为分析鸡PPARγ基因的组织表达特性及其在脂肪细胞增殖和分化过程中的功能, 文章以东北农业大学高、低腹脂双向选择品系肉鸡为实验材料, 利用Western blotting方法, 检测PPARγ基因的组织表达特性及其在高、低脂系肉鸡腹部脂肪组织间的表达差异; 采用RNAi技术, 在鸡原代脂肪细胞中抑制PPARγ基因的表达后, 通过MTT和油红O提取比色的方法, 研究鸡PPARγ基因对脂肪细胞增殖和分化的调控作用; 利用Real-time PCR和Western blotting技术, 分析PPARγ基因表达下调后, 其他脂肪细胞分化转录因子以及与脂肪细胞分化相关的重要基因的表达变化情况。结果表明, PPARγ基因在7周龄高脂系肉鸡腹部脂肪组织、肌胃、脾脏、肾脏组织中表达量较高, 在心脏中表达量较低, 在肝脏、胸肌、腿肌、十二指肠中未检测到表达信号; 与高脂系相比, PPARγ基因在5和7周龄低脂系肉鸡腹部脂肪组织中的表达量较低(P<0.05); PPARγ基因的表达量下降后, 鸡脂肪细胞的增殖能力增强, 分化能力减弱; 同时, C/EBPα、SREBP1、A-FABP、Perilipin1、LPL、IGFBP-2基因的表达量均下降(P<0.05)。由此可见, PPARγ基因的表达可能与肉鸡腹部脂肪的沉积有一定的关系, 该基因可能是调控鸡脂肪细胞增殖与分化的关键因子。

为分析鸡PPARγ基因的组织表达特性及其在脂肪细胞增殖和分化过程中的功能, 文章以东北农业大学高、低腹脂双向选择品系肉鸡为实验材料, 利用Western blotting方法, 检测PPARγ基因的组织表达特性及其在高、低脂系肉鸡腹部脂肪组织间的表达差异; 采用RNAi技术, 在鸡原代脂肪细胞中抑制PPARγ基因的表达后, 通过MTT和油红O提取比色的方法, 研究鸡PPARγ基因对脂肪细胞增殖和分化的调控作用; 利用Real-time PCR和Western blotting技术, 分析PPARγ基因表达下调后, 其他脂肪细胞分化转录因子以及与脂肪细胞分化相关的重要基因的表达变化情况。结果表明, PPARγ基因在7周龄高脂系肉鸡腹部脂肪组织、肌胃、脾脏、肾脏组织中表达量较高, 在心脏中表达量较低, 在肝脏、胸肌、腿肌、十二指肠中未检测到表达信号; 与高脂系相比, PPARγ基因在5和7周龄低脂系肉鸡腹部脂肪组织中的表达量较低(P<0.05); PPARγ基因的表达量下降后, 鸡脂肪细胞的增殖能力增强, 分化能力减弱; 同时, C/EBPα、SREBP1、A-FABP、Perilipin1、LPL、IGFBP-2基因的表达量均下降(P<0.05)。由此可见, PPARγ基因的表达可能与肉鸡腹部脂肪的沉积有一定的关系, 该基因可能是调控鸡脂肪细胞增殖与分化的关键因子。

Peroxisome proliferator-activated receptor-γ ( PPARγ) and CCAAT/enhancer binding protein-α (C/EBPα) are the master regulators of adipogenesis. The regulatory mechanism of PPARγ and C/EBPα gene expression is clear in mammals, however, little is known in chicken. The aim of the present study was to characterize chicken PPARγ promoter and investigate whether PPARγ could be regulated by C/EBPα in chickens. A 2-kb nucleotide sequence upstream of the start codon of chicken PPARγ gene was cloned and characterized by using bioinformatics and experimental approaches. This 2-kb promoter region exhibited strong promoter activity in DF1 cells. The reporter gene assay showed that the chicken C/EBPα could activate PPARγ gene promoter. Further study by electrophoretic mobility shift assay and mutational analysis revealed that the chicken C/EBPα could directly bind to and regulate the PPARγ gene promoter. Our results demonstrate that PPARγ can be directly regulated by C/EBPα in chickens .

The perexiseme proliferator-activated receptors (PPARs) comprise an important subfamily of the nuclear hormone receptor (NHR) superfamily. These ligand-activated transcription factors have been intensively studied for more than a decade and have been implicated in such diverse pathways as lipid and glucose homeostasis, control of cellular proliferation, and differentiation. The name PPAR derives from the initial cloning of one isoform as a target of various xenobiotic compounds that were observed to induce proliferation of peroxisomes in the liver (1). This protein was called the peroxisome proliferator-activated receptor, now known as PPAR#alpha#. Within a few years, the group of PPARs was expanded to include PPAR#gamma# and PPAR#delta# (also referred to as PPAR#beta#, NUC1, and FAa, R) (2-6). This review will focus on PPAR#gamma#.

The nuclear receptor PPAR?? is a master regulator of adipogenesis. PPAR?? is highly expressed in adipose tissues and its expression is markedly induced during adipogenesis. In this review, we describe the current knowledge, as well as future directions, on transcriptional and epigenetic regulation of PPAR?? expression during adipogenesis. Investigating the molecular mechanisms that control PPAR?? expression during adipogenesis is critical for understanding the development of white and brown adipose tissues, as well as pathological conditions such as obesity and diabetes. The robust induction of PPAR?? expression during adipogenesis also serves as an excellent model system for studying transcriptional and epigenetic regulation of cell-type-specific gene expression.

Abstract The nuclear hormone receptor PPAR gamma promotes adipogenesis and macrophage differentiation and is a primary pharmacological target in the treatment of type II diabetes. Here, we show that PPAR gamma gene knockout results in two independent lethal phases. Initially, PPAR gamma deficiency interferes with terminal differentiation of the trophoblast and placental vascularization, leading to severe myocardial thinning and death by E10.0. Supplementing PPAR gamma null embryos with wild-type placentas via aggregation with tetraploid embryos corrects the cardiac defect, implicating a previously unrecognized dependence of the developing heart on a functional placenta. A tetraploid-rescued mutant surviving to term exhibited another lethal combination of pathologies, including lipodystrophy and multiple hemorrhages. These findings both confirm and expand the current known spectrum of physiological functions regulated by PPAR gamma.

Neuroblastoma (NB) is the most common and aggressive pediatric cancer, characterized by a remarkable phenotypic diversity and high malignancy. The heterogeneous clinical behavior, ranging from spontaneous remission to fatal metastatic disease, is attributable to NB biology and genetics. Despite major advances in therapies, NB is still associated with a high morbidity and mortality. Thus, novel diagnostic, prognostic, and therapeutic approaches are required, mainly to improve treatment outcomes of high-risk NB patients. Among neuroepithelial cancers, NB is the most studied tumor as far as PPAR ligands are concerned. PPAR ligands are endowed with antitumoral effects, mainly acting on cancer stem cells, and constitute a possible add-on therapy to antiblastic drugs, in particular for NB with unfavourable prognosis. While discussing clinical background, this review will provide a synopsis of the major studies about PPAR expression in NB, focusing on the potential beneficial effects of hypoglycemic drugs, thiazolidinediones and metformin, to reduce the occurrence of relapses as well as tumor regrowth in NB patients.

