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Sample records for induces adipocyte differentiation

  1. Depletion of white adipocyte progenitors induces beige adipocyte differentiation and suppresses obesity development.

    PubMed

    Daquinag, A C; Tseng, C; Salameh, A; Zhang, Y; Amaya-Manzanares, F; Dadbin, A; Florez, F; Xu, Y; Tong, Q; Kolonin, M G

    2015-02-01

    Overgrowth of white adipose tissue (WAT) in obesity occurs as a result of adipocyte hypertrophy and hyperplasia. Expansion and renewal of adipocytes relies on proliferation and differentiation of white adipocyte progenitors (WAP); however, the requirement of WAP for obesity development has not been proven. Here, we investigate whether depletion of WAP can be used to prevent WAT expansion. We test this approach by using a hunter-killer peptide designed to induce apoptosis selectively in WAP. We show that targeted WAP cytoablation results in a long-term WAT growth suppression despite increased caloric intake in a mouse diet-induced obesity model. Our data indicate that WAP depletion results in a compensatory population of adipose tissue with beige adipocytes. Consistent with reported thermogenic capacity of beige adipose tissue, WAP-depleted mice display increased energy expenditure. We conclude that targeting of white adipocyte progenitors could be developed as a strategy to sustained modulation of WAT metabolic activity. PMID:25342467

  2. Depletion of white adipocyte progenitors induces beige adipocyte differentiation and suppresses obesity development

    PubMed Central

    Daquinag, A C; Tseng, C; Salameh, A; Zhang, Y; Amaya-Manzanares, F; Dadbin, A; Florez, F; Xu, Y; Tong, Q; Kolonin, M G

    2015-01-01

    Overgrowth of white adipose tissue (WAT) in obesity occurs as a result of adipocyte hypertrophy and hyperplasia. Expansion and renewal of adipocytes relies on proliferation and differentiation of white adipocyte progenitors (WAP); however, the requirement of WAP for obesity development has not been proven. Here, we investigate whether depletion of WAP can be used to prevent WAT expansion. We test this approach by using a hunter-killer peptide designed to induce apoptosis selectively in WAP. We show that targeted WAP cytoablation results in a long-term WAT growth suppression despite increased caloric intake in a mouse diet-induced obesity model. Our data indicate that WAP depletion results in a compensatory population of adipose tissue with beige adipocytes. Consistent with reported thermogenic capacity of beige adipose tissue, WAP-depleted mice display increased energy expenditure. We conclude that targeting of white adipocyte progenitors could be developed as a strategy to sustained modulation of WAT metabolic activity. PMID:25342467

  3. Biochemical changes induced by strontium ranelate in differentiating adipocytes.

    PubMed

    Vidal, Christopher; Gunaratnam, Krishanthi; Tong, Jessica; Duque, Gustavo

    2013-04-01

    Low bone formation in osteoporosis is associated with a shift from osteoblastic to adipogenic differentiation of mesenchymal stem cells (MSC) inducing a concomitant lipotoxic milieu within the bone marrow. Strontium ranelate (SrRN), a treatment for osteoporosis, has both anti-resorptive and anabolic effects on bone. The anabolic effect of SrRN has been associated with its effect on both osteoblastogenesis and adipogenesis. However, the effect of SrRN on the potentially lipotoxic factors produced by differentiating marrow adipocytes remains poorly understood. To expand the knowledge on the effect of SrRN treatment on the bone microenvironment, we assessed changes in adipogenic factors and adipokine expression in adipocytic differentiation of MSC in vitro. Primary human MSC were induced to differentiate in adipogenic conditions in the presence or absence of SrRN (1-2 mM). We tested the dose-dependent effects of SrRN on adipocyte differentiation including changes in the expression of adipogenic markers and adipokines. We report that adipogenesis was negatively affected in the presence of SrRN with a concomitant dose-dependent decrease in the expression of adipogenic markers and changes in adipokine profile. Taken together, our data suggests that SrRN induces biochemical changes in differentiating adipocytes that could generate a favorable osteogenic effect within the bone marrow milieu. PMID:23186800

  4. Pathologic endoplasmic reticulum stress induced by glucotoxic insults inhibits adipocyte differentiation and induces an inflammatory phenotype.

    PubMed

    Longo, Michele; Spinelli, Rosa; D'Esposito, Vittoria; Zatterale, Federica; Fiory, Francesca; Nigro, Cecilia; Raciti, Gregory A; Miele, Claudia; Formisano, Pietro; Beguinot, Francesco; Di Jeso, Bruno

    2016-06-01

    Adipocyte differentiation is critical in obesity. By controlling new adipocyte recruitment, adipogenesis contrasts adipocyte hypertrophy and its adverse consequences, such as insulin resistance. Contrasting data are present in literature on the effect of endoplasmic reticulum (ER) stress and subsequent unfolded protein response (UPR) on adipocyte differentiation, being reported to be either necessary or inhibitory. In this study, we sought to clarify the effect of ER stress and UPR on adipocyte differentiation. We have used two different cell lines, the widely used pre-adipocyte 3T3-L1 cells and a murine multipotent mesenchymal cell line, W20-17 cells. A strong ER stress activator, thapsigargin, and a pathologically relevant inducer of ER stress, glucosamine (GlcN), induced ER stress and UPR above those occurring in the absence of perturbation and inhibited adipocyte differentiation. Very low concentrations of 4-phenyl butyric acid (PBA, a chemical chaperone) inhibited only the overactivation of ER stress and UPR elicited by GlcN, leaving unaltered the part physiologically activated during differentiation, and reversed the inhibitory effect of GlcN on differentiation. In addition, GlcN stimulated proinflammatory cytokine release and PBA prevented these effects. An inhibitor of NF-kB also reversed the effects of GlcN on cytokine release. These results indicate that while ER stress and UPR activation is "physiologically" activated during adipocyte differentiation, the "pathologic" part of ER stress activation, secondary to a glucotoxic insult, inhibits differentiation. In addition, such a metabolic insult, causes a shift of the preadipocyte/adipocyte population towards a proinflammatory phenotype. PMID:26940722

  5. Hypoxia induces adipocyte differentiation of adipose-derived stem cells by triggering reactive oxygen species generation.

    PubMed

    Kim, Ji Hye; Kim, Seok-Ho; Song, Seung Yong; Kim, Won-Serk; Song, Sun U; Yi, TacGhee; Jeon, Myung-Shin; Chung, Hyung-Min; Xia, Ying; Sung, Jong-Hyuk

    2014-01-01

    Generation of reactive oxygen species (ROS) by NADPH oxidase 4 (Nox4) induces the proliferation and migration of adipose-derived stem cells (ASCs). However, the functional role of mitochondrial ROS (mtROS) generation in ASCs is unknown. Therefore, we have investigated whether hypoxia induces the differentiation of ASCs via ROS generation. We also have tried to identify the cellular mechanisms of ROS generation underlying adipocyte differentiation. Hypoxia (2%) and ROS generators, such as antimycin and rotenone, induced adipocyte differentiation, which was attenuated by an ROS scavenger. Although Nox4 generates ROS and regulates proliferation of ASCs, Nox4 inhibition or Nox4 silencing did not inhibit adipocyte differentiation; indeed fluorescence intensity of mito-SOX increased in hypoxia, and treatment with mito-CP, a mtROS scavenger, significantly reduced hypoxia-induced adipocyte differentiation. Phosphorylation of Akt and mTOR was induced by hypoxia, while inhibition of these molecules prevented adipocyte differentiation. Thus hypoxia induces adipocyte differentiation by mtROS generation, and the PI3K/Akt/mTOR pathway is involved. PMID:23956071

  6. Small Molecule-Induced Complement Factor D (Adipsin) Promotes Lipid Accumulation and Adipocyte Differentiation.

    PubMed

    Song, No-Joon; Kim, Suji; Jang, Byung-Hyun; Chang, Seo-Hyuk; Yun, Ui Jeong; Park, Ki-Moon; Waki, Hironori; Li, Dean Y; Tontonoz, Peter; Park, Kye Won

    2016-01-01

    Adipocytes are differentiated by various transcriptional cascades integrated on the master regulator, Pparγ. To discover new genes involved in adipocyte differentiation, preadipocytes were treated with three newly identified pro-adipogenic small molecules and GW7845 (a Pparγ agonist) for 24 hours and transcriptional profiling was analyzed. Four genes, Peroxisome proliferator-activated receptor γ (Pparγ), human complement factor D homolog (Cfd), Chemokine (C-C motif) ligand 9 (Ccl9), and GIPC PDZ Domain Containing Family Member 2 (Gipc2) were induced by at least two different small molecules but not by GW7845. Cfd and Ccl9 expressions were specific to adipocytes and they were altered in obese mice. Small hairpin RNA (shRNA) mediated knockdown of Cfd in preadipocytes inhibited lipid accumulation and expression of adipocyte markers during adipocyte differentiation. Overexpression of Cfd promoted adipocyte differentiation, increased C3a production, and led to induction of C3a receptor (C3aR) target gene expression. Similarly, treatments with C3a or C3aR agonist (C4494) also promoted adipogenesis. C3aR knockdown suppressed adipogenesis and impaired the pro-adipogenic effects of Cfd, further suggesting the necessity for C3aR signaling in Cfd-mediated pro-adipogenic axis. Together, these data show the action of Cfd in adipogenesis and underscore the application of small molecules to identify genes in adipocytes. PMID:27611793

  7. Mitochondrial Reactive Oxygen Species Regulate Adipocyte Differentiation of Mesenchymal Stem Cells in Hematopoietic Stress Induced by Arabinosylcytosine

    PubMed Central

    Wang, Weimin; Zhang, Yao; Lu, Wenyi; Liu, Kaiyan

    2015-01-01

    Objective The increase in adipocytes induced by chemotherapeutic drugs may play a negative role in hematopoietic recovery. However, the mechanism underlying adipocyte differentiation of mesenchymal stem cells (MSCs) in hematopoietic stress is still unknown. Hence, the involvement of reactive oxygen species (ROS) in adipocyte differentiation under hematopoietic stress was investigated in vitro and in vivo. Methods The roles of cellular ROS in adipogenesis were investigated in vivo through an adipocyte hyperplasia marrow model under hematopoietic stress induced by arabinosylcytosine (Ara-C) and in vitro via adipocyte differentiation of human MSCs. ROS levels were detected using the CM-H2DCFDA probe and Mito-SOX dye. Adipogenesis was evaluated by histopathology and oil red O staining, whereas detection of mRNA levels of antioxidant enzymes and adipogenesis markers was performed using quantitative real-time polymerase chain reaction analysis. Results ROS were found to play an important role in regulating adipocyte differentiation of MSCs by activating peroxisome proliferator-activated receptor gamma (PPARγ,) while the antioxidant N-acetyl-L-cysteine acts through ROS to inhibit adipocyte differentiation. The elevated ROS levels induced by Ara-C were caused by both over-generation of mitochondrial ROS and reduction of antioxidant enzymes (Cu/Zn Superoxide dismutase and catalase). Our findings suggest that a mitochondrial-targeted antioxidant could diminish adipocyte differentiation. PMID:25768922

  8. Inhibitory effect of leptin on rosiglitazone-induced differentiation of primary adipocytes prepared from TallyHO/Jng mice

    SciTech Connect

    Kim, Ki Young; Kim, Joo Young; Sung, Yoon-Young; Jung, Won Hoon; Kim, Hee-Youn; Park, Ji Seon; Cheon, Hyae Gyeong; Rhee, Sang Dal

    2011-03-25

    Research highlights: {yields} In this study, we investigated the effects of leptin on adipocyte differentiation prepared from subcutaneous fat of TallyHo mice. {yields} Leptin inhibited the adipocytes differentiation at physiological concentration via inhibition of PPAR{gamma} expression. {yields} Inhibitors of ERK and STAT1 restored the leptin's inhibitory activity both in vitro and in vivo. -- Abstract: The effects of leptin on rosiglitazone-induced adipocyte differentiation were investigated in the primary adipocytes prepared from subcutaneous fat of TallyHO/Jng (TallyHO) mouse, a recently developed model animal for type 2 diabetes mellitus (T2DM). The treatment of leptin inhibited the rosiglitazone-induced adipocyte differentiation with a decreased expression of peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) a key adipogenic transcription factor, both in mRNA and protein levels. Leptin (10 nM) was sufficient to inhibit the adipocyte differentiation, which seemed to come from increased expression of leptin receptor genes in the fat of TallyHO mice. The inhibition of adipogenesis by leptin was restored by the treatment of inhibitors for extracellular-signal-regulated kinase (ERK) (PD98059) and signal transducer and activator of transcription-1 (STAT1) (fludarabine). Furthermore, in vivo intraperitoneal administration of PD98059 and fludarabine increased the PPAR{gamma} expression in the subcutaneous fat of TallyHO mice. These data suggest that leptin could inhibit the PPAR{gamma} expression and adipocyte differentiation in its physiological concentration in TallyHO mice.

  9. Aortic preadipocyte differentiation into adipocytes induced by rosiglitazone in an in vitro model.

    PubMed

    Reyes, Miguel R; Lazalde, Brissia

    2007-01-01

    Peroxisome proliferator-activated receptor gamma (PPARgamma) is a key transcription factor for adipocyte differentiation. Preadipocyte differentiation into adipocytes from precursors in blood vessels is an important issue related to atherosclerotic cardiovascular disease; however, it has been poorly studied because of lack of experimental models. Our aim was to evaluate the potential of primary outgrowths derived from rat aortic rings as a model for studying the preadipocyte differentiation from aortic precursors induced by thiazolidinediones, which are exogenous ligands for PPARgamma. Cell outgrowths derived from rat aortic rings were cultured and incubated with rosiglitazone at 1-1,000 nM; presence of lipid droplets was evaluated by oil red O staining. Rosiglitazone at 100 nM exerted a clear adipogenic effect inferred from the cells filled with fine and medium size lipidic droplets; this effect was extreme at 1,000 nM with cells showing lipidic macrodroplets. These results showed that cultures derived from aortic rings are a useful model for studying arterial preadipocyte differentiation. PMID:17564754

  10. Development of a rapid culture method to induce adipocyte differentiation of human bone marrow-derived mesenchymal stem cells

    SciTech Connect

    Ninomiya, Yuichi; Sugahara-Yamashita, Yzumi; Nakachi, Yutaka; Tokuzawa, Yoshimi; Okazaki, Yasushi; Nishiyama, Masahiko

    2010-04-02

    Human mesenchymal stem cells (hMSCs) derived from bone marrow are multipotent stem cells that can regenerate mesenchymal tissues such as adipose, bone or muscle. It is thought that hMSCs can be utilized as a cell resource for tissue engineering and as human models to study cell differentiation mechanisms, such as adipogenesis, osteoblastogenesis and so on. Since it takes 2-3 weeks for hMSCs to differentiate into adipocytes using conventional culture methods, the development of methods to induce faster differentiation into adipocytes is required. In this study we optimized the culture conditions for adipocyte induction to achieve a shorter cultivation time for the induction of adipocyte differentiation in bone marrow-derived hMSCs. Briefly, we used a cocktail of dexamethasone, insulin, methylisobutylxanthine (DIM) plus a peroxisome proliferator-activated receptor {gamma} agonist, rosiglitazone (DIMRo) as a new adipogenic differentiation medium. We successfully shortened the period of cultivation to 7-8 days from 2-3 weeks. We also found that rosiglitazone alone was unable to induce adipocyte differentiation from hMSCs in vitro. However, rosiglitazone appears to enhance hMSC adipogenesis in the presence of other hormones and/or compounds, such as DIM. Furthermore, the inhibitory activity of TGF-{beta}1 on adipogenesis could be investigated using DIMRo-treated hMSCs. We conclude that our rapid new culture method is very useful in measuring the effect of molecules that affect adipogenesis in hMSCs.

  11. Camptothecin and topotecan inhibit adipocyte differentiation by inducing degradation of PPARγ.

    PubMed

    Kim, Jung-Hoon; Jeong, Manhyung; Lee, Sang-sik; Song, Jaewhan

    2015-08-01

    Camptothecin is an anti-cancer drug extracted from Camptotheca acuminata, a tree native to mainland China. Phase III clinical trials for camptothecin have been completed, and it is now used as a chemotherapeutic reagent. We identified a novel function of camptothecin that affects adipocyte differentiation. Following treatment with camptothecin, endogenous or overexpressed PPARγ becomes destabilized; this was prevented in the presence of MG132, a proteasome inhibitor. Our findings suggest that camptothecin is able to induce proteasome-dependent degradation of PPARγ. The ubiquitylation of PPARγ increased in the presence of camptothecin. Adipogenic differentiation of 3T3-L1 cells was prevented by campothecin and topotecan, but not by irinotecan, confirming our initial findings. Our results suggest a possible role for camptothecin analogs in the regulation of PPARγ. PMID:26079886

  12. The endocrine disruptor diethylstilbestrol induces adipocyte differentiation and promotes obesity in mice

    SciTech Connect

    Hao, Chan-Juan; Cheng, Xue-Jia; Xia, Hong-Fei Ma, Xu

    2012-08-15

    Epidemiology studies indicate that exposure to endocrine disruptors during developmental “window” contributes to adipogenesis and the development of obesity. Implication of endocrine disruptor such as diethylstilbestrol (DES) on adipose tissue development has been poorly investigated. Here we evaluated the effects of DES on adipocyte differentiation in vitro and in vivo, and explored potential mechanism involved in its action. DES induced 3T3-L1 preadipocyte differentiation in a dose-dependent manner, and activated the expression of estrogen receptor (ER) and peroxisome proliferator-acivated receptor (PPAR) γ as well as its target genes required for adipogenesis in vitro. ER mediated the enhancement of DES-induced PPARγ activity. Moreover, DES perturbed key regulators of adipogenesis and lipogenic pathway in vivo. In utero exposure to low dose of DES significantly increased body weight, liver weight and fat mass in female offspring at postnatal day (PND) 60. In addition, serum triglyceride and glucose levels were also significantly elevated. These results suggest that perinatal exposure to DES may be expected to increase the incidence of obesity in a sex-dependent manner and can act as a potential chemical stressor for obesity and obesity-related disorders. -- Highlights: ► DES induced adipocyte differentiation in a dose-dependent manner in 3T3-L1 cells. ► DES activated adipogenic critical regulators and markers in vitro and in vivo. ► Perinatal exposure to DES led to the obese phenotype in female offspring. ► DES might be a potential chemical stressor for obesity and obesity-related disorders.

  13. Transforming growth factor beta2 inhibits adipocyte differentiation induced by skeletal unloading in rat bone marrow stroma.

    PubMed

    Ahdjoudj, Souhila; Lasmoles, Françoise; Holy, Xavier; Zerath, Erik; Marie, Pierre J

    2002-04-01

    Skeletal unloading induced by hindlimb suspension in rats reduces bone formation and induces osteopenia, but its effect on adipogenesis is unknown. We assessed the effects of unloading and transforming growth factor (TGF) beta2 on bone marrow stromal cell adipocyte differentiation in relation with osteoblast differentiation. Skeletal unloading rapidly (4-7 days) decreased osteoblast transcription factor Runx2, osteocalcin (OC), and type I collagen messenger RNA (mRNA) levels and reduced bone formation in the long bone metaphysis. Conversely, unloading increased expression of the adipocyte transcription factor peroxisome proliferator-activated receptor gamma2 (PPARgamma2) at 4 days and increased expression of the adipocyte differentiation genes lipoprotein lipase (LPL) and aP2 in the bone marrow stroma at 7 days. Consistently, unloading increased the number and volume of adipocytes in the bone marrow stroma. Continuous (0-7 days) and late (4-7 days) treatments with TGF-beta2 corrected the abnormal expression of Cbfa1/Runx2, OC, and type I collagen mRNAs and normalized bone formation in unloaded metaphyseal bone. Moreover, both TGF-beta2 treatments decreased PPARy2 and C/EBPalpha mRNA levels at 4 days and normalized aP2 and LPL expression and adipocyte number and volume at 7 days. These results show that skeletal unloading increases adipocyte differentiation concomitantly with inhibition of osteoblast differentiation. These abnormalities are prevented and reversed by TGF-beta2, suggesting a role for TGF-beta in the control of adipogenic differentiation in the bone marrow stroma. PMID:11918224

  14. Apolipoprotein E promotes lipid accumulation and differentiation in human adipocytes

    SciTech Connect

    Lasrich, Dorothee; Bartelt, Alexander; Grewal, Thomas; Heeren, Joerg

    2015-09-10

    Several studies in mice indicate a role for apolipoprotein E (APOE) in lipid accumulation and adipogenic differentiation in adipose tissue. However, little is yet known if APOE functions in a similar manner in human adipocytes. This prompted us to compare lipid loading and expression of adipocyte differentiation markers in APOE-deficient and control adipocytes using the differentiated human mesenchymal stem cell line hMSC-Tert as well as primary human and mouse adipocytes as model systems. Differentiated hMSC-Tert were stably transduced with or without siRNA targeting APOE while murine adipocytes were isolated from wild type and Apoe knockout mice. Human APOE knockdown hMSC-Tert adipocytes accumulated markedly less triglycerides compared to control cells. This correlated with strongly decreased gene expression levels of adipocyte markers such as adiponectin (ADIPOQ) and fatty acid binding protein 4 (FABP4) as well as the key transcription factor driving adipocyte differentiation, peroxisome proliferator activator receptor gamma (PPARG), in particular the PPARG2 isoform. Similarly, differentiation of murine Apoe-deficient adipocytes was characterized by reduced gene expression of Adipoq, Fabp4 and Pparg. Interestingly, incubation of APOE-deficient hMSC-Tert adipocytes with conditioned media from APOE3-overexpressing adipocytes or APOE-containing Very Low Density Lipoprotein (VLDL) partially restored triglyceride accumulation, but were unable to induce adipocyte differentiation, as judged by expression of adipocyte markers. Taken together, depletion of endogenous APOE in human adipocytes severely impairs lipid accumulation, which is associated with an inability to initiate differentiation. - Highlights: • Immortalized human mesenchymal stem cells were used to study adipocyte development. • Knockdown of endogenous APOE lead to impaired lipid accumulation and adipogenesis. • APOE supplementation partially restored lipid accumulation but not differentiation.

  15. Interruptin B induces brown adipocyte differentiation and glucose consumption in adipose-derived stem cells

    PubMed Central

    KAEWSUWAN, SIREEWAN; PLUBRUKARN, ANUCHIT; UTSINTONG, MALEERUK; KIM, SEOK-HO; JEONG, JIN-HYUN; CHO, JIN GU; PARK, SANG GYU; SUNG, JONG-HYUK

    2016-01-01

    Interruptin B has been isolated from Cyclosorus terminans, however, its pharamcological effect has not been fully identified. In the present study, the effects of interruptin B, from C. terminans, on brown adipocyte differentiation and glucose uptake in adipose-derived stem cells (ASCs) were investigated. The results revealed that interruptin B dose-dependently enhanced the adipogenic differentiation of ASCs, with an induction in the mRNA expression levels of peroxisome proliferator-activated receptor (PPAR)-α and PPAR-γ. In addition, interruptin B efficiently increased the number and the membrane potential of mitochondria and upregulated the mRNA expression levels of uncoupling protein (UCP)-1 and cyclooxygenase (COX)-2, which are all predominantly expressed in brown adipocytes. Interruptin B increased glucose consumption in differentiated ASCs, accompanied by the upregulation in the mRNA expression levels of glucose transporter (GLUT)-1 and GLUT-4. The computational analysis of molecular docking, a luciferase reporter assay and surface plasmon resonance confirmed the marked binding affinity of interruptin B to PPAR-α and PPAR-γ (KD values of 5.32 and 0.10 µM, respectively). To the best of our knowledge, the present study is the first report to show the stimulatory effects of interruptin B on brown adipocyte differentiation and glucose uptake in ASCs, through its role as a dual PPAR-α and PPAR-γ ligand. Therefore, interruptin B could be further developed as a therapeutic agent for the treatment of diabetes. PMID:26781331

  16. The endocrine disruptor mono-(2-ethylhexyl) phthalate promotes adipocyte differentiation and induces obesity in mice.

    PubMed

    Hao, Chanjuan; Cheng, Xuejia; Xia, Hongfei; Ma, Xu

    2012-12-01

    The environmental obesogen hypothesis proposes that exposure to endocrine disruptors during developmental 'window' contributes to adipogenesis and the development of obesity. MEHP [mono-(2-ethylhexyl) phthalate], a metabolite of the widespread plasticizer DEHP [di-(2-ethylhexyl) phthalate], has been found in exposed organisms and identified as a selective PPARγ (peroxisome-proliferator-activated receptor γ) modulator. However, implication of MEHP on adipose tissue development has been poorly investigated. In the present study, we show the dose-dependent effects of MEHP on adipocyte differentiation and GPDH (glycerol-3-phosphate dehydrogenase) activity in the murine 3T3-L1 cell model. MEHP induced the expression of PPARγ as well as its target genes required for adipogenesis in vitro. Moreover, MEHP perturbed key regulators of adipogenesis and lipogenic pathway in vivo. In utero exposure to a low dose of MEHP significantly increased b.w. (body weight) and fat pad weight in male offspring at PND (postnatal day) 60. In addition, serum cholesterol, TAG (triacylglycerol) and glucose levels were also significantly elevated. These results suggest that perinatal exposure to MEHP may be expected to increase the incidence of obesity in a sex-dependent manner and can act as a potential chemical stressor for obesity and obesity-related disorders. PMID:22953781

  17. Apolipoprotein E promotes lipid accumulation and differentiation in human adipocytes.

    PubMed

    Lasrich, Dorothee; Bartelt, Alexander; Grewal, Thomas; Heeren, Joerg

    2015-09-10

    Several studies in mice indicate a role for apolipoprotein E (APOE) in lipid accumulation and adipogenic differentiation in adipose tissue. However, little is yet known if APOE functions in a similar manner in human adipocytes. This prompted us to compare lipid loading and expression of adipocyte differentiation markers in APOE-deficient and control adipocytes using the differentiated human mesenchymal stem cell line hMSC-Tert as well as primary human and mouse adipocytes as model systems. Differentiated hMSC-Tert were stably transduced with or without siRNA targeting APOE while murine adipocytes were isolated from wild type and Apoe knockout mice. Human APOE knockdown hMSC-Tert adipocytes accumulated markedly less triglycerides compared to control cells. This correlated with strongly decreased gene expression levels of adipocyte markers such as adiponectin (ADIPOQ) and fatty acid binding protein 4 (FABP4) as well as the key transcription factor driving adipocyte differentiation, peroxisome proliferator activator receptor gamma (PPARG), in particular the PPARG2 isoform. Similarly, differentiation of murine Apoe-deficient adipocytes was characterized by reduced gene expression of Adipoq, Fabp4 and Pparg. Interestingly, incubation of APOE-deficient hMSC-Tert adipocytes with conditioned media from APOE3-overexpressing adipocytes or APOE-containing Very Low Density Lipoprotein (VLDL) partially restored triglyceride accumulation, but were unable to induce adipocyte differentiation, as judged by expression of adipocyte markers. Taken together, depletion of endogenous APOE in human adipocytes severely impairs lipid accumulation, which is associated with an inability to initiate differentiation. PMID:26201081

  18. Interruptin B induces brown adipocyte differentiation and glucose consumption in adipose-derived stem cells.

    PubMed

    Kaewsuwan, Sireewan; Plubrukarn, Anuchit; Utsintong, Maleeruk; Kim, Seok-Ho; Jeong, Jin-Hyun; Cho, Jin Gu; Park, Sang Gyu; Sung, Jong-Hyuk

    2016-03-01

    Interruptin B has been isolated from Cyclosorus terminans, however, its pharamcological effect has not been fully identified. In the present study, the effects of interruptin B, from C. terminans, on brown adipocyte differentiation and glucose uptake in adipose‑derived stem cells (ASCs) were investigated. The results revealed that interruptin B dose‑dependently enhanced the adipogenic differentiation of ASCs, with an induction in the mRNA expression levels of peroxisome proliferator‑activated receptor (PPAR)‑α and PPAR‑γ. In addition, interruptin B efficiently increased the number and the membrane potential of mitochondria and upregulated the mRNA expression levels of uncoupling protein (UCP)‑1 and cyclooxygenase (COX)‑2, which are all predominantly expressed in brown adipocytes. Interruptin B increased glucose consumption in differentiated ASCs, accompanied by the upregulation in the mRNA expression levels of glucose transporter (GLUT)‑1 and GLUT‑4. The computational analysis of molecular docking, a luciferase reporter assay and surface plasmon resonance confirmed the marked binding affinity of interruptin B to PPAR‑α and PPAR‑γ (KD values of 5.32 and 0.10 µm, respectively). To the best of our knowledge, the present study is the first report to show the stimulatory effects of interruptin B on brown adipocyte differentiation and glucose uptake in ASCs, through its role as a dual PPAR‑α and PPAR‑γ ligand. Therefore, interruptin B could be further developed as a therapeutic agent for the treatment of diabetes. PMID:26781331

  19. Adipocyte induced arterial calcification is prevented with sodium thiosulfate

    SciTech Connect

    Chen, Neal X.; O’Neill, Kalisha; Akl, Nader Kassis; Moe, Sharon M.

    2014-06-20

    Highlights: • High phosphorus can induce calcification of adipocytes, even when fully differentiated. • Adipocytes can induce vascular calcification in an autocrine manner. • Sodium thiosulfate inhibits adipocyte calcification. - Abstract: Background: Calcification can occur in fat in multiple clinical conditions including in the dermis, breasts and in the abdomen in calciphylaxis. All of these are more common in patients with advanced kidney disease. Clinically, hyperphosphatemia and obesity are risk factors. Thus we tested the hypothesis that adipocytes can calcify in the presence of elevated phosphorus and/or that adipocytes exposed to phosphorus can induce vascular smooth muscle cell (VSMC) calcification. Methods: 3T3-L1 preadipocytes were induced into mature adipocytes and then treated with media containing high phosphorus. Calcification was assessed biochemically and PCR performed to determine the expression of genes for osteoblast and adipocyte differentiation. Adipocytes were also co-cultured with bovine VSMC to determine paracrine effects, and the efficacy of sodium thiosulfate was determined. Results: The results demonstrated that high phosphorus induced the calcification of differentiated adipocytes with increased expression of osteopontin, the osteoblast transcription factor Runx2 and decreased expression of adipocyte transcription factors peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT-enhancer-binding protein α (CEBPα), indicating that high phosphorus led to a phenotypic switch of adipocytes to an osteoblast like phenotype. Sodium thiosulfate, dose dependently decreased adipocyte calcification and inhibited adipocyte induced increase of VSMC calcification. Co-culture studies demonstrated that adipocytes facilitated VSMC calcification partially mediated by changes of secretion of leptin and vascular endothelial growth factor (VEGF) from adipocytes. Conclusion: High phosphorus induced calcification of mature adipocytes, and

  20. Angiotensin II type 2 receptor promotes adipocyte differentiation and restores adipocyte size in high-fat/high-fructose diet-induced insulin resistance in rats.

