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HER2, encoded by the ERBB2 gene, is an important druggable driver of human cancer gaining increasing importance as a therapeutic target in urothelial carcinoma (UC). The genomic underpinnings of HER2 overexpression in ERBB2 nonamplified UC are poorly defined. To address this knowledge gap, we investigated 172 UC tumors from patients treated at the University of California San Francisco, using immunohistochemistry and next-generation sequencing. We found that GATA3 and PPARG copy number gains individually predicted HER2 protein expression independently of ERBB2 amplification. To validate these findings, we interrogated the Memorial Sloan Kettering/The Cancer Genome Atlas (MSK/TCGA) dataset and found that GATA3 and PPARG copy number gains individually predicted ERBB2 mRNA expression independently of ERBB2 amplification. Our findings reveal a potential link between the luminal marker HER2 and the key transcription factors GATA3 and PPARG in UC and highlight the utility of examining GATA3 and PPARG copy number states to identify UC tumors that overexpress HER2 in the absence of ERBB2 amplification. In summary, we found that an increase in copy number of GATA3 and PPARG was independently associated with higher ERBB2 expression in patient samples of UC. This finding provides a potential explanation for HER2 overexpression in UC tumors without ERBB2 amplification and a way to identify these tumors for HER2-targeted therapies.

BackgroundAn imbalance of antiproliferative BMP (bone morphogenetic protein) signaling and proliferative TGF-β (transforming growth factor-β) signaling is implicated in the development of pulmonary arterial hypertension (PAH). The posttranslational modification (eg, phosphorylation and ubiquitination) of TGF-β family receptors, including BMPR2 (bone morphogenetic protein type 2 receptor)/ALK2 (activin receptor-like kinase-2) and TGF-βR2/R1, and receptor-regulated Smads significantly affects their activity and thus regulates the target cell fate. BRCC3 modifies the activity and stability of its substrate proteins through K63-dependent deubiquitination. By modulating the posttranslational modifications of the BMP/TGF-β-PPARγ pathway, BRCC3 may play a role in pulmonary vascular remodeling, hence the pathogenesis of PAH.MethodsBioinformatic analyses were used to explore the mechanism by which BRCC3 deubiquitinates ALK2. Cultured pulmonary artery smooth muscle cells (PASMCs), mouse models, and specimens from patients with idiopathic PAH were used to investigate the rebalance between BMP and TGF-β signaling in regulating ALK2 phosphorylation and ubiquitination in the context of pulmonary hypertension.ResultsBRCC3 was significantly downregulated in PASMCs from patients with PAH and animals with experimental pulmonary hypertension. BRCC3, by de-ubiquitinating ALK2 at Lys-472 and Lys-475, activated receptor-regulated Smad1/5/9, which resulted in transcriptional activation of BMP-regulated PPARγ, p53, and Id1. Overexpression of BRCC3 also attenuated TGF-β signaling by downregulating TGF-β expression and inhibiting phosphorylation of Smad3. Experiments in vitro indicated that overexpression of BRCC3 or the de-ubiquitin-mimetic ALK2-K472/475R attenuated PASMC proliferation and migration and enhanced PASMC apoptosis. In SM22α-BRCC3-Tg mice, pulmonary hypertension was ameliorated because of activation of the ALK2-Smad1/5-PPARγ axis in PASMCs. In contrast, Brcc3-/- mice showed increased susceptibility of experimental pulmonary hypertension because of inhibition of the ALK2-Smad1/5 signaling.ConclusionsThese results suggest a pivotal role of BRCC3 in sustaining pulmonary vascular homeostasis by maintaining the integrity of the BMP signaling (ie, the ALK2-Smad1/5-PPARγ axis) while suppressing TGF-β signaling in PASMCs. Such rebalance of BMP/TGF-β pathways is translationally important for PAH alleviation.

The PPARG gene encodes a member of a nuclear receptor superfamily known as peroxisome proliferator-activated gamma (PPARγ). PPARγ plays an essential role in adipogenesis, stimulating the differentiation of preadipocytes into adipocytes. Loss-of-function pathogenic variants in PPARG reduce the activity of the PPARγ receptor and can lead to severe metabolic consequences associated with familial partial lipodystrophy type 3 (FPLD3). This review focuses on recent scientific data related to FPLD3, including the role of PPARγ in adipose tissue metabolism and the phenotypic and clinical consequences of loss-of-function variants in the PPARG gene. The clinical features of 41 PPARG pathogenic variants associated with FPLD3 patients were reviewed, highlighting the genetic and clinical heterogeneity observed among 91 patients. Most of them were female, and the average age at the onset and diagnosis of lipoatrophy was 21 years and 33 years, respectively. Considering the metabolic profile, hypertriglyceridemia (91.9% of cases), diabetes (77%), hypertension (59.5%), polycystic ovary syndrome (58.2% of women), and metabolic-dysfunction-associated fatty liver disease (87,5%). We also discuss the current treatment for FPLD3. This review provides new data concerning the genetic and clinical heterogeneity in FPLD3 and highlights the importance of further understanding the genetics of this rare disease. [ABSTRACT FROM AUTHOR]

Background: Per- and polyfluoroalkyl substances (PFAS) are human-made chemicals that accumulate in the human body and the environment over time. Humans are primarily exposed to PFAS through drinking water, food, consumer products, and dust. These exposures can have many adverse health effects, including cardiovascular diseases (CVDs) and factors contributing to CVDs. This study identified the molecular mechanisms of CVDs caused by PFAS. Methods: For this purpose, various computational tools, such as the Comparative Toxicogenomic Database, ShinyGO, ChEA3, MIENTURNET, GeneMANIA, STRING, and Cytoscape, were used to conduct in silico analyses. Results: The results showed that 10 genes were common between PFAS and CVDs, and among these common genes, the PPAR signaling pathway, fatty acid metabolic processes, and lipid binding were the most significantly associated gene ontology terms. Among the top 10 transcription factors (TFs) related to these common genes, peroxisome proliferator-activated receptor gamma and androgen receptor were the most prominent. Additionally, hsa-miR-130b-3p, hsa-miR-130a-3p, and hsa-miR-129-5p were featured microRNAs involved in PFAS-induced CVDs. Finally, PPARA and PPARG were identified as core genes involved in PFAS-induced CVDs. Conclusion: These findings may contribute to a better understanding of the molecular mechanisms and reveal new potential targets in PFAS-induced CVDs. [ABSTRACT FROM AUTHOR]

Objective To investigate the effect of ginkgolide B on hepatic ischemia-reperfusion injury in mice with fatty liver disease and its mechanism. Methods A total of 25 male C57BL/6J mice were fed with 60% high-fat diet for 16 weeks, and then they were divided into sham-operation group (laparotomy and suture), model group (occlusion of blood flow in the middle and left lobes of the liver for 1 h, followed by reperfusion for 6 h), ginkgolide B group (intraperitoneal injection of 25 mg/kg ginkgolide B twice, followed by the treatment in the model group), GW9662 group (intraperitoneal injection of 1 mg/kg GW9662 twice, followed by the treatment in the model group), and GW9662+ginkgolide B group (intraperitoneal injection of 25 mg/kg ginkgolide B+1 mg/kg GW9662 twice, followed by the treatment in the model group). At 6 h after administration and surgery, the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured for each group; Western blotting was used to measure the protein expression levels of Bax, Bcl-2, and PPARγ in liver tissue; oil red O staining was used to measure the degree of fatty changes in the liver, and HE staining was used to measure the degree of necrosis in liver tissue. Results Oil red O staining showed that the degree of fatty changes in the liver had reached the level of fatty liver disease in each group. The se-rum enzymatic analysis showed that the ginkgolide B group and the GW9662+ginkgolide B group had significantly lower serum le-vels of ALT and AST than the model group (P

The PPARG gene encodes a member of a nuclear receptor superfamily known as peroxisome proliferator-activated gamma (PPARg). PPARg plays an essential role in adipogenesis, stimulating the differentiation of preadipocytes into adipocytes. Loss-of-function pathogenic variants in PPARG reduce the activity of the PPARg receptor and can lead to severe metabolic consequences associated with familial partial lipodystrophy type 3 (FPLD3). This review focuses on recent scientific data related to FPLD3, including the role of PPARg in adipose tissue metabolism and the phenotypic and clinical consequences of loss-of-function variants in the PPARG gene. The clinical features of 41 PPARG pathogenic variants associated with FPLD3 patients were reviewed, highlighting the genetic and clinical heterogeneity observed among 91 patients. Most of them were female, and the average age at the onset and diagnosis of lipoatrophy was 21 years and 33 years, respectively. Considering the metabolic profile, hypertriglyceridemia (91.9% of cases), diabetes (77%), hypertension (59.5%), polycystic ovary syndrome (58.2% of women), and metabolic-dysfunction-associated fatty liver disease (87,5%). We also discuss the current treatment for FPLD3. This review provides new data concerning the genetic and clinical heterogeneity in FPLD3 and highlights the importance of further understanding the genetics of this rare disease. [ABSTRACT FROM AUTHOR]