As the prevalence of obesity has increased explosively over the last several decades, associated metabolic disorders, including type 2 diabetes, dyslipidemia, hypertension, and cardiovascular diseases, have been also increased. Thus, new strategies for preventing and treating them are needed. The nuclear peroxisome proliferator-activated receptors (PPARs) are involved fundamentally in regulating energy homeostasis; thus, they have been considered attractive drug targets for addressing metabolic disorders. Among the PPARs, PPAR纬 is a master regulator of gene expression for metabolism, inflammation, and other pathways in many cell types, especially adipocytes. It is a physiological receptor of the potent anti-diabetic drugs of the thiazolidinediones (TZDs) class, including rosiglitazone (Avandia). However, TZDs have undesirable and severe side effects, such as weight gain, fluid retention, and cardiovascular dysfunction. Recently, many reports have suggested that PPAR纬 could be modulated by post-translational modifications (PTMs), and modulation of PTM has been considered as novel approaches for treating metabolic disorders with fewer side effects than the TZDs. In this review, we discuss how PTM of PPAR纬 may be regulated and issues to be considered in making novel anti-diabetic drugs that can modulate the PTM of PPAR纬. [BMB Reports 2014; 47(11): 599-608]

Peroxisome proliferator-activated receptor-?? (PPAR??) is a ligand-activated transcription factor belonging to the nuclear receptor superfamily, which plays a central role in regulating lipid and glucose metabolism. However, accumulating evidence demonstrates that PPAR?? agonists have potential to reduce inflammation, influence the balance of immune cells, suppress oxidative stress, and improve endothelial function, which are all involved in the cellular and molecular mechanisms of cardiac fibrosis. Thus, in this review we discuss the role of PPAR?? in various cardiovascular conditions associated with cardiac fibrosis, including diabetes mellitus, hypertension, myocardial infarction, heart failure, ischemia/reperfusion injury, atrial fibrillation, and several other cardiovascular disease (CVD) conditions, and summarize the developmental status of PPAR?? agonists for the clinical management of CVD.

Peroxisome proliferator-activated receptors (PPARs) belong to the nuclear family of ligand activated transcriptional factors and comprise three different isoforms, PPAR-??, PPAR-??/??, and PPAR-??. The main role of PPARs is to regulate the expression of genes involved in lipid and glucose metabolism. Several studies have demonstrated that PPAR agonists improve dyslipidemia and glucose control in animals, supporting their potential as a promising therapeutic option to treat diabetes and dyslipidemia. However, substantial differences exist in the therapeutic or adverse effects of specific drug candidates, and clinical studies have yielded inconsistent data on their cardioprotective effects. This review summarizes the current knowledge regarding the molecular function of PPARs and the mechanisms of the PPAR regulation by posttranslational modification in the heart. We also describe the results and lessons learned from important clinical trials on PPAR agonists and discuss the potential future directions for this class of drugs.

To gain insight into the regulation of expression of peroxisome proliferator-activated receptor (PPAR) isoforms, we have determined the structural organization of the mouse PPAR γ (mPPARγ) gene. This gene extends >105 kb and gives rise to two mRNAs (mPPARγ1 and mPPARγ2) that differ at their 5' ends. The mPPARγ2 cDNA encodes an additional 30 amino acids N-terminal to the first ATG codon of mPPARγ1 and reveals a different 5' untranslated sequence. We show that mPPARγ1 mRNA is encoded by eight exons, whereas the mPPARγ2 mRNA is encoded by seven exons. Most of the 5' untranslated sequence of mPPARγ1 mRNA is encoded by two exons, whereas the 5' untranslated sequence and the extra 30 N-terminal amino acids of mPPARγ2 are encoded by one exon, which is located between the second and third exons coding for mPPARγ1. The last six exons of mPPARγ gene code for identical sequences in mPPARγ1 and mPPARγ2 isoforms. The mPPARγ1 and mPPARγ2 isoforms are transcribed from different promoters. The mPPARγ gene has been mapped to chromosome 6 E3-F1 by in situ hybridization using a biotin-labeled probe. These results establish that at least one of the PPAR genes yields more than one protein product, similar to that encountered with retinoid X receptor and retinoic acid receptor genes. The existence of multiple PPAR isoforms transcribed from different promoters could increase the diversity of ligand and tissue-specific transcriptional responses.

Pancreatic ductal adenocarcinoma(PDAC) is the fourth cause of cancer death with an overall survival of 5% at five years. The development of PDAC is characteristically associated to the accumulation of distinctive genetic mutations and is preceded by the exposure to several risk factors. Epidemiology has demonstrated that PDAC risk factors may be non-modifiable risks(sex, age, presence of genetic mutations, ethnicity) and modifiable and co-morbidity factors related to the specific habits and lifestyle. Recently it has become evident that obesity and diabetes are two important modifiable risk factors for PDAC. Obesity and diabetes are complex systemic and intertwined diseases and, over the years, experimental evidence indicate that insulin-resistance, alteration of adipokines, especially leptin and adiponectin, oxidative stress and inflammation may play a role in PDAC. Peroxisome proliferator activated receptor-纬(PPAR纬) is a nuclear receptor transcription factor that is implicated in the regulation of metabolism, differentiation and inflammation. PPAR纬 is a key regulator of adipocytes differentiation, regulates insulin and adipokines production and secretion, may modulate inflammation, and it is implicated in PDAC. PPAR纬 agonists are used in the treatment of diabetes and oxidative stressassociated diseases and have been evaluated for the treatment of PDAC. PPAR纬 is at the cross-road of diabetes, obesity, and PDAC and it is an interesting target to pharmacologically prevent PDAC in obese and diabetic patients.

Peroxisome proliferator-activated receptor gamma regulates adipogenesis. The genomic structure of the chicken peroxisome proliferator-activated receptor gamma (cPPARγ) gene has not been fully characterized, and only one cPPARγ gene mRNA sequence has been reported in genetic databases. Using 5' rapid amplification of cDNA ends, we identified five different cPPARγ mRNAs that are transcribed from three transcription initiation sites. The open reading frame analysis showed that these five cPPARγ transcript variants (cPPARγ1 to 5) could encode two cPPARγ protein isoforms (cPPARγ1 and cPPARγ2), which differ only in their N-terminal region. Quantitative real-time RT-PCR analysis showed that, of these five cPPARγ transcript variants, cPPARγ1 was ubiquitously highly expressed in various chicken tissues, including adipose tissue, liver, kidney, spleen and duodenal; cPPARγ2 was exclusively highly expressed in adipose tissue; cPPARγ3 was highly expressed in adipose tissue, kidney, spleen and liver; cPPARγ4 and cPPARγ5 were ubiquitously weakly expressed in all the tested tissues, and comparatively, cPPARγ5 was highly expressed in adipose tissue, heart, liver and kidney. The comparison of the expression of the five cPPARγ transcript variants showed that adipose tissue cPPARγ1 expression was significantly higher in the fat line than in the lean line from 2 to 7 wk of age (P<0.05 or P<0.01). Adipose tissue cPPARγ3 expression was significantly higher in the fat line than in the lean line at 3, 5 and 6 wk of age (P<0.01, P<0.05), but lower at 4 wk of age (P<0.05). Adipose tissue cPPARγ5 expression was significantly higher in the fat line than in the lean line at 3, 4, and 6 wk of age (P<0.01) and at 2 and 7 wk of age (P<0.05). This is the first report of transcript variants and protein isoforms of cPPARγ gene. Our findings provided a foundation for future investigations of the function and regulation of cPPARγ gene in adipose tissue development.

Familial partial lipodystrophy (FPLD) results from coding sequence mutations either in LMNA, encoding nuclear lamin A/C, or in PPARG, encoding peroxisome proliferator-activated receptor-gamma (PPARgamma). The LMNA form is called FPLD2 (MIM 151660) and the PPARG form is called FPLD3 (MIM 604367).Our objective was to investigate whether the clinical phenotype of this proband is due to mutation(s) in PPARgamma.This is a case report. Patient and Setting: A 31-yr-old female with the clinical phenotype of FPLD3, i.e. lipodystrophy and early childhood diabetes with extreme insulin resistance and hypertriglyceridemia leading to recurrent pancreatitis, was assessed at an academic medical center.The proband was heterozygous for a novel C-->T mutation in the PPARG gene that led to the substitution of arginine 194 in PPARgamma2 isoform, a conserved residue located in the zinc finger structure involved in DNA binding, by tryptophan (R194W). The mutation was absent from the genomes of 100 healthy Caucasians. In vitro analysis of the mutated protein showed that R194W (and R166W in PPARgamma1 isoform) could not bind to DNA and had no transcriptional activity. Furthermore, R194W had no dominant-negative activity.The R194W mutation in PPARG disrupts its DNA binding activity and through haploinsufficiency leads to clinical manifestation of FPLD3 and the associated metabolic disturbances.