    PubMed

    Shum, Michaël; Pinard, Sandra; Guimond, Marie-Odile; Labbé, Sébastien M; Roberge, Claude; Baillargeon, Jean-Patrice; Langlois, Marie-France; Alterman, Mathias; Wallinder, Charlotta; Hallberg, Anders; Carpentier, André C; Gallo-Payet, Nicole

    2013-01-15

    This study was aimed at establishing whether specific activation of angiotensin II (ANG II) type 2 receptor (AT2R) modulates adipocyte differentiation and function. In primary cultures of subcutaneous (SC) and retroperitoneal (RET) preadipocytes, both AT2R and AT1R were expressed at the mRNA and protein level. Cells were stimulated with ANG II or the AT2R agonist C21/M24, alone or in the presence of the AT1R antagonist losartan or the AT2R antagonist PD123,319. During differentiation, C21/M24 increased PPARγ expression in both RET and SC preadipocytes while the number of small lipid droplets and lipid accumulation solely increased in SC preadipocytes. In mature adipocytes, C21/M24 decreased the mean size of large lipid droplets. Upon abolishment of AT2R expression using AT2R-targeted shRNAs, expressions of AT2R, aP2, and PPARγ remained very low, and cells were unable to differentiate. In Wistar rats fed a 6-wk high-fat/high-fructose (HFHF) diet, a significant shift toward larger adipocytes was observed in RET and SC adipose tissue depots. C21/M24 treatments for 6 wk restored normal adipocyte size distribution in both these tissue depots. Moreover, C21/M24 and losartan decreased hyperinsulinemia and improved insulin sensitivity impaired by HFHF diet. A strong correlation between adipocyte size area and glucose infusion rate during euglycemic-hyperinsulinemic clamp was observed. These results indicate that AT2R is involved in early adipocyte differentiation, while in mature adipocytes and in a model of insulin resistance AT2R activation restores normal adipocyte morphology and improves insulin sensitivity. PMID:23149621

  1. Effect of Wnt-1 inducible signaling pathway protein-2 (WISP-2/CCN5), a downstream protein of Wnt signaling, on adipocyte differentiation

    SciTech Connect

    Inadera, Hidekuni Shimomura, Akiko; Tachibana, Shinjiro

    2009-02-20

    Wnt signaling negatively regulates adipocyte differentiation, and ectopic expression of Wnt-1 in 3T3-L1 cells induces several downstream molecules of Wnt signaling, including Wnt-1 inducible signaling pathway protein (WISP)-2. In this study, we examined the role of WISP-2 in the process of adipocyte differentiation using an in vitro cell culture system. In the differentiation of 3T3-L1 cells, WISP-2 expression was observed in growing cells and declined thereafter. In the mitotic clonal expansion phase of adipocyte differentiation, WISP-2 expression was transiently down-regulated concurrently with up-regulation of CCAAT/enhancer-binding protein {delta} expression. Treatment of 3T3-L1 cells in the differentiation medium with lithium, an activator of Wnt signaling, inhibited the differentiation process with concomitant induction of WISP-2. Treatment of differentiated cells with lithium induced de-differentiation as evidenced by profound reduction of peroxisome proliferator-activator receptor {gamma} expression and concomitant induction of WISP-2. However, de-differentiation of differentiated cells induced by tumor necrosis factor-{alpha} did not induce WISP-2 expression. To directly examine the effect of WISP-2 on adipocyte differentiation, 3T3-L1 cells were infected with a retrovirus carrying WISP-2. Although forced expression of WISP-2 inhibited preadipocyte proliferation, it had no effect on adipocyte differentiation. Thus, although WISP-2 is a downstream protein of Wnt signaling, the role of WISP-2 on adipocyte differentiation may be marginal, at least in this in vitro culture model.

  2. The increasingly complex regulation of adipocyte differentiation.

    PubMed

    Poulos, Sylvia P; Dodson, Michael V; Culver, Melinda F; Hausman, Gary J

    2016-03-01

    Adipose (AD) tissue development and function relies on the ability of adipocytes to proliferate and differentiate into lipid-containing cells that also have endocrine function. Research suggests that certain conditions can induce AD tissue stem cells to differentiate into various cell types and that the microenvironment of the cell, including the extracellular matrix (ECM), is essential in maintaining cell and tissue function. This review provides an overview of factors involved in the proliferation and differentiation of adipocytes. A brief review of the numerous factors that influence PPARγ, the transcription factor thought to be the master regulator of adipocyte differentiation, provides context of established pathways that regulate adipogenesis. Thought provoking findings from research with hypoxia that is supported by earlier research that vascular development is related to adipogenesis are reviewed. Finally, our understanding of the critical role of the ECM and environment in adipogenesis is discussed and compared with studies that suggest that adipocytes may dedifferentiate and can convert into other cell types. PMID:26645953

  3. Trans, trans-farnesol as a mevalonate-derived inducer of murine 3T3-F442A pre-adipocyte differentiation.

    PubMed

    Torabi, Sheida; Mo, Huanbiao

    2016-03-01

    Based on our finding that depletion of mevalonate-derived metabolites inhibits adipocyte differentiation, we hypothesize that trans, trans-farnesol (farnesol), a mevalonate-derived sesquiterpene, induces adipocyte differentiation. Farnesol dose-dependently (25-75 μmol/L) increased intracellular triglyceride content of murine 3T3-F442A pre-adipocytes measured by AdipoRed™ Assay and Oil Red-O staining. Concomitantly, farnesol dose-dependently increased glucose uptake and glucose transport protein 4 (GLUT4) expression without affecting cell viability. Furthermore, quantitative real-time polymerase chain reaction and Western blot showed that farnesol increased the mRNA and protein levels of peroxisome proliferator-activated receptor γ (PPARγ), a key regulator of adipocyte differentiation, and the mRNA levels of PPARγ-regulated fatty acid-binding protein 4 and adiponectin; in contrast, farnesol downregulated Pref-1 gene, a marker of pre-adipocytes. GW9662 (10 µmol/L), an antagonist of PPARγ, reversed the effects of farnesol on cellular lipid content, suggesting that PPARγ signaling pathway may mediate the farnesol effect. Farnesol (25-75 μmol/L) did not affect the mRNA level of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme in the mevalonate pathway. Farnesol may be the mevalonate-derived inducer of adipocyte differentiation and potentially an insulin sensitizer via activation of PPARγ and upregulation of glucose uptake. PMID:26660152

  4. Tributyltin Differentially Promotes Development of a Phenotypically Distinct Adipocyte

    PubMed Central

    Regnier, Shane M.; El-Hashani, Essam; Kamau, Wakanene; Zhang, Xiaojie; Massad, Nicole L.; Sargis, Robert M.

    2015-01-01

    Objective Environmental endocrine disrupting chemicals (EDCs) are increasingly implicated in the pathogenesis of obesity. Evidence implicates various EDCs as being pro-adipogenic, including tributyltin (TBT), which activates the peroxisome proliferator activated receptor-γ (PPARγ). However, the conditions required for TBT-induced adipogenesis and its functional consequences are incompletely known. Methods The co-stimulatory conditions necessary for preadipocyte-to-adipocyte differentiation were compared between TBT and the pharmacological PPARγ agonist troglitazone (Trog) in the 3T3-L1 cell line; basal and insulin-stimulated glucose uptake were assessed using radiolabeled 2-deoxyglucose. Results TBT enhanced expression of the adipocyte marker C/EBPα with co-exposure to either isobutylmethylxanthine or insulin in the absence of other adipogenic stimuli. Examination of several adipocyte-specific proteins revealed that TBT and Trog differentially affected protein expression despite comparable PPARγ stimulation. In particular, TBT reduced adiponectin expression upon maximal adipogenic stimulation. Under submaximal stimulation, TBT and Trog differentially promoted adipocyte-specific gene expression despite similar lipid accumulation. Moreover, TBT attenuated Trog-induced adipocyte gene expression under conditions of co-treatment. Finally, TBT-induced adipocytes exhibited altered glucose metabolism, with increased basal glucose uptake. Conclusions TBT-induced adipocytes are functionally distinct from those generated by a pharmacological PPARγ agonist, suggesting that obesogen-induced adipogenesis may generate dysfunctional adipocytes with the capacity to deleteriously affect global energy homeostasis. PMID:26243053

  5. Dynamics of protein secretion during adipocyte differentiation.

    PubMed

    Ojima, Koichi; Oe, Mika; Nakajima, Ikuyo; Muroya, Susumu; Nishimura, Takanori

    2016-08-01

    The major functions of adipocytes include both lipid storage and the production of secretory factors. However, the type of proteins released from mouse 3T3-L1 cells during adipocyte differentiation remains poorly understood. We examined the dynamics of secreted proteins during adipocyte differentiation using mass spectrometry (MS) combined with an iTRAQ (®) labeling method that enables the simultaneous analysis of relative protein expression levels. A total of 215 proteins were identified and quantified from approximately 10 000 MS/MS spectra. Of these, approximately 38% were categorized as secreted proteins based on gene ontology classification. Adipokine secretion levels were increased with the progression of differentiation. By contrast, levels of fibril collagen components, such as subunits of type I and III collagens, were decreased during differentiation. Basement membrane components attained their peak levels at day 4 when small lipid droplets accumulated in differentiated 3T3-L1 cells. Simultaneously, peak levels of collagen microfibril components that comprise type V and VI collagen subunits were also observed. Our data demonstrated that extracellular matrix components were predominantly released during the early and middle stages of adipocyte differentiation, with a subsequent increase in the secretion of adipokines. This suggests that 3T3-L1 cells secrete adipokines after their ECM is constructed during adipocyte differentiation. PMID:27516960

  6. Sleep Fragmentation in Mice Induces Nicotinamide Adenine Dinucleotide Phosphate Oxidase 2-Dependent Mobilization, Proliferation, and Differentiation of Adipocyte Progenitors in Visceral White Adipose Tissue

    PubMed Central

    Khalyfa, Abdelnaby; Wang, Yang; Zhang, Shelley X.; Qiao, Zhuanhong; Abdelkarim, Amal; Gozal, David

    2014-01-01

    Background: Chronic sleep fragmentation (SF) without sleep curtailment induces increased adiposity. However, it remains unclear whether mobilization, proliferation, and differentiation of adipocyte progenitors (APs) occurs in visceral white adipose tissue (VWAT), and whether nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (Nox2) activity plays a role. Methods: Changes in VWAT depot cell size and AP proliferation were assessed in wild-type and Nox2 null male mice exposed to SF and control sleep (SC). To assess mobilization, proliferation, and differentiation of bone marrow mesenchymal stem cells (BM-MSC), Sca-1+ bone marrow progenitors were isolated from GFP+ or RFP+ mice, and injected intravenously to adult male mice (C57BL/6) previously exposed to SF or SC. Results: In comparison with SC, SF was associated with increased weight accrual at 3 w and thereafter, larger subcutaneous and visceral fat depots, and overall adipocyte size at 8 w. Increased global AP numbers in VWAT along with enhanced AP BrDU labeling in vitro and in vivo emerged in SF. Systemic injections of GFP+ BM-MSC resulted in increased AP in VWAT, as well as in enhanced differentiation into adipocytes in SF-exposed mice. No differences occurred between SF and SC in Nox2 null mice for any of these measurements. Conclusions: Chronic sleep fragmentation (SF) induces obesity in mice and increased proliferation and differentiation of adipocyte progenitors (AP) in visceral white adipose tissue (VWAT) that are mediated by increased Nox2 activity. In addition, enhanced migration of bone marrow mesenchymal stem cells from the systemic circulation into VWAT, along with AP differentiation, proliferation, and adipocyte formation occur in SF-exposed wild-type but not in oxidase 2 (Nox2) null mice. Thus, Nox2 may provide a therapeutic target to prevent obesity in the context of sleep disorders. Citation: Khalyfa A, Wang Y, Zhang SX, Qiao Z, Abdelkarim A, Gozal D. Sleep fragmentation in mice induces

  7. Regulation of human subcutaneous adipocyte differentiation by EID1.

    PubMed

    Vargas, Diana; Shimokawa, Noriaki; Kaneko, Ryosuke; Rosales, Wendy; Parra, Adriana; Castellanos, Ángela; Koibuchi, Noriyuki; Lizcano, Fernando

    2016-02-01

    Increasing thermogenesis in white adipose tissues can be used to treat individuals at high risk for obesity and cardiovascular disease. The objective of this study was to determine the function of EP300-interacting inhibitor of differentiation (EID1), an inhibitor of muscle differentiation, in the induction of beige adipocytes from adipose mesenchymal stem cells (ADMSCs). Subcutaneous adipose tissue was obtained from healthy women undergoing abdominoplasty. ADMSCs were isolated in vitro, grown, and transfected with EID1 or EID1 siRNA, and differentiation was induced after 48 h by administering rosiglitazone. The effects of EID1 expression under the control of the aP2 promoter (aP2-EID1) were also evaluated in mature adipocytes that were differentiated from ADMSCs. Transfection of EID1 into ADMSCs reduced triglyceride accumulation while increasing levels of thermogenic proteins, such as PGC1α, TFAM, and mitochondrial uncoupling protein 1 (UCP1), all of which are markers of energy expenditure and mitochondrial activity. Furthermore, increased expression of the beige phenotype markers CITED1 and CD137 was observed. Transfection of aP2-EID1 transfection induced the conversion of mature white adipocytes to beige adipocytes, as evidenced by increased expression of PGC1α, UCP1, TFAM, and CITED1. These results indicate that EID1 can modulate ADMSCs, inducing a brown/beige lineage. EID1 may also activate beiging in white adipocytes obtained from subcutaneous human adipose tissue. PMID:26643909

  8. Amber Light (590 nm) Induces the Breakdown of Lipid Droplets through Autophagy-Related Lysosomal Degradation in Differentiated Adipocytes

    PubMed Central

    Choi, Min Sik; Kim, Hyoung-June; Ham, Mira; Choi, Dong-Hwa; Lee, Tae Ryong; Shin, Dong Wook

    2016-01-01

    Lipolysis in the adipocytes provides free fatty acids for other tissues in response to the energy demand. With the rapid increase in obesity-related diseases, finding novel stimuli or mechanisms that regulate lipid metabolism becomes important. We examined the effects of visible light (410, 457, 505, 530, 590, and 660 nm) irradiation on lipolysis regulation in adipocytes differentiated from human adipose-derived stem cells (ADSCs). Interestingly, 590 nm (amber) light irradiation significantly reduced the concentration of lipid droplets (LDs). We further investigated the lipolytic signaling pathways that are involved in 590 nm light irradiation-induced breakdown of LDs. Immunoblot analysis revealed that 590 nm light irradiation-induced phosphorylation of hormone-sensitive lipase (HSL) was insufficient to promote reduction of LDs. We observed that 590 nm light irradiation decreased the expression of perilipin 1. We found that 590 nm light irradiation, but not 505 nm, induced conversion of LC3 I to LC3 II, a representative autophagic marker. We further demonstrated that the lysosomal inhibitors leupeptin/NH4Cl inhibited 590 nm light irradiation-induced reduction of LDs in differentiated adipocytes. Our data suggest that 590 nm light irradiation-induced LD breakdown is partially mediated by autophagy-related lysosomal degradation, and can be applied in clinical settings to reduce obesity. PMID:27346059

  9. Amber Light (590 nm) Induces the Breakdown of Lipid Droplets through Autophagy-Related Lysosomal Degradation in Differentiated Adipocytes.

    PubMed

    Choi, Min Sik; Kim, Hyoung-June; Ham, Mira; Choi, Dong-Hwa; Lee, Tae Ryong; Shin, Dong Wook

    2016-01-01

    Lipolysis in the adipocytes provides free fatty acids for other tissues in response to the energy demand. With the rapid increase in obesity-related diseases, finding novel stimuli or mechanisms that regulate lipid metabolism becomes important. We examined the effects of visible light (410, 457, 505, 530, 590, and 660 nm) irradiation on lipolysis regulation in adipocytes differentiated from human adipose-derived stem cells (ADSCs). Interestingly, 590 nm (amber) light irradiation significantly reduced the concentration of lipid droplets (LDs). We further investigated the lipolytic signaling pathways that are involved in 590 nm light irradiation-induced breakdown of LDs. Immunoblot analysis revealed that 590 nm light irradiation-induced phosphorylation of hormone-sensitive lipase (HSL) was insufficient to promote reduction of LDs. We observed that 590 nm light irradiation decreased the expression of perilipin 1. We found that 590 nm light irradiation, but not 505 nm, induced conversion of LC3 I to LC3 II, a representative autophagic marker. We further demonstrated that the lysosomal inhibitors leupeptin/NH4Cl inhibited 590 nm light irradiation-induced reduction of LDs in differentiated adipocytes. Our data suggest that 590 nm light irradiation-induced LD breakdown is partially mediated by autophagy-related lysosomal degradation, and can be applied in clinical settings to reduce obesity. PMID:27346059

  10. Glycine suppresses TNF-α-induced activation of NF-κB in differentiated 3T3-L1 adipocytes.

    PubMed

    Blancas-Flores, Gerardo; Alarcón-Aguilar, Francisco J; García-Macedo, Rebeca; Almanza-Pérez, Julio C; Flores-Sáenz, José L; Román-Ramos, Rubén; Ventura-Gallegos, José L; Kumate, Jesús; Zentella-Dehesa, Alejandro; Cruz, Miguel

    2012-08-15

    Glycine strongly reduces the serum levels of pro-inflammatory cytokines and increases the levels of anti-inflammatory cytokines. Recently, glycine has been shown to decrease the expression and secretion of pro-inflammatory adipokines in monosodium glutamate-induced obese (MSG/Ob) mice. It has been postulated that these effects may be explained by a reduction in nuclear factor kappa B (NF-κB) activation. NF-κB is a transcription factor, which is crucial to the inflammatory response. Hasegawa et al. (2011 and 2012) recently reported a glycine-dependent reduction in NF-κB levels. Here, we have investigated the role of glycine in the regulation of NF-κB in differentiated 3T3-L1 adipocytes. The results revealed that pretreatment with glycine interfered with the activation of NF-κB, which has been shown to be stimulated by tumor necrosis factor-alpha (TNF-α). Glycine alone stimulated NF-κB activation in an unusual way such that the inhibitor κB-β (IκB-β) degradation was more significant than that of the inhibitor κB-α (IκB-α) and led to NF-κB complexes comprised of p50 and p65 subunits; IκB-ε degradation did not affect by glycine. These findings suggest that glycine could be used as an alternative treatment for chronic inflammation, which is a hallmark of obesity and other comorbidities, and is characterized by an elevated production of pro-inflammatory cytokines. PMID:22732655

  11. Silibinin Regulates Lipid Metabolism and Differentiation in Functional Human Adipocytes

    PubMed Central

    Barbagallo, Ignazio; Vanella, Luca; Cambria, Maria T.; Tibullo, Daniele; Godos, Justyna; Guarnaccia, Laura; Zappalà, Agata; Galvano, Fabio; Li Volti, Giovanni

    2016-01-01

    Silibinin, a natural plant flavonolignan is the main active constituent found in milk thistle (Silybum marianum). It is known to have hepatoprotective, anti-neoplastic effect, and suppresses lipid accumulation in adipocytes. Objective of this study was to investigate the effect of silibinin on adipogenic differentiation and thermogenic capacity of human adipose tissue derived mesenchymal stem cells. Silibinin (10 μM) treatment, either at the beginning or at the end of adipogenic differentiation, resulted in an increase of SIRT-1, PPARα, Pgc-1α, and UCPs gene expression. Moreover, silibinin administration resulted in a decrease of PPARγ, FABP4, FAS, and MEST/PEG1 gene expression during the differentiation, confirming that this compound is able to reduce fatty acid accumulation and adipocyte size. Our data showed that silibinin regulated adipocyte lipid metabolism, inducing thermogenesis and promoting a brown remodeling in adipocyte. Taken together, our findings suggest that silibinin increases UCPs expression by stimulation of SIRT1, PPARα, and Pgc-1α, improved metabolic parameters, decreased lipid mass leading to the formation of functional adipocytes. PMID:26834634

  12. Concomitant beige adipocyte differentiation upon induction of mesenchymal stem cells into brown adipocytes.

    PubMed

    Wang, Yung-Li; Lin, Shih-Pei; Hsieh, Patrick C H; Hung, Shih-Chieh

    2016-09-16

    The accumulation of fat, which results in obesity, is related to many metabolic disorders. Besides white and brown adipose tissue, beige adipose tissue has recently been recognized as a new type of accumulated fat. Mesenchymal stem cells (MSCs) have been shown to differentiate into brown adipocytes. Through analyzing levels of mRNA and protein markers associated with beige adipocyte, we found concomitant beige adipocyte differentiation upon induction of MSCs into brown adipocytes in a defined medium containing triiodothyronine, insulin, dexamethasone, and indomethacin. Moreover, we found that protein kinase A (PKA) modulators regulated MSC differentiation into brown or beige adipocytes. Activation of PKA by isobutylmethylxanthine or forskolin increased brown adipocyte differentiation and reduced beige adipocyte differentiation, while inactivation of PKA by KT-5720 or SC-3010 or the knockdown of PKA downstream cAMP response element-binding protein (CREB) decreased brown adipocyte differentiation and increased beige adipocyte differentiation. We also showed that increased brown adipocyte differentiation was accompanied by an increase in mitochondrial mass. In conclusion, we propose a model of beige/brown co-differentiation in MSCs and develop a method for controlling this differentiation via PKA modulation. PMID:27498007

  13. Activation of TRPV2 negatively regulates the differentiation of mouse brown adipocytes.

    PubMed

    Sun, Wuping; Uchida, Kunitoshi; Takahashi, Nobuyuki; Iwata, Yuko; Wakabayashi, Shigeo; Goto, Tsuyoshi; Kawada, Teruo; Tominaga, Makoto

    2016-09-01

    Transient receptor potential vanilloid 2 (TRPV2) acts as a Ca(2+)-permeable non-selective cation channel that has been reported to be sensitive to temperature, mechanical force, and some chemicals. We recently showed that TRPV2 is critical for maintenance of the thermogenic function of brown adipose tissue in mice. However, the involvement of TRPV2 in the differentiation of brown adipocytes remains unexplored. We found that the expression of TRPV2 was dramatically increased during the differentiation of brown adipocytes. Non-selective TRPV2 agonists (2-aminoethoxydiphenyl borate and lysophosphatidylcholine) inhibited the differentiation of brown adipocytes in a dose-dependent manner during the early stage of differentiation of brown adipocytes. The inhibition was rescued by a TRPV2-selective antagonist, SKF96365 (SKF). Mechanical force, which activates TRPV2, also inhibited the differentiation of brown adipocytes in a strength-dependent manner, and the effect was reversed by SKF. In addition, the inhibition of adipocyte differentiation by either TRPV2 ligand or mechanical stimulation was significantly smaller in the cells from TRPV2KO mice. Moreover, calcineurin inhibitors, cyclosporine A and FK506, partially reversed TRPV2 activation-induced inhibition of brown adipocyte differentiation. Thus, we conclude that TRPV2 might be involved in the modulation of brown adipocyte differentiation partially via a calcineurin pathway. PMID:27318696

  14. Mechanism of Regulation of Adipocyte Numbers in Adult Organisms Through Differentiation and Apoptosis Homeostasis

    PubMed Central

    Bozec, Aline; Hannemann, Nicole

    2016-01-01

    Considering that adipose tissue (AT) is an endocrine organ, it can influence whole body metabolism. Excessive energy storage leads to the dysregulation of adipocytes, which in turn induces abnormal secretion of adipokines, triggering metabolic syndromes such as obesity, dyslipidemia, hyperglycemia, hyperinsulinemia, insulin resistance and type 2 diabetes. Therefore, investigating the molecular mechanisms behind adipocyte dysregulation could help to develop novel therapeutic strategies. Our protocol describes methods for evaluating the molecular mechanism affected by hypoxic conditions of the AT, which correlates with adipocyte apoptosis in adult mice. This protocol describes how to analyze AT in vivo through gene expression profiling as well as histological analysis of adipocyte differentiation, proliferation and apoptosis during hypoxia exposure, ascertained through staining of hypoxic cells or HIF-1α protein. Furthermore, in vitro analysis of adipocyte differentiation and its responses to various stimuli completes the characterization of the molecular pathways behind possible adipocyte dysfunction leading to metabolic syndromes. PMID:27284940

  15. Regulation of De Novo Adipocyte Differentiation Through Cross Talk Between Adipocytes and Preadipocytes.

    PubMed

    Challa, Tenagne D; Straub, Leon G; Balaz, Miroslav; Kiehlmann, Elke; Donze, Olivier; Rudofsky, Gottfried; Ukropec, Jozef; Ukropcova, Barbara; Wolfrum, Christian

    2015-12-01

    There are many known adipokines differentially secreted from the different adipose depots; however, their paracrine and autocrine effects on de novo adipocyte formation are not fully understood. By developing a coculture method of preadipocytes with primary subcutaneous and visceral adipocytes or tissue explants, we could show that the total secretome inhibited preadipocyte differentiation. Using a proteomics approach with fractionated secretome samples, we were able to identify a spectrum of factors that either positively or negatively affected adipocyte formation. Among the secreted factors, Slc27a1, Vim, Cp, and Ecm1 promoted adipocyte differentiation, whereas Got2, Cpq, interleukin-1 receptor-like 1/ST2-IL-33, Sparc, and Lgals3bp decreased adipocyte differentiation. In human subcutaneous adipocytes of lean subjects, obese subjects, and obese subjects with type 2 diabetes, Vim and Slc27a1 expression was negatively correlated with adipocyte size and BMI and positively correlated with insulin sensitivity, while Sparc and Got2 showed the opposite trend. Furthermore, we demonstrate that Slc27a1 was increased upon weight loss in morbidly obese patients, while Sparc expression was reduced. Taken together, our findings identify adipokines that regulate adipocyte differentiation through positive or negative paracrine and autocrine feedback loop mechanisms, which could potentially affect whole-body energy metabolism. PMID:26340931

  16. Epidermal Wnt/β-catenin signaling regulates adipocyte differentiation via secretion of adipogenic factors

    PubMed Central

    Donati, Giacomo; Proserpio, Valentina; Lichtenberger, Beate Maria; Natsuga, Ken; Sinclair, Rodney; Fujiwara, Hironobu; Watt, Fiona M.

    2014-01-01

    It has long been recognized that the hair follicle growth cycle and oscillation in the thickness of the underlying adipocyte layer are synchronized. Although factors secreted by adipocytes are known to regulate the hair growth cycle, it is unclear whether the epidermis can regulate adipogenesis. We show that inhibition of epidermal Wnt/β-catenin signaling reduced adipocyte differentiation in developing and adult mouse dermis. Conversely, ectopic activation of epidermal Wnt signaling promoted adipocyte differentiation and hair growth. When the Wnt pathway was activated in the embryonic epidermis, there was a dramatic and premature increase in adipocytes in the absence of hair follicle formation, demonstrating that Wnt activation, rather than mature hair follicles, is required for adipocyte generation. Epidermal and dermal gene expression profiling identified keratinocyte-derived adipogenic factors that are induced by β-catenin activation. Wnt/β-catenin signaling-dependent secreted factors from keratinocytes promoted adipocyte differentiation in vitro, and we identified ligands for the bone morphogenetic protein and insulin pathways as proadipogenic factors. Our results indicate epidermal Wnt/β-catenin as a critical initiator of a signaling cascade that induces adipogenesis and highlight the role of epidermal Wnt signaling in synchronizing adipocyte differentiation with the hair growth cycle. PMID:24706781

  17. c-Jun regulates adipocyte differentiation via the KLF15-mediated mode.

    PubMed

    Lee, Da Som; Choi, Hyeonjin; Han, Baek Soo; Kim, Won Kon; Lee, Sang Chul; Oh, Kyoung-Jin; Bae, Kwang-Hee

    2016-01-15

    Abnormal adipocyte differentiation is implicated in the development of metabolic disorders such as obesity and type II diabetes. Thus, an in-depth understanding of the molecular mechanisms associated with adipocyte differentiation is the first step in overcoming obesity and its related metabolic diseases. Here, we examined the role of c-Jun as a transcription factor in adipocyte differentiation. c-Jun overexpression in murine 3T3-L1 preadipocytes significantly inhibited adipocyte differentiation. In addition, the expression level of KLF15, an upstream effector of the key adipogenic factors C/EBPα and PPARγ, was decreased upon the ectopic expression of c-Jun. We found that c-Jun inhibited basal and glucocorticoid receptor (GR)-induced promoter activities of KLF15. c-Jun directly bound near the glucocorticoid response element (GRE) sites in the KLF15 promoter and inhibited adjacent promoter occupancies of GR. Furthermore, the restoration of KLF15 expression in 3T3-L1 cells with the stable ectopic expression of c-Jun partially rescued adipocyte differentiation. Our results demonstrate that c-Jun can suppress adipocyte differentiation through the down-regulation of KLF15 at the transcriptional level. This study proposes a novel mechanism by which c-Jun regulates adipocyte differentiation. PMID:26692489

  18. Fat intake leads to differential response of rat adipocytes to glucose, insulin and ascorbic acid.

    PubMed

    Garcia-Diaz, Diego F; Campion, Javier; Arellano, Arianna V; Milagro, Fermin I; Moreno-Aliaga, Maria J; Martinez, J Alfredo

    2012-04-01

    Antioxidant-based treatments have emerged as novel and interesting approaches to counteract fat accumulation in obesity and associated metabolic disturbances. Adipocytes from rats that were fed on chow or high-fat diet (HFD) for 50 d were isolated (primary adipocytes) and incubated (72 h) on low (LG; 5.6 mmol/L) or high (HG; 25 mmol/L) glucose levels, in the presence or absence of 1.6 nmol/L insulin and 200 μmol/L vitamin C (VC). Adipocytes from HFD-fed animals presented lower insulin-induced glucose uptake, lower lactate and glycerol release, and lower insulin-induced secretion of some adipokines as compared with controls. HG treatment restored the blunted response to insulin regarding apelin secretion in adipocytes from HFD-fed rats. VC treatment inhibited the levels of nearly all variables, irrespective of the adipocytes' dietary origin. The HG treatment reduced adipocyte viability, and VC protected from this toxic effect, although more drastically in control adipocytes. Summing up, in vivo chow or HFD intake determines a differential response to insulin and glucose treatments that appears to be dependent on the insulin-resistance status of the adipocytes, while VC modifies some responses from adipocytes independently of the previous dietary intake of the animals. PMID:22454546

  19. ATF3 represses PPARγ expression and inhibits adipocyte differentiation

    SciTech Connect

    Jang, Min-Kyung; Jung, Myeong Ho

    2014-11-07

    Highlights: • ATF3 decrease the expression of PPARγ and its target gene in 3T3-L1 adipocytes. • ATF3 represses the promoter activity of PPARγ2 gene. • ATF/CRE (−1537/−1530) is critical for ATF3-mediated downregulation of PPARγ. • ATF3 binds to the promoter region containing the ATF/CRE. • ER stress inhibits adipocyte differentiation through downregulation of PPARγ by ATF3. - Abstract: Activating transcription factor 3 (ATF3) is a stress-adaptive transcription factor that mediates cellular stress response signaling. We previously reported that ATF3 represses CCAAT/enhancer binding protein α (C/EBPα) expression and inhibits 3T3-L1 adipocyte differentiation. In this study, we explored potential role of ATF3 in negatively regulating peroxisome proliferator activated receptor-γ (PPARγ). ATF3 decreased the expression of PPARγ and its target gene in 3T3-L1 adipocytes. ATF3 also repressed the activity of −2.6 Kb promoter of mouse PPARγ2. Overexpression of PPARγ significantly prevented the ATF3-mediated inhibition of 3T3-L1 differentiation. Transfection studies with 5′ deleted-reporters showed that ATF3 repressed the activity of −2037 bp promoter, whereas it did not affect the activity of −1458 bp promoter, suggesting that ATF3 responsive element is located between the −2037 and −1458. An electrophoretic mobility shift assay and chromatin immunoprecipitation assay demonstrated that ATF3 binds to ATF/CRE site (5′-TGACGTTT-3′) between −1537 and −1530. Mutation of the ATF/CRE site abrogated ATF3-mediated transrepression of the PPARγ2 promoter. Treatment with thapsigargin, endoplasmic reticulum (ER) stress inducer, increased ATF3 expression, whereas it decreased PPARγ expression. ATF3 knockdown significantly blocked the thapsigargin-mediated downregulation of PPARγ expression. Furthermore, overexpression of PPARγ prevented inhibition of 3T3-L1 differentiation by thapsigargin. Collectively, these results suggest that ATF3-mediated

  20. MiR-21a-5p suppresses bisphenol A-induced pre-adipocyte differentiation by targeting map2k3 through MKK3/p38/MAPK.

    PubMed

    Xie, Xiuwen; Song, Jia; Li, Gang

    2016-04-22

    Childhood obesity is a metabolic disease characterized by accumulation of excessive fat. Bisphenol A (BPA), a potential obesogen compound, possesses an estrogen mimetic activity and endocrine disruption effect. MicroRNA-21a-5p (miR-21a-5p) is reported to regulate the adipogenic differentiation. Our study showed that miR-21a-5p overexpression significantly decreased the red lipid droplets and triglyceride level in BPA-induced 3T3-L1 cells. BPA induced the mRNA and protein expression levels of PPARγ, C/EBPα and adiponectin, and the induction was inhibited by miR-21a-5p mimics transfection. MiR-21a-5p mimics inhibited the GR activity, GR phosphorylation (S220, S21a-5p2, and S234), and the activation of p38/MAPK pathway, which are elevated by BPA treatment in 3T3-L1 cells. MiR-21a-5p overexpression inhibited the protein level of MKK3, but not in the mRNA level. Luciferase activity assay showed that miR-21a-5p directly targeted map2k3 3'-UTR. MKK3 overexpression attenuated the effect of miR-21a-5p mimics transfection on 3T3-L1 differentiation. We also assessed the body weight, fat mass and the content of serum lipid in rats subcutaneous injected with BPA and miR-21a-5p mimics. MiR-21a-5p overexpression attenuated BPA-induced obesity in vivo. These findings suggested that miR-21a-5p inhibited BPA induced adipocyte differentiation by targeting map2k3 through MKK3/p38/MAPK in 3T3-L1 cells, providing a potential therapeutic strategy for BPA induced obesity. PMID:26996129

  1. Adipocyte-Like Differentiation in a Posttreatment Embryonal Rhabdomyosarcoma

    PubMed Central

    Balitzer, Dana; McCalmont, Timothy H.; Horvai, Andrew E.

