Treatment Of Preadipocytes Research Articles

Methionine sulfoxide suppresses adipogenic differentiation by regulating the mitogen-activated protein kinase signaling pathway.

In this study, methionine sulfoxide (MetO) was identified as an active metabolite that suppresses adipogenesis after screening obese individuals versus the normal population. MetO suppressed the gene and protein expression of CCAAT/enhancer binding protein (C/EBP) α, adipocyte fatty acid binding protein 4 (FABP4), and the nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) during human preadipocyte (HPA) differentiation. Adipogenesis decreased following MetO treatment; however, the preadipocyte number, proliferation, and apoptosis were unaffected. The activity of phosphorylated extracellular signal-related kinase (P-ERK) of the mitogen-activated protein kinase (MAPK) pathway was significantly inhibited in HPA after MetO treatment. Furthermore, treatment of preadipocytes with the selective P-ERK1/2 agonist Ro 67-7476 abolished the effect of MetO against adipogenesis suggesting that MetO function is dependent on the MAPK pathway. The mechanistic insights of adipogenesis suppression by MetO presented in this study shows its potential as an antiobesity drug.

Treatment of preadipocytes with fish oil, mixed oil, or soybean oil-based lipid emulsions have differential effects on the regulation of lipogenic and lipolytic genes in mature 3T3-L1 adipocytes

The key adipose tissue characteristics are established during early development, where lipids play an essential role. Lipid emulsions used in total parenteral nutrition have different omega-(n) 6 to n-3 fatty acid ratios. A lower n-6:n-3 fatty acid decreases lipid accumulation; however, the effects of lipid emulsions with different n-6 to n-3 fatty acid ratios on the programming of preadipocytes to affect lipid accumulation in mature adipocytes is not known. This study compared the effects of Fish oil (FO), Mixed oil (MO), and Soybean oil (SO) based lipid emulsion on genes involved in adipogenesis, lipogenesis, lipolysis, and β-oxidation in 3T3-L1 adipocytes. Preadipocytes were treated with specific lipid emulsions and then differentiated to mature adipocytes in the absence of lipid emulsions. In a separate experiment, mature 3T3-L1 adipocytes were treated with lipid emulsions to investigate the effects on genes involved in lipolysis. Fatty acid composition, triacylglycerol levels, and the mRNA expression of genes involved in adipogenesis, lipogenesis, lipolysis, and β-oxidation were measured. Preadipocytes and mature adipocytes treated with FO showed higher incorporation of n-3 polyunsaturated fatty acids, lower triacylglycerol levels, and decreased mRNA expression of adipogenic and lipogenic genes, followed by MO and SO. FO and MO increased the mRNA expression of carnitine palmitoyltransferase-1, while FO decreased the mRNA expression of lipolytic genes compared to untreated cells. Our findings suggest that FO programs preadipocytes to prevent adipose tissue dysfunction in mature adipocytes; the effects of FO-based lipid emulsion were followed by MO and SO.

Article of Significant Interest in This Issue

The transcription factors C/EBPβ and C/EBPδ are transcriptionally induced soon after the treatment of preadipocytes with an adipogenic cocktail. In turn C/EBPβ and C/EBPδ promote the expression of the master adipocyte transcription factors peroxisome proliferator-activated receptor gamma and C/EBPα. However, the factors responsible for induction of C/EBPβ and C/EBPδ have remained incompletely understood. Lee et al. (e00209-20) show that the MLL3/MLL4/PTIP-associated protein PAGR1 cooperates with CREB and the glucocorticoid receptor to regulate the induction of C/EBPβ and C/EBPδ during the early phase of adipogenesis. This study presents an updated model in which components of the MLL3/MLL4 complex promote adipocyte differentiation by regulating the expression of distinct sets of transcription factors.