There is a strong association of PPAR γ, PPAR α and PPAR δ genes on the susceptibility of T2DM in late onset but not with the early onset of T2DM subjects in North Indian Population: Dual association of PPAR γ was observed with its genotype G/G (Ala/Ala) favoring protection against T2DM and genotype C/C (Pro/Pro) favoring susceptibility to T2DM. Association of intron7 polymorphism of PPAR α and +T294C polymorphism of PPAR δ on the susceptibility to T2DM requires further analysis.

Metabolic syndrome(MetS) and type 2 diabetes mellitus(T2DM) are the serious public health problems worldwide.Moreover,it is estimated that MetS patients have about five-fold greater risk of the T2 DM development compared with people without the syndrome.Peroxisome proliferator-activated receptors are a subgroup of the nuclear hormone receptor superfamily of ligand-activated transcription factors which play an important role in the pathogenesis of MetS and T2 DM.All three members of the peroxisome proliferator-activated receptor(PPAR) nuclear receptor subfamily,PPARα,PPARp/5 and PPARγ are critical in regulating insulin sensitivity,adipogenesis,lipid metabolism,and blood pressure.Recently,more and more studies indicated that the gene polymorphism of PPARs,such as Leu~(162)Val and Val~(227)Ala of PPARα,+294T C of PPARβ/δ,Pro~(12)Ala and C1431 T of PPARγ,are significantly associated with the onset and progressing of MetS and T2 DM in different population worldwide.Furthermore,a large body of evidence demonstrated that the glucose metabolism and lipid metabolism were influenced by gene-gene interaction among PPARs genes.However,given the complexity pathogenesis of metabolic disease,it is unlikely that genetic variation of a single locus would provide an adequate explanation of inter-individual differences which results in diverse clinical syndromes.Thus,gene-gene interactions and gene-environment interactions associated with T2 DM and MetS need future comprehensive studies.

The focus of this review is the lipodystrophy syndrome caused by mutation in the PPARγ nuclear receptor – partial familial lipodystrophy FPLD3. To provide a broader context for how these mutations act to generate the clinical features of partial lipodystrophy we will review the basic biology of PPARγ and also survey the set PPARγ genetic variants that do not cause lipodystrophy, but are nonetheless associated with clinically related syndromes, specifically type 2 diabetes.

Genome-wide association studies have revealed numerous risk loci associated with diverse diseases. However, identification of disease-causing variants within association loci remains a major challenge. Divergence in gene expression due to cis -regulatory variants in noncoding regions is central to disease susceptibility. We show that integrative computational analysis of phylogenetic conservation with a complexity assessment of co-occurring transcription factor binding sites (TFBS) can identify cis -regulatory variants and elucidate their mechanistic role in disease. Analysis of established type 2 diabetes risk loci revealed a striking clustering of distinct homeobox TFBS. We identified the PRRX1 homeobox factor as a repressor of PPARG2 expression in adipose cells and demonstrate its adverse effect on lipid metabolism and systemic insulin sensitivity, dependent on the rs4684847 risk allele that triggers PRRX1 binding. Thus, cross-species conservation analysis at the level of co-occurring TFBS provides a valuable contribution to the translation of genetic association signals to disease-related molecular mechanisms.

ABSTRACT The peroxisome proliferator-activated receptor-gamma (PPARgamma) plays a role in adipocyte differentiation and insulin sensitization. It has been shown that genetic variation in the PPARgamma gene alters body weight control, lipid and insulin homeostasis, and the susceptibility to type 2 diabetes. Four PPARgamma isoforms are generated by alternative splicing and promoter usage. PPARgamma3 is only expressed in adipose tissue, colon, and macrophages and therefore seems to be a good candidate gene for metabolic and cardiovascular-associated diseases. In the present study, we looked for genetic variation in the PPARgamma3 promoter. The proximal PPARgamma3 promoter was sequenced in 20 individuals. We detected a C/G polymorphism at position -681 from exon A2. Interestingly, it was located in a signal transducer and activator of transcription 5B (STAT5B) binding consensus site. In a French population (n=836), the -681G allele was associated with increased height and plasma low-density lipoprotein cholesterol concentrations. In vitro, we showed that the -681G allele completely abolished the binding of STAT5B to the cognate promoter element as well as the transactivation of the PPARgamma3 promoter by the growth hormone/STAT5B pathway. Our results suggest that PPARgamma3 may regulate the control of height and lipid homeostasis via the STAT5B pathway.

PPARγ and C/EBPα are critical transcription factors in adipogenesis, but the precise role of these proteins has been difficult to ascertain because they positively regulate each other's expression. Questions remain about whether these factors operate independently in separate, parallel pathways of differentiation, or whether a single pathway exists. PPARγ can promote adipogenesis in C/EBPα-deficient cells, but the converse has not been tested. We have created an immortalized line of fibroblasts lacking PPARγ, which we use to show that C/EBPα has no ability to promote adipogenesis in the absence of PPARγ. These results indicate that C/EBPα and PPARγ participate in a single pathway of fat cell development with PPARγ being the proximal effector of adipogenesis.

Peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα) are key activators of adipogenesis. They mutually induce the expression of each other and have been reported to cooperate in activation of a few adipocyte genes. Recently, genome-wide profiling revealed a high degree of overlap between PPARγ and C/EBPα binding in adipocytes, suggesting that cooperativeness could be mediated through common binding sites. To directly investigate the interplay between PPARγ and C/EBPα at shared binding sites, we established a fibroblastic model system in which PPARγ and C/EBPα can be independently expressed. Using RNA sequencing, we demonstrate that coexpression of PPARγ and C/EBPα leads to synergistic activation of many key metabolic adipocyte genes. This is associated with extensive C/EBPα-mediated reprogramming of PPARγ binding and vice versa in the vicinity of these genes, as determined by chromatin immunoprecipitation combined with deep sequencing. Our results indicate that this is at least partly mediated by assisted loading involving chromatin remodeling directed by the leading factor. In conclusion, we report a novel mechanism by which the key adipogenic transcription factors, PPARγ and C/EBPα, cooperate in activation of the adipocyte gene program.

78 High-throughput cDNA screening identified TLE3 as a promoter of adipocyte differentiation 78 TLE3 is induced by PPARγ and participates in a feed-forward adipogenic loop 78 TLE3 facilitates PPARγ action on the promoters of adipocyte target genes 78 TLE3 blocks β-catenin action and antagonizes Wnt signaling during adipogenesis

The Ebf (O/E) family of helix-loop-helix transcription factors plays a significant role in B lymphocyte and neuronal development. The three primary members of this family, Ebf1, 2, and 3, are all expressed in adipocytes, and Ebf1 promotes adipogenesis when overexpressed in NIH 3T3 fibroblasts. Here we report that these three proteins have adipogenic potential in multiple cellular models and that peroxisome proliferator-activated receptor gamma (PPAR gamma) is required for this effect, at least in part due to direct activation of the PPAR gamma 1 promoter by Ebf1. Ebf1 also directly binds to and activates the C/EBP alpha promoter, which exerts positive feedback on C/EBP delta expression. Despite this, C/EBP alpha is dispensable for the adipogenic action of Ebf proteins. Ebf1 itself is induced by C/EBP beta and delta, which bind and activate its promoter. Reduction of Ebf1 and Ebf2 proteins by specific short hairpin RNA blocks differentiation of 3T3-L1 cells, suggesting a critical role for these factors and the absence of functional redundancy between members of this family. Altogether, these data place Ebf1 within the known transcriptional cascade of adipogenesis and suggest critical roles for Ebf1 and Ebf2.