    2015-01-01

    We describe a 16-year-old boy with rhabdomyosarcoma, consistent with embryonal subtype, of the lower extremity who received systemic neoadjuvant chemotherapy and subsequent excision. Microscopic sections of the postchemotherapy excision demonstrated diffuse, prominent, and immature adipocyte-like differentiation, in addition to skeletal muscle differentiation. Adipocyte-like differentiation was confirmed by a combination of positive Oil Red O and adipophilin immunohistochemical staining. To our knowledge, this represents the first report of an unusual phenomenon of differentiation of a soft tissue rhabdomyosarcoma into adipocyte-like cells after chemotherapy. PMID:26783483

  2. The effect of dehydroleucodine in adipocyte differentiation.

    PubMed Central

    Galvis, Adriana; Marcano, Adriana; Stefancin, Chad; Villaverde, Nicole; Priestap, Horacio A.; Tonn, Carlos E.; Lopez, Luis A.; Barbieri, Manuel A.

    2012-01-01

    Dehydroleucodine (DhL) is a sesquiterpene lactone of the guaianolide group with gastric citoprotective activity. Recent studies have also demonstrated that DhL inhibits the proliferation of vascular smooth muscle cells. In this study we examined the effect of DhL in the differentiation 3T3-L1 preadipocytes. The addition of DhL significantly inhibited the differentiation 3T3-L1 preadipocytes along with significant decrease in the accumulation of lipid content by a dramatic down regulation of the expression of adipogenic-specific transcriptional factors PPARγ and C-EBPα. However, phosphorylation of AMPKα, Erk1/2 and Akt1 was not inhibited by DhL treatment. Interestingly, we also found that 11,13-dihydro-dehydroleucodine, a derivative of DhL with inactivated α-methylene-γ-lactone function, also inhibited the differentiation 3T3-L1 preadipocytes. Taken together, these data suggest DhL has an important inhibitory effect in cellular pathways regulating adipocyte differentiation by modulating the PPARγ expression, which is known to play a pivotal role during adipogenesis. PMID:21963454

  3. Cell line models of differentiation: preadipocytes and adipocytes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The intense study of adipocyte biology spurred by interest in regulating body composition and metabolism has given rise to a number of in vitro cell models. These in vitro models have been invaluable in determining the mechanisms involved in adipocyte differentiation. In addition in vitro cell sys...

  4. Participation of TNF-α in Inhibitory Effects of Adipocytes on Osteoblast Differentiation.

    PubMed

    Abuna, Robrigo P F; De Oliveira, Fabiola S; Santos, Thiago De S; Guerra, Thais R; Rosa, Adalberto L; Beloti, Marcio M

    2016-01-01

    Mesenchymal stem cells from bone marrow (BM-MSCs) and adipose tissue (AT-MSCs) are attractive tools for cell-based therapies to repair bone tissue. In this study, we investigated the osteogenic and adipogenic potential of BM-MSCs and AT-MSCs as well as the effect of crosstalk between osteoblasts and adipocytes on cell phenotype expression. Rat BM-MSCs and AT-MSCs were cultured either in growth, osteogenic, or adipogenic medium to evaluate osteoblast and adipocyte differentiation. Additionally, osteoblasts and adipocytes were indirectly co-cultured to investigate the effect of adipocytes on osteoblast differentiation and vice versa. BM-MSCs and AT-MSCs exhibit osteogenic and adipogenic potential under non-differentiation-inducing conditions. When exposed to osteogenic medium, BM-MSCs exhibited higher expression of bone markers compared with AT-MSCs. Conversely, under adipogenic conditions, AT-MSCs displayed higher expression of adipose tissue markers compared with BM-MSCs. The presence of adipocytes as indirect co-culture repressed the expression of the osteoblast phenotype, whereas osteoblasts did not exert remarkable effect on adipocytes. The inhibitory effect of adipocytes on osteoblasts was due to the release of tumor necrosis factor alpha (TNF-α) in culture medium by adipocytes. Indeed, the addition of exogenous TNF-α in culture medium repressed the differentiation of BM-MSCs into osteoblasts mimicking the indirect co-culture effect. In conclusion, our study showed that BM-MSCs are more osteogenic while AT-MSCs are more adipogenic. Additionally, we demonstrated the key role of TNF-α secreted by adipocytes on the inhibition of osteoblast differentiation. Thus, we postulate that the higher osteogenic potential of BM-MSCs makes them the first choice for inducing bone repair in cell-based therapies. PMID:26059069

  5. Using a 3D Culture System to Differentiate Visceral Adipocytes In Vitro.

    PubMed

    Emont, Margo P; Yu, Hui; Jun, Heejin; Hong, Xiaowei; Maganti, Nenita; Stegemann, Jan P; Wu, Jun

    2015-12-01

    It has long been recognized that body fat distribution and regional adiposity play a major role in the control of metabolic homeostasis. However, the ability to study and compare the cell autonomous regulation and response of adipocytes from different fat depots has been hampered by the difficulty of inducing preadipocytes isolated from the visceral depot to differentiate into mature adipocytes in culture. Here, we present an easily created 3-dimensional (3D) culture system that can be used to differentiate preadipocytes from the visceral depot as robustly as those from the sc depot. The cells differentiated in these 3D collagen gels are mature adipocytes that retain depot-specific characteristics, as determined by imaging, gene expression, and functional assays. This 3D culture system therefore allows for study of the development and function of adipocytes from both depots in vitro and may ultimately lead to a greater understanding of site-specific functional differences of adipose tissues to metabolic dysregulation. PMID:26425808

  6. Characterization of actions of octanoate on porcine preadipocytes and adipocytes differentiated in vitro.

    PubMed

    Suzuki, Shunichi; Suzuki, Misae; Sembon, Shoichiro; Fuchimoto, Daiichiro; Onishi, Akira

    2013-03-01

    Octanoate is used to induce adipogenic differentiation and/or lipid accumulation in preadipocytes of domestic animals. However, information on detailed actions of octanoate and the characteristics of octanoate-induced adipocytes is limited. The aim of this study was to examine these issues by comparing the outcomes of the effects of octanoate with those of rosiglitazone, which is a well-defined activator of peroxisome proliferator-activated receptor (PPAR)-γ. The adipocytes that were differentiated with 5mM of octanoate had dispersed and diversely sized lipid droplets compared to those that were differentiated with 1 μM of rosiglitazone. The gene expression levels of adiponectin, glycerol-3-phosphate dehydrogenase, perilipin 1, and perilipin 4 were much higher in the adipocytes that were differentiated with rosiglitazone than in those differentiated with octanoate, while the gene expression levels of lipoprotein lipase and perilipin 2 were decreased in rosiglitazone-differentiated adipocytes compared to octanoate-differentiated adipocytes. However, the expressions of aP2 and CD36 genes were comparably induced. Luciferase reporter assays revealed that PPAR and liver-X-receptor activities were upregulated by octanoate more effectively than by rosiglitazone. Overall, these results suggested that the action of octanoate was complicated and may be dependent on the targeted genes and cellular status. PMID:23376076

  7. Differentiation of preadipocytes and mature adipocytes requires PSMB8

    PubMed Central

    Arimochi, Hideki; Sasaki, Yuki; Kitamura, Akiko; Yasutomo, Koji

    2016-01-01

    The differentiation of adipocytes is tightly regulated by a variety of intrinsic molecules and also by extrinsic molecules produced by adjacent cells. Dysfunction of adipocyte differentiation causes lipodystrophy, which impairs glucose and lipid homeostasis. Although dysfunction of immunoproteasomes causes partial lipodystrophy, the detailed molecular mechanisms remain to be determined. Here, we demonstrate that Psmb8, a catalytic subunit for immunoproteasomes, directly regulates the differentiation of preadipocytes and additionally the differentiation of preadipocytes to mature adipocytes. Psmb8−/− mice exhibited slower weight gain than wild-type mice, and this was accompanied by reduced adipose tissue volume and smaller size of mature adipocytes compared with controls. Blockade of Psmb8 activity in 3T3-L1 cells disturbed the differentiation to mature adipocytes. Psmb8−/− mice had fewer preadipocyte precursors, fewer preadipocytes and a reduced ability to differentiate preadipocytes toward mature adipocytes. Our data demonstrate that Psmb8-mediated immunoproteasome activity is a direct regulator of the differentiation of preadipocytes and their ultimate maturation. PMID:27225296

  8. Differentiation of preadipocytes and mature adipocytes requires PSMB8.

    PubMed

    Arimochi, Hideki; Sasaki, Yuki; Kitamura, Akiko; Yasutomo, Koji

    2016-01-01

    The differentiation of adipocytes is tightly regulated by a variety of intrinsic molecules and also by extrinsic molecules produced by adjacent cells. Dysfunction of adipocyte differentiation causes lipodystrophy, which impairs glucose and lipid homeostasis. Although dysfunction of immunoproteasomes causes partial lipodystrophy, the detailed molecular mechanisms remain to be determined. Here, we demonstrate that Psmb8, a catalytic subunit for immunoproteasomes, directly regulates the differentiation of preadipocytes and additionally the differentiation of preadipocytes to mature adipocytes. Psmb8(-/-) mice exhibited slower weight gain than wild-type mice, and this was accompanied by reduced adipose tissue volume and smaller size of mature adipocytes compared with controls. Blockade of Psmb8 activity in 3T3-L1 cells disturbed the differentiation to mature adipocytes. Psmb8(-/-) mice had fewer preadipocyte precursors, fewer preadipocytes and a reduced ability to differentiate preadipocytes toward mature adipocytes. Our data demonstrate that Psmb8-mediated immunoproteasome activity is a direct regulator of the differentiation of preadipocytes and their ultimate maturation. PMID:27225296

  9. Characterization of lipid metabolism in insulin-sensitive adipocytes differentiated from immortalized human mesenchymal stem cells

    SciTech Connect

    Prawitt, Janne; Niemeier, Andreas; Kassem, Moustapha; Beisiegel, Ulrike; Heeren, Joerg

    2008-02-15

    There is a great demand for cell models to study human adipocyte function. Here we describe the adipogenic differentiation of a telomerase-immortalized human mesenchymal stem cell line (hMSC-Tert) that maintains numerous features of terminally differentiated adipocytes even after prolonged withdrawal of the peroxisome proliferator activated receptor {gamma} (PPAR{gamma}) agonist rosiglitazone. Differentiated hMSC-Tert developed the characteristic monolocular phenotype of mature adipocytes. The expression of adipocyte specific markers was highly increased during differentiation. Most importantly, the presence of the PPAR{gamma} agonist rosiglitazone was not required for the stable expression of lipoprotein lipase, adipocyte fatty acid binding protein and perilipin on mRNA and protein levels. Adiponectin expression was post-transcriptionally down-regulated in the absence of rosiglitazone. Insulin sensitivity as measured by insulin-induced phosphorylation of Akt and S6 ribosomal protein was also independent of rosiglitazone. In addition to commonly used adipogenic markers, we investigated further PPAR{gamma}-stimulated proteins with a role in lipid metabolism. We observed an increase of lipoprotein receptor (VLDLR, LRP1) and apolipoprotein E expression during differentiation. Despite this increased expression, the receptor-mediated endocytosis of lipoproteins was decreased in differentiated adipocytes, suggesting that these proteins may have an additional function in adipose tissue beyond lipoprotein uptake.

  10. The adipokine Chemerin induces lipolysis and adipogenesis in bovine intramuscular adipocytes.

    PubMed

    Fu, Yuan-Yuan; Chen, Kun-Lin; Li, Hui-Xia; Zhou, Guang-Hong

    2016-07-01

    The adipokine Chemerin is reported to regulate adipogenesis and glucose homeostasis in vivo and in 3T3-L1 cells. Our team is focused on the role of Chemerin in metabolism and intramuscular adipocyte differentiation because intramuscular fat is the basic material for the formation of marbling in livestock and poultry meat. In this study, bovine intramuscular mature adipocytes were cultured in medium with Chemerin, and the process of lipolysis of mature adipocytes and the adipogenesis of de-differentiated preadipocytes were investigated. The results showed that Chemerin induced significant lipolytic metabolism in intramuscular mature adipocytes, indicated by increased levels of glycerol, FFA, and up-regulated expression of the lipolysis critical factors HSL, LPL, and leptin. Meanwhile, the expressions of adipogenic key factors PPARγ, C/EBPα, and A-FABP were decreased by Chemerin during lipolysis or dedifferentiation in mature adipocytes. The de-differentiated preadipocytes could re-differentiate into mature adipocytes. Intriguingly, the formation of cells' lipid droplets was promoted by Chemerin during preadipocyte differentiation. In addition, mRNA and protein expressions of PPARγ, C/EBPα, and A-FABP were up-regulated by Chemerin during preadipocytes differentiation. These results suggest that Chemerin promotes lipolysis in mature adipocytes and induces adipogenesis during preadipocyte re-differentiation, further indicating a dual role for Chemerin in the deposition of intramuscular fat in ruminant animals. PMID:27260300

  11. Isoform-specific regulation of adipocyte differentiation by Akt/protein kinase B{alpha}

    SciTech Connect

    Yun, Sung-Ji; Kim, Eun-Kyoung; Tucker, David F.; Kim, Chi Dae; Birnbaum, Morris J.; Bae, Sun Sik

    2008-06-20

    The phosphatidylinositol 3-kinase (PI3K)/Akt pathway tightly regulates adipose cell differentiation. Here we show that loss of Akt1/PKB{alpha} in primary mouse embryo fibroblast (MEF) cells results in a defect of adipocyte differentiation. Adipocyte differentiation in vitro and ex vivo was restored in cells lacking both Akt1/PKB{alpha} and Akt2/PKB{beta} by ectopic expression of Akt1/PKB{alpha} but not Akt2/PKB{beta}. Akt1/PKB{alpha} was found to be the major regulator of phosphorylation and nuclear export of FoxO1, whose presence in the nucleus strongly attenuates adipocyte differentiation. Differentiation-induced cell division was significantly abrogated in Akt1/PKB{alpha}-deficient cells, but was restored after forced expression of Akt1/PKB{alpha}. Moreover, expression of p27{sup Kip1}, an inhibitor of the cell cycle, was down regulated in an Akt1/PKB{alpha}-specific manner during adipocyte differentiation. Based on these data, we suggest that the Akt1/PKB{alpha} isoform plays a major role in adipocyte differentiation by regulating FoxO1 and p27{sup Kip1}.

  12. Branched chain amino acid catabolism fuels adipocyte differentiation and lipogenesis

    PubMed Central

    Green, Courtney R.; Wallace, Martina; Divakaruni, Ajit S.; Phillips, Susan A.; Murphy, Anne N.; Ciaraldi, Theodore P.; Metallo, Christian M.

    2015-01-01

    Adipose tissue plays important roles in regulating carbohydrate and lipid homeostasis, though less is known about the regulation of amino acid metabolism in adipocytes. Here we applied isotope tracing to pre–adipocytes and differentiated adipocytes to quantify the contributions of different substrates to tricarboxylic acid metabolism and lipogenesis. In contrast to proliferating cells that use glucose and glutamine for acetyl–coenzyme A (AcCoA) generation, differentiated adipocytes increased branched chain amino acid (BCAA) catabolic flux such that leucine and isoleucine from media and/or protein catabolism accounted for as much as 30% of lipogenic AcCoA pools. Medium cobalamin deficiency caused methylmalonic acid accumulation and odd–chain fatty acid synthesis. B12 supplementation reduced these metabolites and altered the balance of substrates entering mitochondria. Finally, inhibition of BCAA catabolism compromised adipogenesis. These results quantitatively highlight the contribution of BCAAs to adipocyte metabolism and suggest that BCAA catabolism plays a functional role in adipocyte differentiation. PMID:26571352

  13. Phytanic acid, a novel activator of uncoupling protein-1 gene transcription and brown adipocyte differentiation.

    PubMed Central

    Schlüter, Agatha; Barberá, Maria José; Iglesias, Roser; Giralt, Marta; Villarroya, Francesc

    2002-01-01

    Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) is a phytol-derived branched-chain fatty acid present in dietary products. Phytanic acid increased uncoupling protein-1 (UCP1) mRNA expression in brown adipocytes differentiated in culture. Phytanic acid induced the expression of the UCP1 gene promoter, which was enhanced by co-transfection with a retinoid X receptor (RXR) expression vector but not with other expression vectors driving peroxisome proliferator-activated receptor (PPAR)alpha, PPARgamma or a form of RXR devoid of ligand-dependent sensitivity. The effect of phytanic acid on the UCP1 gene required the 5' enhancer region of the gene and the effects of phytanic acid were mediated in an additive manner by three binding sites for RXR. Moreover, phytanic acid activates brown adipocyte differentiation: long-term exposure of brown preadipocytes to phytanic acid promoted the acquisition of the brown adipocyte morphology and caused a co-ordinate induction of the mRNAs for gene markers of brown adipocyte differentiation, such as UCP1, adipocyte lipid-binding protein aP2, lipoprotein lipase, the glucose transporter GLUT4 or subunit II of cytochrome c oxidase. In conclusion, phytanic acid is a natural product of phytol metabolism that activates brown adipocyte thermogenic function. It constitutes a potential nutritional signal linking dietary status to adaptive thermogenesis. PMID:11829740

  14. Amelioration of Mitochondrial Dysfunction-Induced Insulin Resistance in Differentiated 3T3-L1 Adipocytes via Inhibition of NF-κB Pathways

    PubMed Central

    Hafizi Abu Bakar, Mohamad; Sarmidi, Mohamad Roji; Kai, Cheng Kian; Huri, Hasniza Zaman; Yaakob, Harisun

    2014-01-01

    A growing body of evidence suggests that activation of nuclear factor kappa B (NF-κB) signaling pathways is among the inflammatory mechanism involved in the development of insulin resistance and chronic low-grade inflammation in adipose tissues derived from obese animal and human subjects. Nevertheless, little is known about the roles of NF-κB pathways in regulating mitochondrial function of the adipose tissues. In the present study, we sought to investigate the direct effects of celastrol (potent NF-κB inhibitor) upon mitochondrial dysfunction-induced insulin resistance in 3T3-L1 adipocytes. Celastrol ameliorates mitochondrial dysfunction by altering mitochondrial fusion and fission in adipocytes. The levels of oxidative DNA damage, protein carbonylation and lipid peroxidation were down-regulated. Further, the morphology and quantification of intracellular lipid droplets revealed the decrease of intracellular lipid accumulation with reduced lipolysis. Moreover, massive production of the pro-inflammatory mediators tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were markedly depleted. Insulin-stimulated glucose uptake activity was restored with the enhancement of insulin signaling pathways. This study signified that the treatments modulated towards knockdown of NF-κB transcription factor may counteract these metabolic insults exacerbated in our model of synergy between mitochondrial dysfunction and inflammation. These results demonstrate for the first time that NF-κB inhibition modulates mitochondrial dysfunction induced insulin resistance in 3T3-L1 adipocytes. PMID:25474091

  15. Deficiency of angiotensin type 1a receptors in adipocytes reduces differentiation and promotes hypertrophy of adipocytes in lean mice.

    PubMed

    Putnam, Kelly; Batifoulier-Yiannikouris, Frederique; Bharadwaj, Kalyani G; Lewis, Eboni; Karounos, Michael; Daugherty, Alan; Cassis, Lisa A

    2012-10-01

    Adipocytes express angiotensin receptors, but the direct effects of angiotensin II (AngII) stimulating this cell type are undefined. Adipocytes express angiotensin type 1a receptor (AT1aR) and AT2R, both of which have been implicated in obesity. In this study, we determined the effects of adipocyte AT1aR deficiency on adipocyte differentiation and the development of obesity in mice fed low-fat (LF) or high-fat (HF) diets. Mice expressing Cre recombinase under the control of the aP2 promoter were bred with AT1aR-floxed mice to generate mice with adipocyte AT1aR deficiency (AT1aR(aP2)). AT1aR mRNA abundance was reduced significantly in both white and brown adipose tissue from AT1aR(aP2) mice compared with nontransgenic littermates (AT1aR(fl/fl)). Adipocyte AT1aR deficiency did not influence body weight, glucose tolerance, or blood pressure in mice fed either LF or high-fat diets. However, LF-fed AT1aR(aP2) mice exhibited striking adipocyte hypertrophy even though total fat mass was not different between genotypes. Stromal vascular cells from AT1aR(aP2) mice differentiated to a lesser extent to adipocytes compared with controls. Conversely, incubation of 3T3-L1 adipocytes with AngII increased Oil Red O staining and increased mRNA abundance of peroxisome proliferator-activated receptor γ (PPARγ) via AT1R stimulation. These results suggest that reductions in adipocyte differentiation in LF-fed AT1aR(aP2) mice resulted in increased lipid storage and hypertrophy of remaining adipocytes. These results demonstrate that AngII regulates adipocyte differentiation and morphology through the adipocyte AT1aR in lean mice. PMID:22919058

  16. Gallic Acid, the Active Ingredient of Terminalia bellirica, Enhances Adipocyte Differentiation and Adiponectin Secretion.

    PubMed

    Makihara, Hiroko; Koike, Yuka; Ohta, Masatomi; Horiguchi-Babamoto, Emi; Tsubata, Masahito; Kinoshita, Kaoru; Akase, Tomoko; Goshima, Yoshio; Aburada, Masaki; Shimada, Tsutomu

    2016-01-01

    Visceral obesity induces the onset of metabolic disorders such as insulin resistance and diabetes mellitus. Adipose tissue is considered as a potential pharmacological target for treating metabolic disorders. The fruit of Terminalia bellirica is extensively used in Ayurvedic medicine to treat patients with diseases such as diabetes mellitus. We previously investigated the effects of a hot water extract of T. bellirica fruit (TB) on obesity and insulin resistance in spontaneously obese type 2 diabetic mice. To determine the active ingredients of TB and their molecular mechanisms, we focused on adipocyte differentiation using mouse 3T3-L1 cells, which are widely used to study adipocyte physiology. We show here that TB enhanced the differentiation of 3T3-L1 cells to mature adipocytes and that one of the active main components was identified as gallic acid. Gallic acid (10-30 µM) enhanced the expression and secretion of adiponectin via adipocyte differentiation and also that of fatty acid binding protein-4, which is the target of peroxisome proliferator-activated receptor gamma (PPARγ), although it does not alter the expression of the upstream genes PPARγ and CCAAT enhancer binding protein alpha. In the PPARγ ligand assay, the binding of gallic acid to PPARγ was undetectable. These findings indicate that gallic acid mediates the therapeutic effects of TB on metabolic disorders by regulating adipocyte differentiation. Therefore, TB shows promise as a candidate for preventing and treating patients with metabolic syndrome. PMID:27374289

  17. Thioredoxin reductase 1 suppresses adipocyte differentiation and insulin responsiveness

    PubMed Central

    Peng, Xiaoxiao; Giménez-Cassina, Alfredo; Petrus, Paul; Conrad, Marcus; Rydén, Mikael; Arnér, Elias S. J.

    2016-01-01

    Recently thioredoxin reductase 1 (TrxR1), encoded by Txnrd1, was suggested to modulate glucose and lipid metabolism in mice. Here we discovered that TrxR1 suppresses insulin responsiveness, anabolic metabolism and adipocyte differentiation. Immortalized mouse embryonic fibroblasts (MEFs) lacking Txnrd1 (Txnrd1−/−) displayed increased metabolic flux, glycogen storage, lipogenesis and adipogenesis. This phenotype coincided with upregulated PPARγ expression, promotion of mitotic clonal expansion and downregulation of p27 and p53. Enhanced Akt activation also contributed to augmented adipogenesis and insulin sensitivity. Knockdown of TXNRD1 transcripts accelerated adipocyte differentiation also in human primary preadipocytes. Furthermore, TXNRD1 transcript levels in subcutaneous adipose tissue from 56 women were inversely associated with insulin sensitivity in vivo and lipogenesis in their isolated adipocytes. These results suggest that TrxR1 suppresses anabolic metabolism and adipogenesis by inhibition of intracellular signaling pathways downstream of insulin stimulation. PMID:27346647

  18. Brazilein from Caesalpinia sappan L. Antioxidant Inhibits Adipocyte Differentiation and Induces Apoptosis through Caspase-3 Activity and Anthelmintic Activities against Hymenolepis nana and Anisakis simplex

    PubMed Central

    Liang, Chia-Hua; Chan, Leong-Perng; Chou, Tzung-Han; Chiang, Feng-Yu; Yen, Chuan-Min; Chen, Pin-Ju; Ding, Hsiou-Yu

    2013-01-01

    Brazilein, a natural, biologically active compound from Caesalpinia sappan L., has been shown to exhibit anti-inflammatory and antioxidant properties and to inhibit the growth of several cancer cells. This study verifies the antioxidant and antitumor characteristics of brazilein in skin cancer cells and is the first time to elucidate the inhibition mechanism of adipocyte differentiation, cestocidal activities against Hymenolepis nana, and reduction of spontaneous movement in Anisakis simplex. Brazilein exhibits an antioxidant capacity as well as the ability to scavenge DPPH• and ABTS•+ free radicals and to inhibit lipid peroxidation. Brazilein inhibited intracellular lipid accumulation during adipocyte differentiation in 3T3-L1 cells and suppressed the induction of peroxisome proliferator-activated receptor γ (PPARγ), the master regulator of adipogenesis, suggesting that brazilein presents the antiobesity effects. The toxic effects of brazilein were evaluated in terms of cell viability, induction of apoptosis, and the activity of caspase-3 in BCC cells. The inhibition of the growth of skin cancer cells (A431, BCC, and SCC25) by brazilein is greater than that of human skin malignant melanoma (A375) cells, mouse leukemic monocyte macrophage (RAW 264.7 cells), and noncancerous cells (HaCaT and BNLCL2 cells). The anthelmintic activities of brazilein against Hymenolepis nana are better than those of Anisakis simplex. PMID:23554834

  19. Differential Chemokine Signature between Human Preadipocytes and Adipocytes

    PubMed Central

    Ignacio, Rosa Mistica C.; Gibbs, Carla R.; Lee, Eun-Sook

    2016-01-01

    Obesity is characterized as an accumulation of adipose tissue mass represented by chronic, low-grade inflammation. Obesity-derived inflammation involves chemokines as important regulators contributing to the pathophysiology of obesity-related diseases such as cardiovascular disease, diabetes and some cancers. The obesity-driven chemokine network is poorly understood. Here, we identified the profiles of chemokine signature between human preadipocytes and adipocytes, using PCR arrays and qRT-PCR. Both preadipocytes and adipocytes showed absent or low levels in chemokine receptors in spite of some changes. On the other hand, the chemokine levels of CCL2, CCL7-8, CCL11, CXCL1-3, CXCL6 and CXCL10-11 were dominantly expressed in preadipocytes compared to adipocytes. Interestingly, CXCL14 was the most dominant chemokine expressed in adipocytes compared to preadipocytes. Moreover, there is significantly higher protein level of CXCL14 in conditioned media from adipocytes. In addition, we analyzed the data of the chemokine signatures in adipocytes obtained from healthy lean and obese postmenopausal women based on Gene Expression Omnibus (GEO) dataset. Adipocytes from obese individuals had significantly higher levels in chemokine signature as follows: CCL2, CCL13, CCL18-19, CCL23, CCL26, CXCL1, CXCL3 and CXCL14, as compared to those from lean ones. Also, among the chemokine networks, CXCL14 appeared to be the highest levels in adipocytes from both lean and obese women. Taken together, these results identify CXCL14 as an important chemokine induced during adipogenesis, requiring further research elucidating its potential therapeutic benefits in obesity. PMID:27340388

  20. Differential Chemokine Signature between Human Preadipocytes and Adipocytes.

    PubMed

    Ignacio, Rosa Mistica C; Gibbs, Carla R; Lee, Eun-Sook; Son, Deok-Soo

    2016-06-01

    Obesity is characterized as an accumulation of adipose tissue mass represented by chronic, low-grade inflammation. Obesity-derived inflammation involves chemokines as important regulators contributing to the pathophysiology of obesity-related diseases such as cardiovascular disease, diabetes and some cancers. The obesity-driven chemokine network is poorly understood. Here, we identified the profiles of chemokine signature between human preadipocytes and adipocytes, using PCR arrays and qRT-PCR. Both preadipocytes and adipocytes showed absent or low levels in chemokine receptors in spite of some changes. On the other hand, the chemokine levels of CCL2, CCL7-8, CCL11, CXCL1-3, CXCL6 and CXCL10-11 were dominantly expressed in preadipocytes compared to adipocytes. Interestingly, CXCL14 was the most dominant chemokine expressed in adipocytes compared to preadipocytes. Moreover, there is significantly higher protein level of CXCL14 in conditioned media from adipocytes. In addition, we analyzed the data of the chemokine signatures in adipocytes obtained from healthy lean and obese postmenopausal women based on Gene Expression Omnibus (GEO) dataset. Adipocytes from obese individuals had significantly higher levels in chemokine signature as follows: CCL2, CCL13, CCL18-19, CCL23, CCL26, CXCL1, CXCL3 and CXCL14, as compared to those from lean ones. Also, among the chemokine networks, CXCL14 appeared to be the highest levels in adipocytes from both lean and obese women. Taken together, these results identify CXCL14 as an important chemokine induced during adipogenesis, requiring further research elucidating its potential therapeutic benefits in obesity. PMID:27340388

  1. Characterization of actions of octanoate on porcine preadipocytes and adipocytes differentiated in vitro

    SciTech Connect

    Suzuki, Shunichi; Suzuki, Misae; Sembon, Shoichiro; Fuchimoto, Daiichiro; Onishi, Akira

    2013-03-01

    Highlights: ► Octanoate regulated gene expressions in a way distinct from rosiglitasone. ► Octanoate upregulatedPPRE and LXRE reporter activities. ► Octanoate may act on some PPARγ-target genes competitively with other ligands. - Abstract: Octanoate is used to induce adipogenic differentiation and/or lipid accumulation in preadipocytes of domestic animals. However, information on detailed actions of octanoate and the characteristics of octanoate-induced adipocytes is limited. The aim of this study was to examine these issues by comparing the outcomes of the effects of octanoate with those of rosiglitazone, which is a well-defined activator of peroxisome proliferator-activated receptor (PPAR)-γ. The adipocytes that were differentiated with 5 mM of octanoate had dispersed and diversely sized lipid droplets compared to those that were differentiated with 1 μM of rosiglitazone. The gene expression levels of adiponectin, glycerol-3-phosphate dehydrogenase, perilipin 1, and perilipin 4 were much higher in the adipocytes that were differentiated with rosiglitazone than in those differentiated with octanoate, while the gene expression levels of lipoprotein lipase and perilipin 2 were decreased in rosiglitazone-differentiated adipocytes compared to octanoate-differentiated adipocytes. However, the expressions of aP2 and CD36 genes were comparably induced. Luciferase reporter assays revealed that PPAR and liver-X-receptor activities were upregulated by octanoate more effectively than by rosiglitazone. Overall, these results suggested that the action of octanoate was complicated and may be dependent on the targeted genes and cellular status.