Interplay between energy-storing white adipose cells and thermogenic beige adipocytes contributes to obesity and insulin resistance. Irrespective of specialized niche, adipocytes require the activity of the nuclear receptor PPARγ for proper function. Exposure to cold or adrenergic signaling enriches thermogenic cells though multiple pathways that act synergistically with PPARγ; however, the molecular mechanisms by which PPARγ licenses white adipose tissue to preferentially adopt a thermogenic or white adipose fate in response to dietary cues or thermoneutral conditions are not fully elucidated. Here, we show that a PPARγ/long noncoding RNA (lncRNA) axis integrates canonical and noncanonical thermogenesis to restrain white adipose tissue heat dissipation during thermoneutrality and diet-induced obesity. Pharmacologic inhibition or genetic deletion of the lncRNA Lexis enhances uncoupling protein 1-dependent (UCP1-dependent) and -independent thermogenesis. Adipose-specific deletion of Lexis counteracted diet-induced obesity, improved insulin sensitivity, and enhanced energy expenditure. Single-nuclei transcriptomics revealed that Lexis regulates a distinct population of thermogenic adipocytes. We systematically map Lexis motif preferences and show that it regulates the thermogenic program through the activity of the metabolic GWAS gene and WNT modulator TCF7L2. Collectively, our studies uncover a new mode of crosstalk between PPARγ and WNT that preserves white adipose tissue plasticity.

Maintaining healthy adipose tissue is crucial for metabolic health, requiring a deeper understanding of adipocyte development and response to high-calorie diets. This study highlights the importance of TET3 during white adipose tissue (WAT) development and expansion. Selective depletion of Tet3 in adipose precursor cells (APCs) reduces adipogenesis, protects against diet-induced adipose expansion, and enhances whole-body metabolism. Transcriptomic analysis of wild-type and Tet3 knockout (KO) APCs unveiled TET3 target genes, including Pparg and several genes linked to the extracellular matrix, pivotal for adipogenesis and remodeling. DNA methylation profiling and functional studies underscore the importance of DNA demethylation in gene regulation. Remarkably, targeted DNA demethylation at the Pparg promoter restored its transcription. In conclusion, TET3 significantly governs adipogenesis and diet-induced adipose expansion by regulating key target genes in APCs.

Although robustly expressed in the disease-free (DF) breast stroma, CD36 is consistently absent from the stroma surrounding invasive breast cancers (IBCs). In this study, we primarily observed CD36 expression in adipocytes and intralobular capillaries within the DF breast. Larger vessels concentrated in interlobular regions lacked CD36 and were instead marked by the expression of CD31. When evaluated in perilesional capillaries surrounding ductal carcinoma in situ, a nonobligate IBC precursor, CD36 loss was more commonly observed in lesions associated with subsequent IBC. Peroxisome proliferator-activated receptor γ (PPARγ) governs the expression of CD36 and genes involved in differentiation, metabolism, angiogenesis, and inflammation. Coincident with CD36 loss, we observed a dramatic suppression of PPARγ and its target genes in capillary endothelial cells (ECs) and pericytes, which typically surround and support the stability of the capillary endothelium. Factors present in conditioned media from malignant cells repressed PPARγ and its target genes not only in cultured ECs and pericytes but also in adipocytes, which require PPARγ for proper differentiation. In addition, we identified a role for PPARγ in opposing the transition of pericytes toward a tumor-supportive myofibroblast phenotype. In mouse xenograft models, early intervention with rosiglitazone, a PPARγ agonist, demonstrated significant antitumor effects; however, following the development of a palpable tumor, the antitumor effects of rosiglitazone were negated by the repression of PPARγ in the mouse stroma. In summary, PPARγ activity in healthy tissues places several stromal cell types in an antitumorigenic state, directly inhibiting EC proliferation, maintaining adipocyte differentiation, and suppressing the transition of pericytes into tumor-supportive myofibroblasts.

IntroductionHyperoxia-induced lung injury is characterized by acute alveolar injury, disrupted epithelial-mesenchymal signaling, oxidative stress, and surfactant dysfunction, yet currently, there is no effective treatment. Although a combination of aerosolized pioglitazone (PGZ) and a synthetic lung surfactant (B-YL peptide, a surfactant protein B mimic) prevents hyperoxia-induced neonatal rat lung injury, whether it is also effective in preventing hyperoxia-induced adult lung injury is unknown.MethodUsing adult mice lung explants, we characterize the effects of 24 and 72-h (h) exposure to hyperoxia on 1) perturbations in Wingless/Int (Wnt) and Transforming Growth Factor (TGF)-β signaling pathways, which are critical mediators of lung injury, 2) aberrations of lung homeostasis and injury repair pathways, and 3) whether these hyperoxia-induced aberrations can be blocked by concomitant treatment with PGZ and B-YL combination.ResultsOur study reveals that hyperoxia exposure to adult mouse lung explants causes activation of Wnt (upregulation of key Wnt signaling intermediates β-catenin and LEF-1) and TGF-β (upregulation of key TGF-β signaling intermediates TGF-β type I receptor (ALK5) and SMAD 3) signaling pathways accompanied by an upregulation of myogenic proteins (calponin and fibronectin) and inflammatory cytokines (IL-6, IL-1β, and TNFα), and alterations in key endothelial (VEGF-A and its receptor FLT-1, and PECAM-1) markers. All of these changes were largely mitigated by the PGZ + B-YL combination.ConclusionThe effectiveness of the PGZ + B-YL combination in blocking hyperoxia-induced adult mice lung injury ex-vivo is promising to be an effective therapeutic approach for adult lung injury in vivo.

Background Diabetes-induced cardiac fibrosis is one of the main mechanisms of diabetic cardiomyopathy. As a common histone methyltransferase, enhancer of zeste homolog 2 (EZH2) has been implicated in fibrosis progression in multiple organs. However, the mechanism of EZH2 in diabetic myocardial fibrosis has not been clarified. Methods In the current study, rat and mouse diabetic model were established, the left ventricular function of rat and mouse were evaluated by echocardiography and the fibrosis of rat ventricle was evaluated by Masson staining. Primary rat ventricular fibroblasts were cultured and stimulated with high glucose (HG) in vitro. The expression of histone H3 lysine 27 (H3K27) trimethylation, EZH2, and myocardial fibrosis proteins were assayed. Results In STZ-induced diabetic ventricular tissues and HG-induced primary ventricular fibroblasts in vitro, H3K27 trimethylation was increased and the phosphorylation of EZH2 was reduced. Inhibition of EZH2 with GSK126 suppressed the activation, differentiation, and migration of cardiac fibroblasts as well as the overexpression of the fibrotic proteins induced by HG. Mechanical study demonstrated that HG reduced phosphorylation of EZH2 on Thr311 by inactivating AMP-activated protein kinase (AMPK), which transcriptionally inhibited peroxisome proliferator-activated receptor γ (PPAR-γ) expression to promote the fibroblasts activation and differentiation. Conclusion Our data revealed an AMPK/EZH2/PPAR-γ signal pathway is involved in HG-induced cardiac fibrosis.

