be more than doubled from a median of 4.7 months (95% CI, 1.0 to 10.4) to 11.5 months (6.8 to 16.2) in study II, p = 0.00001. Median overall survival of population II was 26 months. Efficacious negative regulation of tumor-associated infl ammation by transcription modulators may result in a steep increase of tumor response and survival.

The effect of cholesterol on fetal rat enterocytes and IEC-6 cells (line originated from normal rat small intestine) was examined. Both cells were cultured in the presence of 20 to 80 µM cholesterol for up to 72 h. Apoptosis was determined by flow cytometric analysis and fluorescence microscopy. The expression of HMG-CoA reductase and peroxisome proliferator-activated receptor gamma (PPARgamma) was measured by RT-PCR. The addition of 20 µM cholesterol reduced enterocyte proliferation as early as 6 h of culture. Reduction of enterocyte proliferation by 28 and 41% was observed after 24 h of culture in the presence and absence of 10% fetal calf serum, respectively, with the effect lasting up to 72 h. Treatment of IEC-6 cells with cholesterol for 24 h raised the proportion of cells with fragmented DNA by 9.7% at 40 µM and by 20.8% at 80 µM. When the culture period was extended to 48 h, the effect of cholesterol was still more pronounced, with the percent of cells with fragmented DNA reaching 53.5% for 40 µM and 84.3% for 80 µM. Chromatin condensation of IEC-6 cells was observed after treatment with cholesterol even at 20 µM. Cholesterol did not affect HMG-CoA reductase expression. A dose-dependent increase in PPARgamma expression in fetal rat enterocytes was observed. The expression of PPAR-gamma was raised by 7- and 40-fold, in the presence and absence of fetal calf serum, respectively, with cholesterol at 80 mM. The apoptotic effect of cholesterol on enterocytes was possibly due to an increase in PPARgamma expression.

Aims To assess the effects of peroxisome proliferator-activated receptor (PPAR) agonists on glucose tolerance and hepatic lipid metabolism in diet-induced obese mice. Main methods Male C57BL/6 mice received a standard chow diet (SC, 10% energy as lipids) or high-fat diet (HF, 50% energy as lipids) for 10 weeks, after which treatment was initiated, forming the groups: SC group, HF group, HF-BZ group (HF + bezafibrate, pan-PPAR agonist), HF-WY group (HF + WY-14643, PPARalpha agonist) and HF-GW group (HF + GW1929, PPARgamma agonist). Treatments lasted for four weeks. Insulin resistance and liver remodeling were evaluated by biochemical and molecular approaches. Key findings The HF and HF-GW mice were overweight. Conversely, the HF-BZ and HF-WY mice presented with body masses equal to those of the SC mice. All treatments restored insulin sensitivity and blood lipid and adiponectin levels. Hepatic steatosis was prevented in the HF-WY and HF-BZ mice as shown by the elevated mRNA levels of PPARalpha and Carnitine palmitoyl transferase-1a in both groups, which favored enhanced beta-oxidation. Marked decreases in liver triacylglycerol levels confirmed these findings. In contrast, the HF-GW mice exhibited increased PPARgamma and fatty acid translocase/CD136 mRNA levels, contributing to enhanced hepatic lipogenesis. Significance The WY14643 and bezafibrate treatments most effectively improved the adverse metabolic and hepatic effects caused by obesity and IR. The results reinforce the central role of PPARalpha, as well as its contrary relationship to PPARgamma in the regulation of metabolic homeostasis and lipolytic pathways in the liver. [ABSTRACT FROM AUTHOR]