  2. Coprinus comatus Cap Inhibits Adipocyte Differentiation via Regulation of PPARγ and Akt Signaling Pathway

    PubMed Central

    Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  3. Coprinus comatus cap inhibits adipocyte differentiation via regulation of PPARγ and Akt signaling pathway.

    PubMed

    Park, Hyoung Joon; Yun, Jisoo; Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  4. Between brown and white: novel aspects of adipocyte differentiation.

    PubMed

    Cinti, Saverio

    2011-03-01

    In all mammals including humans, most white and brown adipocytes are found together in visceral and subcutaneous depots (adipose organ) despite the well known difference in their function, respectively of storing energy and producing heat. A growing body of evidence suggests that the reason for such anatomical arrangement is their plasticity, which under appropriate stimulation allows direct conversion of one cell type into the other. In conditions of chronic cold exposure white-to-brown conversion meets the need for thermogenesis, whereas an obesogenic diet induces brown-to-white conversion to meet the need for storing energy. White-to-brown transdifferentiation is of medical interest, because the brown phenotype of the adipose organ is associated to obesity resistance, and drugs inducing this phenotype curb murine obesity and related disorders. Type 2 diabetes is the most common disorder associated to visceral obesity. Macrophages infiltrating the adipose organ are responsible for the low-grade chronic inflammation related to the removal of dead adipocytes, which leads to insulin resistance and T2 diabetes. Adipocyte death is closely related to their growth up to the critical death size. The critical death size of visceral adipocytes is smaller than that of subcutaneous adipocytes, likely accounting for the greater morbidity related to visceral fat. PMID:21254898

  5. Interferon γ Attenuates Insulin Signaling, Lipid Storage, and Differentiation in Human Adipocytes via Activation of the JAK/STAT Pathway*

    PubMed Central

    McGillicuddy, Fiona C.; Chiquoine, Elise H.; Hinkle, Christine C.; Kim, Roy J.; Shah, Rachana; Roche, Helen M.; Smyth, Emer M.; Reilly, Muredach P.

    2009-01-01

    Recent reports demonstrate T-cell infiltration of adipose tissue in early obesity. We hypothesized that interferon (IFN) γ, a major T-cell inflammatory cytokine, would attenuate human adipocyte functions and sought to establish signaling mechanisms. Differentiated human adipocytes were treated with IFNγ ± pharmacological inhibitors prior to insulin stimulation. [3H]Glucose uptake and AKT phosphorylation were assessed as markers of insulin sensitivity. IFNγ induced sustained loss of insulin-stimulated glucose uptake in human adipocytes, coincident with reduced Akt phosphorylation and down-regulation of the insulin receptor, insulin receptor substrate-1, and GLUT4. Loss of adipocyte triglyceride storage was observed with IFNγ co-incident with reduced expression of peroxisome proliferator-activated receptor γ, adiponectin, perilipin, fatty acid synthase, and lipoprotein lipase. Treatment with IFNγ also blocked differentiation of pre-adipocytes to the mature phenotype. IFNγ-induced robust STAT1 phosphorylation and SOCS1 mRNA expression, with modest, transient STAT3 phosphorylation and SOCS3 induction. Preincubation with a non-selective JAK inhibitor restored glucose uptake and Akt phosphorylation while completely reversing IFNγ suppression of adipogenic mRNAs and adipocyte differentiation. Specific inhibition of JAK2 or JAK3 failed to block IFNγ effects suggesting a predominant role for JAK1-STAT1. We demonstrate that IFNγ attenuates insulin sensitivity and suppresses differentiation in human adipocytes, an effect most likely mediated via sustained JAK-STAT1 pathway activation. PMID:19776010

  6. Stress of endoplasmic reticulum modulates differentiation and lipogenesis of human adipocytes

    SciTech Connect

    Koc, Michal; Mayerová, Veronika; Kračmerová, Jana; Mairal, Aline; Mališová, Lucia; Štich, Vladimír; Langin, Dominique; Rossmeislová, Lenka

    2015-05-08

    Background: Adipocytes are cells specialized for storage of neutral lipids. This storage capacity is dependent on lipogenesis and is diminished in obesity. The reason for the decline in lipogenic activity of adipocytes in obesity remains unknown. Recent data show that lipogenesis in liver is regulated by pathways initiated by endoplasmic reticulum stress (ERS). Thus, we aimed at investigating the effect of ERS on lipogenesis in adipose cells. Methods: Preadipocytes were isolated from subcutaneous abdominal adipose tissue from obese volunteers and in vitro differentiated into adipocytes. ERS was induced pharmacologically by thapsigargin (TG) or tunicamycin (TM). Activation of Unfolded Protein Response pathway (UPR) was monitored on the level of eIF2α phosphorylation and mRNA expression of downstream targets of UPR sensors. Adipogenic and lipogenic capacity was evaluated by Oil Red O staining, measurement of incorporation of radio-labelled glucose or acetic acid into lipids and mRNA analysis of adipogenic/lipogenic markers. Results: Exposition of adipocytes to high doses of TG (100 nM) and TM (1 μg/ml) for 1–24 h enhanced expression of several UPR markers (HSPA5, EDEM1, ATF4, XBP1s) and phosphorylation of eIF2α. This acute ERS substantially inhibited expression of lipogenic genes (DGAT2, FASN, SCD1) and glucose incorporation into lipids. Moreover, chronic exposure of preadipocytes to low dose of TG (2.5 nM) during the early phases of adipogenic conversion of preadipocytes impaired both, lipogenesis and adipogenesis. On the other hand, chronic low ERS had no apparent effect on lipogenesis in mature adipocytes. Conclusions: Acute ERS weakened a capacity of mature adipocytes to store lipids and chronic ERS diminished adipogenic potential of preadipocytes. - Highlights: • High intensity ERS inhibits lipogenic capacity of adipocytes. • ERS impairs adipogenesis when present in early stages of adipogenesis. • Lipogenesis in mature adipocytes is not

  7. Differentiation of human adipocytes at physiological oxygen levels results in increased adiponectin secretion and isoproterenol-stimulated lipolysis

    PubMed Central

    Famulla, Susanne; Schlich, Raphaela; Sell, Henrike; Eckel, Jürgen

    2012-01-01

    Adipose tissue (AT) hypoxia occurs in obese humans and mice. Acute hypoxia in adipocytes causes dysregulation of adipokine secretion with an increase in inflammatory factors and diminished adiponectin release. O2 levels in humans range between 3 and 11% revealing that conventional in vitro culturing at ambient air and acute hypoxia treatment (1% O2) are performed under non-physiological conditions. In this study, we mimicked physiological conditions by differentiating human primary adipocytes under 10% or 5% O2 in comparison to 21% O2. Induction of differentiation markers was comparable between all three conditions. Adipokine release by adipocytes differentiated at lower oxygen levels was altered, with a marked upregulation of adiponectin, IL-6 and DPP4 secretion, and reduced leptin levels compared with adipocytes differentiated at 21% O2. Isoproterenol-induced lipolysis was significantly elevated in adipocytes differentiated at 10% and 5% compared with 21% O2. This effect was accompanied by increased protein expression of β-1 and -2 adrenergic receptor, HSL and perilipin. Conditioned medium (CM) of adipocytes differentiated at the three different conditions was generated for stimulation of human skeletal muscle cells (SkMC) or smooth muscle cells (SMC). CM-induced insulin resistance in SkMC was comparable for the different CMs. However, the SMC proliferative effect of CM from adipocytes differentiated at 10% O2 was significantly reduced compared with 21% O2. This study demonstrates that oxygen levels during adipogenesis are important factors altering adipocyte functionality such as adipokine release, in particular adiponectin secretion, as well as the hormone-induced lipolytic pathway. PMID:23700522

  8. Glial-like differentiation potential of human mature adipocytes.

    PubMed

    Poloni, Antonella; Maurizi, Giulia; Foia, Federica; Mondini, Eleonora; Mattiucci, Domenico; Ambrogini, Patrizia; Lattanzi, Davide; Mancini, Stefania; Falconi, Massimo; Cinti, Saverio; Olivieri, Attilio; Leoni, Pietro

    2015-01-01

    The potential ability to differentiate dedifferentiated adipocytes into a neural lineage is attracting strong interest as an emerging method of producing model cells for the treatment of a variety of neurological diseases. Here, we describe the efficient conversion of dedifferentiated adipocytes into a neural-like cell population. These cells grew in neurosphere-like structures and expressed a high level of the early neuroectodermal marker Nestin. These neurospheres could proliferate and express stemness genes, suggesting that these cells could be committed to the neural lineage. After neural induction, NeuroD1, Sox1, Double Cortin, and Eno2 were not expressed. Patch clamp data did not reveal different electrophysiological properties, indicating the inability of these cells to differentiate into mature neurons. In contrast, the differentiated cells expressed a high level of CLDN11, as demonstrated using molecular method, and stained positively for the glial cell markers CLDN11 and GFAP, as demonstrated using immunocytochemistry. These data were confirmed by quantitative results for glial cell line-derived neurotrophic factor production, which showed a higher secretion level in neurospheres and the differentiated cells compared with the untreated cells. In conclusion, our data demonstrate morphological, molecular, and immunocytochemical evidence of initial neural differentiation of mature adipocytes, committing to a glial lineage. PMID:25007949

  9. Downregulation of Runx2 by 1,25-Dihydroxyvitamin D3 Induces the Transdifferentiation of Osteoblasts to Adipocytes

    PubMed Central

    Kim, Jung Ha; Seong, Semun; Kim, Kabsun; Kim, Inyoung; Jeong, Byung-Chul; Kim, Nacksung

    2016-01-01

    1,25-Dihydroxyvitamin D3 (1,25(OH)2D3) indirectly stimulates bone formation, but little is known about its direct effect on bone formation. In this study, we observed that 1,25(OH)2D3 enhances adipocyte differentiation, but inhibits osteoblast differentiation during osteogenesis. The positive role of 1,25(OH)2D3 in adipocyte differentiation was confirmed when murine osteoblasts were cultured in adipogenic medium. Additionally, 1,25(OH)2D3 enhanced the expression of adipocyte marker genes, but inhibited the expression of osteoblast marker genes in osteoblasts. The inhibition of osteoblast differentiation and promotion of adipocyte differentiation mediated by 1,25(OH)2D3 were compensated by Runx2 overexpression. Our results suggest that 1,25(OH)2D3 induces the transdifferentiation of osteoblasts to adipocytes via Runx2 downregulation in osteoblasts. PMID:27213351

  10. Inhibition of Wnt/β-catenin signaling by dexamethasone promotes adipocyte differentiation in mesenchymal progenitor cells, ROB-C26.

    PubMed

    Naito, Masako; Omoteyama, Kazuki; Mikami, Yoshikazu; Takahashi, Tomihisa; Takagi, Minoru

    2012-12-01

    Dexamethasone (Dex) stimulates the differentiation of mesenchymal progenitor cells into adipocytes and osteoblasts. However, the mechanisms underlying Dex-induced differentiation have not been clearly elucidated. We examined the effect of Dex on the expression and activity of Wnt/β-catenin signal-related molecules in a clonal mesenchymal progenitor cell line, ROB-C26 (C26). Dex induced the mRNA expression of Wnt antagonists, dickkopf-1 (Dkk-1), and Wnt inhibitory factor (WIF)-1. Immunocytochemical analysis showed that the downregulation of β-catenin protein expression by Dex occured concomitantly with the increased expression of the PPARγ protein. Dex decreased phosphorylation of Ser9-GSK3β and expression of active β-catenin protein. To examine the effects of Dex on Wnt/β-catenin activity, we used immunocytochemistry to analyze TCF/LEF-mediated transcription during Dex-induced adipogenesis in Wnt indicator (TOPEGFP) C26 cells. Our results demonstrated that Dex repressed TCF/LEF-mediated transcription, but induced adipocyte differentiation. Treatment with a GSK3β inhibitor attenuated Dex-induced inhibition of TCF/LEF-mediated transcriptional activity, but suppressed Dex-induced adipocyte differentiation, indicating that adipocyte differentiation and inhibition of Wnt/β-catenin activity by Dex are mediated by GSK3β activity. Furthermore, β-catenin knockdown not only suppressed Dex-induced ALP-positive osteoblasts differentiation but also promoted Dex-induced adipocytes differentiation. These results suggest that inhibition of β-catenin expression by Dex promotes the differentiation of mesenchymal progenitor cells into adipocytes. PMID:22886144

  11. Regulatory circuits controlling white versus brown adipocyte differentiation

    PubMed Central

    Hansen, Jacob B.; Kristiansen, Karsten

    2006-01-01

    Adipose tissue is a major endocrine organ that exerts a profound influence on whole-body homoeostasis. Two types of adipose tissue exist in mammals: WAT (white adipose tissue) and BAT (brown adipose tissue). WAT stores energy and is the largest energy reserve in mammals, whereas BAT, expressing UCP1 (uncoupling protein 1), can dissipate energy through adaptive thermogenesis. In rodents, ample evidence supports BAT as an organ counteracting obesity, whereas less is known about the presence and significance of BAT in humans. Despite the different functions of white and brown adipocytes, knowledge of factors differentially influencing the formation of white and brown fat cells is sparse. Here we summarize recent progress in the molecular understanding of white versus brown adipocyte differentiation, including novel insights into transcriptional and signal transduction pathways. Since expression of UCP1 is the hallmark of BAT and a key factor determining energy expenditure, we also review conditions associated with enhanced energy expenditure and UCP1 expression in WAT that may provide information on processes involved in brown adipocyte differentiation. PMID:16898874

  12. miR-155 regulates differentiation of brown and beige adipocytes via a bistable circuit

    PubMed Central

    Chen, Yong; Siegel, Franziska; Kipschull, Stefanie; Haas, Bodo; Fröhlich, Holger; Meister, Gunter; Pfeifer, Alexander

    2013-01-01

    Brown adipocytes are a primary site of energy expenditure and reside not only in classical brown adipose tissue but can also be found in white adipose tissue. Here we show that microRNA 155 is enriched in brown adipose tissue and is highly expressed in proliferating brown preadipocytes but declines after induction of differentiation. Interestingly, microRNA 155 and its target, the adipogenic transcription factor CCAAT/enhancer-binding protein β, form a bistable feedback loop integrating hormonal signals that regulate proliferation or differentiation. Inhibition of microRNA 155 enhances brown adipocyte differentiation and induces a brown adipocyte-like phenotype (‘browning’) in white adipocytes. Consequently, microRNA 155-deficient mice exhibit increased brown adipose tissue function and ‘browning’ of white fat tissue. In contrast, transgenic overexpression of microRNA 155 in mice causes a reduction of brown adipose tissue mass and impairment of brown adipose tissue function. These data demonstrate that the bistable loop involving microRNA 155 and CCAAT/enhancer-binding protein β regulates brown lineage commitment, thereby, controlling the development of brown and beige fat cells. PMID:23612310

  13. TMEM120A and B: Nuclear Envelope Transmembrane Proteins Important for Adipocyte Differentiation

    PubMed Central

    Batrakou, Dzmitry G.; de las Heras, Jose I.; Czapiewski, Rafal; Mouras, Rabah; Schirmer, Eric C.

    2015-01-01

    Recent work indicates that the nuclear envelope is a major signaling node for the cell that can influence tissue differentiation processes. Here we present two nuclear envelope trans-membrane proteins TMEM120A and TMEM120B that are paralogs encoded by the Tmem120A and Tmem120B genes. The TMEM120 proteins are expressed preferentially in fat and both are induced during 3T3-L1 adipocyte differentiation. Knockdown of one or the other protein altered expression of several genes required for adipocyte differentiation, Gata3, Fasn, Glut4, while knockdown of both together additionally affected Pparg and Adipoq. The double knockdown also increased the strength of effects, reducing for example Glut4 levels by 95% compared to control 3T3-L1 cells upon pharmacologically induced differentiation. Accordingly, TMEM120A and B knockdown individually and together impacted on adipocyte differentiation/metabolism as measured by lipid accumulation through binding of Oil Red O and coherent anti-Stokes Raman scattering microscopy (CARS). The nuclear envelope is linked to several lipodystrophies through mutations in lamin A; however, lamin A is widely expressed. Thus it is possible that the TMEM120A and B fat-specific nuclear envelope transmembrane proteins may play a contributory role in the tissue-specific pathology of this disorder or in the wider problem of obesity. PMID:26024229

  14. Differential regulation of pro- and antiapoptotic proteins in fish adipocytes during hypoxic conditions.

    PubMed

    Ekambaram, Padmini; Parasuraman, Parimala; Jayachandran, Tharani

    2016-06-01

    Worldwide, the frequencies and magnitudes of hypoxic events in estuarine waters have increased considerably over the past two decades. Fish populations are suitable indicators for the assessment of quality of aquatic ecosystems and often comprise a variety of adaptation systems by triggering oxidants, antioxidants and hypoxia-responsive signaling proteins. Signaling pathway may lead to cell survival or cell death which is fine-tuned by both positive and negative factors, which includes hypoxia-inducible factor-1α (HIF1α), heat-shock protein-70 (HSP70), phospho-c-Jun N-terminal kinase 1/2 (p-JNK1/2) and apoptosis signal-regulating kinase-1 (ASK1). In the present study, we attempt to determine stress-mediated signaling changes and molecular mechanism behind the cell survival by comparing adipocytes of fish from field hypoxic condition and laboratory-induced hypoxic condition (in vitro hypoxia). Comparison of field and laboratory studies in fish adipocytes showed differential expression of HIF1α, HSP70, p-JNK1/2 and ASK1 with altered oxidants and antioxidants. Further, the results also suggest that in vitro hypoxic conditions mimic field hypoxic conditions. Trends of hypoxia response were same in in vitro hypoxia of control adipocytes as in Ennore estuary, and hypoxia response was more pronounced in the test adipocytes under in vitro hypoxic condition. Results of the present work suggest that hypoxia is the major crusade of water pollutants affecting fish by differential regulation of pro- and antiapoptotic proteins probably through HSP70. This may play a vital role by providing cytoprotection in pollutant-induced stressed fish adipocytes substantiated by the in vitro hypoxic studies. PMID:26744268

  15. Disruption of cell-matrix interactions by heparin enhances mesenchymal progenitor adipocyte differentiation

    SciTech Connect

    Luo Weijun; Shitaye, Hailu; Friedman, Michael; Bennett, Christina N.; Miller, Joshua; MacDougald, Ormond A.; Hankenson, Kurt D.

    2008-11-01

    Differentiation of marrow-derived mesenchymal progenitors to either the osteoblast or adipocyte lineage is reciprocally regulated. Factors that promote osteoblastogenesis inhibit adipogenesis, while adipogenic factors are inhibitory to osteoblast differentiation. Heparin, a soluble glycosaminoglycan, inhibits bone formation in vivo and osteoblast cell differentiation and function in vitro, and has been shown to promote adipocyte differentiation. To elucidate the role that heparin plays in the adipogenic induction of murine mesenchymal progenitors, we studied immortalized marrow stromal cells (IM-MSC), the MSC cell line, ST2, and 3T3L1 pre-adipocytes. Heparin alone was not sufficient to induce adipogenesis, but enhanced the induction under a variety of adipogenic cocktails. This effect was both dose- and time-dependent. Heparin showed a positive effect at concentrations > 0. 1 {mu}g/ml when applied before day 3 during the induction course. Heparin's effect on adipogenesis was independent of cell proliferation, cell density, and extracellular lipid. This effect is likely related to the unique structure of heparin because another polyanionic glycosaminoglycan, dextran sulfate, did not promote adipogenic differentiation. Heparin treatment altered morphology and adhesion characteristics of progenitor cells, resulting in cell rounding and aggregation. As well, heparin counteracted the known inhibitory effect of fibronectin on adipogenesis and decreased basal focal adhesion kinase and paxillin phosphorylation. We conclude that heparin-mediated disruption of cell-matrix adhesion enhances adipogenic potential.

  16. Ursodeoxycholic Acid but Not Tauroursodeoxycholic Acid Inhibits Proliferation and Differentiation of Human Subcutaneous Adipocytes

    PubMed Central

    Mališová, Lucia; Kováčová, Zuzana; Koc, Michal; Kračmerová, Jana; Štich, Vladimír; Rossmeislová, Lenka

    2013-01-01

    Stress of endoplasmic reticulum (ERS) is one of the molecular triggers of adipocyte dysfunction and chronic low inflammation accompanying obesity. ERS can be alleviated by chemical chaperones from the family of bile acids (BAs). Thus, two BAs currently used to treat cholestasis, ursodeoxycholic and tauroursodeoxycholic acid (UDCA and TUDCA), could potentially lessen adverse metabolic effects of obesity. Nevertheless, BAs effects on human adipose cells are mostly unknown. They could regulate gene expression through pathways different from their chaperone function, namely through activation of farnesoid X receptor (FXR) and TGR5, G-coupled receptor. Therefore, this study aimed to analyze effects of UDCA and TUDCA on human preadipocytes and differentiated adipocytes derived from paired samples of two distinct subcutaneous adipose tissue depots, abdominal and gluteal. While TUDCA did not alter proliferation of cells from either depot, UDCA exerted strong anti-proliferative effect. In differentiated adipocytes, acute exposition to neither TUDCA nor UDCA was able to reduce effect of ERS stressor tunicamycin. However, exposure of cells to UDCA during whole differentiation process decreased expression of ERS markers. At the same time however, UDCA profoundly inhibited adipogenic conversion of cells. UDCA abolished expression of PPARγ and lipogenic enzymes already in the early phases of adipogenesis. This anti-adipogenic effect of UDCA was not dependent on FXR or TGR5 activation, but could be related to ability of UDCA to sustain the activation of ERK1/2 previously linked with PPARγ inactivation. Finally, neither BAs did lower expression of chemokines inducible by TLR4 pathway, when UDCA enhanced their expression in gluteal adipocytes. Therefore while TUDCA has neutral effect on human preadipocytes and adipocytes, the therapeutic use of UDCA different from treating cholestatic diseases should be considered with caution because UDCA alters functions of human adipose cells

  17. Vitisin A inhibits adipocyte differentiation through cell cycle arrest in 3T3-L1 cells

    SciTech Connect

    Kim, Soon-hee; Park, Hee-Sook; Lee, Myoung-su; Cho, Yong-Jin; Kim, Young-Sup; Hwang, Jin-Taek; Sung, Mi Jeong; Kim, Myung Sunny; Kwon, Dae Young

    2008-07-18

    Inhibition of adipocyte differentiation is one approach among the anti-obesity strategies. This study demonstrates that vitisin A, a resveratrol tetramer, inhibits adipocyte differentiation most effectively of 18 stilbenes tested. Fat accumulation and PPAR{gamma} expression were decreased by vitisin A in a dose-dependent manner. Vitisin A significantly inhibited preadipocyte proliferation and consequent differentiation within the first 2 days of treatment, indicating that the anti-adipogenic effect of vitisin A was derived from anti-proliferation. Based on cell cycle analysis, vitisin A blocked the cell cycle at the G1-S phase transition, causing cells to remain in the preadipocyte state. Vitisin A increased p21 expression, while the Rb phosphorylation level was reduced. Therefore, vitisin A seems to induce G1 arrest through p21- and consequent Rb-dependent suppression of transcription. On the other hand, ERK and Akt signaling pathways were not involved in the anti-mitotic regulation by vitisin A. Taken together, these results suggest that vitisin A inhibits adipocyte differentiation through preadipocyte cell cycle arrest.

  18. Impacts of Alternative Splicing Events on the Differentiation of Adipocytes

    PubMed Central

    Lin, Jung-Chun

    2015-01-01

    Alternative splicing was found to be a common phenomenon after the advent of whole transcriptome analyses or next generation sequencing. Over 90% of human genes were demonstrated to undergo at least one alternative splicing event. Alternative splicing is an effective mechanism to spatiotemporally expand protein diversity, which influences the cell fate and tissue development. The first focus of this review is to highlight recent studies, which demonstrated effects of alternative splicing on the differentiation of adipocytes. Moreover, use of evolving high-throughput approaches, such as transcriptome analyses (RNA sequencing), to profile adipogenic transcriptomes, is also addressed. PMID:26389882

  19. Induction of Adipocyte Differentiation by Polybrominated Diphenyl Ethers (PBDEs) in 3T3-L1 Cells

    PubMed Central

    Tung, Emily W. Y.; Boudreau, Adèle; Wade, Michael G.; Atlas, Ella

    2014-01-01

    Polybrominated diphenyl ethers (PBDEs) are a class of brominated flame retardants that were extensively used in commercial products. PBDEs are ubiquitous environmental contaminants that are both lipophilic and bioaccumulative. Effects of PBDEs on adipogenesis were studied in the 3T3-L1 preadipocyte cell model in the presence and absence of a known adipogenic agent, dexamethasone (DEX). A PBDE mixture designed to mimic body burden of North Americans was tested, in addition to the technical mixture DE-71 and the individual congener BDE-47. The mixture, DE-71, and BDE-47 all induced adipocyte differentiation as assessed by markers for terminal differentiation [fatty acid binding protein 4 (aP2) and perilipin] and lipid accumulation. Characterization of the differentiation process in response to PBDEs indicated that adipogenesis induced by a minimally effective dose of DEX was enhanced by these PBDEs. Moreover, C/EBPα, PPARγ, and LXRα were induced late in the differentiation process. Taken together, these data indicate that adipocyte differentiation is induced by PBDEs; they act in the absence of glucocorticoid and enhance glucocorticoid-mediated adipogenesis. PMID:24722056

  20. AMP-Activated Kinase (AMPK) Activation by AICAR in Human White Adipocytes Derived from Pericardial White Adipose Tissue Stem Cells Induces a Partial Beige-Like Phenotype

    PubMed Central

    Abdul-Rahman, Omar; Kristóf, Endre; Doan-Xuan, Quang-Minh; Vida, András; Nagy, Lilla; Horváth, Ambrus; Simon, József; Maros, Tamás; Szentkirályi, István; Palotás, Lehel; Debreceni, Tamás; Csizmadia, Péter; Szerafin, Tamás; Fodor, Tamás; Szántó, Magdolna; Tóth, Attila; Kiss, Borbála; Bacsó, Zsolt; Bai, Péter

    2016-01-01

    Beige adipocytes are special cells situated in the white adipose tissue. Beige adipocytes, lacking thermogenic cues, morphologically look quite similar to regular white adipocytes, but with a markedly different response to adrenalin. White adipocytes respond to adrenergic stimuli by enhancing lipolysis, while in beige adipocytes adrenalin induces mitochondrial biogenesis too. A key step in the differentiation and function of beige adipocytes is the deacetylation of peroxisome proliferator-activated receptor (PPARγ) by SIRT1 and the consequent mitochondrial biogenesis. AMP-activated protein kinase (AMPK) is an upstream activator of SIRT1, therefore we set out to investigate the role of AMPK in beige adipocyte differentiation using human adipose-derived mesenchymal stem cells (hADMSCs) from pericardial adipose tissue. hADMSCs were differentiated to white and beige adipocytes and the differentiation medium of the white adipocytes was supplemented with 100 μM [(2R,3S,4R,5R)-5-(4-Carbamoyl-5-aminoimidazol-1-yl)-3,4-dihydroxyoxolan-2-yl]methyl dihydrogen phosphate (AICAR), a known activator of AMPK. The activation of AMPK with AICAR led to the appearance of beige-like morphological properties in differentiated white adipocytes. Namely, smaller lipid droplets appeared in AICAR-treated white adipocytes in a similar fashion as in beige cells. Moreover, in AICAR-treated white adipocytes the mitochondrial network was more fused than in white adipocytes; a fused mitochondrial system was characteristic to beige adipocytes. Despite the morphological similarities between AICAR-treated white adipocytes and beige cells, functionally AICAR-treated white adipocytes were similar to white adipocytes. We were unable to detect increases in basal or cAMP-induced oxygen consumption rate (a marker of mitochondrial biogenesis) when comparing control and AICAR-treated white adipocytes. Similarly, markers of beige adipocytes such as TBX1, UCP1, CIDEA, PRDM16 and TMEM26 remained the same when

  1. Differentiation of Pre-Adipocytes in Modelled Microgravity

    NASA Astrophysics Data System (ADS)

    Coinu, R.; Postiglione, I.; Meloni, M. A.; Galleri, G.; Pippia, P.; Palumbo, G.

    2008-06-01

    It has been demonstrated that microgravity affects biological and biochemical functions of cells including: morphology, cytoskeleton and embryogenesis [1]; proliferation, reduction of DNA, protein synthesis and glucose transport [2]; signalling, reduction of EGF-dependant c-fos and c-jun expression [3]; gene expression, reduction of IL2 expression and release by activated T-cells [4]. Moreover it has be found that peroxisome proliferators activated receptor γ (PPARγ2), which is known to be important for adipocyte differentiation, adipsin, leptin, and glucose transporter-4, are highly expressed in response to modelled microgravity [5]. These findings prompted us to investigate the effects of microgravity on cellular differentiation rate using a well characterized model. Such model consists in murine pre-adipocyte cells (3T3-L1) properly stimulated with insulin, dexamethazone and isobuthylmethyl-xantine (DMI protocol). The adipogenic program is completed within a short time. The entire process requires coordinated and temporarily beated molecular events. Early events. Growth arrest at confluence; Clonal expansion (this process involves synchronous entry of cells into S phase of the cell cycle, leading to one or two rounds of mitosis); Early expression of C/EBPβ and C/EBPδ. Late events. Expression of PPARγ and C/EBPα Assumption of rounded morphology and accumulation of lipid droplets.