Background: Diabetes-induced cardiac fibrosis is one of the main mechanisms of diabetic cardiomyopathy. As a common histone methyltransferase, enhancer of zeste homolog 2 (EZH2) has been implicated in fibrosis progression in multiple organs. However, the mechanism of EZH2 in diabetic myocardial fibrosis has not been clarified. Methods: In the current study, rat and mouse diabetic model were established, the left ventricular function of rat and mouse were evaluated by echocardiography and the fibrosis of rat ventricle was evaluated by Masson staining. Primary rat ventricular fibroblasts were cultured and stimulated with high glucose (HG) in vitro. The expression of histone H3 lysine 27 (H3K27) trimethylation, EZH2, and myocardial fibrosis proteins were assayed. Results: In STZ-induced diabetic ventricular tissues and HG-induced primary ventricular fibroblasts in vitro, H3K27 trimethylation was increased and the phosphorylation of EZH2 was reduced. Inhibition of EZH2 with GSK126 suppressed the activation, differentiation, and migration of cardiac fibroblasts as well as the overexpression of the fibrotic proteins induced by HG. Mechanical study demonstrated that HG reduced phosphorylation of EZH2 on Thr311 by inactivating AMP-activated protein kinase (AMPK), which transcriptionally inhibited peroxisome proliferator-activated receptor γ (PPAR-γ) expression to promote the fibroblasts activation and differentiation. Conclusion: Our data revealed an AMPK/EZH2/PPAR-γ signal pathway is involved in HG-induced cardiac fibrosis. [ABSTRACT FROM AUTHOR]

Obesity has become a pandemic, as currently more than half a billion people worldwide are obese. The etiology of obesity is multifactorial, and combines a contribution of hereditary and behavioral factors, such as nutritional inadequacy, along with the influences of environment and reduced physical activity. Two types of adipose tissue widely known are white and brown. While white adipose tissue functions predominantly as a key energy storage, brown adipose tissue has a greater mass of mitochondria and expresses the uncoupling protein 1 (UCP1) gene, which allows thermogenesis and rapid catabolism. Even though white and brown adipocytes are of different origin, activation of the brown adipocyte differentiation program in white adipose tissue cells forces them to transdifferentiate into "beige" adipocytes, characterized by thermogenesis and intensive lipolysis. Nowadays, researchers in the field of small molecule medicinal chemistry and gene therapy are making efforts to develop new drugs that effectively overcome insulin resistance and counteract obesity. Here, we discuss various aspects of white-to-beige conversion, adipose tissue catabolic re-activation, and non-shivering thermogenesis. [ABSTRACT FROM AUTHOR]

Purpose of Review: This review summarizes evidence on osteocyte support of extramedullary and bone marrow adipocyte development and discusses the role of endogenous osteocyte activities of nuclear receptors peroxisome proliferator-activated receptor gamma (PPARG) and alpha (PPARA) in this support. Recent Findings: PPARG and PPARA proteins, key regulators of glucose and fatty acid metabolism, are highly expressed in osteocytes. They play significant roles in the regulation of osteocyte secretome and osteocyte bioenergetics; both activities contributing to the levels of systemic energy metabolism in part through an effect on metabolic function of extramedullary and bone marrow adipocytes. The PPARs-controlled osteocyte endocrine/paracrine activities, including sclerostin expression, directly regulate adipocyte function, while the PPARs-controlled osteocyte fuel utilization and oxidative phosphorylation contribute to the skeletal demands for glucose and fatty acids, whose availability is under the control of adipocytes. Summary: Bone is an inherent element of systemic energy metabolism with PPAR nuclear receptors regulating osteocyte-adipocyte metabolic axes. [ABSTRACT FROM AUTHOR]

The PPARG gene encodes a member of a nuclear receptor superfamily known as peroxisome proliferator-activated gamma (PPARγ). PPARγ plays an essential role in adipogenesis, stimulating the differentiation of preadipocytes into adipocytes. Loss-of-function pathogenic variants in PPARG reduce the activity of the PPARγ receptor and can lead to severe metabolic consequences associated with familial partial lipodystrophy type 3 (FPLD3). This review focuses on recent scientific data related to FPLD3, including the role of PPARγ in adipose tissue metabolism and the phenotypic and clinical consequences of loss-of-function variants in the PPARG gene. The clinical features of 41 PPARG pathogenic variants associated with FPLD3 patients were reviewed, highlighting the genetic and clinical heterogeneity observed among 91 patients. Most of them were female, and the average age at the onset and diagnosis of lipoatrophy was 21 years and 33 years, respectively. Considering the metabolic profile, hypertriglyceridemia (91.9% of cases), diabetes (77%), hypertension (59.5%), polycystic ovary syndrome (58.2% of women), and metabolic-dysfunction-associated fatty liver disease (87,5%). We also discuss the current treatment for FPLD3. This review provides new data concerning the genetic and clinical heterogeneity in FPLD3 and highlights the importance of further understanding the genetics of this rare disease.

Inflammatory bowel disease, comprising Crohn's disease (CD) and ulcerative colitis (UC), is often debilitating. The disease etiology is multifactorial, involving genetic susceptibility, microbial dysregulation, abnormal immune activation, and environmental factors. Currently, available drug therapies are associated with adverse effects when used long-term. Therefore, the search for new drug candidates to treat IBD is imperative. The peroxisome proliferator-activated receptor-γ (PPARγ) is highly expressed in the colon. PPARγ plays a vital role in regulating colonic inflammation. 1,8-cineole, also known as eucalyptol, is a monoterpene oxide present in various aromatic plants which possess potent anti-inflammatory activity. Molecular docking and dynamics studies revealed that 1,8-cineole binds to PPARγ and if it were an agonist, that would explain the anti-inflammatory effects of 1,8-cineole. Therefore, we investigated the role of 1,8-cineole in colonic inflammation, using both in vivo and in vitro experimental approaches. Dextran sodium sulfate (DSS)-induced colitis was used as the in vivo model, and tumor necrosis factor-α (TNFα)-stimulated HT-29 cells as the in vitro model. 1,8-cineole treatment significantly decreased the inflammatory response in DSS-induced colitis mice. 1,8-cineole treatment also increased nuclear factor erythroid 2-related factor 2 (Nrf2) translocation into the nucleus to induce potent antioxidant effects. 1,8-cineole also increased colonic PPARγ protein expression. Similarly, 1,8-cineole decreased proinflammatory chemokine production and increased PPARγ protein expression in TNFα-stimulated HT-29 cells. 1,8-cineole also increased PPARγ promoter activity time-dependently. Because of its potent anti-inflammatory effects, 1,8-cineole may be valuable in treating IBD.

Abstract Interleukin-33 (IL-33), an emerging cytokine within the IL-1 family, assumes a pivotal function in the control of obesity. However, the specific mechanism of its regulation of obesity formation remains unclear. In this study, we found that the expression level of IL-33 increased in visceral adipose tissue in mice fed with a high-fat diet (HFD) compared with that in mice fed with a normal diet (ND). In vitro, we also found the expression level of IL-33 was upregulated during the adipogenesis of 3T3-L1 cells. Functional test results showed that knockdown of IL-33 in 3T3-L1 cells differentiation could promote the accumulation of lipid droplets, the content of triglyceride and the expression of adipogenic–related genes (i.e. PPAR-γ, C/EBPα, FABP4, LPL, Adipoq and CD36). In contrast, overexpression of IL-33 inhibits adipogenic differentiation. Meanwhile, the above tests were repeated after over-differentiation of 3T3-L1 cells induced by oleic acid, and the results showed that IL-33 played a more significant role in the regulation of adipogenesis. To explore the mechanism, transcriptome sequencing was performed and results showed that IL-33 regulated the PPAR signaling pathway in 3T3-L1 cells. Further, Western blot and confocal microscopy showed that the inhibition of IL-33 could promote PPAR-γ expression by inhibiting the Wnt/β-catenin signal in 3T3-L1 cells. This study demonstrated that IL-33 was an important regulator of preadipocyte differentiation and inhibited adipogenesis by regulating the Wnt/β-catenin/PPAR-γ signaling pathway, which provided a new insight for further research on IL-33 as a new intervention target for metabolic disorders.

PURPOSE: Recent studies have shown that two-dimensional (2D) culture of primary rabbit and immortalized human meibomian gland epithelial cells (iHMGEC) do not recapitulate normal meibocyte differentiation and fail to express critical enzymes necessary for synthesis of meibum lipids. The purpose of this study was to test the hypothesis that 3D-spheroid culture of iHMGEC can facilitate meibocyte differentiation and induce the expression of acyl-CoA wax-alcohol acyltransferase 2 (AWAT2), shown to be required for synthesis of meibum wax esters. METHODS: iHMGEC were suspended in matrigel/basement membrane matrix and grown in proliferation media to form distinct cell clusters or spheroids. Cells were then treated with serum-free, differentiation media (advanced DMEM/F12) with and without FGF10 and synthetic agonists for the nuclear lipid receptor, peroxisome proliferator activator receptor gamma (PPARγ). Cells were then evaluated for differentiation markers using western blotting, immunocytochemistry (ICC) and real-time PCR. Control cells were grown in standard 2D culture systems. RESULTS: Under proliferative conditions, 3D culture induced the formation of KRT5+ spheroids that contained a Ki67+/P63+ undifferentiated, basal cell population. When spheroids were switched to differentiation media containing PPARγ agonists, two different organoid populations were detected, a KRT6low population that was AWAT2+/PPARγ+ and a KRT6high population that was AWAT2-/PPARγ-, suggesting that iHMGEC exhibit a dual differentiation potential toward either a ductal or meibocyte organoid phenotype. CONCLUSION: The 3D culturing of iHMGEC can induce the formation of both meibocyte and ductal organoids and may thus serve as a better in vitro model system for studying the regulatory mechanisms controlling meibomian gland function.