When preadipocytes reenter the cell cycle, PPARγ expression is induced, coincident with an increase in DNA synthesis, suggesting the involvement of the E2F family of cell cycle regulators. We show here that E2F1 induces PPARγ transcription during clonal expansion, whereas E2F4 represses PPARγ expression during terminal adipocyte differentiation. Using a combination of in vivo experiments with knockout and chimeric animals and in vitro experiments, we demonstrate that the absence of E2F1 impairs, whereas depletion of E2F4 stimulates, adipogenesis. E2Fs hence represent the link between proliferative signaling pathways, triggering clonal expansion, and terminal adipocyte differentiation through regulation of PPARγ expression. This underscores the complex role of the E2F protein family in the control of both cell proliferation and differentiation.

Adipogenesis is the process of differentiation from preadipocytes to adipocytes and is orchestrated by various transcription factors, such as the peroxisome proliferator-activated receptor gamma (PPARγ) and the CCAAT-enhancer-binding protein alpha (C/EBPα). Oxidative stress is also a crucial factor in adipogenesis, and adipocyte differentiation is affected by the cellular redox status. The nuclear factor E2-related factor 2 (Nrf2), which is a basic leucine zipper (bZIP) transcription factor, acts as a regulator of cellular oxidative stress. Although several previous studies examined the function of Nrf2 in adipogenesis, their results were controversial. In this study, we investigated whether the suppression of Nrf2 in 3T3-L1 cells affected adipogenesis. We found that adipogenesis master regulator genes, such as PPARγ and C/EBPα , were downregulated during the differentiation stage in Nrf2-knockdown 3T3-L1 cells. Moreover, the fibroblast growth factor 21 (FGF21) and manganese superoxide dismutase (MnSOD) were markedly downregulated in Nrf2-knockdown 3T3-L1 cells. Taken together, the results of the present study suggest that the suppression of Nrf2 attenuates adipogenesis and decreases FGF21 expression through PPARγ in 3T3-L1 cells.

PPARγ and Wnt signaling are central positive and negative regulators of adipogenesis, respectively. Here we identified that, eicosapentaenoic acid (EPA) could effectively induce the transdifferentiation of myoblasts into adipocytes through modulation of both PPARγ expression and Wnt signaling. During the early stage of transdifferentiation, EPA activates PPARδ and PPARγ1, which in turn targets β-catenin to degradation and down-regulates Wnt/β-catenin signaling, such that the myogenic fate of myoblasts could be switched to adipogenesis. In addition, EPA up-regulates the expression of PPARγ1 by activating RXRα, then PPARγ1 binds to the functional peroxisome proliferator responsive element (PPRE) in the promoter of adipocyte-specific PPARγ2 to continuously activate the expression of PPARγ2 throughout the transdifferentiation process. Our data indicated that EPA acts as a dual-function stimulator of adipogenesis that both inhibits Wnt signaling and induces PPARγ2 expression to facilitate the transdifferentiation program, and the transcriptional activation of PPARγ2 by PPARγ1 is not only the key factor for the transdifferentiation of myoblasts to adipocytes, but also the crucial evidence for successful transdifferentiation. The present findings provided insight for the first time as to how EPA induces the transdifferentiation of myoblasts to adipocytes, but also provide new clues for strategies to prevent and treat some metabolic diseases.

Signal transducer and activator of transcription 5 () mediates of many cytokines and . Here we show that functions as an initial mediator of adipogenesis. The preadipocyte line 3T3-L1 undergoes upon appropriate hormonal induction. We found that Stat5A and Stat5B were strongly activated at an early stage of 3T3-L1 differentiation. To investigate physiological roles of in adipogenesis, we have constructed 3T3-L1 lines in which either an exogenous wild type (wt) or dominant negative (dn) form of Stat5A expression was controlled under the -regulatable promoter. Precocious induction of wt-Stat5A in promoted accumulation of within the . In contrast, induction of dn-Stat5A attenuated accumulation. Northern blot analyses revealed that the expression of proadipogenic transcription factors was influenced in a complementary fashion by ectopic expression of either wt- or dn-Stat5A. Notably, regulated expression of , which plays crucial roles in adipogenesis. We have also generated in which dn-Stat5A is expressed in an adipose tissue-specific fashion and found attenuation of and of many adipocyte-related genes. These results highlight a novel role of in .

A detailed understanding of the processes governing adipose tissue formation will be instrumental in combating the obesity epidemic. Much progress has been made in the last two decades in defining transcriptional events controlling the differentiation of mesenchymal stem cells into adipocytes. A complex network of transcription factors and cell-cycle regulators, in concert with specific transcriptional coactivators and corepressors, respond to extracellular stimuli to activate or repress adipocyte differentiation. This review summarizes advances in this field, which constitute a framework for potential antiobesity strategies.

Kr眉ppel-like factors (KLFs) recognize CACCC and GC-rich sequences in gene regulatory elements. Here, we describe the disruption of the murine basic Kr眉ppel-like factor gene (Bklf or Klf3). Klf3 knockout mice have less white adipose tissue, and their fat pads contain smaller and fewer cells. Adipocyte differentiation is altered in murine embryonic fibroblasts from Klf3 knockouts. Klf3 expression was studied in the 3T3-L1 cellular system. Adipocyte differentiation is accompanied by a decline in Klf3 expression, and forced overexpression of Klf3 blocks 3T3-L1 differentiation. Klf3 represses transcription by recruiting C-terminal binding protein (CtBP) corepressors. CtBPs bind NADH and may function as metabolic sensors. A Klf3 mutant that does not bind CtBP cannot block adipogenesis. Other KLFs, Klf2, Klf5, and Klf15, also regulate adipogenesis, and functional CACCC elements occur in key adipogenic genes, including in the C/ebpalpha promoter. We find that C/ebpalpha is derepressed in Klf3 and Ctbp knockout fibroblasts and adipocytes from Klf3 knockout mice. Chromatin immunoprecipitations confirm that Klf3 binds the C/ebpalpha promoter in vivo. These results implicate Klf3 and CtBP in controlling adipogenesis.

<P>Krüppel-like factor 7 (KLF7) has been extensively studied in mammalian species, but its role in birds is still unclear. In the current study, cloning and sequencing showed that the full-length coding region of chicken KLF7 (Gallus gallus KLF7, gKLF7) was 891 bp long, encoding 296 amino acids. In addition, real-time RT-PCR analysis showed that gKLF7 was broadly expressed in all 15 chicken tissues selected, and its expression was significantly different in spleen, proventriculus, abdominal fat, brain, leg muscle, gizzard and heart between fat and lean broilers at 7 weeks of age. Additionally, one novel single nucleotide polymorphism (SNP), XM_426569.3: c. A141G, was identified in the second exon of gKLF7. Association analysis showed that this locus was significantly associated with fatness traits in Arbor Acres broiler random population and the eighth generation of Northeast Agricultural University broiler lines divergently selected for abdominal fat content (NEAUHLF) population (P65<650.05). These results suggest that gKLF7 might be a candidate gene for chicken fatness traits.</P>

Obesity is an important public health problem associated with a number of disease states such as diabetes and arteriosclerosis. As such, an understanding of the mechanisms governing adipose tissue differentiation and function is of considerable importance. We recently reported that the Kr眉ppel-like zinc finger transcription factor KLF15 can induce adipocyte maturation and GLUT4 expression. In this study, we identify that a second family member, KLF2/Lung Kr眉ppel-like factor (LKLF), as a negative regulator of adipocyte differentiation. KLF2 is highly expressed in adipose tissue, and studies in cell lines and primary cells demonstrate that KLF2 is expressed in preadipocytes but not mature adipocytes. Constitutive overexpression of KLF2 but not KLF15 potently inhibits peroxisome proliferator-activated receptor-gamma (PPARgamma) expression with no effect on the upstream regulators C/EBPbeta and C/EBPdelta. However, the expression of C/EBPalpha and SREBP1c/ADD1 (adipocyte determination and differentiation factor-1/sterol regulatory element-binding protein-1), two factors that feedback in a positive manner to enhance PPARgamma function, was also markedly reduced. In addition, transient transfection studies show that KLF2 directly inhibits PPARgamma2 promoter activity (70% inhibition; p < 0.001). Using a combination of promoter mutational analysis and gel mobility shift assays, we have identified a binding site within the PPARgamma2 promoter, which mediates this inhibitory effect. These data identify a novel role for KLF2 as a negative regulator of adipogenesis.