  2. CBX7 gene expression plays a negative role in adipocyte cell growth and differentiation.

    PubMed

    Forzati, Floriana; Federico, Antonella; Pallante, Pierlorenzo; Colamaio, Marianna; Esposito, Francesco; Sepe, Romina; Gargiulo, Sara; Luciano, Antonio; Arra, Claudio; Palma, Giuseppe; Bon, Giulia; Bucher, Stefania; Falcioni, Rita; Brunetti, Arturo; Battista, Sabrina; Fedele, Monica; Fusco, Alfredo

    2014-01-01

    We have recently generated knockout mice for the Cbx7 gene, coding for a polycomb group protein that is downregulated in human malignant neoplasias. These mice develop liver and lung adenomas and carcinomas, which confirms a tumour suppressor role for CBX7. The CBX7 ability to downregulate CCNE1 expression likely accounts for the phenotype of the Cbx7-null mice. Unexpectedly, Cbx7-knockout mice had a higher fat tissue mass than wild-type, suggesting a role of CBX7 in adipogenesis. Consistently, we demonstrate that Cbx7-null mouse embryonic fibroblasts go towards adipocyte differentiation more efficiently than their wild-type counterparts, and this effect is Cbx7 dose-dependent. Similar results were obtained when Cbx7-null embryonic stem cells were induced to differentiate into adipocytes. Conversely, mouse embryonic fibroblasts and human adipose-derived stem cells overexpressing CBX7 show an opposite behaviour. These findings support a negative role of CBX7 in the control of adipocyte cell growth and differentiation. PMID:25190058

  3. CBX7 gene expression plays a negative role in adipocyte cell growth and differentiation

    PubMed Central

    Forzati, Floriana; Federico, Antonella; Pallante, Pierlorenzo; Colamaio, Marianna; Esposito, Francesco; Sepe, Romina; Gargiulo, Sara; Luciano, Antonio; Arra, Claudio; Palma, Giuseppe; Bon, Giulia; Bucher, Stefania; Falcioni, Rita; Brunetti, Arturo; Battista, Sabrina; Fedele, Monica; Fusco, Alfredo

    2014-01-01

    ABSTRACT We have recently generated knockout mice for the Cbx7 gene, coding for a polycomb group protein that is downregulated in human malignant neoplasias. These mice develop liver and lung adenomas and carcinomas, which confirms a tumour suppressor role for CBX7. The CBX7 ability to downregulate CCNE1 expression likely accounts for the phenotype of the Cbx7-null mice. Unexpectedly, Cbx7-knockout mice had a higher fat tissue mass than wild-type, suggesting a role of CBX7 in adipogenesis. Consistently, we demonstrate that Cbx7-null mouse embryonic fibroblasts go towards adipocyte differentiation more efficiently than their wild-type counterparts, and this effect is Cbx7 dose-dependent. Similar results were obtained when Cbx7-null embryonic stem cells were induced to differentiate into adipocytes. Conversely, mouse embryonic fibroblasts and human adipose-derived stem cells overexpressing CBX7 show an opposite behaviour. These findings support a negative role of CBX7 in the control of adipocyte cell growth and differentiation. PMID:25190058

  4. Suppression of PPARγ through MKRN1-mediated ubiquitination and degradation prevents adipocyte differentiation

    PubMed Central

    Kim, J-H; Park, K W; Lee, E-W; Jang, W-S; Seo, J; Shin, S; Hwang, K-A; Song, J

    2014-01-01

    The central regulator of adipogenesis, PPARγ, is a nuclear receptor that is linked to obesity and metabolic diseases. Here we report that MKRN1 is an E3 ligase of PPARγ that induces its ubiquitination, followed by proteasome-dependent degradation. Furthermore, we identified two lysine sites at 184 and 185 that appear to be targeted for ubiquitination by MKRN1. Stable overexpression of MKRN1 reduced PPARγ protein levels and suppressed adipocyte differentiation in 3T3-L1 and C3H10T1/2 cells. In contrast, MKRN1 depletion stimulated adipocyte differentiation in these cells. Finally, MKRN1 knockout MEFs showed an increased capacity for adipocyte differentiation compared with wild-type MEFs, with a concomitant increase of PPARγ and adipogenic markers. Together, these data indicate that MKRN1 is an elusive PPARγ E3 ligase that targets PPARγ for proteasomal degradation by ubiquitin-dependent pathways, and further depict MKRN1 as a novel target for diseases involving PPARγ. PMID:24336050

  5. Extract of Chaga mushroom (Inonotus obliquus) stimulates 3T3-L1 adipocyte differentiation.

    PubMed

    Joo, Jeong In; Kim, Dong Hyun; Yun, Jong Won

    2010-11-01

    Chaga mushroom (Inonotus obliquus) has long been used as a folk medicine due to its numerous biological functions such as antibacterial, antiallergic, antiinflammatory and antioxidative activities. In the present study, it was found that the I. obliquus hot water extract (IOWE) activated adipogenesis of 3T3-L1 preadipocytes. Even in the absence of adipogenic stimuli by insulin, the IOWE strongly induced adipogenesis of 3T3-L1 preadipocytes. The major constituent of IOWE was glucose-rich polysaccharides with a molecular mass of 149  kDa. IOWE enhanced the differentiation of 3T3-L1 preadipocytes, increasing TG (triacylglycerol) accumulation that is critical for acquisition of the adipocyte phenotype, in a dose-dependent manner. IOWE stimulated gene expression of C/EBPα (CCAAT/enhancer-binding protein α) and PPARγ (peroxisome proliferator-activated receptors γ) during adipocyte differentiation, and induced the expression of PPARγ target genes such as aP2 (adipocyte protein 2), LPL (lipoprotein lipase) and CD36 (fatty acid translocase). Immunoblot analysis revealed that IOWE increased the expression of adipogenic makers such as PPARγ and GLUT4 (glucose transporter 4). The luciferase reporter assay demonstrated that IOWE did not exhibit PPARγ ligand activity. Although these results require further investigation, the ability of natural mushroom product to increase PPARγ transcriptional activities may be expected to be therapeutic targets for dyslipidemia and type 2 diabetes. PMID:21031614

  6. Atypical antipsychotics induce both proinflammatory and adipogenic gene expression in human adipocytes in vitro

    SciTech Connect

    Sárvári, Anitta K.; Veréb, Zoltán; Uray, Iván P.; Fésüs, László; Balajthy, Zoltán

    2014-08-08

    Highlights: • Antipsychotics modulate the expression of adipogenic genes in human adipocytes. • Secretion of proinflammatory cytokine IL8 and MCP-1 is induced by antipsychotics. • Adipocyte-dependent inflammatory abnormality could develop during chronic treatment. • Infiltrated macrophages would further enhance proinflammatory cytokine production. - Abstract: Schizophrenia requires lifelong treatment, potentially causing systemic changes in metabolic homeostasis. In the clinical setting, antipsychotic treatment may differentially lead to weight gain among individual patients, although the molecular determinants of such adverse effects are currently unknown. In this study, we investigated changes in the expression levels of critical regulatory genes of adipogenesis, lipid metabolism and proinflammatory genes during the differentiation of primary human adipose-derived stem cells (ADSCs). These cells were isolated from patients with body mass indices <25 and treated with the second-generation antipsychotics olanzapine, ziprasidone, clozapine, quetiapine, aripiprazole and risperidone and the first-generation antipsychotic haloperidol. We found that antipsychotics exhibited a marked effect on key genes involved in the regulation of cell cycle, signal transduction, transcription factors, nuclear receptors, differentiation markers and metabolic enzymes. In particular, we observed an induction of the transcription factor NF-KB1 and NF-KB1 target genes in adipocytes in response to these drugs, including the proinflammatory cytokines TNF-α, IL-1β, IL-8 and MCP-1. In addition, enhanced secretion of both IL8 and MCP-1 was observed in the supernatant of these cell cultures. In addition to their remarkable stimulatory effects on proinflammatory gene transcription, three of the most frequently prescribed antipsychotic drugs, clozapine, quetiapine and aripiprazole, also induced the expression of essential adipocyte differentiation genes and the adipocyte hormones leptin

  7. Nrf2 activation diminishes during adipocyte differentiation of ST2 cells.

    PubMed

    Chartoumpekis, Dionysios V; Ziros, Panos G; Sykiotis, Gerasimos P; Zaravinos, Apostolos; Psyrogiannis, Agathoklis I; Kyriazopoulou, Venetsana E; Spandidos, Demetrios A; Habeos, Ioannis G

    2011-11-01

    Adipocyte differentiation (adipogenesis) is a highly controlled process known to be affected, among other factors, by the redox status of the cell. Nrf2 (NFE2-related factor 2) is a transcription factor that orchestrates the expression of a battery of antioxidant and detoxification genes under both basal and stress conditions. The present study investigated the activation of Nrf2 during adipocyte differentiation using as a model the mouse bone marrow-derived ST2 cell line. Treatment of ST2 cells with a differentiation cocktail containing IBMX, indomethacin, hydrocortisone and insulin induced differentiation into adipocytes over 5 days. During adipogenesis, the intracellular glutathione redox potential, which is an indicator of oxidative stress levels, became steadily more oxidized, as shown by real-time measurement in differentiating ST2 cells stably transfected with a redox-sensitive Grx1-roGFP2 fusion protein. The nuclear abundance of Nrf2 was assessed by Western immunoblotting and its DNA binding activity by EMSA (electrophoretic mobility shift assay) performed on nuclear protein extracts prepared every 24 h. The nuclear abundance of Nrf2 continuously decreased during adipogenesis in ST2 cells. Its DNA binding activity reached a nadir during the first two days of differentiation, after which it increased slightly without approaching its initial level. The pattern of Nrf2 DNA binding corresponded to its transcriptional activity as assessed in ST2 cells stably transfected with a reporter construct bearing a Nrf2 bind site upstream of the luciferase gene. In conclusion, the activation of Nrf2 decreased significantly during adipogenesis. The observed changes might lead to increased oxidative stress levels that could facilitate the differentiation process. These findings could shed new light on the pathogenesis of obesity, in which the adipose tissue and oxidative stress play prominent roles. PMID:21805027

  8. Macrophage-secreted factors induce adipocyte inflammation and insulin resistance

    SciTech Connect

    Permana, Paska A. . E-mail: Paska.Permana@med.va.gov; Menge, Christopher; Reaven, Peter D.

    2006-03-10

    Macrophage infiltration into adipose tissue increases with obesity, a condition associated with low-grade inflammation and insulin resistance. We investigated the direct effects of macrophage-secreted factors on adipocyte inflammation and insulin resistance. 3T3-L1 adipocytes incubated with media conditioned by RAW264.7 macrophages (RAW-CM) showed dramatically increased transcription of several inflammation-related genes, greater nuclear factor kappa B (NF-{kappa}B) activity, and enhanced binding of U937 monocytes. All of these effects were prevented by co-incubation with pyrrolidinedithiocarbamate, an NF-{kappa}B inhibitor. Adipocytes incubated with RAW-CM also released more non-esterified fatty acids and this increased lipolysis was not suppressed by insulin. In addition, RAW-CM treatment decreased insulin-stimulated glucose uptake in adipocytes. Taken together, these results indicate that macrophage-secreted factors induce inflammatory responses and reduce insulin responsiveness in adipocytes. These effects of macrophage-secreted factors on adipocytes may contribute significantly to the systemic inflammation and insulin resistance associated with obesity.

  9. Glucagon Like Peptide-1 Promotes Adipocyte Differentiation via the Wnt4 Mediated Sequestering of Beta-Catenin

    PubMed Central

    Liu, Rui; Li, Na; Lin, Yi; Wang, Mei; Peng, Yongde; Lewi, Keidren; Wang, Qinghua

    2016-01-01

    Glucagon-like peptide-1 (GLP-1) plays a role in the regulation of adipogenesis; however, the precise underlying molecular mechanism has not been fully defined. Wnt was recently identified as an important regulator of adipogenesis. This study aimed to investigate the involvement of the Wnt signaling pathway in the effects of GLP-1 on adipocyte differentiation. 3T3-L1 cells were induced to differentiate. The changes in the expression levels of adipogenic transcription factors and Wnts and the phosphorylation level and subcellular localization of β-catenin were quantified after GLP-1 treatment. GLP-1 stimulated adipocyte differentiation and lipid accumulation, which were accompanied by the expression of adipocyte marker genes. The expression of Wnt4 was upregulated in the process of adipocyte differentiation, which was further enhanced by treatment with GLP-1. β-catenin, an important mediator of the Wnt pathway, was immediately dephosphorylated and translocated from cytoplasm to nucleus when differentiation was induced. In the presence of GLP-1, however, β-catenin was redirected to the cell plasma membrane leading to its decreased accumulation in the nucleus. Knockdown of Wnt4 blocked the effect of GLP-1 on the cellular localization of β-catenin and expression level of adipogenic transcription factors. Our findings showed that GLP-1 promoted adipogenesis through the modulation of the Wnt4/β-catenin signaling pathway, suggesting that the GLP-1-Wntβ-catenin system might be a new target for the treatment of metabolic disease. PMID:27504979

  10. Silica nanoparticles inhibit brown adipocyte differentiation via regulation of p38 phosphorylation

    NASA Astrophysics Data System (ADS)

    Son, Min Jeong; Kim, Won Kon; Kwak, Minjeong; Oh, Kyoung-Jin; Chang, Won Seok; Min, Jeong-Ki; Lee, Sang Chul; Song, Nam Woong; Bae, Kwang-Hee

    2015-10-01

    Nanoparticles are of great interest due to their wide variety of biomedical and bioengineering applications. However, they affect cellular differentiation and/or intracellular signaling when applied and exposed to target organisms or cells. The brown adipocyte is a cell type important in energy homeostasis and thus closely related to obesity. In this study, we assessed the effects of silica nanoparticles (SNPs) on brown adipocyte differentiation. The results clearly showed that brown adipocyte differentiation was significantly repressed by exposure to SNPs. The brown adipocyte-specific genes as well as mitochondrial content were also markedly reduced. Additionally, SNPs led to suppressed p38 phosphorylation during brown adipocyte differentiation. These effects depend on the size of SNPs. Taken together, these results lead us to suggest that SNP has anti-brown adipogenic effect in a size-dependent manner via regulation of p38 phosphorylation.

  11. Effects of homocysteine on adipocyte differentiation and CD36 gene expression in 3T3-L1 adipocytes.

    PubMed

    Mentese, Ahmet; Alver, Ahmet; Sumer, Aysegul; Demir, Selim

    2016-03-01

    The aim of this study was to investigate the effects of homocysteine (Hcy), a risk factor for cardiovascular diseases, hypertension, stroke and obesity, on expression of CD36 that regulates uptake of oxidized low-density lipoprotein (Ox-LDL) by adipocytes and differentiation of 3T3-L1 cells to adipocytes. Cell viability was determined using MTT assay, and density of triglycerides were measured with Oil Red O staining. The expression levels of CD36 were analyzed using SYBR green assay by quantitative RT-PCR. Our results showed that the addition of Hcy inhibited differentiation of 3T3-L1 preadipocytes in a dose-dependent manner without a significant cell toxicity (p < 0.05). Percentage CD36 gene expression increased in the Hcy treatment groups, but not statistically significantly (p > 0.05) compared to differentiated adipocytes. Hcy reduced adipocyte differentiation, but had no effect on the expression level of CD36 in vitro conditions. The effect of Hcy on uptake and clearance of Ox-LDL by adipose tissue now needs to be investigated in vivo. PMID:26691520

  12. Effect of Glucagon-like Peptide-1 on the Differentiation of Adipose-derived Stem Cells into Osteoblasts and Adipocytes

    PubMed Central

    Lee, Hye Min; Joo, Bo Sun; Lee, Chang Hoon; Kim, Heung Yeol

    2015-01-01

    Objectives Glucagon-like peptide-1 (GLP-1) is an intestinally secreted hormone and it plays an important role in the regulation of glucose homeostasis. However, the possible role of GLP-1 in the differentiation of adipose-derived stem cells (ADSCs) remains unknown. Therefore this study investigated the effect of GLP-1 on the differentiation of ADSCs into osteoblasts and adipocytes. Methods ADSCs were isolated from human adipose tissues of the abdomens, cultured and characterized by flow cytometry and multi-lineage potential assay. ADSCs were induced in osteogenic and adipogenic media treated with two different doses (10 and 100 nM) of GLP-1, and then the effect of GLP-1 on differentiation of ADSCs into osteoblast and adipocyte was examined. The signaling pathway involved in these processes was also examined. Results Isolated human ADSCs expressed mesenchymal stem cell (MSC) specific markers as well as GLP-1 receptor (GLP-1R) proteins. They also showed multiple-lineage potential of MSC. GLP-1 was upregulated the activity and mRNA expression of osteoblast-specific marker, alkaline phosphatase and the mineralization of calcium. In contrast, GLP-1 significantly suppressed the expression of adipocyte-specific markers, peroxisome proliferator-activated receptor gamma (PPAR-γ), lipoprotein lipase (LPL) and adipocyte protein 2 (AP2). This decreased expression of adipocyte specific markers caused by GLP-1 was significantly reversed by the treatment of extracellular signal-regulated kinase (ERK) inhibitor, PD98059 (P < 0.05). Conclusion This result demonstrates that GLP-1 stimulates osteoblast differentiation in ADSCs, whereas it inhibits adipocyte differentiation. The ERK signaling pathway seems to be involved in these differentiation processes mediated by GLP-1. PMID:26357647

  13. Cellular origins of cold-induced brown adipocytes in adult mice

    PubMed Central

    Lee, Yun-Hee; Petkova, Anelia P.; Konkar, Anish A.; Granneman, James G.

    2015-01-01

    This work investigated how cold stress induces the appearance of brown adipocytes (BAs) in brown and white adipose tissues (WATs) of adult mice. In interscapular brown adipose tissue (iBAT), cold exposure increased proliferation of endothelial cells and interstitial cells expressing platelet-derived growth factor receptor, α polypeptide (PDGFRα) by 3- to 4-fold. Surprisingly, brown adipogenesis and angiogenesis were largely restricted to the dorsal edge of iBAT. Although cold stress did not increase proliferation in inguinal white adipose tissue (ingWAT), the percentage of BAs, defined as multilocular adipocytes that express uncoupling protein 1, rose from undetectable to 30% of total adipocytes. To trace the origins of cold-induced BAs, we genetically tagged PDGFRα+ cells and adipocytes prior to cold exposure, using Pdgfra-Cre recombinase estrogen receptor T2 fusion protein (CreERT2) and adiponectin-CreERT2, respectively. In iBAT, cold stress triggered the proliferation and differentiation of PDGFRα+ cells into BAs. In contrast, all newly observed BAs in ingWAT (5207 out of 5207) were derived from unilocular adipocytes tagged by adiponectin-CreERT2-mediated recombination. Surgical denervation of iBAT reduced cold-induced brown adipogenesis by >85%, whereas infusion of norepinephrine (NE) mimicked the effects of cold in warm-adapted mice. NE-induced de novo brown adipogenesis in iBAT was eliminated in mice lacking β1-adrenergic receptors. These observations identify a novel tissue niche for brown adipogenesis in iBAT and further define depot-specific mechanisms of BA recruitment.—Lee, Y.-H., Petkova, A. P., Konkar, A. A., Granneman, J. G. Cellular origins of cold-induced brown adipocytes in adult mice. PMID:25392270

  14. Mesenchymal Stromal Cells Differentiating to Adipocytes Accumulate Autophagic Vesicles Instead of Functional Lipid Droplets.

    PubMed

    Gruia, Alexandra T; Suciu, Maria; Barbu-Tudoran, Lucian; Azghadi, Seyed Mohammad Reza; Cristea, Mirabela I; Nica, Dragos V; Vaduva, Adrian; Muntean, Danina; Mic, Ani Aurora; Mic, Felix A

    2016-04-01

    Adult bone marrow mesenchymal stromal cells (BMSCs) can easily be differentiated into a variety of cells. In vivo transplantation of BMSCs-differentiated cells has had limited success, suggesting that these cells may not be fully compatible with the cells they are intended to replace in vivo. We investigated the structural and functional features of BMSCs-derived adipocytes as compared with adipocytes from adipose tissue, and the structure and functionality of lipid vesicles formed during BMSCs differentiation to adipocytes. Gas chromatography-mass spectrometry showed fatty acid composition of BMSCs-derived adipocytes and adipocytes from the adipose tissue to be very different, as is the lipid rafts composition, caveolin-1 expression, caveolae distribution in their membranes, and the pattern of expression of fatty acid elongases. Confocal microscopy confirmed the absence from BMSCs-derived adipocytes of markers of lipid droplets. BMSCs-derived adipocytes cannot convert deuterated glucose into deuterated species of fatty acids and cannot uptake the deuterated fatty acid-bovine serum albumin complexes from the culture medium, suggesting that intra-cellular accumulation of lipids does not occur by lipogenesis. We noted that BMSCs differentiation to adipocytes is accompanied by an increase in autophagy. Autophagic vesicles accumulate in the cytoplasm of BMSCs-derived adipocytes and their size and distribution resembles that of Nile Red-stained lipid vesicles. Stimulation of autophagy in BMSCs triggers the intra-cellular accumulation of lipids, while inhibition of autophagy prevents this accumulation. In conclusion, differentiation of BMSCs-derived adipocytes leads to intra-cellular accumulation of autophagic vesicles rather than functional lipid droplets, suggesting that these cells are not authentic adipocytes. J. Cell. Physiol. 231: 863-875, 2016. © 2015 Wiley Periodicals, Inc. PMID:26332160

  15. Isolation and Culture of Pig Spermatogonial Stem Cells and Their in Vitro Differentiation into Neuron-Like Cells and Adipocytes

    PubMed Central

    Wang, Xiaoyan; Chen, Tingfeng; Zhang, Yani; Li, Bichun; Xu, Qi; Song, Chengyi

    2015-01-01

    Spermatogonial stem cells (SSCs) renew themselves throughout the life of an organism and also differentiate into sperm in the adult. They are multipopent and therefore, can be induced to differentiate into many cells types in vitro. SSCs from pigs, considered an ideal animal model, are used in studies of male infertility, regenerative medicine, and preparation of transgenic animals. Here, we report on a culture system for porcine SSCs and the differentiation of these cells into neuron-like cells and adipocytes. SSCs and Sertoli cells were isolated from neonatal piglet testis by differential adhesion and SSCs were cultured on a feeder layer of Sertoli cells. Third-generation SSCs were induced to differentiate into neuron-like cells by addition of retinoic acid, β-mercaptoethanol, and 3-isobutyl-1-methylxanthine (IBMX) to the induction media and into adipocytes by the addition of hexadecadrol, insulin, and IBMX to the induction media. The differentiated cells were characterized by biochemical staining, qRT-PCR, and immunocytochemistry. The cells were positive for SSC markers, including alkaline phosphatase and SSC-specific genes, consistent with the cells being undifferentiated. The isolated SSCs survived on the Sertoli cells for 15 generations. Karyotyping confirmed that the chromosomal number of the SSCs were normal for pig (2n = 38, n = 19). Pig SSCs were successfully induced into neuron-like cells eight days after induction and into adipocytes 22 days after induction as determined by biochemical and immunocytochemical staining. qPCR results also support this conclusion. The nervous tissue markers genes, Nestin and β-tubulin, were expressed in the neuron-like cells and the adipocyte marker genes, PPARγ and C/EBPα, were expressed in the adipocytes. PMID:26556335

  16. Isolation and Culture of Pig Spermatogonial Stem Cells and Their in Vitro Differentiation into Neuron-Like Cells and Adipocytes.

    PubMed

    Wang, Xiaoyan; Chen, Tingfeng; Zhang, Yani; Li, Bichun; Xu, Qi; Song, Chengyi

    2015-01-01

    Spermatogonial stem cells (SSCs) renew themselves throughout the life of an organism and also differentiate into sperm in the adult. They are multipopent and therefore, can be induced to differentiate into many cells types in vitro. SSCs from pigs, considered an ideal animal model, are used in studies of male infertility, regenerative medicine, and preparation of transgenic animals. Here, we report on a culture system for porcine SSCs and the differentiation of these cells into neuron-like cells and adipocytes. SSCs and Sertoli cells were isolated from neonatal piglet testis by differential adhesion and SSCs were cultured on a feeder layer of Sertoli cells. Third-generation SSCs were induced to differentiate into neuron-like cells by addition of retinoic acid, β-mercaptoethanol, and 3-isobutyl-1-methylxanthine (IBMX) to the induction media and into adipocytes by the addition of hexadecadrol, insulin, and IBMX to the induction media. The differentiated cells were characterized by biochemical staining, qRT-PCR, and immunocytochemistry. The cells were positive for SSC markers, including alkaline phosphatase and SSC-specific genes, consistent with the cells being undifferentiated. The isolated SSCs survived on the Sertoli cells for 15 generations. Karyotyping confirmed that the chromosomal number of the SSCs were normal for pig (2n = 38, n = 19). Pig SSCs were successfully induced into neuron-like cells eight days after induction and into adipocytes 22 days after induction as determined by biochemical and immunocytochemical staining. qPCR results also support this conclusion. The nervous tissue markers genes, Nestin and β-tubulin, were expressed in the neuron-like cells and the adipocyte marker genes, PPARγ and C/EBPα, were expressed in the adipocytes. PMID:26556335

  17. Aristolochia Manshuriensis Kom Inhibits Adipocyte Differentiation by Regulation of ERK1/2 and Akt Pathway

    PubMed Central

    Kwak, Dong Hoon; Lee, Ji-Hye; Kim, Taesoo; Ahn, Hyo Sun; Cho, Won-Kyung; Ha, Hyunil; Hwang, Youn-Hwan; Ma, Jin Yeul

    2012-01-01

    Aristolochia manshuriensis Kom (AMK) is a traditional medicinal herb used for the treatment of arthritis, rheumatism, hepatitis, and anti-obesity. Because of nephrotoxicity and carcinogenicity of AMK, there are no pharmacological reports on anti-obesity potential of AMK. Here, we showed AMK has an inhibitory effect on adipocyte differentiation of 3T3-L1 cells along with significantly decrease in the lipid accumulation by downregulating several adipocyte-specific transcription factors including peroxisome proliferation-activity receptor γ (PPAR-γ), CCAAT/enhancer binding protein α (C/EBP-α) and C/EBP-β, which are critical for adipogenesis in vitro. AMK also markedly activated the extracellular signal-regulated protein kinase 1/2 (ERK1/2) pathway including Ras, Raf1, and mitogen-activated protein kinase kinase 1 (MEK1), and significantly suppressed Akt pathway by inhibition of phosphoinositide-dependent kinase 1 (PDK1). Aristolochic acid (AA) and ethyl acetate (EtOAc) fraction of AMK with AA were significantly inhibited TG accumulation, and regulated two pathway (ERK1/2 and Akt) during adipocyte differentiation, and was not due to its cytotoxicity. These two pathways were upstream of PPAR-γ and C/EBPα in the adipogenesis. In addition, gene expressions of secreting factors such as fatty acid synthase (FAS), adiponectin, lipopreotein lipase (LPL), and aP2 were significantly inhibited by treatment of AMK during adipogenesis. We used the high-fat diet (HFD)-induced obesity mouse model to determine the inhibitory effects of AMK on obesity. Oral administration of AMK (62.5 mg/kg/day) significantly decreased the fat tissue weight, total cholesterol (TC), and low density lipoprotein-cholesterol (LDL-C) concentration in the blood. The results of this study suggested that AMK inhibited lipid accumulation by the down-regulation of the major transcription factors of the adipogensis pathway including PPAR-γ and C/EBP-α through regulation of Akt pathway and ERK 1

  18. Cholesteryl ester transfer protein gene expression during differentiation of human preadipocytes to adipocytes in primary culture.

    PubMed

    Gauthier, B; Robb, M; McPherson, R

    1999-02-01

    The expression pattern of the CETP gene in relationship to that of LPL, adipsin, PPARgamma, C/EBPalpha, ADD1/SREBPI and actin was examined by RT-PCR during differentiation of human fibroblastic preadipocytes to adipocytes in primary culture. Preadipocytes were isolated from subcutaneous fat obtained from healthy female subjects undergoing mammary reduction procedures, and induced to differentiate in culture. Morphologically, adipogenesis was confirmed by the accumulation of lipid droplets in cells. We show that the gene encoding CETP is expressed in preadipocytes and is present throughout differentiation as compared to LPL and adipsin which were detected in the majority of samples by day 2 or 3 of adipogenesis. The transcription factors, PPARgamma, ADD1/SREBP1 and C/EBPalpha were expressed by day 2, concomitant with the appearance of LPL and adipsin but subsequent to the appearance of CETP. CETP mRNA was not detectable in human skin fibroblasts. These studies demonstrate that CETP. expression is induced at an early stage of commitment to the adipocyte lineage and may be activated by transcription factor(s), which are not members of the PPAR, ADD1/SREBP1 or C/EBP families. PMID:10030381

  19. A Cannabinoid Receptor Agonist N-Arachidonoyl Dopamine Inhibits Adipocyte Differentiation in Human Mesenchymal Stem Cells

    PubMed Central

    Ahn, Seyeon; Yi, Sodam; Seo, Won Jong; Lee, Myeong Jung; Song, Young Keun; Baek, Seung Yong; Yu, Jinha; Hong, Soo Hyun; Lee, Jinyoung; Shin, Dong Wook; Jeong, Lak Shin; Noh, Minsoo

    2015-01-01

    Endocannabinoids can affect multiple cellular targets, such as cannabinoid (CB) receptors, transient receptor potential cation channel, subfamily V, member 1 (TRPV1) and peroxisome proliferator-activated receptor γ (PPARγ). The stimuli to induce adipocyte differentiation in hBM-MSCs increase the gene transcription of the CB1 receptor, TRPV1 and PPARγ. In this study, the effects of three endocannabinoids, N-arachidonoyl ethanolamine (AEA), N-arachidonoyl dopamine (NADA) and 2-arachidonoyl glycerol (2-AG), on adipogenesis in hBM-MSCs were evaluated. The adipocyte differentiation was promoted by AEA whereas inhibited by NADA. No change was observed by the treatment of non-cytotoxic concentrations of 2-AG. The difference between AEA and NADA in the regulation of adipogenesis is associated with their effects on PPARγ transactivation. AEA can directly activate PPARγ. The effect of AEA on PPARγ in hBM-MSCs may prevail over that on the CB1 receptor mediated signal transduction, giving rise to the AEA-induced promotion of adipogenesis. In contrast, NADA had no effect on the PPARγ activity in the PPARγ transactivation assay. The inhibitory effect of NADA on adipogenesis in hBM-MSCs was reversed not by capsazepine, a TRPV1 antagonist, but by rimonabant, a CB1 antagonist/inverse agonist. Rimonabant by itself promoted adipogenesis in hBM-MSCs, which may be interpreted as the result of the inverse agonism of the CB1 receptor. This result suggests that the constantly active CB1 receptor may contribute to suppress the adipocyte differentiation of hBM-MSCs. Therefore, the selective CB1 agonists that are unable to affect cellular PPARγ activity inhibit adipogenesis in hBM-MSCs. PMID:25995819

  20. Early soy exposure via maternal diet regulates rat mammary epithelial differentiation by paracrine signaling from stromal adipocytes.

    PubMed

    Su, Ying; Shankar, Kartik; Simmen, Rosalia C M

    2009-05-01

    Diet-mediated changes in transcriptional programs that promote the early differentiation of the mammary gland may lead to reduced breast cancer risk. The disparity in adult breast cancer incidence between Asian women and Western counterparts is attributed partly to high soy food intake. Here, we conducted genome-wide profiling of mammary tissues of weanling rats exposed to soy protein isolate (SPI) or control casein (CAS) via maternal diet to evaluate the contribution of early exposure on mammary gene expression. Of the identified 18 up- and 39 downregulated genes with SPI relative to CAS, a subset was associated with lipid metabolic pathways, consistent with reduced mammary adipocyte size and suggesting stromal adipocyte-specific genomic changes. Female offspring of rats fed SPI tended to have fewer terminal end buds (P = 0.06) and had significantly lower body weight and abdominal fat mass. To demonstrate the functional consequence of SPI-mediated adipocyte metabolic changes on neighboring mammary epithelium, the expression of in vivo regulated genes in 3T3-L1 adipocytes treated with soy isoflavone genistein and effects of the resultant conditioned medium (CM) on the differentiation of HC11 mammary epithelial cells were evaluated by quantitative RT-PCR and/or Western immunoblots. In differentiated 3T3-L1, genistein decreased fatty acid synthase and stearoyl-CoA desaturase and increased hydroxysteroid 11-beta dehydrogenase 1 expression. CM from genistein-treated adipocytes had higher adiponectin levels and augmented prolactin-induced, glucocorticoid-regulated beta-casein levels. These findings suggest that soy-associated components, by targeting mammary adipocytes, alter paracrine signaling to enhance mammary epithelial differentiation, with important implications for the prevention of breast cancer associated with obesity and obesity-related diseases. PMID:19321580

  1. Unexpected blockade of adipocyte differentiation by K-7174: Implication for endoplasmic reticulum stress

    SciTech Connect

    Shimada, Tsuyoshi; Hiramatsu, Nobuhiko; Okamura, Maro; Hayakawa, Kunihiro; Kasai, Ayumi; Yao, Jian; Kitamura, Masanori

    2007-11-16

    Preadipocytes constitutively express GATA-2 and GATA-3 that are required to halt the cells at the undifferentiated stage. However, we unexpectedly found that K-7174, a GATA-specific inhibitor, did not induce but rather inhibited differentiation of 3T3-L1 preadipocytes. It was associated with lack of lipid accumulation, blunted expression of adipocyte markers including adiponectin and peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}), and sustained expression of a preadipocyte marker monocyte chemoattractant protein 1 (MCP-1). Subsequent experiments revealed that K-7174 had the potential to induce endoplasmic reticulum (ER) stress evidenced by induction of GRP78 and CHOP. Other inducers of ER stress completely reproduced the effects of K-7174 including suppression of lipid accumulation, blockade of induction of adiponection and PPAR{gamma} and maintenance of MCP-1 expression. These results indicated a possibility that ER stress suppresses adipocyte differentiation and that GATA inhibitor K-7174 has the potential for interfering with adipogenesis through induction of ER stress.