Estradiol signaling interacts with Sirt1 in the regulation of autophagy and adipogenesis

Adiposity is regulated by estradiol signaling and the nutrient sensor Sirt1. It has been recognized that autophagy plays a key role in adipogenesis and metabolism. However, it remains largely unknown whether and how estradiol signaling interacts with Sirt1 in the regulation of autophagy. Here we showed that activation and overexpression of Sirt1 suppressed autophagy in 3T3L1 cells, which was associated with suppressed adipocyte differentiation. Treatment of preadipocytes with estradiol induced Sirt1 and dampened autophagy, thereby suppressing adipogenesis. Consistently, knockdown of Sirt1 induced autophagy and adipocyte differentiation, which was recapitulated by estrogen receptor inhibition or knockout. In addition, knockdown of Sirt1 abolished the effects of estradiol on autophagy and adipogenesis. The modulation of autophagy and adipogenesis by estradiol and Sirt1 was ascribed at least in part to mTOR‐ULK1 cascade. Protein analysis did not show significant changes in the acetylation levels of autophagy regulators (e.g., Atg5, Atg7, or LC3) during Sirt1 modulation. Taken together, our data reveals an estradiol‐Sirt1‐autophagy axis that regulates adipogenesis. Future study will be of interest and significance to determine the potential role of the estradiol‐Sirt1‐autophagy axis in sex‐dependent fat distribution.Support or Funding InformationAmerican Heart Association Grant 18TPA34230082 (Z.C.) and USDA National Institute of Food and Agriculture Hatch Project 1020373 (Z.C.).

PCB126 blocks the thermogenic beiging response of adipocytes.

Subcutaneous white adipose tissue is capable of becoming thermogenic in a process that is referred to as "beiging." Beiging is associated with activation of the uncoupling protein, UCP1, and is known to be important for preventing adipose hypertrophy and development of insulin resistance. Polychlorinated biphenyls (PCBs) accumulate in fat, and it is hypothesized that disruption of adipogenesis and adipocyte function by PCBs may be causative in the development of obesity and diabetes. We developed immortal human subcutaneous preadipocytes that, when differentiated, are capable of beiging. Preadipocytes that were treated with polychlorinated biphenyl congener 126 (PCB126), followed by differentiation, were suppressed for their ability to activate UCP1 upon β-adrenergic stimulation with norepinephrine (NE), demonstrating a block in the beiging response. Treatment of preadipocytes with another known endogenous AhR agonist, indoxyl sulfate (IS), followed by differentiation also blocked the NE-stimulated upregulation of UCP1. Knockdown of the aryl hydrocarbon receptor (AhR) caused the preadipocytes to be refractory to PCB126 and IS effects. The chemical AhR antagonist, CH223191, was effective at preventing the effects of PCB126 but not IS, indicating AhR ligand specificity of CH223191. Repression of NE-induced UCP1 upregulation was also observed when already-differentiated mature adipocytes were treated with PCB126 but not IS. These results indicate that exposure of preadipocytes to endogenous (IS) or exogenous (PCB126) AhR agonists is effective at blocking them from becoming functional adipocytes that are capable of the beiging response. Mature adipocytes may have differential responses. This finding suggests a mechanism by which dioxin-like PCBs such as PCB126 could lead to disruption in energy homeostasis, potentially leading to obesity and diabetes.

A novel peptide suppresses adipogenic differentiation through activation of the AMPK pathway

Obesity rates have risen rapidly over the past several decades and obesity is now a global public health challenge. The reduction of excessive adipogenesis is thought to be an effective intervention for obesity and obesity-related metabolic diseases such as type 2 diabetes. In this study, a novel peptide PDBSN was identified that functions to suppress adipogenesis. In both human preadipocytes and mouse adipose-derived stem cells (ADSCs), PDBSN exhibited a suppressive effect on the accumulation of lipids and the expression of genes as well as their corresponding proteins (CCAAT/enhancer binding protein (C/EBP)β, C/EBPα and nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ)) relevant to adipogenic cell differentiation. Although adipogenesis decreased, the preadipocyte number and proliferation were not influenced by the PDBSN treatment. Apoptosis and the cell cycle were also determined to not have a role in the action of PDBSN. Mechanistically, the activity of the AMPK (adenosine 5ʹ-monophosphate-activated protein kinase) pathway was markedly increased upon PDBSN treatment. Moreover, treatment of preadipocytes with compound C, a selective AMPK inhibitor, abolished the effect of PDBSN in anti-adipogenesis, suggesting that the function of PDBSN relied on the AMPK pathway. These results suggest an effective role for PDBSN in suppressing adipogenesis and show potential for anti-obesity drug discovery.