Interleukin-33 (IL-33), an emerging cytokine within the IL-1 family, assumes a pivotal function in the control of obesity. However, the specific mechanism of its regulation of obesity formation remains unclear. In this study, we found that the expression level of IL-33 increased in visceral adipose tissue in mice fed with a high-fat diet (HFD) compared with that in mice fed with a normal diet (ND). In vitro, we also found the expression level of IL-33 was upregulated during the adipogenesis of 3T3-L1 cells. Functional test results showed that knockdown of IL-33 in 3T3-L1 cells differentiation could promote the accumulation of lipid droplets, the content of triglyceride and the expression of adipogenic–related genes (i.e. PPAR-γ, C/EBPα, FABP4, LPL, Adipoq and CD36). In contrast, overexpression of IL-33 inhibits adipogenic differentiation. Meanwhile, the above tests were repeated after over-differentiation of 3T3-L1 cells induced by oleic acid, and the results showed that IL-33 played a more significant role in the regulation of adipogenesis. To explore the mechanism, transcriptome sequencing was performed and results showed that IL-33 regulated the PPAR signaling pathway in 3T3-L1 cells. Further, Western blot and confocal microscopy showed that the inhibition of IL-33 could promote PPAR-γ expression by inhibiting the Wnt/β-catenin signal in 3T3-L1 cells. This study demonstrated that IL-33 was an important regulator of preadipocyte differentiation and inhibited adipogenesis by regulating the Wnt/β-catenin/PPAR-γ signaling pathway, which provided a new insight for further research on IL-33 as a new intervention target for metabolic disorders. [ABSTRACT FROM AUTHOR]

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The nuclear receptor peroxisome proliferator activated receptor-gamma (PPARg) controls adipocyte differentiation, as well as lipid metabolism and insulin sensitivity. As a result, PPARg is a promising candidate gene for a variety of human diseases, such as obesity and type 2 diabetes. Hyperglycemia associated with the metabolic syndrome and type 2 diabetes is treated with agonists of the nuclear receptor PPARg. The PPARg agonists of the thiazolidinedione type, despite being effective in normalizing blood glucose levels, have serious side effects, making the discovery of novel ligands extremely important. Natural products have historically proven to be a promising source of structures for drug development, and recent research has focused on the PPARg-activating potential of a wide range of natural products derived from traditionally used medicinal plants or dietary sources. This review highlights recent studies that have advanced our understanding of this receptor's role in metabolism, with a focus on the effects of compounds in the plant.

Decades of work have elucidated cytokine signalling and transcriptional pathways that control T cell differentiation and have led the way to targeted biologic therapies that are effective in a range of autoimmune, allergic and inflammatory diseases. Recent evidence indicates that obesity and metabolic disease can also influence the immune system1-7, although the mechanisms and effects on immunotherapy outcomes remain largely unknown. Here, using two models of atopic dermatitis, we show that lean and obese mice mount markedly different immune responses. Obesity converted the classical type 2 T helper (TH2)-predominant disease associated with atopic dermatitis to a more severe disease with prominent TH17 inflammation. We also observed divergent responses to biologic therapies targeting TH2 cytokines, which robustly protected lean mice but exacerbated disease in obese mice. Single-cell RNA sequencing coupled with genome-wide binding analyses revealed decreased activity of nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ) in TH2 cells from obese mice relative to lean mice. Conditional ablation of PPARγ in T cells revealed that PPARγ is required to focus the in vivo TH response towards a TH2-predominant state and prevent aberrant non-TH2 inflammation. Treatment of obese mice with a small-molecule PPARγ agonist limited development of TH17 pathology and unlocked therapeutic responsiveness to targeted anti-TH2 biologic therapies. These studies reveal the effects of obesity on immunological disease and suggest a precision medicine approach to target the immune dysregulation caused by obesity.

BackgroundCholangiocarcinoma is a devastating cancer with a poor prognosis. Previous reports have presented conflicting results on the role of transforming growth factor-β-induced protein (TGFBI) in malignant cancers. Currently, our understanding of the role of TGFBI in cholangiocarcinoma is ambiguous. The aim of the present study was to investigate the role of TGFBI in human cholangiocarcinoma.MethodsIterative patient partitioning (IPP) scoring and consecutive elimination methods were used to select prognostic biomarkers. mRNA and protein expression levels were determined using Gene Expression Omnibus (GEO), Western blot and ELISA analyses. Biological activities of selected biomarkers were examined using both in vitro and in vivo assays. Prognostic values were assessed using Kaplan-Meier and Liptak's z score analyses.ResultsTGFBI was selected as a candidate cholangiocarcinoma biomarker. GEO database analysis revealed significantly higher TGFBI mRNA expression levels in cholangiocarcinoma tissues compared to matched normal tissues. TGFBI protein was specifically detected in a soluble form in vitro and in vivo. TGFBI silencing evoked significant anti-cancer effects in vitro. Soluble TGFBI treatment aggravated the malignancy of cholangiocarcinoma cells both in vitro and in vivo through activation of the integrin beta-1 (ITGB1) dependent PPARγ signalling pathway. High TGFBI expression was associated with a poor prognosis in patients with cholangiocarcinoma.ConclusionsOur data suggest that TGFBI may serve as a promising prognostic biomarker and therapeutic target for cholangiocarcinoma.

Cannabidiol (CBD) is a non-psychoactive phytocannabinoid found in the Cannabis sativa plant. Human exposure to CBD can be through recreational marijuana use, commercially available CBD-containing products, and medical treatments. Previous studies found that cannabidiol may activate the master regulator of adipogenesis, peroxisome proliferator activated receptor gamma (PPARγ). Here we investigated the effects of CBD on adipogenesis in human and mouse multipotent mesenchymal stromal stem cells (MSCs). We tested the effects of CBD on nuclear receptor activation and adipogenic potential to demonstrate the mechanism of CBD effects and employed the in vitro MSC differentiation models to assess adipogenic effects of CBD.Using transient transfection assays, we demonstrated that CBD activated mouse and human PPARγ, but not its heterodimeric partner, the retinoid 'X' receptor, RXR. Our results showed that CBD increased lipid accumulation and the expression of adipogenic genes in mouse and human MSCs in vitro. Adipogenic differentiation induced by CBD was significantly decreased by the PPARγ antagonist T0070907, supporting the hypothesis that CBD promoted differentiation via PPARγ. Taken together, our results indicate that in humans and in mice, CBD induced adipogenic differentiation in MSCs through a PPARγ-dependent mechanism.

A computational platform, Boolean network explorer (BoNE), has recently been developed to infuse AI-enhanced precision into drug discovery; it enables invariant Boolean Implication Networks of disease maps for prioritizing high-value targets. Here we used BoNE to query an Inflammatory Bowel Disease (IBD)-map and prioritize a therapeutic strategy that involves dual agonism of two nuclear receptors, PPARα/γ. Balanced agonism of PPARα/γ was predicted to modulate macrophage processes, ameliorate colitis, 'reset' the gene expression network from disease to health. Predictions were validated using a balanced and potent PPARα/γ-dual-agonist (PAR5359) in Citrobacter rodentium- and DSS-induced murine colitis models. Using inhibitors and agonists, we show that balanced-dual agonism promotes bacterial clearance efficiently than individual agonists, both in vivo and in vitro. PPARα is required and sufficient to induce the pro-inflammatory cytokines and cellular ROS, which are essential for bacterial clearance and immunity, whereas PPARγ-agonism blunts these responses, delays microbial clearance; balanced dual agonism achieved controlled inflammation while protecting the gut barrier and 'reversal' of the transcriptomic network. Furthermore, dual agonism reversed the defective bacterial clearance observed in PBMCs derived from IBD patients. These findings not only deliver a macrophage modulator for use as barrier-protective therapy in IBD, but also highlight the potential of BoNE to rationalize combination therapy.