Studies in mammalian species showed that Krüppel-like factor 2 (KLF2) regulates adipogenesis. However, its role in birds is unclear. The objective of the current study was to explore the expression and function of KLF2 in chicken adipogenesis. Results showed that chicken KLF2 (Gallus gallus KLF2 [gKLF2]) was greatly expressed in abdominal adipose tissue, and its transcripts fluctuated during adipose tissue development. In addition, gKLF2 transcripts in abdominal adipose tissue of lean broilers were greater at 1 wk of age but lower at 3, 5, and 8 wk of age than those in fat broilers (P < 0.05). The gKLF2 was more greatly expressed in preadipocytes than in mature adipocytes (P < 0.05), and its expression level decreased during the preadipocyte differentiation in vitro (P < 0.05). The functional analysis showed that gKLF2 overexpression inhibited chicken preadipocyte differentiation (P < 0.05), accompanied by the reduced expression of CCAAT/enhancer binding protein α (C/EBPα) and peroxisome proliferator-activated receptor γ (PPARγ) and the elevated expression of GATA binding protein 2 (GATA2). Additionally, the luciferase reporter assays showed that gKLF2 overexpression suppressed the promoter activities of chicken C/EBPα and PPARγ (P < 0.05). In conclusion, our results indicated that gKLF2 inhibits chicken adipogenesis, at least in part, through inhibition of PPARγ and C/EBPα expression.

Abstract The transcriptional factor FoxO1 plays an important role in metabolic homeostasis. Herein we identify a novel transrepressional function that converts FoxO1 from an activator of transcription to a promoter-specific repressor of peroxisome proliferator-activated receptor gamma (PPARgamma) target genes that regulate adipocyte biology. FoxO1 transrepresses PPARgamma via direct protein-protein interactions; it is recruited to PPAR response elements (PPRE) on PPARgamma target genes by PPARgamma bound to PPRE and interferes with promoter DNA occupancy of the receptor. The FoxO1 transrepressional function, which is independent and dissectible from the transactivational effects, does not require a functional FoxO1 DNA binding domain, but dose require an evolutionally conserved 31 amino acids LXXLL-containing domain. Insulin induces FoxO1 phosphorylation and nuclear exportation, which prevents FoxO1-PPARgamma interactions and rescues transrepression. Adipocytes from insulin resistant mice show reduced phosphorylation and increased nuclear accumulation of FoxO1, which is coupled to lowered expression of endogenous PPARgamma target genes. Thus the innate FoxO1 transrepression function enables insulin to augment PPARgamma activity, which in turn leads to insulin sensitization, and this feed-forward cycle represents positive reinforcing connections between insulin and PPARgamma signaling.

Olfactory receptors (ORs) comprise the largest G protein-coupled receptor gene superfamily. Recent studies indicate that ORs are also expressed in non-olfactory organs, including metabolically active tissues, although their biological functions in these tissues are largely unknown. In this study, OR1A1 expression was detected in HepG2 liver cells. OR1A1 activation by (61)-carvone, a known OR1A1 ligand, increased the cyclic adenosine monophosphate (cAMP), but not intracellular Ca2+ concentration, thereby inducing protein kinase A (PKA) activity with subsequent phosphorylation of cAMP response element-binding protein (CREB) and upregulation of the CREB-responsive gene hairy and enhancer of split (HES)-1, a corepressor of peroxisome proliferator-activated receptor-γ (PPAR-γ) in hepatocytes. In (61)-carvone-stimulated cells, the repression of PPAR-γ reduced the expression of the target gene, mitochondrial glycerol-3-phosphate acyltransferase, which encodes a key enzyme involved in triglyceride synthesis. Intracellular triglyceride level and lipid accumulation were reduced in cells stimulated with (61)-carvone, effects that were diminished following the loss of OR1A1 function. These results indicate that OR1A1 may function as a non-redundant receptor in hepatocytes that regulates the PKA-CREB-HES-1 signaling axis and thereby modulates hepatic triglyceride metabolism.

Differentiation of adipocytes from preadipocytes contributes to adipose tissue expansion in obesity. Impaired adipogenesis may underlie the development of metabolic diseases such as insulin resistance and type 2 diabetes. Mechanistically, a well defined transcriptional network coordinates adipocyte differentiation. The family of paired-related homeobox transcription factors, which includes Prrx1a, Prrx1b, and Prrx2, is implicated with regulation of mesenchymal cell fate, including myogenesis and skeletogenesis; however, whether these proteins impact adipogenesis remains to be addressed. In this study, we identify Prrx1a and Prrx1b as negative regulators of adipogenesis. We show that Prrx1a and Prrx1b are down-regulated during adipogenesis in vitro and in vivo. Stable knockdown of Prrx1a/b enhances adipogenesis, with increased expression of peroxisome proliferator-activated receptor-gamma, CCAAT/enhancer-binding protein-alpha and FABP4 and increased secretion of the adipokines adiponectin and chemerin. Although stable low-level expression of Prrx1a, Prrx1b, or Prrx2 does not affect 3T3-L1 adipogenesis, transient overexpression of Prrx1a or Prrx1b inhibits peroxisome proliferator-activated receptor-gamma activity. Prrx1 knockdown decreases expression of Tgfb2 and Tgfb3, and inhibition of TGF beta signaling during adipogenesis mimics the effects of Prrx1 knockdown. These data support the hypothesis that endogenous Prrx1 restrains adipogenesis by regulating expression of TGF beta ligands and thereby activating TGF beta signaling. Finally, we find that expression of Prrx1a or Prrx1b in adipose tissue increases during obesity and strongly correlates with Tgfb3 expression in BL6 mice. These observations suggest that increased Prrx1 expression may promote TGF beta activity in adipose tissue and thereby contribute to aberrant adipocyte function during obesity.

Obesity, which is a key risk for the development of hyperglycemia, hypertension, hyperlipidemia and insulin resistance and is totally referred to as the metabolic disorders, has aroused people090005s great attention because of its alarming increase rate around the world. It is widely known that the occurrence of obesity can be attributed to both environmental and genetic factors. Peroxisome proliferators- activated receptor (PPAR), a member of ligand-dependent receptor, is one of the important genetic factors. PPAR includes three isoforms: PPAR-02±, PPAR- 0205 and PPAR- 0206, all of which are exerting critical influences on the maintenance of the metabolism of saccharides, lipids and proteins. PPAR-0206 is of great importance in the regulation of adipogenesis; in addition, it is essential in the prevention of adiposis and the treatment of 2-diabetes mellitus. In this review, we focus on giving a brief introduction about PPAR family, the indispensible function of PPAR-0206 in adipogenesis and the inseparable relationship between PPAR-0206 and obesity, deriving from the understanding of how these receptors activated will provide windows of opportunities for the treatment of obesity and associated metabolism syndromes.

In the course of an effort to identify the regulators for peroxisome proliferator-activated receptor y (PPARγ)-dependent perilipin gene expression, we found that tribbles homolog 3 (TRB3), containing a single kinase domain without enzymatic activity, downregulates PPAR

Epigenetic control of gene expression in children remains poorly understood, but new technologies can help elucidate the relationship between expression and DNA methylation. Here, we utilized the ...