  2. The Acute Phase Protein Serum Amyloid A Induces Lipolysis and Inflammation in Human Adipocytes through Distinct Pathways

    PubMed Central

    Faty, Aurélie; Ferré, Pascal; Commans, Stéphane

    2012-01-01

    Background The acute phase response (APR) is characterized by alterations in lipid and glucose metabolism leading to an increased delivery of energy substrates. In adipocytes, there is a coordinated decrease in Free Fatty acids (FFAs) and glucose storage, in addition to an increase in FFAs mobilization. Serum Amyloid A (SAA) is an acute phase protein mainly associated with High Density Lipoproteins (HDL). We hypothesized that enrichment of HDL with SAA, during the APR, could be implicated in the metabolic changes occurring in adipocytes. Methodology/Principal Findings In vitro differentiated human adipocytes (hMADS) were treated with SAA enriched HDL or recombinant SAA and the metabolic phenotype of the cells analyzed. In hMADS, SAA induces an increased lipolysis through an ERK dependent pathway. At the molecular level, SAA represses PPARγ2, C/EBPα and SREBP-1c gene expression, three transcription factors involved in adipocyte differentiation or lipid synthesis. In addition, the activation of the NF-κB pathway by SAA leads to the induction of pro-inflammatory cytokines and chemokines, as in the case of immune cells. These latter findings were replicated in freshly isolated mature human adipocytes. Conclusions/Significance Besides its well-characterized role in cholesterol metabolism, SAA has direct metabolic effects on human adipocytes. These metabolic changes could be at least partly responsible for alterations of adipocyte metabolism observed during the APR as well as during pathophysiological conditions such as obesity and conditions leading to insulin resistant states. PMID:22532826

  3. Obesity-associated Inflammation Induces microRNA-155 Expression in Adipocytes and Adipose Tissue: Outcome on Adipocyte Function.

    PubMed

    Karkeni, Esma; Astier, Julien; Tourniaire, Franck; El Abed, Mouna; Romier, Béatrice; Gouranton, Erwan; Wan, Lin; Borel, Patrick; Salles, Jérôme; Walrand, Stéphane; Ye, Jianping; Landrier, Jean-François

    2016-04-01

    miR-155 expression is induced in adipocytes and adipose tissue submitted to inflammatory conditions in obesity context in murine and human models and participate to a pro-inflammatory loop by targeting PPARg. PMID:26829440

  4. Transcriptional and Epigenetic Mechanisms Underlying Enhanced in Vitro Adipocyte Differentiation by the Brominated Flame Retardant BDE-47

    PubMed Central

    2015-01-01

    Recent studies suggest that exposure to endocrine-disrupting compounds (EDCs) may play a role in the development of obesity. EDCs such as the flame retardant 2,2′,4,4′-tetrabrominated diphenyl ether (BDE-47) have been shown to enhance adipocyte differentiation in the murine 3T3-L1 model. The mechanisms by which EDCs direct preadipocytes to form adipocytes are poorly understood. Here, we examined transcriptional and epigenetic mechanisms underlying the induction of in vitro adipocyte differentiation by BDE-47. Quantitative high content microscopy revealed concentration-dependent enhanced adipocyte differentiation following exposure to BDE-47 or the antidiabetic drug troglitazone (TROG). BDE-47 modestly activated the key adipogenic transcription factor peroxisome proliferator-activated receptor gamma (PPARγ) in COS7 cells, transiently transfected with a GAL4 reporter construct. Increased gene expression was observed for Pparγ2, leptin (Lep), and glucose-6-phophatase catalytic subunit (G6pc) in differentiated 3T3-L1 cells after BDE-47 exposure compared to TROG. Methylation-sensitive high resolution melting (MS-HRM) revealed significant demethylation of three CpG sites in the Pparγ2 promoter after exposure to both BDE-47 and TROG in differentiated 3T3-L1 cells. This study shows the potential of BDE-47 to induce adipocyte differentiation through various mechanisms that include Pparγ2 gene induction and promoter demethylation accompanied by activation of PPARγ, and possible disruption of glucose homeostasis and IGF1 signaling. PMID:24559133

  5. Actions of β-Apo-Carotenoids in Differentiating Cells: Differential Effects in P19 Cells and 3T3-L1 Adipocytes

    PubMed Central

    Wang, Cynthia X.; Jiang, Hongfeng; Yuen, Jason J.; Lee, Seung-Ah; Narayanasamy, Sureshbabu; Curley, Robert W.; Harrison, Earl H.; Blaner, William S.

    2015-01-01

    β-Apo-carotenoids, including β-apo-13-carotenone and β-apo-14′-carotenal, are potent retinoic acid receptor (RAR) antagonists in transactivation assays. We asked how these influence RAR-dependent processes in living cells. Initially, we explored the effects of β-apo-13-carotenone and β-apo-14′-carotenal on P19 cells, a mouse embryonal carcinoma cell line that differentiates into neurons when treated with all-trans-retinoic acid. Treatment of P19 cells with either compound failed to block all-trans-retinoic acid induced differentiation. Liquid chromatography tandem mass spectrometry studies, however, established that neither of these β-apo-carotenoids accumulates in P19 cells. All-trans-retinoic acid accumulated to high levels in P19 cells. This suggests that the uptake and metabolism of β-apo-carotenoids by some cells does not involve the same processes used for retinoids and that these may be cell type specific. We also investigated the effects of two β-apo-carotenoids on 3T3-L1 adipocyte marker gene expression during adipocyte differentiation. Treatment of 3T3-L1 adipocytes with either β-apo-13-carotenone or β-apo-10′-carotenoic acid, which lacks RAR antagonist activity, stimulated adipocyte marker gene expression. Neither blocked the inhibitory effects of a relatively large dose of exogenous all-trans-retinoic acid on adipocyte differentiation. Our data suggest that in addition to acting as transcriptional antagonists, some β-apo-carotenoids act through other mechanisms to influence 3T3-L1 adipocyte differentiation. PMID:25602703

  6. The Glucose Sensor ChREBP Links De Novo Lipogenesis to PPARγ Activity and Adipocyte Differentiation.

    PubMed

    Witte, Nicole; Muenzner, Matthias; Rietscher, Janita; Knauer, Miriam; Heidenreich, Steffi; Nuotio-Antar, Alli M; Graef, Franziska A; Fedders, Ronja; Tolkachov, Alexander; Goehring, Isabel; Schupp, Michael

    2015-11-01

    Reduced de novo lipogenesis in adipose tissue, often observed in obese individuals, is thought to contribute to insulin resistance. Besides trapping excess glucose and providing for triglycerides and energy storage, endogenously synthesized lipids can function as potent signaling molecules. Indeed, several specific lipids and their molecular targets that mediate insulin sensitivity have been recently identified. Here, we report that carbohydrate-response element-binding protein (ChREBP), a transcriptional inducer of glucose use and de novo lipogenesis, controls the activity of the adipogenic master regulator peroxisome proliferator-activated receptor (PPAR)γ. Expression of constitutive-active ChREBP in precursor cells activated endogenous PPARγ and promoted adipocyte differentiation. Intriguingly, ChREBP-constitutive-active ChREBP expression induced PPARγ activity in a fatty acid synthase-dependent manner and by trans-activating the PPARγ ligand-binding domain. Reducing endogenous ChREBP activity by either small interfering RNA-mediated depletion, exposure to low-glucose concentrations, or expressing a dominant-negative ChREBP impaired differentiation. In adipocytes, ChREBP regulated the expression of PPARγ target genes, in particular those involved in thermogenesis, similar to synthetic PPARγ ligands. In summary, our data suggest that ChREBP controls the generation of endogenous fatty acid species that activate PPARγ. Thus, increasing ChREBP activity in adipose tissue by therapeutic interventions may promote insulin sensitivity through PPARγ. PMID:26181104

  7. Epigenetic Priming Confers Direct Cell Trans-Differentiation From Adipocyte to Osteoblast in a Transgene-Free State.

    PubMed

    Cho, Young-Dan; Bae, Han-Sol; Lee, Dong-Seol; Yoon, Won-Joon; Woo, Kyung-Mi; Baek, Jeong-Hwa; Lee, Gene; Park, Joo-Cheol; Ku, Young; Ryoo, Hyun-Mo

    2016-07-01

    The bone marrow of healthy individuals is primarily composed of osteoblasts and hematopoietic cells, while that of osteoporosis patients has a larger portion of adipocytes. There is evidence that the epigenetic landscape can strongly influence cell differentiation. We have shown that it is possible to direct the trans-differentiation of adipocytes to osteoblasts by modifying the epigenetic landscape with a DNA methyltransferase inhibitor (DNMTi), 5'-aza-dC, followed by Wnt3a treatment to signal osteogenesis. Treating 3T3-L1 adipocytes with 5'-aza-dC induced demethylation in the hypermethylated CpG regions of bone marker genes; subsequent Wnt3a treatment drove the cells to osteogenic differentiation. When old mice with predominantly adipose marrow were treated with both 5'-aza-dC and Wnt3a, decreased fatty tissue and increased bone volume were observed. Together, our results indicate that epigenetic modification permits direct programming of adipocytes into osteoblasts in a mouse model of osteoporosis, suggesting that this approach could be useful in bone tissue-engineering applications. PMID:26335354

  8. Isoflavones in Chickpeas Inhibit Adipocyte Differentiation and Prevent Insulin Resistance in 3T3-L1 Cells.

    PubMed

    Gao, Yue; Yao, Yang; Zhu, Yinging; Ren, Guixing

    2015-11-11

    Diabetes mellitus is a metabolic disease characterized by hyperglycemia arising from defects in insulin secretion. This study investigated the effects of isoflavones in chickpea sprouts germinated in light (IGL) and isoflavones in chickpea seeds (ICS) on insulin resistance through their role in suppression of 3T3-L1 adipocyte differentiation. Results showed that IGL and ICS inhibit the differentiation of 3T3-L1 pre-adipocytes induced by differentiation medium in a dose-dependent manner, and the suppressive effect of IGL was stronger (p < 0.05) than that of ICS, evidenced by a decrease of Oil Red O staining and intracellular triacylglycerol content in the mature adipocytes. IGL and ICS also stimulated glucose uptake significantly (p < 0.05). Besides, IGL and ICS treatment caused a significant decrease in mRNA and protein expression levels of adipogenesis-related transcription factors peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT-enhancer-binding protein α (C/EBPα). Furthermore, the mRNA and protein expression levels of adipocyte fatty acid-binding protein (ap2), lipoprotein lipase (LPL), uncoupling protein-2 (UCP-2), and glucose transporter 4 (Glut4) in 3T3-L1 cells were also markedly down-regulated (p < 0.05). PMID:26494490

  9. Oleanolic acid reduces markers of differentiation in 3T3-L1 adipocytes.

    PubMed

    Sung, Hye-Young; Kang, Sang-Wook; Kim, Jung-Lye; Li, Jing; Lee, Eun-Sook; Gong, Ju-Hyun; Han, Seoung Jun; Kang, Young-Hee

    2010-12-01

    Oleanolic acid is a triterpenoid compound that is widely present in vegetables, medicinal herbs, and other plants and has potent antioxidant and antiinflammatory properties. However, the potential of oleanolic acid to offset obesity is not clear. This study tested the hypothesis that oleanolic acid suppresses the differentiation of 3T3-L1 adipocytes by downregulating cellular induction of peroxisome proliferators-activated receptor γ (PPARγ) and cytidine-cytidine-adenosine-adenosine-thymidine (CCAAT) enhancer binding protein α (C/EBPα). The 3T3-L1 adipocytes were cultured and differentiated in Dulbecco modified Eagle medium containing 10% fetal bovine serum for 6 to 8 days in the absence and presence of 1 to 25 μmol/L oleanolic acid according to differentiating protocols. Nontoxic oleanolic acid, at 25 μmol/L or less, dose-dependently attenuated lipid accumulation in differentiated adipocytes as evidenced by Oil Red O staining. Western blot analysis showed that the induction of PPARγ and C/EBPα was markedly attenuated in differentiated and oleanolic acid-treated adipocytes at their transcriptional messenger RNA levels. Furthermore, this study examined whether oleanolic acid dampened the induction of visfatin, a proinflammatory and visceral fat-specific adipokine expressed in adipocytes. Visfatin expression was inhibited in differentiated adipocytes exposed to a PPARγ inhibitor GW9662. In addition, the visfatin production was significantly repressed in 25 μmol/L oleanolic acid-treated adipocytes, possibly through blocking PPARγ activation. These results demonstrate that oleanolic acid may be a promising agent to disturb adipocyte differentiation and suppress obesity-associated inflammation. PMID:21147366

  10. Cellular origins of cold-induced brown adipocytes in adult mice.

    PubMed

    Lee, Yun-Hee; Petkova, Anelia P; Konkar, Anish A; Granneman, James G

    2015-01-01

    This work investigated how cold stress induces the appearance of brown adipocytes (BAs) in brown and white adipose tissues (WATs) of adult mice. In interscapular brown adipose tissue (iBAT), cold exposure increased proliferation of endothelial cells and interstitial cells expressing platelet-derived growth factor receptor, α polypeptide (PDGFRα) by 3- to 4-fold. Surprisingly, brown adipogenesis and angiogenesis were largely restricted to the dorsal edge of iBAT. Although cold stress did not increase proliferation in inguinal white adipose tissue (ingWAT), the percentage of BAs, defined as multilocular adipocytes that express uncoupling protein 1, rose from undetectable to 30% of total adipocytes. To trace the origins of cold-induced BAs, we genetically tagged PDGFRα(+) cells and adipocytes prior to cold exposure, using Pdgfra-Cre recombinase estrogen receptor T2 fusion protein (CreER(T2)) and adiponectin-CreER(T2), respectively. In iBAT, cold stress triggered the proliferation and differentiation of PDGFRα(+) cells into BAs. In contrast, all newly observed BAs in ingWAT (5207 out of 5207) were derived from unilocular adipocytes tagged by adiponectin-CreER(T2)-mediated recombination. Surgical denervation of iBAT reduced cold-induced brown adipogenesis by >85%, whereas infusion of norepinephrine (NE) mimicked the effects of cold in warm-adapted mice. NE-induced de novo brown adipogenesis in iBAT was eliminated in mice lacking β1-adrenergic receptors. These observations identify a novel tissue niche for brown adipogenesis in iBAT and further define depot-specific mechanisms of BA recruitment. PMID:25392270

  11. DYNAMICS OF LIPID DROPLET-ASSOCIATED PROTEINS DURING HORMONALLY STIMULATED LIPOLYSIS IN ENGINEERED ADIPOCYTES: STABILIZATION AND LIPID DROPLET BINDING OF ADIPOCYTE DIFFERENTIATION-RELATED PROTEIN/ADIPOPHILIN

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In mature adipocytes, triglyceride is stored within lipid droplets, which are coated with the protein perilipin, which functions to regulate lipolysis by controlling lipase access to the droplet in a hormone-regulatable fashion. Adipocyte-differentiation related protein (ADRP) is a widely expressed ...

  12. Impact of elvitegravir on human adipocytes: Alterations in differentiation, gene expression and release of adipokines and cytokines.

    PubMed

    Moure, Ricardo; Domingo, Pere; Gallego-Escuredo, José M; Villarroya, Joan; Gutierrez, Maria Del Mar; Mateo, Maria G; Domingo, Joan C; Giralt, Marta; Villarroya, Francesc

    2016-08-01

    Elvitegravir is a recently developed integrase inhibitor used for antiretroviral treatment of HIV infection. Secondary effects, including disturbances in lipid metabolism and, ultimately, in adipose tissue distribution and function, are common concerns associated with antiretroviral treatments. Here, we provide the first study of the effects of elvitegravir (in comparison with efavirenz, a non-nucleoside analog inhibitor of reverse transcriptase; and raltegravir, another integrase inhibitor) on human adipocyte differentiation, gene expression and secretion of adipokines and cytokines. Elvitegravir impaired adipogenesis and adipocyte metabolism in human SGBS adipocytes in a concentration-dependent manner (delaying acquisition of adipocyte morphology and reducing the expression of adipogenesis marker genes such as PPARγ, glucose transporter GLUT4, lipoprotein lipase, and the adipokines adiponectin and leptin). Compared with efavirenz, the effects of elvitegravir were similar but tended to occur at higher concentrations than those elicited by efavirenz, or were somewhat less intense than those caused by efavirenz at similar concentration. Elvitegravir tended to cause a more moderate induction of pro-inflammatory cytokines than efavirenz. Efavirenz induced a marked concentration-dependent increase in interleukin-8 expression and release whereas elvitregravir had little effect. Raltegravir had totally neutral actions of adipogenesis, adipocyte metabolism-related gene expression and release of adipokines and cytokines. In conclusion, elvitegravir alters adipocyte differentiation and function and promotes induction of pro-inflammatory cytokines similarly to efavirenz, but several effects were less intense. Further assessment of lipid metabolism and adipose tissue function in patients administered elvitegravir-based regimes is advisable considering that totally neutral effects of elvitegravir on lipid homeostasis cannot be anticipated from the current study in

  13. Hedgehog associated to microparticles inhibits adipocyte differentiation via a non-canonical pathway

    PubMed Central

    Fleury, Audrey; Hoch, Lucile; Martinez, M. Carmen; Faure, Hélène; Taddei, Maurizio; Petricci, Elena; Manetti, Fabrizio; Girard, Nicolas; Mann, André; Jacques, Caroline; Larghero, Jérôme; Ruat, Martial; Andriantsitohaina, Ramaroson; Le Lay, Soazig

    2016-01-01

    Hedgehog (Hh) is a critical regulator of adipogenesis. Extracellular vesicles are natural Hh carriers, as illustrated by activated/apoptotic lymphocytes specifically shedding microparticles (MP) bearing the morphogen (MPHh+). We show that MPHh+ inhibit adipocyte differentiation and orientate mesenchymal stem cells towards a pro-osteogenic program. Despite a Smoothened (Smo)-dependency, MPHh+ anti-adipogenic effects do not activate a canonical Hh signalling pathway in contrast to those elicited either by the Smo agonist SAG or recombinant Sonic Hedgehog. The Smo agonist GSA-10 recapitulates many of the hallmarks of MPHh+ anti-adipogenic effects. The adipogenesis blockade induced by MPHh+ and GSA-10 was abolished by the Smo antagonist LDE225. We further elucidate a Smo/Lkb1/Ampk axis as the non-canonical Hh pathway used by MPHh+ and GSA-10 to inhibit adipocyte differentiation. Our results highlight for the first time the ability of Hh-enriched MP to signal via a non-canonical pathway opening new perspectives to modulate fat development. PMID:27010359

  14. SREBP-1c/MicroRNA 33b Genomic Loci Control Adipocyte Differentiation.

    PubMed

    Price, Nathan L; Holtrup, Brandon; Kwei, Stephanie L; Wabitsch, Martin; Rodeheffer, Matthew; Bianchini, Laurence; Suárez, Yajaira; Fernández-Hernando, Carlos

    2016-04-01

    White adipose tissue (WAT) is essential for maintaining metabolic function, especially during obesity. The intronic microRNAs miR-33a and miR-33b, located within the genes encoding sterol regulatory element-binding protein 2 (SREBP-2) and SREBP-1, respectively, are transcribed in concert with their host genes and function alongside them to regulate cholesterol, fatty acid, and glucose metabolism. SREBP-1 is highly expressed in mature WAT and plays a critical role in promoting in vitro adipocyte differentiation. It is unknown whether miR-33b is induced during or involved in adipogenesis. This is in part due to loss of miR-33b in rodents, precluding in vivo assessment of the impact of miR-33b using standard mouse models. This work demonstrates that miR-33b is highly induced upon differentiation of human preadipocytes, along with SREBP-1. We further report that miR-33b is an important regulator of adipogenesis, as inhibition of miR-33b enhanced lipid droplet accumulation. Conversely, overexpression of miR-33b impaired preadipocyte proliferation and reduced lipid droplet formation and the induction of peroxisome proliferator-activated receptor γ (PPARγ) target genes during differentiation. These effects may be mediated by targeting of HMGA2, cyclin-dependent kinase 6 (CDK6), and other predicted miR-33b targets. Together, these findings demonstrate a novel role of miR-33b in the regulation of adipocyte differentiation, with important implications for the development of obesity and metabolic disease. PMID:26830228

  15. Human induced pluripotent stem cells: A new source for brown and white adipocytes

    PubMed Central

    Hafner, Anne-Laure; Dani, Christian

    2014-01-01

    Mesenchymal stem cells (MSCs) derived from human induced pluripotent stem cells (hiPSCs) provide a novel source for generating adipocytes, thus opening new avenues for fundamental research and clinical medicine. We present the adipogenic potential of hiPSCs and the various methods to derive hiPSC-MSCs. We discuss the main characteristic of hiPSC-MSCs, which is their low adipogenic capacity as compared to adult-MSCs. Finally, we propose several hypotheses to explanation this feature, underlying a potential critical role of the micro-environment. We favour the hypothesis that the range of factors or culture conditions required to induce adipocyte differentiation of MSCs derived from adult tissues and from embryonic-like cells could differ. PMID:25258668

  16. Proliferation and differentiation of brown adipocytes from interstitial cells during cold acclimation

    SciTech Connect

    Bukowiecki, L.J.; Geloeen, A.; Collet, A.J.

    1986-06-01

    The mechanisms of brown adipocyte proliferation and differentiation during cold acclimation (and/or adaptation to hyperphagia) have been studied by quantitative photonic radioautography. (/sup 3/H)thymidine was injected to warm-acclimated (25/sup 0/C) rats and to animals exposed to 5/sup 0/C for 2 days. Samples of interscapular brown adipose tissue were collected for quantitative analysis of mitotic frequencies at various periods of time (4 h-15 days) after the injection of (/sup 3/H)thymidine, the rats being maintained at the temperatures to which they were initially exposed. It was found that cold exposure for 2 days markedly enhanced mitotic activity in endothelial cells, interstitial cells, and brown preadipocytes rather than in fully differentiated brown adipocytes. The total tissue labeling index (percent of labeled nuclei) increased approx.70 times over control values. The authors now report that cellular labeling progressively increased in mature brown adipocytes during cold acclimation, whereas it correspondingly decreased in interstitial cells and brown preadipocytes. This indicates that the sequence of events for cellular differentiation is interstitial cells ..-->.. brown preadipocytes ..-->.. mature brown adipocytes. Remarkable, labeling frequency did not change in endothelial cells during cold acclimation demonstrating that these cells cannot be considered as progenitors of brown adipocytes. It is suggested that brown adipocyte proliferation and differentiation from interstitial cells represent the fundamental phenomena explaining the enhanced capacity of cold-acclimated and/or hyperphagic rats to respond calorigenically to catecholamines.

  17. Transient hypoxia reprograms differentiating adipocytes for enhanced insulin sensitivity and triglyceride accumulation

    PubMed Central

    Lu, Hongyun; Gao, Zhanguo; Zhao, Zhiyun; Weng, Jianping; Ye, Jianping

    2015-01-01

    Objective To investigate the impact of transient (2-4 h) hypoxia on metabolic reprogramming of adipocytes. Methods The impact of transient hypoxia on metabolic reprogramming was investigated in 3T3-L1 cells before and after differentiation. Glucose uptake, fatty acid oxidation, lipolysis, and mitochondria were examined to determine the hypoxia effects. Preadipocytes were exposed to transient hypoxia (4h/day) in the course of differentiation. Insulin sensitivity and TG accumulation was examined in the cells at the end of differentiation to determine the reprogramming effects. AMPK activity and gene expression were determined by quantitative RT-PCR and Western blotting in search for mechanism of the reprogramming. Results In acute response to hypoxia, adipocytes exhibited an increase in insulin-dependent and -independent glucose uptake. Fatty acid β-oxidation and pyruvate dehydrogenase (PDH) activity were decreased. Multiple exposures of differentiating adipocytes to transient hypoxia enhanced insulin signaling, TG accumulation, expression of antioxidant genes in differentiated adipocytes in the absence of hypoxia. The metabolic memory was associated with elevated AMPK activity and gene expression (GLUT1, PGC-1α, PPARγ, SREBP, NRF-1, ESRRα, LPL). The enhanced insulin sensitivity was blocked by an AMPK inhibitor. Conclusions Repeated exposure of differentiating adipocytes to transient hypoxia is able to reprogram the cells for increased TG accumulation and enhanced insulin sensitivity. The metabolic alterations were observed in post-differentiated cells under normoxia. The reprogramming involves AMPK activation and gene expression in the metabolic pathways in cytosol and mitochondria. PMID:26219415

  18. Mitogen-Dependent Regulation of DUSP1 Governs ERK and p38 Signaling During Early 3T3-L1 Adipocyte Differentiation.

    PubMed

    Ferguson, Bradley S; Nam, Heesun; Stephens, Jacqueline M; Morrison, Ron F

    2016-07-01

    Knowledge concerning mechanisms that control proliferation and differentiation of preadipocytes is essential to our understanding of adipocyte hyperplasia and the development of obesity. Evidence has shown that temporal regulation of mitogen-activated protein kinase (MAPK) phosphorylation and dephosphorylation is critical for coupling extracellular stimuli to cellular growth and differentiation. Using differentiating 3T3-L1 preadipocytes as a model of adipocyte hyperplasia, we examined a role for dual-specificity phosphatase 1 (DUSP1) on the timely modulation of MAPK signaling during states of growth arrest, proliferation, and differentiation. Using real-time reverse transcription PCR (qRT-PCR), we report that DUSP1 is induced during early preadipocyte proliferation concomitant with ERK and p38 dephosphorylation. As deactivation of ERK and p38 is essential for the progression of adipocyte differentiation, we further showed that de novo mRNA synthesis was required for ERK and p38 dephosphorylation, suggesting a role for "inducible" phosphatases in regulating MAPK signaling. Pharmacological and genetic inhibition of DUSP1 markedly increased ERK and p38 phosphorylation during early adipocyte differentiation. Based on these findings, we postulated that loss of DUSP1 would block adipocyte hyperplasia. However, genetic loss of DUSP1 was not sufficient to prevent preadipocyte proliferation or differentiation, suggesting a role for other phosphatases in the regulation of adipogenesis. In support of this, qRT-PCR identified several MAPK-specific DUSPs induced during early (DUSP2, -4, -5, & -6), mid (DUSP4 & -16) and late (DUSP9) stages of adipocyte differentiation. Collectively, these data suggest an important role for DUSPs in regulating MAPK dephosphorylation, with an emphasis on DUSP1, during early adipogenesis. PMID:26566083

  19. Branched-chain amino acid catabolism fuels adipocyte differentiation and lipogenesis.

    PubMed

    Green, Courtney R; Wallace, Martina; Divakaruni, Ajit S; Phillips, Susan A; Murphy, Anne N; Ciaraldi, Theodore P; Metallo, Christian M

    2016-01-01

    Adipose tissue plays important roles in regulating carbohydrate and lipid homeostasis, but less is known about the regulation of amino acid metabolism in adipocytes. Here we applied isotope tracing to pre-adipocytes and differentiated adipocytes to quantify the contributions of different substrates to tricarboxylic acid (TCA) metabolism and lipogenesis. In contrast to proliferating cells, which use glucose and glutamine for acetyl-coenzyme A (AcCoA) generation, differentiated adipocytes showed increased branched-chain amino acid (BCAA) catabolic flux such that leucine and isoleucine from medium and/or from protein catabolism accounted for as much as 30% of lipogenic AcCoA pools. Medium cobalamin deficiency caused methylmalonic acid accumulation and odd-chain fatty acid synthesis. Vitamin B12 supplementation reduced these metabolites and altered the balance of substrates entering mitochondria. Finally, inhibition of BCAA catabolism compromised adipogenesis. These results quantitatively highlight the contribution of BCAAs to adipocyte metabolism and suggest that BCAA catabolism has a functional role in adipocyte differentiation. PMID:26571352

  20. Atypical antipsychotics induce both proinflammatory and adipogenic gene expression in human adipocytes in vitro.

    PubMed

    Sárvári, Anitta K; Veréb, Zoltán; Uray, Iván P; Fésüs, László; Balajthy, Zoltán

    2014-08-01

    Schizophrenia requires lifelong treatment, potentially causing systemic changes in metabolic homeostasis. In the clinical setting, antipsychotic treatment may differentially lead to weight gain among individual patients, although the molecular determinants of such adverse effects are currently unknown. In this study, we investigated changes in the expression levels of critical regulatory genes of adipogenesis, lipid metabolism and proinflammatory genes during the differentiation of primary human adipose-derived stem cells (ADSCs). These cells were isolated from patients with body mass indices <25 and treated with the second-generation antipsychotics olanzapine, ziprasidone, clozapine, quetiapine, aripiprazole and risperidone and the first-generation antipsychotic haloperidol. We found that antipsychotics exhibited a marked effect on key genes involved in the regulation of cell cycle, signal transduction, transcription factors, nuclear receptors, differentiation markers and metabolic enzymes. In particular, we observed an induction of the transcription factor NF-KB1 and NF-KB1 target genes in adipocytes in response to these drugs, including the proinflammatory cytokines TNF-α, IL-1β, IL-8 and MCP-1. In addition, enhanced secretion of both IL8 and MCP-1 was observed in the supernatant of these cell cultures. In addition to their remarkable stimulatory effects on proinflammatory gene transcription, three of the most frequently prescribed antipsychotic drugs, clozapine, quetiapine and aripiprazole, also induced the expression of essential adipocyte differentiation genes and the adipocyte hormones leptin and adiponectin, suggesting that both glucose and fat metabolism may be affected by these drugs. These data further suggest that antipsychotic treatments in patients alter the gene expression patterns in adipocytes in a coordinated fashion and priming them for a low-level inflammatory state. PMID:25019983

  1. Adipocyte and leptin accumulation in tumor-induced thymic involution.

    PubMed

    Lamas, Alejandro; Lopez, Elena; Carrio, Roberto; Lopez, Diana M

    2016-01-01

    Cell-mediated immunity is an important defense mechanism against pathogens and developing tumor cells. The thymus is the main lymphoid organ involved in the formation of the cell-mediated immune response by the maturation and differentiation of lymphocytes that travel from the bone marrow, through the lymphatic ducts, to become T lymphocytes. Thymic involution has been associated with aging; however, other factors such as obesity, viral infection and tumor development have been shown to increase the rate of shrinkage of this organ. The heavy infiltration of adipocyte fat cells has been reported in the involuted thymuses of aged mice. In the present study, the possible accumulation of such cells in the thymus during tumorigenesis was examined by immunohistochemistry. A significant number of adipocytes around and infiltrating the thymuses of tumor-bearing mice was observed. Leptin is a pro-inflammatory adipocytokine that enhances thymopoiesis and modulates T cell immune responses. The levels of leptin and adiponectin, another adipocytokine that has anti-inflammatory properties, were examined by western blot analysis. While no changes were observed in the amounts of adiponectin present in the thymuses of the normal and tumor-bearing mice, significantly higher levels of leptin were detected in the thymocytes of the tumor-bearing mice. This correlated with an increase in the expression of certain cytokines, such as interleukin (IL)-2, interferon (IFN)-γ and granulocyte-macrophage colony-stimulating factor (GM-CSF). The co-culture of thymocytes isolated from normal mice with ex vivo isolated adipocytes from tumor-bearing mice yielded similar results. Our findings suggest that the infiltration and accumulation of adipocytes in the thymuses of tumor-bearing mice play an important role in their altered morphology and functions. PMID:26530443

  2. Cardiac mesenchymal progenitors differentiate into adipocytes via Klf4 and c-Myc

    PubMed Central

    Kami, D; Kitani, T; Kawasaki, T; Gojo, S

    2016-01-01

    Direct reprogramming of differentiated cells to pluripotent stem cells has great potential to improve our understanding of developmental biology and disorders such as cancers, and has implications for regenerative medicine. In general, the effects of transcription factors (TFs) that are transduced into cells can be influenced by pre-existing transcriptional networks and epigenetic modifications. However, previous work has identified four key TFs, Oct4, Sox2, Klf4 and c-Myc, which can reprogram various differentiated cells to generate induced pluripotent stem cells. Here, we show that in the heart, the transduction of cardiac mesenchymal progenitors (CMPs) with Klf4 and c-Myc (KM) was sufficient to drive the differentiation of these cells into adipocytes without the use of adipogenic stimulation cocktail, that is, insulin, 3-isobutyl-1-methylxanthine (IBMX) and dexamethasone. KM-transduced CMPs exhibited a gradually increased expression of adipogenic-related genes, such as C/Ebpα, Pparγ and Fabp4, activation of the peroxisome proliferator-activated receptor (PPAR) signaling pathway, inactivation of the cell cycle-related pathway and formation of cytoplasmic lipid droplets within 10 days. In contrast, NIH3T3 fibroblasts, 3T3-L1 preadipocytes, and bone marrow-derived mesenchymal stem cells transduced with KM did not differentiate into adipocytes. Both in vitro and in vivo cardiac ischemia reperfusion injury models demonstrated that the expression of KM genes sharply increased following a reperfusion insult. These results suggest that ectopic adipose tissue formation in the heart following myocardial infarction results from CMPs that express KM following a stress response. PMID:27077806

  3. Effects of quercetin, a natural phenolic compound, in the differentiation of human mesenchymal stem cells (MSC) into adipocytes and osteoblasts.