BackgroundThe comparative evolutionary genomics analysis was used to study the functions of novel Ka/Ks-predicted human exons in a zebrafish model. The Yulink (MIOS, Entrez Gene: 54,468), a conserved gene from zebrafish to human with WD40 repeats at N-terminus, was identified and found to encode an 875 amino acid in human. The biological function of this Yulink gene in cardiomyocytes remains unexplored. The purpose of this study is to determine the involvement of Yulink in the functions of cardiomyocytes and to investigate its molecular regulatory mechanism.MethodsKnockdown of Yulink was performed using morpholino or shRNA in zebrafish, mouse HL-1 cardiomyocytes, and human iPSC-derived cardiomyocytes. The expression levels of mRNA and protein were quantified by qPCR and western blots. Other methods including DNA binding, ligand uptake, agonists treatment and Ca2+ imaging assays were used to study the molecular regulatory mechanism by Yulink. Statistical data were shown as mean ± SD or mean ± standard error.ResultsThe knockdown of yulink with three specific morpholinos in zebrafish resulted in cardiac dysfunctions with pericardial edema, decreased heart beats and cardiac output. The Yulink knockdown in mouse HL-1 cardiomyocytes disrupted Ca2+ cycling, reduced DNA binding activity of PPARγ (peroxisome proliferator-activated receptor gamma) and resulted in a reduction of Serca2 (sarcoplasmic reticulum Ca2+ ATPase 2) expression. Expression of Serca2 was up-regulated by PPARγ agonists and down-regulated by PPARγ-shRNA knockdown, suggesting that Yulink regulates SERCA2 expression through PPARγ in mouse HL-1 cardiomyocytes. On the other hand, YULINK, PPARγ or SERCA2 over-expression rescued the phenotypes of Yulink KD cells. In addition, knockdown of YULINK in human iPSC-derived cardiomyocytes also disrupted Ca2+ cycling via decreased SERCA2 expression.ConclusionsOverall, our data showed that Yulink is an evolutionarily conserved gene from zebrafish to human. Mechanistically Yulink regulated Serca2 expression in cardiomyocytes, presumably mediated through PPARγ nuclear entry. Deficiency of Yulink in mouse and human cardiomyocytes resulted in irregular Ca2+ cycling, which may contribute to arrhythmogenesis.

To investigate the targeted differentiation ability of mouse bone marrow derived mesenchymal stem cells (BM-MSCs) and adipose-derived mesenchymal stem cells (AD-MSCs). Methods BM-MSCs and AD-MSCs were isolated and cultured from bone marrow of femur and white adipose tissue of groin of C57BL/6J mice respectively, and the two types of cells were induced by osteogenic, chondrogenic and adipogenic differentiation medium respectively. Alizarin red, alcian blue and oil red O staining were used to detect the differentiated degree of osteogenic, chondrogenic and lipogenic differentiation. Real-time fluorescence quantitative PCR (qPCR) was used to identify MSCs and detected expression levels of directed differentiation-related genes Runx2, Sp7 (osteoblast), Sox9, Col2a1 (chondroblast), Pparg and Cebpa (lipogenesis) to determine the directed differentiation ability of cells. Based on gene expression profiles of mouse and human BM-MSCs and AD-MSCs in GEO database GSE43804 and GSE122778, the differentially expressed genes and their enrichment signal pathways were analyzed. Results The cell morphology of BM-MSCs and AD-MSCs obtained by isolation and culture was different, and spindle-shaped morphology was more obvious in AD-MSCs. Both cells expressed CD29, CD44 and CD90, but did not express CD34 and CD45. AD-MSCs showed higher osteogenic and lipogenic differentiation than those of BM-MSCs after directed induction, while chondrogenic differentiation was lower in AD-MSCs than that of BM-MSCs (P＜0.05). After directional induction, expression levels of Runx2, Pparg and Cebpa mRNA were higher in AD-MSCs than those in BMMSCs, and Sox9 mRNA expression levels were lower than those in BM-MSCs (P＜0.05). Highly expressed genes of ADMSCs in mice and human were enriched in PPAR and WNT signaling pathways. Highly expressed genes of BM-MSCs were enriched in cartilage and bone developmental signaling pathways. Conclusion The osteogenic and adipogenic differentiation ability of mouse AD-MSCs is stronger than those of BM-MSCs, while the chondrogenic differentiation ability AD-MSCs is weaker than that of BM-MSCs. The activation status of PPAR, WNT, cartilages and skeletal system development signaling pathways plays an important regulatory role in determining the different directional differentiation potential of AD-MSCs and BM-MSCs. [ABSTRACT FROM AUTHOR]

Adipose tissue plays a major role in maintaining organismal metabolic equilibrium. Control over the fate decision from mesenchymal stem cells (MSCs) to adipocyte differentiation involves coordinated command of phosphorylation. Protein phosphatase 2A plays an important role in Wnt pathway and adipocyte development, yet how PP2A complexes actively respond to adipocyte differentiation signals and acquire specificity in the face of the promiscuous activity of its catalytic subunit remains unknown. Here, we report the PP2A phosphatase B subunit B56α is specifically induced during adipocyte differentiation and mediates PP2A to dephosphorylate GSK3β, thereby blocking Wnt activity and driving adipocyte differentiation. Using an inducible B56α knock-out mouse, we further demonstrate that B56α is essential for gonadal adipose tissue development in vivo and required for the fate decision of adipocytes over osteoblasts. Moreover, we show B56α expression is driven by the adipocyte transcription factor PPARγ thereby establishing a novel link between PPARγ signaling and Wnt blockade. Overall, our results reveal B56α is a necessary part of the machinery dictating the transition from pre-adipocyte to mature adipocyte and provide fundamental insights into how PP2A complex specifically and actively regulates unique signaling pathway in biology.

ObjectiveTo examine whether subcutaneous (SC) abdominal adipose stem cell differentiation into adipocytes in vitro predicts insulin sensitivity (Si) in vivo in normal-weight women with polycystic ovary syndrome (PCOS) and controls.DesignProspective cohort study.SettingAcademic medical center.Patient(s)Eight normal-weight women with PCOS and 8 age- and body mass index-matched controls.Intervention(s)Women underwent circulating hormone/metabolic determinations, intravenous glucose tolerance testing, total-body dual-energy x-ray absorptiometry, and SC abdominal fat biopsy.Main outcome measure(s)PPARγ and CEBPa gene expression and lipid content of adipocytes matured in vitro were compared between women with PCOS and control women, and correlated with patient characteristics, systemic Si, and adipose insulin resistance (adipose-IR).Result(s)Serum androgen levels, adipose-IR, and percentage of android fat were greater in women with PCOS than control women. Stem cell PPARγ and CEBPa gene expression increased maximally by day 12 without a female-type effect. In control cells, gene expression positively correlated with fasting serum insulin levels (both genes) and adipose-IR (CEBPa) and negatively correlated with Si (CEBPa). Conversely, CEBPa gene expression in PCOS cells negatively correlated with adipose-IR and serum free testosterone, whereas total lipid accumulation in these cells positively corelated with Si.ConclusionIn normal-weight women with PCOS, accelerated SC abdominal adipose stem cell differentiation into adipocytes in vitro favors Si in vivo, suggesting a role for hyperandrogenism in the evolution of metabolic thrift to enhance fat storage through increased cellular glucose uptake.

Activation of non-shivering thermogenesis is considered a promising approach to lower body weight in obesity. p62 deficiency in adipocytes reduces systemic energy expenditure but its role in sustaining mitochondrial function and thermogenesis remains unresolved. NBR1 shares a remarkable structural similarity with p62 and can interact with p62 through their respective PB1 domains. However, the physiological relevance of NBR1 in metabolism, as compared to that of p62, was not clear. Here we show that whole-body and adipocyte-specific ablation of NBR1 reverts the obesity phenotype induced by p62 deficiency by restoring global energy expenditure and thermogenesis in brown adipose tissue. Impaired adrenergic-induced browning of p62-deficient adipocytes is rescued by NBR1 inactivation, unveiling a negative role of NBR1 in thermogenesis under conditions of p62 loss. We demonstrate that upon p62 inactivation, NBR1 represses the activity of PPARγ, establishing an unexplored p62/NBR1-mediated paradigm in adipocyte thermogenesis that is critical for the control of obesity.

BackgroundA non-synonymous single nucleotide polymorphism of the ATG16L1 gene, T300A, is a major Crohn's disease (CD) susceptibility allele, and is known to be associated with increased apoptosis induction in the small intestinal crypt base in CD subjects and mouse models. We hypothesized that ATG16L1 T300A genotype also correlates with increased tumor apoptosis and therefore could lead to superior clinical outcome in cancer subjects.MethodsT300A genotyping by Taqman assay was performed for gastric carcinoma subjects who underwent resection from two academic medical centers. Transcriptomic analysis was performed by RNA-seq on formalin-fixed paraffin-embedded cancerous tissue. Tumor apoptosis and autophagy were determined by cleaved caspase-3 and p62 immunohistochemistry, respectively. The subjects' genotypes were correlated with demographics, various histopathologic features, transcriptome, and clinical outcome.FindingsOf the 220 genotyped subjects, 163 (74%) subjects carried the T300A allele(s), including 55 (25%) homozygous and 108 (49%) heterozygous subjects. The T300A/T300A subjects had superior overall survival than the other groups. Their tumors were associated with increased CD-like lymphoid aggregates and increased tumor apoptosis without concurrent increase in tumor mitosis or defective autophagy. Transcriptomic analysis showed upregulation of WNT/β-catenin signaling and downregulation of PPAR, EGFR, and inflammatory chemokine pathways in tumors of T300A/T300A subjects.InterpretationGastric carcinoma of subjects with the T300A/T300A genotype is associated with repressed EGFR and PPAR pathways, increased tumor apoptosis, and improved overall survival. Genotyping gastric cancer subjects may provide additional insight for clinical stratification.