Background Adipose tissues serve not only as a store for energy in the form of lipid, but also as endocrine tissues that regulates metabolic activities of the organism by secreting various kinds of hormones. Peroxisome proliferator activated receptor ?? (PPAR??) is a key regulator of adipocyte differentiation that induces the expression of adipocyte-specific genes in preadipocytes and mediates their differentiation into adipocytes. Furthermore, PPAR?? has an important role to maintain the physiological function of mature adipocyte by controlling expressions of various genes properly. Therefore, any reduction in amount and activity of PPAR?? is linked to the pathogenesis of metabolic syndrome. Results In this study, we investigated the contribution of epigenetic transcriptional regulatory mechanisms, such as DNA methylation, to the expression of the PPAR?? gene, and further evaluated the contribution of such epigenetic regulatory mechanisms to the pathogenesis of metabolic syndrome. In 3T3-L1 preadipocytes, the promoter of the PPAR??2 gene was hypermethylated, but was progressively demethylated upon induction of differentiation, which was accompanied by an increase of mRNA expression. Moreover, treatment of cells with 5'-aza-cytideine, an inhibitor of DNA methylation, increased expression of the PPAR?? gene in a dose-dependent manner. Methylation in vitro of a PPAR?? promoter-driven reporter construct also repressed the transcription of a downstream reporter gene. These results suggest that the expression of the PPAR?? gene is inhibited by methylation of its promoter. We next compared the methylation status of the PPAR?? promoters in adipocytes from wild-type (WT) mice with those from two diabetic mouse models: +Leprdb/+Leprdb and diet-induced obesity mice. Interestingly, we found increased methylation of the PPAR?? promoter in visceral adipose tissues (VAT) of the mouse models of diabetes, compared to that observed in wild-type mice. We observed a concomitant decrease in the level of PPAR?? mRNA in the diabetic mice compared to the WT mice. Conclusion We conclude that the expression of PPAR?? gene is regulated by DNA methylation of its promoter region and propose that reduced expression of PPAR?? owing to DNA methylation in adipocytes of the VAT may contribute to the pathogenesis of metabolic syndrome.

Prostate cancer is one of the most common non-cutaneous malignancies among men worldwide. Epigenetic aberrations, including changes in DNA methylation patterns and/or histone modifications, are key drivers of prostate carcinogenesis. These epigenetic defects might be due to deregulated function and/or expression of the epigenetic machinery, affecting the expression of several important genes. Remarkably, epigenetic modifications are reversible and numerous compounds that target the epigenetic enzymes and regulatory proteins were reported to be effective in cancer growth control. In fact, some of these drugs are already being tested in clinical trials. This review discusses the most important epigenetic alterations in prostate cancer, highlighting the role of epigenetic modulating compounds in pre-clinical and clinical trials as potential therapeutic agents for prostate cancer management.

Bladder cancer is one of the most common cancers worldwide, with a high rate of recurrence and poor outcomes as a result of relapse. Bladder cancer patients require lifelong invasive monitoring and treatment, making bladder cancer one of the most expensive malignancies. Lines of evidence increasingly point to distinct genetic and epigenetic alteration patterns in bladder cancer, even between the different stages and grades of disease. In addition, genetic and epigenetic alterations have been demonstrated to play important roles during bladder tumorigenesis. This review will focus on bladder cancer-associated genomic and epigenomic alterations, which are common in bladder cancer and provide potential diagnostic markers and therapeutic targets for bladder cancer treatment.

Abstract: Enhancers play a central role in cell-type-specific gene expression and are marked by H3K4me1/2. Active enhancers are further marked by H3K27ac. However, the methyltransferases responsible for H3K4me1/2 on enhancers remain elusive. Furthermore, how these enzymes function on enhancers to regulate cell-type-specific gene expression is unclear. Here we identify MLL4 (KMT2D) as a major mammalian H3K4 mono- and di-methyltransferase with partial functional redundancy with MLL3 (KMT2C). Using adipogenesis and myogenesis as model systems, we show that MLL4 exhibits cell-type- and differentiation-stage-specific genomic binding and is predominantly localized on enhancers. MLL4 co-localizes with lineage-determining transcription factors (TFs) on active enhancers during differentiation. Deletion of MLL4 markedly decreases H3K4me1/2, H3K27ac, Polymerase II and Mediator levels on enhancers and leads to severe defects in cell-type-specific gene expression and cell differentiation. Together, these findings identify MLL4 as a major mammalian H3K4 mono- and di-methyltransferase essential for enhancer activation during cell differentiation.

PPAR promotes adipogenesis while Wnt proteins inhibit adipogenesis. However, the mechanisms that control expression of these positive and negative master regulators of adipogenesis remain incompletely understood. By genome-wide histone methylation profiling in preadipocytes, we find that among gene loci encoding adipogenesis regulators, histone methyltransferase (HMT) G9a-mediated repressive epigenetic mark H3K9me2 is selectively enriched on the entire PPAR locus. H3K9me2 and G9a levels decrease during adipogenesis, which correlates inversely with induction of PPAR. Removal of H3K9me2 by G9a deletion enhances chromatin opening and binding of the early adipogenic transcription factor C/EBP尾 to PPAR promoter, which promotes PPAR expression. Interestingly, G9a represses PPAR expression in an HMT activity-dependent manner but facilitates Wnt10a expression independent of its enzymatic activity. Consistently, deletion of G9a or inhibiting G9a HMT activity promotes adipogenesis. Finally, deletion of G9a in mouse adipose tissues increases adipogenic gene expression and tissue weight. Thus, by inhibiting PPAR expression and facilitating Wnt10a expression, G9a represses adipogenesis.

Wnt/β-catenin signaling inhibits adipogenesis. Genome-wide profiling studies have revealed the enrichment of histone H3K27 methyltransferase Ezh2 on Wnt genes. However, the functional significance of such a direct link between the two types of developmental regulators in mammalian cells, and the role of Ezh2 in adipogenesis, remain unclear. Here we show Ezh2 and its H3K27 methyltransferase activity are required for adipogenesis. Ezh2 directly represses Wnt1, -6, -10a, and -10b genes in preadipocytes and during adipogenesis. Deletion of Ezh2 eliminates H3K27me3 on Wnt promoters and derepresses Wnt expression, which leads to activation of Wnt/β-catenin signaling and inhibition of adipogenesis. Ectopic expression of the wild-type (WT) Ezh2, but not the enzymatically inactive F667I mutant, prevents the loss of H3K27me3 and the defects in adipogenesis in Ezh2 -/- preadipocytes. The adipogenesis defects in Ezh2 -/- cells can be rescued by expression of adipogenic transcription factors PPARγ, C/EBPα, or inhibitors of Wnt/β-catenin signaling. Interestingly, Ezh2 -/- cells show marked increase of H3K27 acetylation globally as well as on Wnt promoters. These results indicate that H3K27 methyltransferase Ezh2 directly represses Wnt genes to facilitate adipogenesis and suggest that acetylation and trimethylation on H3K27 play opposing roles in regulating Wnt expression.