Expression analysis of a cholecystokinin system in human and rat white adipose tissue

AimCholecystokinin (CCK) participates in the storage of dietary triglycerides in white adipose tissue (WAT). Our goal was to characterize, both in subcutaneous (Sc-WAT) and visceral WAT (Vis-WAT), the functional expression of the two known CCK receptors, CCK-1 (CCK-1R) and CCK-2 (CCK-2R), as well as of CCK. Main methodsGene and protein expression was assessed in different cell types of rat and human WAT by means of RT-PCR and western-blot, respectively. The functionality of CCK-Rs was tested by quantifying protein kinase B (Akt) phosphorylation after treatment of pre-adipocytes with the bioactive fragment of CCK, CCK-8. The CCK receptor subtype involved in Akt phosphorylation was investigated by using selective CCK-1R (SR-27,897) and CCK-2R antagonists (L-365,260). Key findingsIn rats, CCK-1R (Cckar) and CCK-2R (Cckbr) gene expression was detected in the two types of WAT analyzed as well as in isolated adipocytes, mesenchymal stem cells and pre-adipocytes. CCK-1R and CCK-2R proteins were identified in adipocytes and, to a minor extent, in pre-adipocytes. In addition, CCK-2R were detected in subcutaneous mesenchymal stem cells. Gene expression of the CCK precursor preproCCK as well as CCK immunoreactivity were also found in Sc-WAT and Vis-WAT. In human WAT, CCK gene expression as well as CCK-2Rs and CCK were also identified. CCK-8 evoked Akt phosphorylation in rat pre-adipocytes, and this effect was antagonized by SR-27,897 and L-365,260. SignificanceOur data show that both human and rat WAT express a complete CCK system, and suggest that CCK may have an autocrine/paracrine role in regulating adipose tissue biology.

A delayed proinflammatory response of human preadipocytes to PCB126 is dependent on the aryl hydrocarbon receptor.

Inflammation in adipose tissue is recognized as a causative factor in the development of type II diabetes. Adipocyte hypertrophy as well as bacterial and environmental factors have been implicated in causing inflammation in mature adipocytes. Exposure to persistent organic pollutants such as polychlorinated biphenyls (PCBs) has been associated with the development of type II diabetes. We show here that PCB126, a dioxin-like PCB, activates a robust proinflammatory state in fat cell precursors (preadipocytes). The response was found to be dependent on aryl hydrocarbon receptor (AhR) activation, although induction of the response was delayed compared to upregulation of CYP1A1, a classic AhR-responsive gene. Treatment of preadipocytes with a nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) inhibitor partially attenuated the PCB126-induced inflammatory response and partly, but not completely, ameliorated disruption of adipogenesis caused by PCB126. Our results indicate a role for PCB126 in mediating an inflammatory response through AhR in preadipocytes that interferes with adipogenesis.

Combined metformin and insulin treatment reverses metabolically impaired omental adipogenesis and accumulation of 4-hydroxynonenal in obese diabetic patients

ObjectiveObesity-associated impaired fat accumulation in the visceral adipose tissue can lead to ectopic fat deposition and increased risk of insulin resistance and type 2 diabetes mellitus (T2DM). This study investigated whether impaired adipogenesis of omental (OM) adipose tissues and elevated 4-hydroxynonenal (4-HNE) accumulation contribute to this process, and if combined metformin and insulin treatment in T2DM patients could rescue this phenotype. MethodsOM adipose tissues were obtained from forty clinically well characterized obese individuals during weight reduction surgery. Levels of 4-HNE protein adducts, adipocyte size and number of macrophages were determined within these tissues by immunohistochemistry. Adipogenic capacity and gene expression profiles were assessed in preadipocytes derived from these tissues in relation to insulin resistance and in response to 4-HNE, metformin or combined metformin and insulin treatment. ResultsPreadipocytes isolated from insulin resistant (IR) and T2DM individuals exhibited lower adipogenesis, marked by upregulation of anti-adipogenic genes, compared to preadipocytes derived from insulin sensitive (IS) individuals. Impaired adipogenesis was also associated with increased 4-HNE levels, smaller adipocytes and greater macrophage presence in the adipose tissues. Within the T2DM group, preadipocytes from combined metformin and insulin treated subset showed better in vitro adipogenesis compared to metformin alone, which was associated with less presence of macrophages and 4-HNE in the adipose tissues. Treatment of preadipocytes in vitro with 4-HNE reduced their adipogenesis and increased proliferation, even in the presence of metformin, which was partially rescued by the presence of insulin. ConclusionThis study reveals involvement of 4-HNE in the impaired OM adipogenesis-associated with insulin resistance and T2DM and provides a proof of concept that this impairment can be reversed by the synergistic action of insulin and metformin. Further studies are needed to evaluate involvement of 4-HNE in metabolically impaired abdominal adipogenesis and to confirm benefits of combined metformin-insulin therapy in T2DM patients.