Although obesity is one of the strongest risk factors for esophageal adenocarcinoma, the molecular mechanisms underlying this association remain unclear. We recently identified four esophageal adenocarcinoma-specific master regulator transcription factors (MRTF) ELF3, KLF5, GATA6, and EHF. In this study, gene-set enrichment analysis of both esophageal adenocarcinoma patient samples and cell line models unbiasedly underscores fatty acid synthesis as the central pathway downstream of three MRTFs (ELF3, KLF5, GATA6). Further characterizations unexpectedly identified a transcriptional feedback loop between MRTF and fatty acid synthesis, which mutually activated each other through the nuclear receptor, PPARG. MRTFs cooperatively promoted PPARG transcription by directly regulating its promoter and a distal esophageal adenocarcinoma-specific enhancer, leading to PPARG overexpression in esophageal adenocarcinoma. PPARG was also elevated in Barrett's esophagus, a recognized precursor to esophageal adenocarcinoma, implying that PPARG might play a role in the intestinal metaplasia of esophageal squamous epithelium. Upregulation of PPARG increased de novo synthesis of fatty acids, phospholipids, and sphingolipids as revealed by mass spectrometry-based lipidomics. Moreover, ChIP-seq, 4C-seq, and a high-fat diet murine model together characterized a novel, noncanonical, and cancer-specific function of PPARG in esophageal adenocarcinoma. PPARG directly regulated the ELF3 super-enhancer, subsequently activating the transcription of other MRTFs through an interconnected regulatory circuitry. Together, elucidation of this novel transcriptional feedback loop of MRTF/PPARG/fatty acid synthesis advances our understanding of the mechanistic foundation for epigenomic dysregulation and metabolic alterations in esophageal adenocarcinoma. More importantly, this work identifies a potential avenue for prevention and early intervention of esophageal adenocarcinoma by blocking this feedback loop. SIGNIFICANCE: These findings elucidate a transcriptional feedback loop linking epigenomic dysregulation and metabolic alterations in esophageal adenocarcinoma, indicating that blocking this feedback loop could be a potential therapeutic strategy in high-risk individuals.

5-Aminosalicylic acid (5-ASA), a peroxisome proliferator-activated receptor gamma (PPAR-γ) agonist, is a widely used first-line medication for the treatment of ulcerative colitis, but its anti-inflammatory mechanism is not fully resolved. Here, we show that 5-ASA ameliorates colitis in dextran sulfate sodium (DSS)-treated mice by activating PPAR-γ signaling in the intestinal epithelium. DSS-induced colitis was associated with a loss of epithelial hypoxia and a respiration-dependent luminal expansion of Escherichia coli, which could be ameliorated by treatment with 5-ASA. However, 5-ASA was no longer able to reduce inflammation, restore epithelial hypoxia, or blunt an expansion of E. coli in DSS-treated mice that lacked Pparg expression specifically in the intestinal epithelium. These data suggest that the anti-inflammatory activity of 5-ASA requires activation of epithelial PPAR-γ signaling, thus pointing to the intestinal epithelium as a potential target for therapeutic intervention in ulcerative colitis.IMPORTANCE An expansion of Enterobacterales in the fecal microbiota is a microbial signature of dysbiosis that is linked to many noncommunicable diseases, including ulcerative colitis. Here, we used Escherichia coli, a representative of the Enterobacterales, to show that its dysbiotic expansion during colitis can be remediated by modulating host epithelial metabolism. Dextran sulfate sodium (DSS)-induced colitis reduced mitochondrial activity in the colonic epithelium, thereby increasing the amount of oxygen available to fuel an E. coli expansion through aerobic respiration. Activation of epithelial peroxisome proliferator-activated receptor gamma (PPAR-γ) signaling with 5-aminosalicylic acid (5-ASA) was sufficient to restore mitochondrial activity and blunt a dysbiotic E. coli expansion. These data identify the host's epithelial metabolism as a potential treatment target to remediate microbial signatures of dysbiosis, such as a dysbiotic E. coli expansion in the fecal microbiota.

Infections are a major complication of obesity, but the mechanisms responsible for impaired defense against microbes are not well understood. Here, we found that adipocyte progenitors were lost from the dermis during diet-induced obesity (DIO) in humans and mice. The loss of adipogenic fibroblasts from mice resulted in less antimicrobial peptide production and greatly increased susceptibility to Staphylococcus aureus infection. The decrease in adipocyte progenitors in DIO mice was explained by expression of transforming growth factor-β (TGFβ) by mature adipocytes that then inhibited adipocyte progenitors and the production of cathelicidin in vitro. Administration of a TGFβ receptor inhibitor or a peroxisome proliferator-activated receptor-γ agonist reversed this inhibition in both cultured adipocyte progenitors and in mice and subsequently restored the capacity of obese mice to defend against S. aureus skin infection. Together, these results explain how obesity promotes dysfunction of the antimicrobial function of reactive dermal adipogenesis and identifies potential therapeutic targets to manage skin infection associated with obesity.

Efferocytosis is critical for tissue homeostasis, as its deregulation is associated with several autoimmune pathologies. While engulfing apoptotic cells, phagocytes activate transcription factors, such as peroxisome proliferator-activated receptors (PPAR) or liver X receptors (LXR) that orchestrate metabolic, phagocytic, and inflammatory responses towards the ingested material. Coordination of these transcription factors in efferocytotic human macrophages is not fully understood. In this study, we evaluated the transcriptional profile of macrophages following the uptake of apoptotic Jurkat T cells using RNA-seq analysis. Results indicated upregulation of PPAR and LXR pathways but downregulation of sterol regulatory element-binding proteins (SREBP) target genes. Pharmacological inhibition and RNA interference pointed to LXR and PPARδ as relevant transcriptional regulators, while PPARγ did not substantially contribute to gene regulation. Mechanistically, lysosomal digestion and lysosomal acid lipase (LIPA) were required for PPAR and LXR activation, while PPARδ activation also demanded an active lysosomal phospholipase A2 (PLA2G15). Pharmacological interference with LXR signaling attenuated ABCA1-dependent cholesterol efflux from efferocytotic macrophages, but suppression of inflammatory responses following efferocytosis occurred independently of LXR and PPARδ. These data provide mechanistic details on LXR and PPARδ activation in efferocytotic human macrophages.

Inflammatory bowel diseases (IBD) are chronic inflammatory disorders with increasing incidence and prevalence worldwide. Here, we investigated thymoquinone (TQ), a naturally occurring phytochemical present in Nigella sativa, for anti-inflammatory effects in colonic inflammation. To address this, we used in vivo (mice) and in vitro (HT-29 cells) models in this investigation. Our results showed that TQ treatment significantly reduced the disease activity index (DAI), myeloperoxidase (MPO) activity, and protected colon microscopic architecture. In addition, TQ also reduced the expression of proinflammatory cytokines and mediators at both the mRNA and protein levels. Further, TQ decreased phosphorylation of the activated mitogen-activated protein kinase (MAPK) signaling pathway and nuclear factor kappa B (NF-κB) proteins and enhanced colon epithelial PPAR-γ transcription factor expression. TQ significantly decreased proinflammatory chemokines (CXCL-1 and IL-8), and mediator (COX-2) mRNA expression in HT-29 cells treated with TNF-α. TQ also increased HT-29 PPAR-γ mRNA, PPAR-γ protein expression, and PPAR-γ promoter activity. These results indicate that TQ inhibits MAPK and NF-κB signaling pathways and transcriptionally regulates PPAR-γ expression to induce potent anti-inflammatory activity in vivo and in vitro models of colon inflammation.