    PubMed

    Casado-Díaz, Antonio; Anter, Jaouad; Dorado, Gabriel; Quesada-Gómez, José Manuel

    2016-06-01

    Natural phenols may have beneficial properties against oxidative stress, which is associated with aging and major chronic aging-related diseases, such as loss of bone mineral mass (osteoporosis) and diabetes. The main aim of this study was to analyze the effect of quercetin, a major nutraceutical compound present in the "Mediterranean diet", on mesenchymal stem-cell (MSC) differentiation. Such cells were induced to differentiate into osteoblasts or adipocytes in the presence of two quercetin concentrations (0.1 and 10μM). Several physiological parameters and the expression of osteoblastogenesis and adipogenesis marker genes were monitored. Quercetin (10μM) inhibited cell proliferation, alkaline phosphatase (ALPL) activity and mineralization, down-regulating the expression of ALPL, collagen type I alpha 1 (COL1A1) and osteocalcin [bone gamma-carboxyglutamate protein (BGLAP)] osteoblastogenesis-related genes in MSC differentiating into osteoblasts. Moreover, in these cultures, CCAAT/enhancer-binding protein alpha (CEBPA) and peroxisome proliferator-activated receptor gamma 2 (PPARG2) adipogenic genes were induced, and cells differentiated into adipocytes were observed. Quercetin did not affect proliferation, but increased adipogenesis, mainly at 10-μM concentration in MSC induced to differentiate to adipocytes. β- and γ-catenin (plakoglobin) nuclear levels were reduced and increased, respectively, in quercetin-treated cultures. This suggests that the effect of high concentration of quercetin on MSC osteoblastic and adipogenic differentiation is mediated via Wnt/β-catenin inhibition. In conclusion, quercetin supplementation inhibited osteoblastic differentiation and promoted adipogenesis at the highest tested concentration. Such possible adverse effects of high quercetin concentrations should be taken into account in nutraceutical or pharmaceutical strategies using such flavonol. PMID:27142748

  4. Methylation of miR-145a-5p promoter mediates adipocytes differentiation.

    PubMed

    Du, Jingjing; Cheng, Xiao; Shen, Linyuan; Tan, Zhendong; Luo, Jia; Wu, Xiaoqian; Liu, Chendong; Yang, Qiong; Jiang, Yanzhi; Tang, Guoqing; Li, Xuewei; Zhang, Shunhua; Zhu, Li

    2016-06-17

    MicroRNAs (miRNAs, miR) play important roles in adipocyte development. Recent studies showed that the expression of several miRNAs is closely related with promoter methylation. However, it is not known whether miRNA mediates adipocytes differentiation by means of DNA methylation. Here, we showed that miR-145a-5p was poorly expressed in adipose tissue from mice fed a high fat diet (HFD). Overexpression or inhibition of miR-145a-5p was unfavorable or beneficial, respectively, for adipogenesis, and these effects were achieved by regulating adipocyte-specific genes involved in lipogenic transcription, fatty acid synthesis, and fatty acid transportation. Particularly, we first suggested that miR-145a-5p mimics or inhibitors promoted or repressed adipocytes proliferation by regulating p53 and p21, which act as cell cycle regulating factors. Surprisingly, the miR-145a-5p-repressed adipocyte differentiation was enhanced or rescued when cells treated with 5-Aza-dC were transfected with miR-145a-5p mimics or inhibitors, respectively. These data indicated that, as a new mean to positively regulate adipocyte proliferation, the process of miR-145a-5p-inhibited adipogenesis may be regulated by DNA methylation. PMID:27179777

  5. Luteolin suppresses TCDD-induced wasting syndrome in a cultured adipocyte model.

    PubMed

    Ashida, Hitoshi; Harada, Kiyonari; Mishima, Sakiho; Mitani, Takakazu; Yamashita, Yoko; Matsumura, Fumio

    2015-05-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various toxic effects, including wasting syndrome, through activation of an aryl hydrocarbon receptor (AhR). Our previous report demonstrated that certain flavonoids inhibit the activation of AhR and suppress its DNA binding activity. In this study, we searched for an active compound among 13 flavonoids that suppressed TCDD-induced loss of lipid accumulation using 3T3-L1 adipocytes as a cell culture model for wasting syndrome. Two flavonoids, luteolin and epigallocatechin gallate, suppressed TCDD-induced loss of lipid accumulation in this model. We further investigated luteolin to clarify the underlying molecular mechanism and confirmed that luteolin inhibited nuclear translocation of AhR caused by TCDD. Luteolin also inhibited the TCDD-driven decrease in protein expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα). Although TCDD alone did not change protein expression of C/EBPβ and C/EBPδ, luteolin and TCDD up-regulated C/EBPδ expression in a dose-dependent manner. On the other hand, TCDD significantly decreased DNA binding of C/EBPβ and C/EBPδ, and luteolin completely canceled TCDD-decreased DNA binding of them. We conclude that luteolin suppresses the TCDD-induced loss of lipid accumulation in 3T3-L1 adipocytes by preventing a decrease in protein expression of PPARγ and C/EBPα, the master regulators of adipocyte differentiation and in DNA binding of C/EBPβ and C/EBPδ. Moreover, luteolin was rapidly incorporated and accumulated in 3T3-L1 adipocytes. Thus, luteolin is an attractive compound for the prevention of TCDD-induced wasting syndrome. PMID:25987215

  6. Differentiation of rat brown adipocytes during late foetal development: role of insulin-like growth factor I.

    PubMed Central

    Teruel, T; Valverde, A M; Alvarez, A; Benito, M; Lorenzo, M

    1995-01-01

    Rat brown adipocytes at day 22 of foetal development showed greater size, higher mitochondria content and larger amounts of lipids, as determined by flow cytometry, than 20-day foetal cells. Simultaneously, an inhibition on the percentage of brown adipocytes into S+G2/M phases of the cell cycle was observed between days 20 and 22 of foetal development. The expression of several adipogenesis-related genes, such as fatty acid synthase, malic enzyme, glucose-6-phosphate dehydrogenase and insulin-regulated glucose transporter, increased at the end of foetal life in brown adipose tissue. In addition, the lipogenic enzyme activities and the lipogenic flux increased during late foetal development, resulting in mature brown adipocytes showing a multilocular fat droplet phenotype. Concurrently, brown adipocytes induced the expression of the uncoupling protein (UP) mRNA and UP protein, as visualized by immunofluorescence. The three isoforms of CCAAT enhancer-binding proteins (C/EBPs) were expressed at the mRNA level in brown adipose tissue at day 20. C/EBP alpha decreased and C/EBP beta and delta increased their expression between days 20 and 22 of foetal development, respectively. Brown adipose tissue constitutively expressed insulin-like growth factor I (IGF-I) and IGF-I receptor (IGF-IR) mRNAs. Moreover, IGF-IR mRNA content increased between days 20 and 22 in parallel with the occurrence of tissue differentiation. Images Figure 2 Figure 3 Figure 4 PMID:7575409

  7. PKC-ALDH2 Pathway Plays a Novel Role in Adipocyte Differentiation

    PubMed Central

    Yu, Yu-Hsiang; Liao, Pei-Ru; Guo, Chien-Jung; Chen, Che-Hong; Mochly-Rosen, Daria; Chuang, Lee-Ming

    2016-01-01

    The ALDH2 gene encodes the mitochondrial aldehyde dehydrogenase 2 (ALDH2), a critical enzyme involved in ethanol clearance through acetaldehyde metabolism. ALDH2 also catalyzes the metabolism of other bioreactive aldehydes, including propionaldehyde, butyraldehyde, and 4-hydroxykenals (4-HNE). Increased levels of 4-HNE in adipose tissue positively correlate with obesity and insulin resistance. However, it remains unclear whether ALDH2 is involved in regulation of adipocyte differentiation. Here, we found that ALDH2 protein levels were lower in white adipose tissue of high-fat diet-fed mice and ob/ob mice relative to lean mice. Knockdown of ALDH2 expression in 3T3-L1 preadipocytes caused an increase in intracellular 4-HNE, thereby attenuated adipocyte differentiation. By contrast, an ALDH2 activator, Alda-1, significantly accelerated adipogenesis, which was accompanied by an increase in adipogenic gene expression. Consistently, adipogenesis was reduced when protein kinase C ε (PKCε), an ALDH2 phosphorylating activator, was silenced in 3T3-L1 preadipocytes, whereas treatment with a PKCε agonist in 3T3-L1 preadipocytes enhanced adipogenesis. Whole-genome microarray profiling of Alda-1-treated cells demonstrated several upregulated transcripts encoding proteins involved in metabolism and the majority of these transcripts are for proteins involved in PPAR signaling pathways. Furthermore, PKCε-ALDH2 interaction alleviates 4-HNE induced aberrant PPARγ regulation on adipogenesis. Taken together, these results demonstrate that ALDH2 activation enhances adipogenesis and signaling pathways involving PPARγ. Thus, activation of PKCε-ALDH2 regulatory axis may be a therapeutic target for treating obesity and type 2 diabetes. PMID:27575855

  8. PKC-ALDH2 Pathway Plays a Novel Role in Adipocyte Differentiation.

    PubMed

    Yu, Yu-Hsiang; Liao, Pei-Ru; Guo, Chien-Jung; Chen, Che-Hong; Mochly-Rosen, Daria; Chuang, Lee-Ming

    2016-01-01

    The ALDH2 gene encodes the mitochondrial aldehyde dehydrogenase 2 (ALDH2), a critical enzyme involved in ethanol clearance through acetaldehyde metabolism. ALDH2 also catalyzes the metabolism of other bioreactive aldehydes, including propionaldehyde, butyraldehyde, and 4-hydroxykenals (4-HNE). Increased levels of 4-HNE in adipose tissue positively correlate with obesity and insulin resistance. However, it remains unclear whether ALDH2 is involved in regulation of adipocyte differentiation. Here, we found that ALDH2 protein levels were lower in white adipose tissue of high-fat diet-fed mice and ob/ob mice relative to lean mice. Knockdown of ALDH2 expression in 3T3-L1 preadipocytes caused an increase in intracellular 4-HNE, thereby attenuated adipocyte differentiation. By contrast, an ALDH2 activator, Alda-1, significantly accelerated adipogenesis, which was accompanied by an increase in adipogenic gene expression. Consistently, adipogenesis was reduced when protein kinase C ε (PKCε), an ALDH2 phosphorylating activator, was silenced in 3T3-L1 preadipocytes, whereas treatment with a PKCε agonist in 3T3-L1 preadipocytes enhanced adipogenesis. Whole-genome microarray profiling of Alda-1-treated cells demonstrated several upregulated transcripts encoding proteins involved in metabolism and the majority of these transcripts are for proteins involved in PPAR signaling pathways. Furthermore, PKCε-ALDH2 interaction alleviates 4-HNE induced aberrant PPARγ regulation on adipogenesis. Taken together, these results demonstrate that ALDH2 activation enhances adipogenesis and signaling pathways involving PPARγ. Thus, activation of PKCε-ALDH2 regulatory axis may be a therapeutic target for treating obesity and type 2 diabetes. PMID:27575855

  9. Investigating the role of class-IA PI 3-kinase isoforms in adipocyte differentiation

    SciTech Connect

    Kim, Ji Eun; Shepherd, Peter R. Chaussade, Claire

    2009-02-20

    PI 3-kinases, in particular class-IA, are key signalling molecules controlling many cellular processes including growth, proliferation, migration and differentiation. In this study, we have used a collection of isoform selective PI 3-kinase inhibitors to determine whether attenuation of signalling through class-IA PI 3-kinase isoforms will impact adipocyte differentiation. First, we analysed the expression profiles and found that fibroblastic pre-adipocytes express detectable levels of p110{alpha} and p110{delta} and that after differentiation, p110{delta} levels fall while p110{alpha} levels rise, together with C/EBP{alpha} and PPAR{gamma}. When using specific inhibitors during the differentiation process, we observed that neither p110{beta} nor p110{delta} inhibition, had any significant effect. In contrast PIK-75, a selective p110{alpha} inhibitor completely abolished adipocyte differentiation as assessed by morphology, transcript and protein levels of adipocyte markers. These results indicate that long term treatment with p110{alpha} inhibitors could potentially have a severe impact on fat cell numbers in vivo.

  10. ATF3 inhibits adipocyte differentiation of 3T3-L1 cells

    SciTech Connect

    Jang, Min Kyung; Kim, Cho Hee; Seong, Je Kyung; Jung, Myeong Ho

    2012-04-27

    Highlights: Black-Right-Pointing-Pointer Overexpression of ATF3 inhibits adipocyte differentiation in 3T3-L1 cells. Black-Right-Pointing-Pointer Overexpression of ATF3 represses C/EBP{alpha} expression. Black-Right-Pointing-Pointer ATF3 directly binds to mouse C/EBP{alpha} promoter spanning from -1928 to -1907. Black-Right-Pointing-Pointer ATF3 may play a role in hypoxia-mediated inhibition of adipocyte differentiation. -- Abstract: ATF3 is a stress-adaptive gene that regulates proliferation or apoptosis under stress conditions. However, the role of ATF3 is unknown in adipocyte cells. Therefore, in this study, we investigated the functional role of ATF3 in adipocytes. Both lentivirus-mediated overexpression of ATF3 and stably-overexpressed ATF3 inhibited adipocyte differentiation in 3T3-L1 cells, as revealed by decreased lipid staining with oil red staining and reduction in adipogenic genes. Thapsigargin treatment and overexpression of ATF3 decreased C/EBP{alpha} transcript and repressed the activity of the 3.6-kb mouse C/EBP{alpha} promoter, demonstrating that ATF3 downregulates C/EBP{alpha} expression. Transfection studies using mutant constructs containing 5 Prime -deletions in the C/EBP{alpha} promoter revealed that a putative ATF/CRE element, GGATGTCA, is located between -1921 and -1914. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay demonstrated that ATF3 directly binds to mouse C/EBP{alpha} promoter spanning from -1928 to -1907. Both chemical hypoxia-mimetics or physical hypoxia led to reduce the C/EBP{alpha} mRNA and repress the promoter activity of the C/EBP{alpha} gene, whereas increase ATF3 mRNA, suggesting that ATF3 may contribute to the inhibition of adipocyte differentiation in hypoxia through downregulation of C/EBP{alpha} expression. Collectively, these results demonstrate that ATF3 represses the C/EBP{alpha} gene, resulting in inhibition of adipocyte differentiation, and thus plays a role in hypoxia-mediated inhibition

  11. Mitochondrial (Dys)function in Adipocyte (De)differentiation and Systemic Metabolic Alterations

    PubMed Central

    De Pauw, Aurélia; Tejerina, Silvia; Raes, Martine; Keijer, Jaap; Arnould, Thierry

    2009-01-01

    In mammals, adipose tissue, composed of BAT and WAT, collaborates in energy partitioning and performs metabolic regulatory functions. It is the most flexible tissue in the body, because it is remodeled in size and shape by modifications in adipocyte cell size and/or number, depending on developmental status and energy fluxes. Although numerous reviews have focused on the differentiation program of both brown and white adipocytes as well as on the pathophysiological role of white adipose tissues, the importance of mitochondrial activity in the differentiation or the dedifferentiation programs of adipose cells and in systemic metabolic alterations has not been extensively reviewed previously. Here, we address the crucial role of mitochondrial functions during adipogenesis and in mature adipocytes and discuss the cellular responses of white adipocytes to mitochondrial activity impairment. In addition, we discuss the increase in scientific knowledge regarding mitochondrial functions in the last 10 years and the recent suspicion of mitochondrial dysfunction in several 21st century epidemics (ie, obesity and diabetes), as well as in lipodystrophy found in HIV-treated patients, which can contribute to the development of new therapeutic strategies targeting adipocyte mitochondria. PMID:19700756

  12. Functional characterization of retromer in GLUT4 storage vesicle formation and adipocyte differentiation.

    PubMed

    Yang, Zhe; Hong, Lee Kian; Follett, Jordan; Wabitsch, Martin; Hamilton, Nicholas A; Collins, Brett M; Bugarcic, Andrea; Teasdale, Rohan D

    2016-03-01

    Insulin-stimulated translocation of glucose transporter 4 (GLUT4) storage vesicles (GSVs), the specialized intracellular compartments within mature adipocytes, to the plasma membrane (PM) is a fundamental cellular process for maintaining glucose homeostasis. Using 2 independent adipocyte cell line models, human primary Simpson-Golabi-Behmel syndrome and mouse 3T3-L1 fibroblast cell lines, we demonstrate that the endosome-associated protein-sorting complex retromer colocalizes with GLUT4 on the GSVs by confocal microscopy in mature adipocytes. By use of both confocal microscopy and differential ultracentrifugation techniques, retromer is redistributed to the PM of mature adipocytes upon insulin stimulation. Furthermore, stable knockdown of the retromer subunit-vacuolar protein-sorting 35, or the retromer-associated protein sorting nexin 27, by lentivirus-delivered small hairpin RNA impaired the adipogenesis process when compared to nonsilence control. The knockdown of retromer decreased peroxisome proliferator activated receptor γ expression during differentiation, generating adipocytes with decreased levels of GSVs, lipid droplet accumulation, and insulin-stimulated glucose uptake. In conclusion, our study demonstrates a role for retromer in the GSV formation and adipogenesis. PMID:26581601

  13. miR-140-5p regulates adipocyte differentiation by targeting transforming growth factor-β signaling.

    PubMed

    Zhang, Xin; Chang, Ailing; Li, Yongmei; Gao, Yifei; Wang, Haixiao; Ma, Zhongshu; Li, Xiaoxia; Wang, Baoli

    2015-01-01

    Recent emerging studies of miRNAs in adipocyte commitment provide new insights to understand the molecular basis of adipogenesis. The current study indicated that miR-140-5p was altered in primary cultured marrow stromal cells and established progenitor lines after adipogenic and/or osteogenic treatment. miR-140-5p was increased in adipose tissue in db/db obese mice vs. lean mice. Supplementing miR-140-5p activity induced stromal cell ST2 and preadipocyte 3T3-L1 to differentiate into mature adipocytes. Conversely, inhibition of the endogenous miR-140-5p repressed ST2 and 3T3-L1 to fully differentiate. By contrast, knockdown of the endogenous miR-140-5p enhanced osteoblast differentiation. Transforming growth factor-β receptor I (Tgfbr1) was shown to be a direct target of miR-140-5p. Supplementing miR-140-5p in ST2 reduced the level of TGFBR1 protein, while suppression of endogenous miR-140-5p increased TGFBR1. Overexpression of Tgfbr1 inhibited, whereas knockdown of Tgfbr1 promoted adipogenic differentiation of ST2 cells. Further investigation of mechanisms that control miR-140-5p expression revealed that C/EBPα induced transcriptional activity of the miR-140-5p promoter. Removal of the putative response element of C/EBP from the promoter abolished the enhancement of the promoter activity by C/EBPα, suggesting that C/EBPα transcriptionally controls miR-140-5p expression. Taken together, our study provides evidences that miR-140-5p regulates adipocyte differentiation through a C/EBP/miR-140-5p/TGFBR1 regulatory feedback loop. PMID:26657345

  14. Cocoa tea (Camellia ptilophylla) water extract inhibits adipocyte differentiation in mouse 3T3-L1 preadipocytes

    PubMed Central

    Li, Kai Kai; Liu, Chuek Lun; Shiu, Hoi Ting; Wong, Hing Lok; Siu, Wing Sum; Zhang, Cheng; Han, Xiao Qiang; Ye, Chuang Xing; Leung, Ping Chung; Ko, Chun Hay

    2016-01-01

    Cocoa tea (Camellia ptilophylla) is a naturally decaffeinated tea plant. Previously we found that cocoa tea demonstrated a beneficial effect against high-fat diet induced obesity, hepatic steatosis, and hyperlipidemia in mice. The present study aimed to investigate the anti-adipogenic effect of cocoa tea in vitro using preadipocytes 3T3-L1. Adipogenic differentiation was confirmed by Oil Red O stain, qPCR and Western blot. Our results demonstrated that cocoa tea significantly inhibited triglyceride accumulation in mature adipocytes in a dose-dependent manner. Cocoa tea was shown to suppress the expressions of key adipogenic transcription factors, including peroxisome proliferator-activated receptor gamma (PPAR γ) and CCAAT/enhancer binding protein (C/EBP α). The tea extract was subsequently found to reduce the expressions of adipocyte-specific genes such as sterol regulatory element binding transcription factor 1c (SREBP-1c), fatty acid synthase (FAS), Acetyl-CoA carboxylase (ACC), fatty acid translocase (FAT) and stearoylcoenzyme A desaturase-1 (SCD-1). In addition, JNK, ERK and p38 phosphorylation were inhibited during cocoa tea inhibition of 3T3-L1 adipogenic differentiation. Taken together, this is the first study that demonstrates cocoa tea has the capacity to suppress adipogenesis in pre-adipocyte 3T3-L1 similar to traditional green tea PMID:26833256

  15. Cocoa tea (Camellia ptilophylla) water extract inhibits adipocyte differentiation in mouse 3T3-L1 preadipocytes.

    PubMed

    Li, Kai Kai; Liu, Chuek Lun; Shiu, Hoi Ting; Wong, Hing Lok; Siu, Wing Sum; Zhang, Cheng; Han, Xiao Qiang; Ye, Chuang Xing; Leung, Ping Chung; Ko, Chun Hay

    2016-01-01

    Cocoa tea (Camellia ptilophylla) is a naturally decaffeinated tea plant. Previously we found that cocoa tea demonstrated a beneficial effect against high-fat diet induced obesity, hepatic steatosis, and hyperlipidemia in mice. The present study aimed to investigate the anti-adipogenic effect of cocoa tea in vitro using preadipocytes 3T3-L1. Adipogenic differentiation was confirmed by Oil Red O stain, qPCR and Western blot. Our results demonstrated that cocoa tea significantly inhibited triglyceride accumulation in mature adipocytes in a dose-dependent manner. Cocoa tea was shown to suppress the expressions of key adipogenic transcription factors, including peroxisome proliferator-activated receptor gamma (PPAR γ) and CCAAT/enhancer binding protein (C/EBP α). The tea extract was subsequently found to reduce the expressions of adipocyte-specific genes such as sterol regulatory element binding transcription factor 1c (SREBP-1c), fatty acid synthase (FAS), Acetyl-CoA carboxylase (ACC), fatty acid translocase (FAT) and stearoylcoenzyme A desaturase-1 (SCD-1). In addition, JNK, ERK and p38 phosphorylation were inhibited during cocoa tea inhibition of 3T3-L1 adipogenic differentiation. Taken together, this is the first study that demonstrates cocoa tea has the capacity to suppress adipogenesis in pre-adipocyte 3T3-L1 similar to traditional green tea. PMID:26833256

  16. Control of Adipocyte Differentiation in Different Fat Depots; Implications for Pathophysiology or Therapy

    PubMed Central

    Ma, Xiuquan; Lee, Paul; Chisholm, Donald J.; James, David E.

    2015-01-01

    Adipocyte differentiation and its impact on restriction or expansion of particular adipose tissue depots have physiological and pathophysiological significance in view of the different functions of these depots. Brown or “beige” fat [brown adipose tissue (BAT)] expansion can enhance thermogenesis, lipid oxidation, insulin sensitivity, and glucose tolerance; conversely expanded visceral fat [visceral white adipose tissue (VAT)] is associated with insulin resistance, low grade inflammation, dyslipidemia, and cardiometabolic risk. The largest depot, subcutaneous white fat [subcutaneous white adipose tissue (SAT)], has important beneficial characteristics including storage of lipid “out of harms way” and secretion of adipokines, especially leptin and adiponectin, with positive metabolic effects including lipid oxidation, energy utilization, enhanced insulin action, and an anti-inflammatory role. The absence of these functions in lipodystrophies leads to major metabolic disturbances. An ability to expand white adipose tissue adipocyte differentiation would seem an important defense mechanism against the detrimental effects of energy excess and limit harmful accumulation of lipid in “ectopic” sites, such as liver and muscle. Adipocyte differentiation involves a transcriptional cascade with PPARγ being most important in SAT but less so in VAT, with increased angiogenesis also critical. The transcription factor, Islet1, is fairly specific to VAT and in vitro inhibits adipocyte differentiation. The physiological importance of Islet1 requires further study. Basic control of differentiation is similar in BAT but important differences include the effect of PGC-1α on mitochondrial biosynthesis and upregulation of UCP1; also PRDM16 plays a pivotal role in expression of the BAT phenotype. Modulation of the capacity or function of these different adipose tissue depots, by altering adipocyte differentiation or other means, holds promise for interventions that can be

  17. Phloretin promotes adipocyte differentiation in vitro and improves glucose homeostasis in vivo

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Adipocyte dysfunction is associated with many metabolic diseases such as obesity, insulin resistance and diabetes. Previous studies found that phloretin promotes 3T3-L1 cells differentiation, but the underlying mechanisms for phloretin's effects on adipogenesis remain unclear. In this study, we demo...

  18. Sclerostin Enhances Adipocyte Differentiation in 3T3-L1 Cells.

    PubMed

    Ukita, Mayumi; Yamaguchi, Taihiko; Ohata, Noboru; Tamura, Masato

    2016-06-01

    Sclerostin, a secreted protein encoded by the Sost gene, is produced by osteocytes and is inhibited by osteoblast differentiation and bone formation. Recently, a functional association between bone and fat tissue has been suggested, and a correlation between circulating sclerostin levels and lipid metabolism has been reported in humans. However, the effects of sclerostin on adipogenesis remain unexplored. In the present study, we examined the role of sclerostin in regulating adipocyte differentiation using 3T3-L1 preadipocytes. In these cells, sclerostin enhanced adipocyte-specific gene expression and the accumulation of lipid deposits. Sclerostin also upregulated CCAAT/enhancer binding protein β expression but not cell proliferation and caspase-3/7 activities. Sclerostin also attenuated canonical Wnt3a-inhibited adipocyte differentiation. Recently, the transcriptional modulator TAZ has been involved in the canonical Wnt signaling pathway. Sclerostin reduced TAZ-responsive transcriptional activity and TAZ-responsive gene expression. Transfection of 3T3-L1 cells with TAZ siRNA increased the lipid deposits and adipogenic gene expression. These results show that sclerostin upregulates adipocyte differentiation in 3T3-L1 cells, suggesting a possible role for the osteocyte-derived sclerostin as a regulator of fat metabolism and as a reciprocal regulator of bone and adipose tissues metabolism. J. Cell. Biochem. 117: 1419-1428, 2016. © 2015 Wiley Periodicals, Inc. PMID:26553151

  19. Hsp90 chaperones PPARγ and regulates differentiation and survival of 3T3-L1 adipocytes

    PubMed Central

    Nguyen, M T; Csermely, P; Sőti, C

    2013-01-01

    Adipose tissue dysregulation has a major role in various human diseases. The peroxisome proliferator-activated receptor-γ (PPARγ) is a key regulator of adipocyte differentiation and function, as well as a target of insulin-sensitizing drugs. The Hsp90 chaperone stabilizes a diverse set of signaling ‘client' proteins, thereby regulates various biological processes. Here we report a novel role for Hsp90 in controlling PPARγ stability and cellular differentiation. Specifically, we show that the Hsp90 inhibitors geldanamycin and novobiocin efficiently impede the differentiation of murine 3T3-L1 preadipocytes. Geldanamycin at higher concentrations also inhibits the survival of both developing and mature adipocytes, respectively. Further, Hsp90 inhibition disrupts an Hsp90-PPARγ complex, leads to the destabilization and proteasomal degradation of PPARγ, and inhibits the expression of PPARγ target genes, identifying PPARγ as an Hsp90 client. A similar destabilization of PPARγ and a halt of adipogenesis also occur in response to protein denaturing stresses caused by a single transient heat-shock or proteasome inhibition. Recovery from stress restores PPARγ stability and adipocyte differentiation. Thus, our findings reveal Hsp90 as a critical stress-responsive regulator of adipocyte biology and offer a potential therapeutic target in obesity and the metabolic syndrome. PMID:24096869

  20. Insulin/glucose induces natriuretic peptide clearance receptor in human adipocytes: a metabolic link with the cardiac natriuretic pathway.