78 Chromatin state maps were generated throughout murine and human adipogenesis 78 Most active cis-regulatory elements differ between species 78 Many differences are due to evolutionary turnover of transcription factor motifs 78 Motif enrichment predicted that PLZF and SRF play a repressive role in adipogenesis

Abstract Bone marrow-derived multipotent stromal cells (BM-MSCs) display a broad range of therapeutically valuable properties, including the capacity to form skeletal tissues and dampen immune system responses. However, to use BM-MSCs in a clinical setting, amplification is required, which may introduce epigenetic changes that affect biological properties. Here we used chromatin immunoprecipitation to compare post-translationally modified histones at a subset of gene promoters associated with developmental and environmental plasticity in BM-MSCs from multiple donors following culture expansion. At many locations, we observed localization of both transcriptionally permissive (H3K4me3) and repressive (H3K27me3) histone modifications. These chromatin signatures were consistent among BM-MSCs from multiple donors. Since promoter activity depends on the relative levels of H3K4me3 and H3K27me3, we examined the ratio of H3K4me3 to H3K27me3 (K4/K27) at promoters during culture expansion. The H3K4me3 to H3K27me3 ratios were maintained at most assayed promoters over time. The exception was the adipose-tissue specific promoter for the PPAR-γ2 isoform of PPAR- γ, which is a critical positive regulator of adipogenesis. At PPAR-γ2 , we observed a change in K4/K27 levels favoring the repressed chromatin state during culture. This change correlated with diminished promoter activity in late passage cells exposed to adipogenic stimuli. In contrast to BM-MSCs and osteoblasts, lineage-restricted preadipocytes exhibited levels of H3K4me3 and H3K27me3 that favored the permissive chromatin state at PPAR- γ2. These results demonstrate that locus-specific changes in H3K4me3 and H3K27me3 levels can occur during BM-MSC culture that may affect their properties. S tem C ells 2015;33:2169–2181

Peroxisome proliferator-activated receptor gamma (PPARγ), a ligand-regulated nuclear hormone receptor, plays critical roles in metabolism and adipogenesis. PPARγ ligands such as thiazolidinediones (TZDs) exert insulin sensitizing and anti-inflammatory effects primarily through action on adipocytes, and are thus widely used to treat metabolic syndrome, especially type II diabetes. A number of PPARγ interacting partners have been identified, many of which are known epigenetic regulators, including enzymes for histone acetylation/deacetylation and histone methylation/demethylation. However, their functional roles in the PPARγ transcriptional pathway are not well defined. Recent advances in ChIP-based and deep sequencing technology are revealing previously underappreciated epigenomic mechanisms and therapeutic potentials of this nuclear receptor pathway.

The transcriptional mechanisms by which temporary exposure to developmental signals instigates adipocyte differentiation are unknown. During early adipogenesis, we find transient enrichment of the glucocorticoid receptor (GR), CCAAT/enhancer-binding protein beta (CEBPbeta), p300, mediator subunit 1, and histone H3 acetylation near genes involved in cell proliferation, development, and differentiation, including the gene encoding the master regulator of adipocyte differentiation, peroxisome proliferator-activated receptor gamma2 (PPARgamma2). Occupancy and enhancer function are triggered by adipogenic signals, and diminish upon their removal. GR, which is important for adipogenesis but need not be active in the mature adipocyte, functions transiently with other enhancer proteins to propagate a new program of gene expression that includes induction of PPARgamma2, thereby providing a memory of the earlier adipogenic signal. Thus, the conversion of preadipocyte to adipocyte involves the formation of an epigenomic transition state that is not observed in cells at the beginning or end of the differentiation process.

Worldwide obesity is a growing health problem, associated with increased risk of chronic disease. Understanding the molecular basis of adipogenesis and fat cell development in obesity is essential to identify new biomarkers and therapeutic targets for the development of anti-obesity drugs. microRNAs (miRNAs) appear to play regulatory roles in many biological processes associated with obesity, including adipocyte differentiation, insulin action and fat metabolism. Recent studies show miRNAs are dysregulated in obese adipose tissue. During adipogenesis miRNAs can accelerate or inhibit adipocyte differentiation and hence regulate fat cell development. In addition miRNAs may regulate adipogenic lineage commitment in multipotent stem cells and hence govern fat cell numbers. Recent findings suggest miR-519d may be associated with human obesity, but larger case-control studies are needed. Few miRNA targets have been experimentally validated in adipocytes but interestingly both miR-27 and miR-519d target PPAR family members, which are well established regulators of fat cell development. In this review recent advances in our understanding of the role of miRNAs in fat cell development and obesity are discussed. The potential of miRNA based therapeutics targeting obesity is highlighted as well as recommendations for future research which could lead to a breakthrough in the treatment of obesity. <br/> <br/> <br/>

The present study explored the involvement and role of miR-301a in the adipose tissues. For the first time we identified the expression of miR-301a in the white adipose tissues of mice. A decreased level of miR-301a was correlated to increased chronic inflammation in the 3T3-L1 cells and circula- tion in an obese mouse model. Mechanistically, we demonstrated that miR-301a attenuated saturated free fatty acid-induced activation of peroxisome proliferator-activated receptor gamma (PPARy) and production of proinflammatory cytokines in 3T3-L1 cells. Target gene reporter assays showed that miR-301a directly targeted the 3'-untranslated region (3'UTR) of PPAR纬, resulting in a decrease of PPARy protein expression. The miR-301a inhibition of adipocyte differentiation was reversed by PPAR纬 overexpression.

The present study explored the involvement of miR-302a in adipocyte differentiation via interaction with 3′-untranslated region of peroxisome proliferator-activated receptor gamma (PPARγ) mRNA. In differentiating 3T3-L1 adipocytes, expression of miR-302a was negatively correlated with that of the adipogenic gene aP2 and PPARγ. Overexpression of miR-302a inhibited adipogenic differentiation with lipid accumulation, and inversely anti-miR-302a increased the differentiation. In silico analysis revealed a complementary region of miR-302a seed sequence in 3′-UTR of PPARγ mRNA. Luciferase assay showed the direct interaction of miR-302a with PPARγ at the cellular level. The miR-302a inhibition of adipocyte differentiation was reversed by PPARγ overexpression. These findings suggest that miR-302a might be a negative regulator of adipocyte differentiation and that the dysregulation of miR-302a should lead to metabolic disorders.

Adipose tissue development is tightly regulated by altering gene expression. MicroRNAs are strong posttranscriptional regulators of mammalian differentiation. We hypothesized that microRNAs might influence human adipogenesis by targeting specific adipogenic factors. We identified microRNAs that showed varying abundance during the differentiation of human preadipocytes into adipocytes. Among them, miR-130 strongly affected adipocyte differentiation, as overexpressing miR-130 impaired adipogenesis and reducing miR-130 enhanced adipogenesis. A key effector of miR-130 actions was the protein peroxisome proliferator-activated receptor γ (PPARγ), a major regulator of adipogenesis. Interestingly, miR-130 potently repressed PPARγ expression by targeting both the PPARγ mRNA coding and 3′ untranslated regions. Adipose tissue from obese women contained significantly lower miR-130 and higher PPARγ mRNA levels than that from nonobese women. Our findings reveal that miR-130 reduces adipogenesis by repressing PPARγ biosynthesis and suggest that perturbations in this regulation is linked to human obesity.

We have shown previously that microvesicle (MV)-delivered miR-130b (miR-130b-MV) is able to target PPAR-γ and subsequently reduce the lipid accumulationin vitro. However, thein vivoeffect of miR-130b on fat deposition and glucose homeostasis remains unknown. Three-week-old C57BL/6 mice were fed a high-fat diet for 802weeks and then intravenously injected with MV-packaged scrambled control microRNA (miRNA) or miR-130b every other day for 1002days. Glucose tolerance test was performed and body weight, epididymal fat weight, as well as the expression of lipid metabolic genes were determined. We showed that mice fed on high-fat diet for 802weeks demonstrated significantly higher body weight, elevated blood glucose and impaired glucose tolerance. miR-130b-MV injection significantly reduced body weight and epididymal fat weight and partly restored glucose tolerance. miR-130b expression was significantly increased in the epididymal fat after miR-130b-MV injection while the protein content of its target gene PPAR-γ was significantly suppressed, together with a significant up-regulation of the lipolysis genes, hormone sensitive lipase, monoglyceride lipase and leptin. Moreover, miR-130b-MV injection increased the expression of miR-378a and miR-378-3p that are reported to participate in the regulation of fat deposition. Our results indicate that miR-130b-MV is able to reduce the epididymal fat deposition and partly restore glucose tolerance, through translational repression of PPAR-γ in a high-fat diet-induced obese mouse model.