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that, upon activation by ligands, heterodimerizes with retinoid X receptor (RXR), binds to PPAR response elements (PPREs), and activates transcription of downstream genes. As PPARγ plays a central role in adipogenesis, fatty acid storage, and glucose metabolism, PPARγ-specific pharmaceuticals (e.g., thiazolidinediones) have been developed to treat Type II diabetes and obesity within human populations. However, to our knowledge, no prior studies have concurrently assessed the effects of PPARγ ligand exposure on genome-wide PPARγ binding as well as effects on the transcriptome and lipidome within human cells at biologically active, non-cytotoxic concentrations. In addition to quantifying concentration-dependent effects of ciglitazone (a reference PPARγ agonist) and GW 9662 (a reference PPARγ antagonist) on human hepatocarcinoma (HepG2) cell viability, PPARγ abundance in situ, and neutral lipids, HepG2 cells were exposed to either vehicle (0.1% DMSO), ciglitazone, or GW 9662 for up to 24 h, and then harvested for 1) chromatin immunoprecipitation-sequencing (ChIP-seq) to identify PPARγ-bound regions across the entire genome, 2) mRNA-sequencing (mRNA-seq) to identify potential impacts on the transcriptome, and 3) lipidomics to identify potential alterations in lipid profiles. Following exposure to ciglitazone and GW 9662, we found that PPARγ levels were not significantly different after 2-8 h of exposure. While ciglitazone and GW 9662 resulted in a concentration-dependent increase in neutral lipids, the magnitude and localization of PPARγ-bound regions across the genome (as identified by ChIP-seq) did not vary by treatment. However, mRNA-seq and lipidomics revealed that exposure of HepG2 cells to ciglitazone and GW 9662 resulted in significant, treatment-specific effects on the transcriptome and lipidome. Overall, our findings suggest that exposure of human cells to PPARγ ligands at biologically active, non-cytotoxic concentrations results in toxicity that may be driven by a combination of both PPARγ-dependent and PPARγ-independent mechanisms.

Worldwide, three-quarters of a million babies are born extremely preterm (<28 weeks gestation) with devastating outcomes: 20% die in the newborn period, a further 35% develop bronchopulmonary dysplasia (BPD), and 10% suffer from cerebral palsy. Pioglitazone, a Peroxisome Proliferator Activated Receptor Gamma (PPARγ) agonist, may reduce the incidence of BPD and improve neurodevelopment in extreme preterm babies. Pioglitazone exerts an anti-inflammatory action mediated through Nuclear Factor-kappa B repression. PPARγ signalling is underactive in preterm babies as adiponectin remains low during the neonatal period. In newborn animal models, pioglitazone has been shown to be protective against BPD, necrotising enterocolitis, and lipopolysaccharide-induced brain injury. Single Nucleotide Polymorphisms of PPARγ are associated with inhibited preterm brain development and impaired neurodevelopment. Pioglitazone was well tolerated by the foetus in reproductive toxicology experiments. Bladder cancer, bone fractures, and macular oedema, seen rarely in adults, may be avoided with a short treatment course. The other effects of pioglitazone, including improved glycaemic control and lipid metabolism, may provide added benefit in the context of prematurity. Currently, there is no formulation of pioglitazone suitable for administration to preterm babies. A liquid formulation of pioglitazone needs to be developed before clinical trials. The potential benefits are likely to outweigh any anticipated safety concerns. [ABSTRACT FROM AUTHOR]

Intrauterine growth restriction (IUGR) leads to offspring obesity. In a maternal food restriction (MFR) during pregnancy-related IUGR rat model, bone marrow stem cells showed enhanced adipogenic programming; however, the effect of IUGR on white adipose tissue (WAT) progenitors is unknown. Here, by mRNA and functional profiling, we determined sex-specific adipogenic programming of WAT progenitors isolated from pups on the postnatal day (PND) 1 and 21. On PND1, PPARγ and Pref-1 expression was significantly downregulated in preadipocytes of both MFR males and females; however, at PND21, preadipocytes of MFR males showed upregulation in these genes. Even following adipogenic induction, both male and female MFR adipocytes exhibited lower PPARγ, ADRP, and adiponectin levels at PND1; however, at PND21 MFR male adipocytes showed an upward trend in the expression of these genes. An adipogenesis-specific RT-PCR array showed that male MFR adipocytes were programmed to exhibit stronger adipogenic propensity than females. Last, serum sex hormone and adipocyte estrogen/testosterone receptor expression profiles provide preliminary insights into the possible mechanism underlying sex-specific adipogenic programming in the IUGR offspring. In summary, IUGR programs WAT preadipocytes to greater adipogenic potential in males. Although the altered adipogenic programming following MFR was detectable at PND1, the changes were more pronounced at PND21, suggesting a potential role of postnatal nutrition in facilitating the sex-specific adipogenic programming in the IUGR offspring.

Heavy metals released into the environment have a significant effect on respiratory health. Lung macrophages are important in mounting an inflammatory response to injury, but they are also involved in repair of injury. Macrophages develop mixed phenotypes in complex pathological conditions and polarize to a predominant phenotype depending on the duration and stage of injury and/or repair. Little is known about the reprogramming required for lung macrophages to switch between these divergent functions; therefore, understanding the mechanism(s) by which macrophages promote metabolic reprogramming to regulate lung injury is essential. Here, we show that lung macrophages polarize to a pro-inflammatory, classically activated phenotype after cadmium-mediated lung injury. Because metabolic adaptation provides energy for the diverse macrophage functions, these classically activated macrophages show metabolic reprogramming to glycolysis. RNA-Seq revealed up-regulation of glycolytic enzymes and transcription factors regulating glycolytic flux in lung macrophages from cadmium-exposed mice. Moreover, cadmium exposure promoted increased macrophage glycolytic function with enhanced extracellular acidification rate, glycolytic metabolites, and lactate excretion. These observations suggest that cadmium mediates the persistence of classically activated lung macrophages to exacerbate lung injury.

Substance abuse disorders are linked to alteration of circadian rhythms, although the molecular and neuronal pathways implicated have not been fully elucidated. Addictive drugs, such as cocaine, induce a rapid increase of dopamine levels in the brain. Here, we show that acute administration of cocaine triggers reprogramming in circadian gene expression in the striatum, an area involved in psychomotor and rewarding effects of drugs. This process involves the activation of peroxisome protein activator receptor gamma (PPARγ), a nuclear receptor involved in inflammatory responses. PPARγ reprogramming is altered in mice with cell-specific ablation of the dopamine D2 receptor (D2R) in the striatal medium spiny neurons (MSNs) (iMSN-D2RKO). Administration of a specific PPARγ agonist in iMSN-D2RKO mice elicits substantial rescue of cocaine-dependent control of circadian genes. These findings have potential implications for development of strategies to treat substance abuse disorders.

The clinical spectra of irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) intersect to form a scantily defined overlap syndrome, termed pre-IBD. We show that increased Enterobacteriaceae and reduced Clostridia abundance distinguish the fecal microbiota of pre-IBD patients from IBS patients. A history of antibiotics in individuals consuming a high-fat diet was associated with the greatest risk for pre-IBD. Exposing mice to these risk factors resulted in conditions resembling pre-IBD and impaired mitochondrial bioenergetics in the colonic epithelium, which triggered dysbiosis. Restoring mitochondrial bioenergetics in the colonic epithelium with 5-amino salicylic acid, a PPAR-γ (peroxisome proliferator-activated receptor gamma) agonist that stimulates mitochondrial activity, ameliorated pre-IBD symptoms. As with patients, mice with pre-IBD exhibited notable expansions of Enterobacteriaceae that exacerbated low-grade mucosal inflammation, suggesting that remediating dysbiosis can alleviate inflammation. Thus, environmental risk factors cooperate to impair epithelial mitochondrial bioenergetics, thereby triggering microbiota disruptions that exacerbate inflammation and distinguish pre-IBD from IBS.