Progesterone dose-dependently modulates hepatocyte growth factor production in 3T3-L1 mouse preadipocytes.

It is well documented that estrogen is predominant inducer of hepatocyte growth factor (HGF) in a variety of cell types. However, the effect of progesterone (P) remains to be elusive. Thus, in the present study, we examined the effect of P and combined effect of P and 17β-estradiol (E2) on HGF expression and production in 3T3-L1 fibroblastic preadipocytes and mature adipocytes, as a model of stromal cells. Northern blot analysis showed that hgf mRNA expressed in preadipocytes was notably higher than that of mature adipocytes, and increased by treatment of preadipocytes with E2 or 10 nM P, but not with 1,000 nM P. The E2-induced hgf mRNA expression was enhanced by 10 nM P, but suppressed by 1,000 nM P. Western blot analysis revealed that biological active forms of HGF protein was found in the preadipocyte culture medium, while the lesser amount of HGF precursor protein was detected in the mature adipocyte culture medium. The amounts of HGF were changed dependently on the hgf mRNA expression levels. These results indicate that HGF production is intricately regulated by E2 and P at the transcriptional levels in 3T3-L1 cells, and may explain the changes in the HGF production during the mammary gland development, especially decrease in HGF expression during pregnancy when P concentration is high.

Skipped BSCL2 Transcript in Celia's Encephalopathy (PELD): New Insights on Fatty Acids Involvement, Senescence and Adipogenesis.

ObjectivePELD (Progressive Encephalopathy with or without Lipodystrophy or Celia’s Encephalopathy) is a fatal and rare neurodegenerative syndrome associated with the BSCL2 mutation c.985C>T, that results in an aberrant transcript without the exon 7 (Celia seipin). The aim of this study was to evaluate both the process of cellular senescence and the effect of unsaturated fatty acids on preadipocytes from a homozygous c.985C>T patient. Also, the role of aberrant seipin isoform on adipogenesis was studied in adipose-derived human mesenchymal stem cells.Material and methodsCellular senescence was evaluated using β-galactosidase staining of preadipocytes obtained from a homozygous c.985C>T patient. Moreover, these cells were cultured during 24 hours with Intralipid, a soybean oil-based commercial lipid emulsion. The expression of the different BSCL2 transcripts was measured by qPCR. Adipose-derived human mesenchymal stem cells were differentiated to a fat lineage using StemPRO adipogenesis kit, and the expression of BSCL2 transcripts and several adipogenesis-related genes was measured by qPCR.Resultsthe treatment of preadipocytes with unsaturated fatty acids significantly reduced the expression of the BSCL2 transcript without exon 7 by 34 to 63%. On the other hand, at least in preadipocytes, this mutation does not disturb cellular senescence rate. Finally, during adipocyte differentiation of adipose-derived human mesenchymal stem cells, the expression of adipogenic genes (PPARG, LPIN1, and LPL) increased significantly over 14 days, and noteworthy is that the BSCL2 transcript without exon 7 was differentially expressed by 332 to 723% when compared to day 0, suggesting an underlying role in adipogenesis.Conclusionsour results suggest that Celia seipin is probably playing an underestimated role in adipocyte maturation, but not in senescence, and its expression can be modified by exogenous factors as fatty acids.

Bisphenol S Induces Adipogenesis in Primary Human Preadipocytes From Female Donors.