We investigated the regulation and involvement of microRNAs (miRNAs) in fat cell development and obesity. Using miRNA microarrays, we profiled the expression of >370 miRNAs during adipogenesis of preadipocyte 3T3-L1 cells and adipocytes from leptin deficientob/oband diet-induced obese mice. Changes in key miRNAs were validated by RT-PCR. We further assessed the contribution of the chronic inflammatory environment in obese adipose tissue to the dysregulated miRNA expression by tumor necrosis factor (TNF)-伪 treatment of adipocytes. We functionally characterized two adipocyte-enriched miRNAs, miR-103 and miR-143, by a gain-of-function approach. Similar miRNAs were differentially regulated during in vitro and in vivo adipogenesis. Importantly, miRNAs that were induced during adipogenesis were downregulated in adipocytes from both types of obese mice and vice versa. These changes are likely associated with the chronic inflammatory environment, since they were mimicked by TNF-伪 treatment of differentiated adipocytes. Ectopic expression of miR-103 or miR-143 in preadipocytes accelerated adipogenesis, as measured both by the upregulation of many adipogenesis markers and by an increase in triglyceride accumulation at an early stage of adipogenesis. Our results provide the first experimental evidence for miR-103 function in adipose biology. The remarkable inverse regulatory pattern for many miRNAs during adipogenesis and obesity has important implications for understanding adipose tissue dysfunction in obese mice and humans and the link between chronic inflammation and obesity with insulin resistance.

Obesity is a risk factor for metabolic diseases, while preadipocyte differentiation or adipogenesis is closely related to obesity occurrence. Long noncoding RNAs (lncRNAs) are a unique class of transcripts in regulation of a variety of biological processes. Using cDNA microarray, we found lncRNA U90926 is negatively correlated with 3T3-L1 preadipocyte differentiation. The aim of this study was to explore the role of lncRNA U90926 (lnc-U90926) in adipogenesis and the underlying mechanisms. Quantitative real-time PCR (qPCR) was performed to determine lnc-U90926 expression in 3T3-L1 preadipocytes, differentiated adipocytes, and in adipose tissues form mice. RNA fluorescent in situ hybridization (FISH) was performed to determine the localization of lnc-U90926 in3T3-L1 preadipocytes. The effects of lnc-U90926 on 3T3-L1 adipogenesis were analyzed with lentivirus-mediated gain-and loss-of-function experiments. Lipid accumulation was evaluated by oil red O staining; several adipogenesis makers were analyzed by qPCR and Western Blotting. Dual luciferase assay was applied to explore the trans-activation of target genes modulated by lnc-U90926. All measurements were performed at least for three times. Lnc-U90926 expression decreased along the differentiation of 3T3-L1 preadipocytes. In mice, lnc-U90926 is predominantly expressed in adipose tissue. Obese mice have lower lnc-U90926 expression in subcutaneous and visceral adipose tissue than non-obese mice. FISH results showed that lnc-U90926 was mainly located in the cytoplasm. Overexpression lnc-U90926 attenuated 3T3-L1 adipocyte differentiation as evidenced by its ability to inhibit lipid accumulation, to decrease the mRNA levels of peroxisome proliferator-activated receptor gamma 2 (PPAR纬2), fatty acid binding protein 4 (FABP4) and adiponectin (AdipoQ) as well as to reduce the protein levels of PPAR纬 and FABP4 (P<0.05). Knockdown of lnc-U90926 showed opposite effects, which increased mRNA expression of PPAR纬2, FABP4, CCAAT/enhancer binding protein伪 (C/EBP伪) and AdipoQ. Lnc-U90926 attenuates 3T3-L1 adipocyte differentiation via inhibiting the transactivation of PPAR纬2 or PPAR纬.International Journal of Obesity accepted article preview online, 26 October 2016. doi:10.1038/ijo.2016.189.

Long non-coding (lnc) RNAs serve a multitude of functions in cells. NEAT1 RNA is a highly abundant 4 kb lncRNA in nuclei, and coincides with paraspeckles, nuclear domains that control sequestration of paraspeckle proteins. We examined NEAT1 RNA levels and its function in 3T3-L1 cells during differentiation to adipocytes. Levels of NEAT1 transcript, measured by RT-PCR, fluctuated in a temporal manner over the course of differentiation that suggested its role in alternative splicing of PPARγ mRNA, the major transcription factor driving adipogenesis. When cells were induced to differentiate by a media cocktail of insulin, dexamethasone, and isobutylmethyxanthine (IBMX) on Day 0, NEAT1 levels dropped on Day 4, when the PPARγ2 variant was spliced and when terminal differentiation occurs The appearance of PPARγ2 coordinates with the PPARγ1 variant to drive differentiation of adipocytes. SiRNA used to deplete NEAT1 resulted in the inability of cells to phosphorylate the serine/arginine-rich splicing protein, SRp40. SiRNA treatment for SRp40 resulted in dysregulation of PPARγ1 and, primarily, PPARγ2 mRNA levels. SRp40 associated with NEAT1, as shown by RNA-IP on days 0 and 8, but decreased on day 4, and concentrations increased over that of IgG control. Overexpression of SRp40 increased PPARγ2, but not γ1. Although lncRNA MALAT1 has been investigated in SR protein function, NEAT1 has not been shown to bind SR proteins for phosphorylation such that alternative splicing results. The ability of cells to increase phosphorylated SR proteins for PPARγ2 splicing suggests that fluxes in NEAT1 levels during adipogenesis regulate alternative splicing events.

ObjectivePeripheral lower body fat is associated with lower cardiometabolic risk. Physiological differences in gluteal compared with abdominal subcutaneous (sc) adipocyte functions are known but the molecular basis for depot differences in adipocyte function is poorly understood. Our goal is to identify novel gene regulatory pathways that underlie the heterogeneity of human fat distribution.MethodsAbdominal and gluteal adipose tissue aspirates obtained from 35 subjects (age = 30 卤 1.6 years; BMI = 27.3 卤 1.3 kg/m2) were analyzed using Illumina microarrays and confirmed by RT-PCR. The HOTAIR gene was stably transfected into primary cultured human abdominal sc preadipocytes using a lentivirus and effects on adipogenic differentiation were analyzed.ResultsA long noncoding RNA, HOTAIR that was expressed in gluteal but not in Abd sc adipose tissue was identified. This difference was retained throughout in vitro differentiation and was maximal at day 4. Ectopic expression of HOTAIR in abdominal preadipocytes produced an increase in differentiation as reflected by a higher percentage of differentiated cells, and increased expression of key adipogenic genes including PPAR纬 and LPL.ConclusionsHOTAIR is expressed in gluteal adipose and may regulate key processes in adipocyte differentiation. The role of this lncRNA in determining the metabolic properties of gluteal compared with abdominal adipocytes merits further study.

Cellular development requires reprogramming of the genome to modulate the gene program of the undifferentiated cell and allow expression of the gene program unique to differentiated cells. A number of key transcription factors involved in this reprogramming of preadipocytes to adipocytes have been identified; however, it is not until recently that we have begun to understand how these factors act at a genome-wide scale. In a recent publication we have mapped the genome-wide changes in chromatin structure during differentiation of 3T3-L1 preadipocytes and shown that a major reorganization of the chromatin landscape occurs within few hours following the addition of the adipogenic cocktail. In addition, we have mapped the genome-wide profiles of several of the early adipogenic transcription factors and shown that they act in a highly cooperative manner to drive this dramatic remodeling process.

Chronic heart failure (HF) is a leading clinical and public problem affected higher risk of morbidity and mortality in different population. HF appears to be in both phenotypic forms: HF with reduced left ventricular ejection fraction (HFrEF) and HF with preserved left ventricular ejection fraction (HFpEF). Although both HF phenotypes are distinguished in clinical features, co-morbidity status, prediction score, and treatment, the clinical outcomes in patients with HFrEF and HFpEF are similar. In this context investigation of various molecular and cellular mechanisms leading to development and progression of both HF phenotypes are very important. There is emerging evidence regarding that the epigenetic regulation may have a clue in the pathogenesis of HF. The review is represented current available evidence regarding an implication of epigenetic modifications in development of different HF phenotypes and perspectives of epigenetic-based therapies of HF.