PurposeThe purpose of this study was to access the ability of the natural PPAR agonist, eicosapentaenoic acid (EPA), to activate PPAR gamma (γ) signaling leading to meibocyte differentiation in human meibomian gland epithelial cell (hMGEC).MethodsHMGEC were exposed to EPA, alone and in combination with the specific PPARγ antagonist, T0070907, to selectively block PPARγ signaling. Expression of PPARγ response genes were evaluated by qPCR. Effect on cell cycle was evaluated using Ki-67 labelling and western blots. During differentiation, autophagy was monitored using the Autophagy Tandem Sensor (ATS) and LysoTracker. Lipid accumulation was characterized by Stimulated Raman Scattering microscopy (SRS) and neutral lipid staining in combination with ER-Tracker, LysoTracker, and ATS. Autophagy was also investigated using western blotting. Seahorse XF analysis was performed to monitor mitochondrial function.ResultsEPA specifically upregulated expression of genes related to lipid synthesis and induced cell cycle exit through reduced cyclin D1 expression and increased p21 and p27 expression. EPA also induced accumulation of lipid droplets in a time and dose dependent manner (P

G protein-coupled receptor 120 (GPR120) and PPARγ agonists each have insulin sensitizing effects. But whether these two pathways functionally interact and can be leveraged together to markedly improve insulin resistance has not been explored. Here, we show that treatment with the PPARγ agonist rosiglitazone (Rosi) plus the GPR120 agonist Compound A leads to additive effects to improve glucose tolerance and insulin sensitivity, but at lower doses of Rosi, thus avoiding its known side effects. Mechanistically, we show that GPR120 is a PPARγ target gene in adipocytes, while GPR120 augments PPARγ activity by inducing the endogenous ligand 15d-PGJ2 and by blocking ERK-mediated inhibition of PPARγ. Further, we used macrophage- (MKO) or adipocyte-specific GPR120 KO (AKO) mice to show that GRP120 has anti-inflammatory effects via macrophages while working with PPARγ in adipocytes to increase insulin sensitivity. These results raise the prospect of a safer way to increase insulin sensitization in the clinic.

Background & aimsDevelopment of liver fibrosis is associated with activation of quiescent hepatic stellate cells (HSCs) into collagen type I-producing myofibroblasts (activated HSCs). Cessation of liver injury often results in fibrosis resolution and inactivation of activated HSCs/myofibroblasts into a quiescent-like state (inactivated HSCs). We aimed to identify molecular features of phenotypes of HSCs from mice and humans.MethodsWe performed studies with LratCre, Ets1-floxed, Nf1-floxed, Pparγ-floxed, Gata6-floxed, Rag2-/-γc-/-, and C57/Bl6 (control) mice. Some mice were given carbon tetrachloride (CCl4) to induce liver fibrosis, with or without a peroxisome proliferator-activated receptor-γ (PPARγ) agonist. Livers from mice were analyzed by immunohistochemistry. Quiescent, activated, and inactivated HSCs were isolated from livers of Col1α1YFP mice and analyzed by chromatin immunoprecipitation and sequencing. Human HSCs were isolated from livers denied for transplantation. We compared changes in gene expression patterns and epigenetic modifications (histone H3 lysine 4 dimethylation and histone H3 lysine 27 acetylation) in primary mouse and human HSCs. Transcription factors were knocked down with small hairpin RNAs in mouse HSCs.ResultsMotif enrichment identified E26 transcription-specific transcription factors (ETS) 1, ETS2, GATA4, GATA6, interferon regulatory factor (IRF) 1, and IRF2 transcription factors as regulators of the mouse and human HSC lineage. Small hairpin RNA-knockdown of these transcription factors resulted in increased expression of genes that promote fibrogenesis and inflammation, and loss of HSC phenotype. Disruption of Gata6 or Ets1, or Nf1 or Pparγ (which are regulated by ETS1), increased the severity of CCl4-induced liver fibrosis in mice compared to control mice. Only mice with disruption of Gata6 or Pparγ had defects in fibrosis resolution after CCl4 administration was stopped, associated with persistent activation of HSCs. Administration of a PPARγ agonist accelerated regression of liver fibrosis after CCl4 administration in control mice but not in mice with disruption of Pparγ.ConclusionsPhenotypes of HSCs from humans and mice are regulated by transcription factors, including ETS1, ETS2, GATA4, GATA6, IRF1, and IRF2. Activated mouse and human HSCs can revert to a quiescent-like, inactivated phenotype. We found GATA6 and PPARγ to be required for inactivation of human HSCs and regression of liver fibrosis in mice.

Insulin resistance is a major factor in obesity-linked type 2 diabetes. PPARγ is a master regulator of adipogenesis, and small molecule agonists, termed thiazolidinediones, are potent therapeutic insulin sensitizers. Here, we studied the role of transcriptional co-activator with PDZ-binding motif (TAZ) as a transcriptional co-repressor of PPARγ. We found that adipocyte-specific TAZ knockout (TAZ AKO) mice demonstrate a constitutively active PPARγ state. Obese TAZ AKO mice show improved glucose tolerance and insulin sensitivity compared to littermate controls. PPARγ response genes are upregulated in adipose tissue from TAZ AKO mice and adipose tissue inflammation was also decreased. In vitro and in vivo mechanistic studies revealed that the TAZ-PPARγ interaction is partially dependent on ERK-mediated Ser112 PPARγ phosphorylation. As adipocyte PPARγ Ser112 phosphorylation is increased in obesity, repression of PPARγ activity by TAZ could contribute to insulin resistance. These results identify TAZ as a new factor in the development of obesity-induced insulin resistance.

BACKGROUND/AIM:Cisplatinum (CDDP) is a first-line drug in osteosarcoma treatment and the acquisition of resistance to CDDP is associated with a poor prognosis. Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear hormone receptor that plays important roles in cell proliferation, differentiation, development, metabolism and cell death. Recently, PPARγ was reported to enhance the efficacy, overcome resistance, and decrease the toxicity of CDDP in various human cancers. In this study we tested whether pioglitazone (PIO), a PPARγ agonist, could overcome CDDP resistance in osteosarcoma. MATERIALS AND METHODS:In this study, we used a human osteosarcoma cell line and a patient-derived orthotopic xenograft (PDOX) models of osteosarcoma. We measured cell viability of 143B human osteosarcoma cells when treated with CDDP and PIO. We randomized PDOX models of osteosarcoma into four treatment groups: Group 1, Untreated control; Group 2, PIO alone; Group 3, CDDP alone; Group 4, a combination of CDDP and PIO. Each group comprised six mice. Mice were treated for 14 days and tumor size and body weight were measured. RESULTS:Cell viability of 143B human osteosarcoma cells was significantly reduced when PIO (50 μmol/l) was combined with CDDP compared to CDDP alone. PDOX osteosarcoma tumors treated with the CDDP-PIO combination showed the strongest tumor growth inhibition compared to other treatment groups. PDOX osteosarcoma tumors treated with the CDDP-PIO combination had the least cancer cells and the most necrosis in histological section. CONCLUSION:These results suggest that combining PIO along with CDDP could be an effective treatment strategy for osteosarcoma and has important clinical potential for patients.

BackgroundMaternal smoking and/or exposure to environmental tobacco smoke continue to be significant factors in fetal and childhood morbidity and are a serious public health issue worldwide. Nicotine passes through the placenta easily with minimal biotransformation, entering fetal circulation, where it results in many harmful effects on the developing offspring, especially on the developing respiratory system.ObjectivesRecently, in a rat model, electroacupuncture (EA) at maternal acupoints ST 36 has been shown to block perinatal nicotine-induced pulmonary damage; however, the underlying mechanism and the specificity of ST 36 acupoints for this effect are unknown. Here, we tested the hypothesis that compared with EA at ST 36, EA at LU 5 acupoints, which are on lung-specific meridian, will be equally or more effective in preventing perinatal nicotine-induced pulmonary changes.MethodsTwenty-four pregnant rat dams were randomly divided into 4 groups: saline ("S"), nicotine ("N"), nicotine + ST 36 (N + ST 36), and nicotine + LU 5 (N + LU 5) groups. Nicotine (1 mg/kg, subcutaneously) and EA (at ST 36 or LU 5 acupoints, bilaterally) were administered from embryonic day 6 to postnatal day 21 once daily. The "S" group was injected saline. As needed, using ELISA, western analysis, q-RT-PCR, lung histopathology, maternal and offspring hypothalamic pituitary adrenal axes, offspring key lung developmental markers, and lung morphometry were determined.ResultsWith nicotine exposure, alveolar count decreased, but mean linear intercept and septal thickness increased. It also led to a decrease in pulmonary function and PPARγ and an increase of β-catenin and glucocorticoid receptor expression in lung tissue and corticosterone in the serum of offspring rats. Electroacupuncture at ST 36 normalized all of these changes, whereas EA at LU 5 had no obvious effect.ConclusionElectroacupuncture applied to ST 36 acupoints provided effective protection against perinatal nicotine-induced lung changes, whereas EA applied at LU 5 acupoints was ineffective, suggesting mechanistic specificity and HPA axis' involvement in mediating EA at ST 36 acupoints' effects in mitigating perinatal nicotine-induced pulmonary phenotype. This opens the possibility that other acupoints, besides ST 36, can have similar or even more robust beneficial effects on the developing lung against the harmful effect of perinatal nicotine exposure. The approach proposed by us is simple, cheap, quick, easy to administer, and is devoid of any significant side effects.