Human exposure to bisphenol A has been associated with negative health outcomes in humans and its use is now regulated in a number of countries. Bisphenol S (BPS) is increasingly used as a replacement for bisphenol A; however, its effects on cellular metabolism and potential role as an endocrine disruptor have not been fully characterized. In the current study, we evaluated the effect of BPS on adipogenesis in primary human preadipocytes. The effect of BPS on the differentiation of human preadipocytes was determined after treatment with BPS at concentrations ranging from 0.1 nM to 25 μM by quantifying lipid accumulation and mRNA and protein levels of key adipogenic markers. Treatment of preadipocytes with 25 μM BPS induced lipid accumulation and increased the mRNA and protein levels of several adipogenic markers including lipoprotein lipase and adipocyte protein 2 (aP2). Cotreatment of cells with the estrogen receptor antagonist ICI-182,780 significantly inhibited BPS-induced lipid accumulation and affected aP2 but not lipoprotein lipase protein levels. Cotreatment of cells with the glucocorticoid receptor antagonist RU486 had no effect on BPS-induced lipid accumulation or protein levels. Furthermore, reporter gene assays using a synthetic promoter containing peroxisome proliferator-activated receptor-γ (PPARG)-response elements and a PPARG-responsive human aP2 promoter region showed that BPS was able to activate PPARG. To our knowledge, this study is the first to show that BPS induces lipid accumulation and differentiation of primary human preadipocytes, and this effect may be mediated through a PPARG pathway.

Identification of carboxypeptidase X (CPX)-1 as a positive regulator of adipogenesis.

Adipose tissue expansion occurs through a combination of hypertrophy of existing adipocytes and generation of new adipocytes via the process of hyperplasia, which involves the proliferation and subsequent differentiation of preadipocytes. Deficiencies in hyperplasia contribute to adipose tissue dysfunction and the association of obesity with chronic cardiometabolic diseases. Thus, increased understanding of hyperplastic pathways may be expected to afford novel therapeutic strategies. We have reported that fibroblast growth factor (FGF)-1 promotes proliferation and differentiation of human preadipocytes and recently demonstrated that bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) is a central, proximal effector. Herein, we describe the identification and characterization of carboxypeptidase X (CPX)-1, a secreted collagen-binding glycoprotein, as a novel downstream effector in human primary and Simpson-Golabi-Behmel syndrome preadipocytes. CPX-1 expression increased after treatment of preadipocytes with FGF-1, BAMBI knockdown, or induction of differentiation. CPX-1 knockdown compromised preadipocyte differentiation coincident with reduced collagen expression. Furthermore, preadipocytes differentiated on matrix derived from CPX-1 knockdown cells exhibited reduced Glut4 expression and insulin-stimulated glucose uptake. Finally, CPX-1 expression was increased in adipose tissue from obese mice and humans. Collectively, these findings establish CPX-1 as a positive regulator of adipogenesis situated downstream of FGF-1/BAMBI that may contribute to hyperplastic adipose tissue expansion via affecting extracellular matrix remodeling.-Kim, Y.-H., Barclay, J. L., He, J., Luo, X., O'Neill, H. M., Keshvari, S., Webster, J. A., Ng, C., Hutley, L. J., Prins, J. B., Whitehead, J. P. Identification of carboxypeptidase X (CPX)-1 as a positive regulator of adipogenesis.

The Gustatory Signaling Pathway and Bitter Taste Receptors Affect the Development of Obesity and Adipocyte Metabolism in Mice.

Intestinal chemosensory signaling pathways involving the gustatory G-protein, gustducin, and bitter taste receptors (TAS2R) have been implicated in gut hormone release. Alterations in gut hormone profiles may contribute to the success of bariatric surgery. This study investigated the involvement of the gustatory signaling pathway in the development of diet-induced obesity and the therapeutic potential of targeting TAS2Rs to induce body weight loss. α-gustducin-deficient (α-gust-/-) mice became less obese than wild type (WT) mice when fed a high-fat diet (HFD). White adipose tissue (WAT) mass was lower in α-gust-/- mice due to increased heat production as a result of increases in brown adipose tissue (BAT) thermogenic activity, involving increased protein expression of uncoupling protein 1. Intra-gastric treatment of obese WT and α-gust-/- mice with the bitter agonists denatonium benzoate (DB) or quinine (Q) during 4 weeks resulted in an α-gustducin-dependent decrease in body weight gain associated with a decrease in food intake (DB), but not involving major changes in gut peptide release. Both WAT and 3T3-F442A pre-adipocytes express TAS2Rs. Treatment of pre-adipocytes with DB or Q decreased differentiation into mature adipocytes. In conclusion, interfering with the gustatory signaling pathway protects against the development of HFD-induced obesity presumably through promoting BAT activity. Intra-gastric bitter treatment inhibits weight gain, possibly by directly affecting adipocyte metabolism.

MFAP5is related to obesity-associated adipose tissue and extracellular matrix remodeling and inflammation

Low-grade inflammation is involved in adipose tissue (AT) and extracellular matrix (ECM) remodeling and induces deposition of ECM proteins in AT. We have previously shown that MFAP5 (microfibrillar-associated protein 5) expression decreases in AT after weight loss. The aim of this study was to investigate MFAP5 localization in human AT and gene expression in adipocytes and the role of MFAP5 in adipocyte metabolism and inflammation. MFAP5 protein localization and gene expression were studied with immunohistochemistry and quantitative reverse transcriptase PCR (RT-qPCR) in human subcutaneous AT and cultured Simpson-Golabi-Behmel syndrome (SGBS) adipocytes, respectively. The effect of MFAP5 knock-down by siRNA on gene expression and insulin action was examined with RT-qPCR, western blot, and insulin-stimulated glucose uptake. The effect of different cytokines on MFAP5 gene and protein expression was investigated in cultured human SGBS preadipocytes. MFAP5 protein was highly expressed in AT, and gene expression decreased during adipocyte differentiation in SGBS cells. Treatment of preadipocytes with TNFα and TGFβ1 increased MFAP5 gene and protein expression. Furthermore, MFAP5 knock-down decreased the expression of genes involved in inflammation. Our results demonstrate that factors involving low-grade inflammation modulate MFAP5 expression and that the modified expression of MFAP5 may further regulate AT inflammation.

Ethnic differences in pre-adipocyte intracellular lipid accumulation and alkaline phosphatase activity

Alkaline phosphatase (ALP) increases lipid accumulation in human pre-adipocytes. This study was performed to assess whether ethnic differences in the prevalence of obesity in African and European females are related to differences in pre-adipocyte lipid accretion and ALP activity. Pre-adipocytes were isolated from 13 black and 14 white females. Adipogenesis was quantified using the lipid dye, Oil red O, whilst ALP activity was assayed in cell extracts on day zero and 12days after initiating adipogenesis. Lipid levels (OD units/mg protein) were lower in pre-adipocytes from white than black females on day 0 (0.36±0.05 versus 0.44±0.03, respectively; p<0.0005) and day 12 (1.18±0.14 versus 1.80±0.22, respectively; p<0.0005), as was ALP activity (mU/mg protein) on day zero (36.5±5.8 versus 136.4±10.9, respectively; p<0.0005) and day 12 (127±16 versus 278±27, respectively; p<0.0005). Treatment of pre-adipocytes with histidine, an ALP inhibitor, blocked lipid accumulation. Thus, lipid uptake is higher in pre-adipocytes isolated from black compared to white females which parallels the obesity prevalence rates in these population groups. The reason for higher fat accumulation in pre-adipocytes isolated from black females may be related to higher ALP activity.

Epigenetic Modifications and Canonical Wingless/int-1 Class (WNT) Signaling Enable Trans-differentiation of Nonosteogenic Cells into Osteoblasts

Mesenchymal cells alter and retain their phenotype during skeletal development through activation or suppression of signaling pathways. For example, we have shown that Wnt3a only stimulates osteoblast differentiation in cells with intrinsic osteogenic potential (e.g. MC3T3-E1 pre-osteoblasts) and not in fat cell precursors or fibroblasts (3T3-L1 pre-adipocytes or NIH3T3 fibroblasts, respectively). Wnt3a promotes osteogenesis in part by stimulating autocrine production of the osteoinductive ligand Bmp2. Here, we show that the promoter regions of the genes for Bmp2 and the osteoblast marker Alp are epigenetically locked to prevent their expression in nonosteogenic cells. Both genes have conserved CpG islands that exhibit increased CpG methylation, as well as decreased acetylation and increased methylation of histone H3 lysine 9 (H3-K9) specifically in nonosteogenic cells. Treatment of pre-adipocytes or fibroblasts with the CpG-demethylating agent 5'-aza-2'-deoxycytidine or the histone deacetylase inhibitor trichostatin-A renders Bmp2 and Alp responsive to Wnt3a. Hence, drug-induced epigenetic activation of Bmp2 gene expression contributes to Wnt3a-mediated direct trans-differentiation of pre-adipocytes or fibroblasts into osteoblasts. We propose that direct conversion of nonosteogenic cells into osteoblastic cell types without inducing pluripotency may improve prospects for novel epigenetic therapies to treat skeletal afflictions.

Bisphenol A induces differentiation of human preadipocytes in the absence of glucocorticoid and is inhibited by an estrogen-receptor antagonist

Background:Obesity is a major health concern in the developed world, and increasing evidence suggests that exposures to common environmental substances may enhance the risk for the development of this disease.Objectives:The current study examines the effect of the ubiquitous plastic monomer bisphenol A (BPA) on the differentiation of primary human preadipocytes in vitro and the role of the estrogen and glucocorticoid receptors.Methods:In this study, the mechanism of BPA-induced adipogenesis in preadipocytes from donors with healthy body mass index in the absence of exogenous glucocorticoid was evaluated. The effects of estradiol, the estrogen-receptor (ER) antagonist ICI and the glucocorticoid receptor (GR) antagonist RU486 on BPA-induced adipogenesis were examined. The expression levels of key adipogenic factors were assessed.Results:Treatment of preadipocytes with 1–50 μM BPA induced a dose-dependent increase in differentiation and lipid accumulation as determined by lipid staining and triacylglyceride quantification. BPA also induced expression of the adipogenic markers aP2, adipsin, peroxisome proliferator-activated receptor γ and the CCAAT-enhancer-binding proteins α and β. Co-treatment of cells with ICI inhibited the BPA-induced increase in aP2 levels, while treatment with ICI or estradiol alone had no effect. Treatment of cells with the GR antagonist RU486 had no effect on BPA-induced differentiation as evaluated by aP2 levels.Conclusions:This study is one of the first to show that BPA induces human adipocyte differentiation in the absence of exogenous glucocorticoid through a non-classical ER pathway rather than through GR activation. These studies add to the growing evidence that endocrine-disrupting chemicals such as BPA have the potential to modulate adipogenesis and impact human biology.

Role of sphingosine kinases and sphingosine 1-phosphate in mediating adipogenesis

Recent Background: Development of obesity involves promotion of preadipocyte differrentiation. This study investigated the role that sphingosine kinases (SPHK) and ceramide-derived sphingosine 1-phosphate (S1P) play in adipocyte terminal differentiation. Materials and Methods: The mouse 3T3-L1 cell line was used as a model for adipogenesis. Cells were harvested at specific time points after initation of differentiation, and SPHK activity was measured. 3T3-L1 cells were treated with S1P and expression of early adipogenesis transcription markers was measured by real time PCR. The expression of S1P-receptors (S1PRs) during differentiation was measured. Results: SPHK activity is induced when 3T3-L1 cells are treated with insulin, dexamethasone, and isobutylmethylxanthine to induce differentiation. SPHK1 is active in preadipocytes and early in the differentiation process. Both SPHK1 and SPHK2 isozymes contribute to activity in differentiated adipocytes. Inhibition of SPHK1 attenuates adipocyte differentiation; however, extracellular S1P does not rescue the effect of SPHK1 inhibition on adipogenesis. Although treatment of preadipocytes with S1P induced message expression of the early adipogenesis transcription factor CC AAT/ binding proteinalpha, continued treatment did not fully support the development of differentiated adipocytes. Sphingosine 1-phosphate receptors (S1PRs) are expressed in preadipocytes and message expression declines markedly during adipocyte differentiation. Conclusion: These results demonstrate that the contribution of SPHK and S1P to adipogenesis is mediated primarily through biphasic activation of SPHK1 and 2 with extracellular S1P and S1PRs playing little role during preadipocyte differentiation.

Tyk2 and Stat3 Regulate Brown Adipose Tissue Differentiation and Obesity

Mice lacking the Jak tyrosine kinase member Tyk2 become progressively obese due to aberrant development of Myf5+ brown adipose tissue (BAT). Tyk2 RNA levels in BAT and skeletal muscle, which shares a common progenitor with BAT, are dramatically decreased in mice placed on a high-fat diet and in obese humans. Expression of Tyk2 or the constitutively active form of the transcription factor Stat3 (CAStat3) restores differentiation in Tyk2(-/-) brown preadipocytes. Furthermore, Tyk2(-/-) mice expressing CAStat3 transgene in BAT also show improved BAT development, normal levels of insulin, and significantly lower body weights. Stat3 binds to PRDM16, a master regulator of BAT differentiation, and enhances the stability of PRDM16 protein. These results define Tyk2 and Stat3 as critical determinants of brown fat lineage and suggest that altered levels of Tyk2 are associated with obesity in both rodents and humans.