    PubMed

    Bordicchia, M; Ceresiani, M; Pavani, M; Minardi, D; Polito, M; Wabitsch, M; Cannone, V; Burnett, J C; Dessì-Fulgheri, P; Sarzani, R

    2016-07-01

    Cardiac natriuretic peptides (NP) are involved in cardiorenal regulation and in lipolysis. The NP activity is largely dependent on the ratio between the signaling receptor NPRA and the clearance receptor NPRC. Lipolysis increases when NPRC is reduced by starving or very-low-calorie diet. On the contrary, insulin is an antilipolytic hormone that increases sodium retention, suggesting a possible functional link with NP. We examined the insulin-mediated regulation of NP receptors in differentiated human adipocytes and tested the association of NP receptor expression in visceral adipose tissue (VAT) with metabolic profiles of patients undergoing renal surgery. Differentiated human adipocytes from VAT and Simpson-Golabi-Behmel Syndrome (SGBS) adipocyte cell line were treated with insulin in the presence of high-glucose or low-glucose media to study NP receptors and insulin/glucose-regulated pathways. Fasting blood samples and VAT samples were taken from patients on the day of renal surgery. We observed a potent insulin-mediated and glucose-dependent upregulation of NPRC, through the phosphatidylinositol 3-kinase pathway, associated with lower lipolysis in differentiated adipocytes. No effect was observed on NPRA. Low-glucose medium, used to simulate in vivo starving conditions, hampered the insulin effect on NPRC through modulation of insulin/glucose-regulated pathways, allowing atrial natriuretic peptide to induce lipolysis and thermogenic genes. An expression ratio in favor of NPRC in adipose tissue was associated with higher fasting insulinemia, HOMA-IR, and atherogenic lipid levels. Insulin/glucose-dependent NPRC induction in adipocytes might be a key factor linking hyperinsulinemia, metabolic syndrome, and higher blood pressure by reducing NP effects on adipocytes. PMID:27101299

  1. White Tea extract induces lipolytic activity and inhibits adipogenesis in human subcutaneous (pre)-adipocytes

    PubMed Central

    Söhle, Jörn; Knott, Anja; Holtzmann, Ursula; Siegner, Ralf; Grönniger, Elke; Schepky, Andreas; Gallinat, Stefan; Wenck, Horst; Stäb, Franz; Winnefeld, Marc

    2009-01-01

    Background The dramatic increase in obesity-related diseases emphasizes the need to elucidate the cellular and molecular mechanisms underlying fat metabolism. To investigate how natural substances influence lipolysis and adipogenesis, we determined the effects of White Tea extract on cultured human subcutaneous preadipocytes and adipocytes. Methods For our in vitro studies we used a White Tea extract solution that contained polyphenols and methylxanthines. Utilizing cultured human preadipocytes we investigated White Tea extract solution-induced inhibition of triglyceride incorporation during adipogenesis and possible effects on cell viability. In vitro studies on human adipocytes were performed aiming to elucidate the efficacy of White Tea extract solution to stimulate lipolytic activity. To characterize White Tea extract solution-mediated effects on a molecular level, we analyzed gene expression of essential adipogenesis-related transcription factors by qRT-PCR and determined the expression of the transcription factor ADD1/SREBP-1c on the protein level utilizing immunofluorescence analysis. Results Our data show that incubation of preadipocytes with White Tea extract solution significantly decreased triglyceride incorporation during adipogenesis in a dose-dependent manner (n = 10) without affecting cell viability (n = 10). These effects were, at least in part, mediated by EGCG (n = 10, 50 μM). In addition, White Tea extract solution also stimulated lipolytic activity in adipocytes (n = 7). Differentiating preadipocytes cultivated in the presence of 0.5% White Tea extract solution showed a decrease in PPARγ, ADD1/SREBP-1c, C/EBPα and C/EBPδ mRNA levels. Moreover, the expression of the transcription factor ADD1/SREBP-1c was not only decreased on the mRNA but also on the protein level. Conclusion White Tea extract is a natural source that effectively inhibits adipogenesis and stimulates lipolysis-activity. Therefore, it can be utilized to modulate different

  2. Cathepsin S controls adipocytic and osteoblastic differentiation, bone turnover, and bone microarchitecture.

    PubMed

    Rauner, M; Föger-Samwald, U; Kurz, M F; Brünner-Kubath, C; Schamall, D; Kapfenberger, A; Varga, P; Kudlacek, S; Wutzl, A; Höger, H; Zysset, P K; Shi, G P; Hofbauer, L C; Sipos, W; Pietschmann, P

    2014-07-01

    Cathepsin S is a cysteine protease that controls adipocyte differentiation and has been implicated in vascular and metabolic complications of obesity. Considering the inverse relation of osteoblasts and adipocytes and their mutual precursor cell, we hypothesized that cathepsin S may also affect osteoblast differentiation and bone remodeling. Thus, the fat and bone phenotypes of young (3 months old) and aged (12 or 18 months old) cathepsin S knock-out (KO) and wild-type (WT) mice were determined. Cathepsin S KO mice had a normal body weight at both ages investigated, even though the amount of subscapular and gonadal fat pads was reduced by 20%. Further, cathepsin S deficiency impaired adipocyte formation (-38%, p<0.001), which was accompanied by a lower expression of adipocyte-related genes and a reduction in serum leptin, IL-6 and CCL2 (p<0.001). Micro-CT analysis revealed an unchanged trabecular bone volume fraction and density, while tissue mineral density was significantly lower in cathepsin S KO mice at both ages. Aged KO mice further had a lower cortical bone mass (-2.3%, p<0.05). At the microarchitectural level, cathepsin S KO mice had thinner trabeculae (-8.3%), but a better connected trabecular network (+24%). Serum levels of the bone formation marker type 1 procollagen amino-terminal-propeptide and osteocalcin were both 2-3-fold higher in cathepsin S KO mice as was the mineralized surface. Consistently, osteogenic differentiation was increased 2-fold along with an increased expression of osteoblast-specific genes. Interestingly, serum levels of C-terminal telopeptide of type I collagen were also higher (+43%) in cathepsin S KO mice as were histological osteoclast parameters and ex vivo osteoclast differentiation. Thus, cathepsin S deficiency alters the balance between adipocyte and osteoblast differentiation, increases bone turnover, and changes bone microarchitecture. Therefore, bone and fat metabolisms should be monitored when using cathepsin S

  3. Critical role for cytosolic group IVA phospholipase A2 in early adipocyte differentiation and obesity.

    PubMed

    Peña, Lucía; Meana, Clara; Astudillo, Alma M; Lordén, Gema; Valdearcos, Martín; Sato, Hiroyasu; Murakami, Makoto; Balsinde, Jesús; Balboa, María A

    2016-09-01

    Adipogenesis is the process of differentiation of immature mesenchymal stem cells into adipocytes. Elucidation of the mechanisms that regulate adipocyte differentiation is key for the development of novel therapies for the control of obesity and related comorbidities. Cytosolic group IVA phospholipase A2 (cPLA2α) is the pivotal enzyme in receptor-mediated arachidonic acid (AA) mobilization and attendant eicosanoid production. Using primary multipotent cells and cell lines predetermined to become adipocytes, we show here that cPLA2α displays a proadipogenic function that occurs very early in the adipogenic process. Interestingly, cPLA2α levels decrease during adipogenesis, but cPLA2α-deficient preadipocytes exhibit a reduced capacity to differentiate into adipocytes, which affects early and terminal adipogenic transcription factors. Additionally, the absence of the phospholipase alters proliferation and cell-cycle progression that takes place during adipogenesis. Preconditioning of preadipocytes with AA increases the adipogenic capacity of these cells. Moreover, animals deficient in cPLA2α show resistance to obesity when fed a high fat diet that parallels changes in the expression of adipogenic transcription factors of the adipose tissue. Collectively, these results show that preadipocyte cPLA2α activation is a hitherto unrecognized factor for adipogenesis in vitro and in vivo. PMID:27317983

  4. Cucurbitacin B and cucurbitacin I suppress adipocyte differentiation through inhibition of STAT3 signaling.

    PubMed

    Seo, Cho-Rong; Yang, Dong Kwon; Song, No-Joon; Yun, Ui Jeong; Gwon, A-Ryeong; Jo, Dong-Gyu; Cho, Jae Youl; Yoon, Keejung; Ahn, Jee-Yin; Nho, Chu Won; Park, Woo Jin; Yang, Seung Yul; Park, Kye Won

    2014-02-01

    Cucurbitacin B, a member of the cucurbitaceae family, can act as a STAT3 signaling inhibitor to regulate the growth of hepatocellular carcinoma. STAT3 signaling has been shown to inhibit adipocyte differentiation through C/EBPα and PPARγ. Based on these studies, we hypothesized that cucurbitacin B would prevent PPARγ mediated adipocyte differentiation through STAT3 signaling. To test this hypothesis, mesenchymal C3H10T1/2 and 3T3-L1 preadipocyte cells were treated with a sub-cytotoxic concentration of cucurbitacin B. Cucurbitacin B treatment inhibits lipid accumulation and expression of adipocyte markers including PPARγ and its target genes in a dose-dependent manner. Cucurbitacin B treatment impairs STAT3 signaling as manifested by reduced phosphorylation of STAT3 and suppression of STAT3 target gene expression in preadipocytes. The anti-adipogenic effects of cucurbitacin B are significantly blunted in cells with STAT3 silenced by introducing small interfering RNA. Finally, our data show that cucurbitacin I, another cucurbitacin family member, also inhibits adipocyte differentiation by suppressing STAT3 signaling. Together, our data suggest the possibility of utilizing cucurbitacins as a new strategy to treat metabolic diseases and implicate STAT3 as a new target for the development of functional foods and drugs. PMID:24316209

  5. Nobiletin enhances differentiation and lipolysis of 3T3-L1 adipocytes

    SciTech Connect

    Saito, Takeshi; Abe, Daigo; Sekiya, Keizo . E-mail: ksekiya@affrc.go.jp

    2007-06-01

    Nobiletin is a polymethoxylated flavone found in certain citrus fruits. Here we demonstrate that nobiletin enhance differentiation of 3T3-L1 preadipocytes. Nobiletin dose-dependently increased accumulation of lipid droplets in adipocytes. Quantitative RT-PCR analyses indicated that nobiletin increased the expression of genes critical for acquisition of the adipocyte phenotype. Some of them were known peroxisome proliferator activated receptor {gamma} (PPAR{gamma}) targets and PPAR{gamma} itself, however, nobiletin did not exhibit PPAR{gamma} ligand activity. We observed the expression of CCAAT/enhancer binding protein {beta} (C/EBP{beta}), a transcription factor for PPAR{gamma}, was increased by nobiletin. The activation of cAMP-responsive element binding protein (CREB) and extracellular signal-regulated kinase (ERK), which play important roles in C/EBP{beta} expression were also potentiated by nobiletin. Furthermore, nobiletin stimulated lipolysis in differentiated adipocytes, which is known to be stimulated by cAMP pathway. These results suggested that nobiletin enhanced both differentiation and lipolysis of adipocyte through activation of signaling cascades mediated by cAMP/CREB.

  6. Characterization of human mesenchymal stem cell secretome at early steps of adipocyte and osteoblast differentiation

    PubMed Central

    Chiellini, Chiara; Cochet, Olivia; Negroni, Luc; Samson, Michel; Poggi, Marjorie; Ailhaud, Gérard; Alessi, Marie-Christine; Dani, Christian; Amri, Ez-Zoubir

    2008-01-01

    Background It is well established that adipose tissue plays a key role in energy storage and release but is also a secretory organ and a source of stem cells. Among different lineages, stem cells are able to differentiate into adipocytes and osteoblasts. As secreted proteins could regulate the balance between both lineages, we aimed at characterizing the secretome of human multipotent adipose-derived stem cell (hMADS) at an early step of commitment to adipocytes and osteoblasts. Results A proteomic approach, using mono-dimensional electrophoresis and tandem mass spectrometry, allowed us to identify a total of 73 proteins at day 0 and day 3 of adipocyte and osteoblast differentiation. Analysis of identified proteins showed that 52 % corresponded to classical secreted proteins characterized by a signal peptide, that 37 % previously described in the extracellular compartment were devoid of signal peptide and that 11 % neither exhibited a signal peptide nor had been previously described extracellularly. These proteins were classified into 8 clusters according to their function. Quantitative analysis has been performed for 8 candidates: PAI-1, PEDF, BIGH3, PTX3, SPARC, ENO1, GRP78 and MMP2. Among them, PAI-1 was detected at day 0 and day 3 of osteoblast differentiation but never in adipocyte secretome. Furthermore we showed that PAI-1 mRNA was down-regulated in the bone of ovariectomized mice. Conclusion Given its regulation during the early events of hMADS cell differentiation and its status in ovariectomized mice, PAI-1 could play a role in the adipocyte/osteoblast balance and thus in bone diseases such as osteoporosis. PMID:18302751

  7. Modulation of adipogenesis-related gene expression by estrogen-related receptor gamma during adipocytic differentiation.

    PubMed

    Kubo, Mayumi; Ijichi, Nobuhiro; Ikeda, Kazuhiro; Horie-Inoue, Kuniko; Takeda, Satoru; Inoue, Satoshi

    2009-02-01

    Estrogen-related receptor gamma (ERRgamma) is an orphan nuclear receptor that regulates cellular energy metabolism by modulating gene expression involved in oxidative metabolism and mitochondrial biogenesis in brown adipose tissue and heart. However, the physiological role of ERRgamma in adipogenesis and the development of white adipose tissue has not been well studied. Here we show that ERRgamma was up-regulated in murine mesenchyme-derived cells, especially in ST2 and C3H10T1/2 cells, at mRNA levels under the adipogenic differentiation condition including the inducer of cAMP, glucocorticoid, and insulin. The up-regulation of ERRgamma mRNA was also observed in inguinal white adipose and brown adipose tissues of mice fed a high-fat diet. Gene knockdown by ERRgamma-specific siRNA results in mRNA down-regulation of adipogenic marker genes including fatty acid binding protein 4, PPARgamma, and PGC-1beta in a preadipocyte cell line 3T3-L1 preadipocytes and mesenchymal ST2 and C3H10T1/2 cells in the adipogenesis medium. In contrast, stable expression of ERRgamma in 3T3-L1 cells resulted in up-regulation of these adipogenic marker genes under the adipogenic condition. These results suggest that ERRgamma positively regulate the adipocyte differentiation with modulating the expression of various adipogenesis-related genes. PMID:18809516

  8. Synthesis of mitochondrial uncoupling protein in brown adipocytes differentiated in cell culture

    SciTech Connect

    Kopecky, J.; Baudysova, M.; Zanotti, F.; Janikova, D.; Pavelka, S.; Houstek, J. )

    1990-12-25

    In order to characterize the biogenesis of unique thermogenic mitochondria of brown adipose tissue, differentiation of precursor cells isolated from mouse brown adipose tissue was studied in cell culture. Synthesis of mitochondrial uncoupling protein (UCP), F1-ATPase, and cytochrome oxidase was examined by L-(35S)methionine labeling and immunoblotting. For the first time, synthesis of physiological amounts of the UCP, a key and tissue-specific component of thermogenic mitochondria, was observed in cultures at about confluence (day 6), indicating that a complete differentiation of brown adipocytes was achieved in vitro. In postconfluent cells (day 8) the content of UCP decreased rapidly, in contrast to some other mitochondrial proteins (beta subunit of F1-ATPase, cytochrome oxidase). In these cells, it was possible, by using norepinephrine, to induce specifically the synthesis of the UCP but not of F1-ATPase or cytochrome oxidase. The maximal response was observed at 0.1 microM norepinephrine and the synthesis of UCP remained activated for at least 24 h. Detailed analysis revealed a major role of the beta-adrenergic receptors and elevated intracellular concentration of cAMP in stimulation of UCP synthesis. A quantitative recovery of the newly synthesized UCP in the mitochondrial fraction indicated completed biogenesis of functionally competent thermogenic mitochondria.

  9. Estrogen-related receptor {alpha} modulates the expression of adipogenesis-related genes during adipocyte differentiation

    SciTech Connect

    Ijichi, Nobuhiro; Ikeda, Kazuhiro; Horie-Inoue, Kuniko; Yagi, Ken; Okazaki, Yasushi; Inoue, Satoshi . E-mail: INOUE-GER@h.u-tokyo.ac.jp

    2007-07-06

    Estrogen-related receptor {alpha} (ERR{alpha}) is an orphan nuclear receptor that regulates cellular energy metabolism by modulating gene expression involved in fatty acid oxidation and mitochondrial biogenesis in brown adipose tissue. However, the physiological role of ERR{alpha} in adipogenesis and white adipose tissue development has not been well studied. Here, we show that ERR{alpha} and ERR{alpha}-related transcriptional coactivators, peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) coactivator-1{alpha} (PGC-1{alpha}) and PGC-1{beta}, can be up-regulated in 3T3-L1 preadipocytes at mRNA levels under the adipogenic differentiation condition including the inducer of cAMP, glucocorticoid, and insulin. Gene knockdown by ERR{alpha}-specific siRNA results in mRNA down-regulation of fatty acid binding protein 4, PPAR{gamma}, and PGC-1{alpha} in 3T3-L1 cells in the adipogenesis medium. ERR{alpha} and PGC-1{beta} mRNA expression can be also up-regulated in another preadipocyte lineage DFAT-D1 cells and a pluripotent mesenchymal cell line C3H10T1/2 under the differentiation condition. Furthermore, stable expression of ERR{alpha} in 3T3-L1 cells up-regulates adipogenic marker genes and promotes triglyceride accumulation during 3T3-L1 differentiation. These results suggest that ERR{alpha} may play a critical role in adipocyte differentiation by modulating the expression of various adipogenesis-related genes.

  10. MicroRNA-23a regulates 3T3-L1 adipocyte differentiation.

    PubMed

    Shen, Linyuan; Zhang, Yi; Du, Jingjing; Chen, Li; Luo, Jia; Li, Xuewei; Li, Mingzhou; Tang, Guoqing; Zhang, Shunhua; Zhu, Li

    2016-01-10

    MicroRNAs (miRNAs) are small, non-coding RNAs, which are involved in regulation of a variety of biological processes. Since previous studies regarding the role of miRNAs in the regulation of adipogenic differentiation have shown that miRNA-27a, one member of miRNA-23a∼27a∼24 cluster, could suppress adipogenesis. We now investigated whether miRNA-23a regulates adipogenic differentiation. In the present study, we showed that the expression of miRNA-23a is decreased during the process of adipogenic differentiation. Over-expression of miRNA-23a decreased lipid accumulation and triglyceride content in 3T3-L1 adipocytes. Our results also demonstrated that miRNA-23a decreases mRNA levels of adipocyte-specific genes involved in lipogenic transcription, fatty acid synthesis and fatty acid transport. These findings suggested miRNA-23a to be a new type of adipogenic depressor and to play an important role in regulating adipocyte differentiation. PMID:26415879

  11. The Fto Gene Regulates the Proliferation and Differentiation of Pre-Adipocytes in Vitro

    PubMed Central

    Jiao, Yang; Zhang, Jingying; Lu, Lunjie; Xu, Jiaying; Qin, Liqiang

    2016-01-01

    The highly regulated differentiation and proliferation of pre-adipocytes play a key role in the initiation of obesity. Fat mass and obesity associated (FTO) is a novel gene strongly associated with the risk of obesity. A deficiency of FTO may cause growth retardation in addition to fat mass and adipocyte size reduction in vivo. To investigate the potential role of Fto gene on the proliferation and differentiation of pre-adipocytes, we generated Fto-knockdown and overexpressed 3T3-L1 cells. Using numerous proliferation assays our results suggest that Fto knockdown leads to suppression of proliferation, lower mitochondrial membrane potential, less cellular ATP, and decreased and smaller intracellular lipid droplets compared with controls (p < 0.05). Western blot analysis demonstrated that Fto knockdown can significantly suppress peroxisome proliferator-activated receptor gamma (PPARγ) and glucose transporter type 4 (GLUT4) expression and inhibit Akt phosphorylation. By contrast, overexpression of Fto had the opposing effect on proliferation, mitochondrial membrane potential, ATP generation, in vitro differentiation, Akt phosphorylation, and PPARγ and GLUT4 expression. Moreover, we demonstrated that Wortmannin, a phosphoinositide 3-kinase (PI3K) inhibitor, could inhibit phospho-Akt in Fto overexpressed 3T3-L1 cells. Taken together, the results suggest that Fto regulates the proliferation and differentiation of 3T3-L1 cells via multiple mechanisms, including PPARγ and PI3K/Akt signaling. PMID:26907332

  12. The Fto Gene Regulates the Proliferation and Differentiation of Pre-Adipocytes in Vitro.

    PubMed

    Jiao, Yang; Zhang, Jingying; Lu, Lunjie; Xu, Jiaying; Qin, Liqiang

    2016-02-01

    The highly regulated differentiation and proliferation of pre-adipocytes play a key role in the initiation of obesity. Fat mass and obesity associated (FTO) is a novel gene strongly associated with the risk of obesity. A deficiency of FTO may cause growth retardation in addition to fat mass and adipocyte size reduction in vivo. To investigate the potential role of Fto gene on the proliferation and differentiation of pre-adipocytes, we generated Fto-knockdown and overexpressed 3T3-L1 cells. Using numerous proliferation assays our results suggest that Fto knockdown leads to suppression of proliferation, lower mitochondrial membrane potential, less cellular ATP, and decreased and smaller intracellular lipid droplets compared with controls (p < 0.05). Western blot analysis demonstrated that Fto knockdown can significantly suppress peroxisome proliferator-activated receptor gamma (PPARγ) and glucose transporter type 4 (GLUT4) expression and inhibit Akt phosphorylation. By contrast, overexpression of Fto had the opposing effect on proliferation, mitochondrial membrane potential, ATP generation, in vitro differentiation, Akt phosphorylation, and PPARγ and GLUT4 expression. Moreover, we demonstrated that Wortmannin, a phosphoinositide 3-kinase (PI3K) inhibitor, could inhibit phospho-Akt in Fto overexpressed 3T3-L1 cells. Taken together, the results suggest that Fto regulates the proliferation and differentiation of 3T3-L1 cells via multiple mechanisms, including PPARγ and PI3K/Akt signaling. PMID:26907332

  13. Dynamics of lipid droplet-associated proteins during hormonally stimulated lipolysis in engineered adipocytes: stabilization and lipid droplet binding of adipocyte differentiation-related protein/adipophilin.

    PubMed

    Gross, Danielle N; Miyoshi, Hideaki; Hosaka, Toshio; Zhang, Hui-Hong; Pino, Elizabeth C; Souza, Sandra; Obin, Martin; Greenberg, Andrew S; Pilch, Paul F

    2006-02-01

    In mature adipocytes, triglyceride is stored within lipid droplets, which are coated with the protein perilipin, which functions to regulate lipolysis by controlling lipase access to the droplet in a hormone-regulatable fashion. Adipocyte differentiation-related protein (ADRP) is a widely expressed lipid droplet binding protein that is coexpressed with perilipin in differentiating fat cells but is minimally present in fully differentiated cultured adipocytes. We find that fibroblasts ectopically expressing C/EBPalpha (NIH-C/EBPalpha cells) differentiate into mature adipocytes that simultaneously express perilipin and ADRP. In response to isoproterenol, perilipin is hyperphosphorylated, lipolysis is enhanced, and subsequently, ADRP expression increases coincident with it surrounding intracellular lipid droplets. In the absence of lipolytic stimulation, inhibition of proteasomal activity with MG-132 increased ADRP levels to those of cells treated with 10 mum isoproterenol, but ADRP does not surround the lipid droplet in the absence of lipolytic stimulation. We overexpressed a perilipin A construct in NIH-C/EBPalpha cells where the six serine residues known to be phosphorylated by protein kinase A were changed to alanine (Peri A Delta1-6). These cells show no increase in ADRP expression in response to isoproterenol. We propose that ADRP can replace perilipin on existing lipid droplets or those newly formed as a result of fatty acid reesterification, under dynamic conditions of hormonally stimulated lipolysis, thus preserving lipid droplet morphology/structure. PMID:16239256

  14. Anthraquinones from Morinda officinalis roots enhance adipocyte differentiation in 3T3-L1 cells.

    PubMed

    Liu, Qing; Kim, Seon Beom; Ahn, Jong Hoon; Hwang, Bang Yeon; Kim, Sung Yeon; Lee, Mi Kyeong

    2012-01-01

    To search for anti-diabetic and insulin-sensitising natural products, the effect on adipocyte differentiation was investigated by assessing fat accumulation in 3T3-L1 preadipocytes using Oil Red O staining. Fractionation and separation of n-hexane and CHCl₃ fractions of Morinda officinalis (Rubiaceae) using several chromatographic methods led to the isolation of three anthraquinones, 1,2-dimethoxyanthraquinone (1), alizarin-2-methyl ether (2) and rubiadin-1-methyl ether (3). Among them, alizarin-2-methyl ether (2) showed the strongest enhancing activity, followed by rubiadin-1-methyl ether (3) and 1,2-dimethoxyanthraquinone (1). At a concentration of 100 µM, alizarin-2-methyl ether (2) enhanced adipocyte differentiation by up to 131% (compared to insulin-treated cells). Thus, these compounds could be beneficial in the treatment of diabetes. PMID:22008000

  15. Adipocyte differentiation-related protein promotes lipid accumulation in goat mammary epithelial cells.

    PubMed

    Shi, H B; Yu, K; Luo, J; Li, J; Tian, H B; Zhu, J J; Sun, Y T; Yao, D W; Xu, H F; Shi, H P; Loor, J J

    2015-10-01

    Milk fat originates from the secretion of cytosolic lipid droplets (CLD) synthesized within mammary epithelial cells. Adipocyte differentiation-related protein (ADRP; gene symbol PLIN2) is a CLD-binding protein that is crucial for synthesis of mature CLD. Our hypothesis was that ADRP regulates CLD production and metabolism in goat mammary epithelial cells (GMEC) and thus plays a role in determining milk fat content. To understand the role of ADRP in ruminant milk fat metabolism, ADRP (PLIN2) was overexpressed or knocked down in GMEC using an adenovirus system. Immunocytochemical staining revealed that ADRP localized to the surface of CLD. Supplementation with oleic acid (OA) enhanced its colocalization with CLD surface and enhanced lipid accumulation. Overexpression of ADRP increased lipid accumulation and the concentration of triacylglycerol in GMEC. In contrast, morphological examination revealed that knockdown of ADRP decreased lipid accumulation even when OA was supplemented. This response was confirmed by the reduction in mass of cellular TG when ADRP was knocked down. The fact that knockdown of ADRP did not completely eliminate lipid accumulation at a morphological level in GMEC without OA suggests that some other compensatory factors may also aid in the process of CLD formation. The ADRP reversed the decrease of CLD accumulation induced by adipose triglyceride lipase. This is highly suggestive of ADRP promoting triacylglycerol stability within CLD by preventing access to adipose triglyceride lipase. Collectively, these data provide direct in vitro evidence that ADRP plays a key role in CLD formation and stability in GMEC. PMID:26298750

  16. Endothelial differentiation in multipotent cells derived from mouse and human white mature adipocytes.

    PubMed

    Jumabay, Medet; Abdmaulen, Raushan; Urs, Sumithra; Heydarkhan-Hagvall, Sepideh; Chazenbalk, Gregorio D; Jordan, Maria C; Roos, Kenneth P; Yao, Yucheng; Boström, Kristina I

    2012-12-01

    White mature adipocytes give rise to multipotent cells, so-called de-differentiated fat (DFAT) cells, when losing their fat in culture. The objective of this study was to examine the ability of DFAT cells to give rise to endothelial cells (ECs) in vitro and vivo. We demonstrate that mouse and human DFAT cells, derived from adipose tissue and lipospirate, respectively, initially lack expression of CD34, CD31, CD146, CD45 and pericyte markers, distinguishing them from progenitor cells previously identified in adipose stroma. The DFAT cells spontaneously differentiate into vascular ECs in vitro, as determined by real-time PCR, fluorescence activated cell sorting, immunostaining, and formation of tube structures. Treatment with bone morphogenetic protein (BMP)4 and BMP9, important in regulating angiogenesis, significantly enhances the EC differentiation. Furthermore, adipocyte-derived cells from Green Fluorescent Protein-transgenic mice were detected in the vasculature of infarcted myocardium up to 6 weeks after ligation of the left anterior descending artery in mice. We conclude that adipocyte-derived multipotent cells are able to spontaneously give rise to ECs, a process that is promoted by BMPs and may be important in cardiovascular regeneration and in physiological and pathological changes in fat and other tissues. PMID:22999861

  17. Puerarin enhances adipocyte differentiation, adiponectin expression, and antioxidant response in 3T3-L1 cells.

    PubMed

    Lee, Ok-Hwan; Seo, Dong-Ho; Park, Cheon-Seok; Kim, Young-Cheul

    2010-01-01

    Puerarin, a major isoflavone glycoside from Kudzu root (Pueraria lobata), has been reported to exert antihyperglycemic and antioxidant effects and thus have pharmacological actions in the treatment of diabetes and cardiovascular diseases. We investigated the effects of puerarin on the changes of key gene expression associated with adipocyte differentiation and insulin sensitivity and link to cellular antioxidant response pathways. Puerarin treatment significantly enhanced differentiation of 3T3-L1 preadipocytes accompanying increased lipid accumulation and glucose-6-phosphate dehydrogenase (G6PDH) activity. At a molecular level, puerarin upregulated mRNA expression of peroxisome proliferator-activated receptor γ (PPARγ) and its target genes, an adipocyte-specific fatty acid binding protein (aP2) and GLUT4. Puerarin also caused a significant increase in mRNA level of adiponectin, an important insulin-sensitizing adipocytokine that is downregulated in insulin-resistant and diabetic states. In addition, treatment with puerarin was found to upregulate mRNA levels of G6PDH, glutathione reductase, and catalase, all of which are important for endogenous antioxidant responses. These data suggest that the hypoglycemic effects of puerarin can be attributed to the upregulation of PPARγ and its downstream target genes, GLUT4 and adiponectin expression, leading to increased glucose utilization. Puerarin may also be effective in preventing the rise of oxidative stress during adipocyte differentiation by increasing endogenous antioxidant responses. PMID:20806284

  18. Regulation of Adipocyte Differentiation by Distinct Subcellular Pools of Protein Kinase B (PKB/Akt)*

    PubMed Central

    Maiuri, Tamara; Ho, Jason; Stambolic, Vuk

    2010-01-01

    The phosphatidylinositol 3-kinase (PI3K)-protein kinase B (PKB)/Akt-PTEN signal transduction pathway orchestrates a variety of fundamental cell processes and its deregulation is implicated in many human diseases. Although the importance of this pathway to many cellular functions is well established, the mechanisms by which it achieves context-specific physiological outcomes in response to a variety of stimuli, using a relatively limited pool of effectors, remain largely unknown. Spatial restriction of signaling events is one means by which cells coordinate specific responses using common molecules. To investigate the subcellular location-specific roles of the major PI3K effector PKB/Akt in various cell processes, we have developed a novel experimental system employing cellular compartment-directed PKB/Akt pseudosubstrate inhibitors. Subcellular location-restricted PKB/Akt inhibition in the 3T3L1 adipocyte differentiation model revealed that nuclear and plasma membrane, but not cytoplasmic, PKB/Akt activity is required for terminal adipocyte differentiation. Nuclear and plasma membrane pools of PKB/Akt were found to contribute to distinct stages of adipocyte differentiation, revealing that PKB/Akt activity impacts multiple points of this program. Our work establishes the use of localized pseudosubstrate PKB/Akt inhibitors as an effective method for the dissection of PKB/Akt signaling. PMID:20223817

  19. Isoliquiritigenin impairs insulin signaling and adipocyte differentiation through the inhibition of protein-tyrosine phosphatase 1B oxidation in 3T3-L1 preadipocytes.

    PubMed

    Park, Sun-Ji; Choe, Young-Geun; Kim, Jung-Hak; Chang, Kyu-Tae; Lee, Hyun-Shik; Lee, Dong-Seok

    2016-07-01

    Isoliquritigenin (ISL) is an abundant dietary flavonoid with a chalcone structure, which is an important constituent in Glycyrrhizae Radix (GR). ISL exhibits anti-oxidant activity, and this activity has been shown to play a beneficial role in various health conditions. However, it is unclear whether the anti-oxidant activity of ISL affects insulin signaling pathway and lipid accumulation of adipocytes. We sought to investigate the effects and molecular mechanisms of ISL on insulin-stimulated adipogenesis in 3T3-L1 cells. We investigated whether ISL attenuates insulin-induced Reactive Oxygen Species (ROS) generation, and whether ISL inhibits the lipid accumulation and the expression of adipogenic-genes during the differentiation of 3T3-L1 cells. ISL blocked the ROS generation, suppressed the lipid accumulation and the expression of adipocyte-specific proteins, which are increased in response to insulin stimulation during adipocyte differentiation of 3T3-L1 cells. We also investigated whether the anti-oxidant capacity of ISL is involved in regulating the molecular events of insulin-signaling cascade in 3T3-L1 adipocytes. ISL restores PTP1B activity by inhibiting PTP1B oxidation and IR/PI3K/AKT phosphorylation during the early stages of insulin-induced adipogenesis. Our findings show that the anti-oxidant capacity of ISL attenuated insulin IR/PI3K/AKT signaling through inhibition of PTP1B oxidation, and ultimately attenuated insulin-induced adipocyte differentiation of 3T3-L1 cells. PMID:27117918

  20. Proinflammatory cytokines differentially regulate adipocyte mitochondrial metabolism, oxidative stress, and dynamics

    PubMed Central

    Hahn, Wendy S.; Kuzmicic, Jovan; Burrill, Joel S.; Donoghue, Margaret A.; Foncea, Rocio; Jensen, Michael D.; Lavandero, Sergio; Arriaga, Edgar A.

    2014-01-01

    Proinflammatory cytokines differentially regulate adipocyte mitochondrial metabolism, oxidative stress, and dynamics. Macrophage infiltration of adipose tissue and the chronic low-grade production of inflammatory cytokines have been mechanistically linked to the development of insulin resistance, the forerunner of type 2 diabetes mellitus. In this study, we evaluated the chronic effects of TNFα, IL-6, and IL-1β on adipocyte mitochondrial metabolism and morphology using the 3T3-L1 model cell system. TNFα treatment of cultured adipocytes led to significant changes in mitochondrial bioenergetics, including increased proton leak, decreased ΔΨm, increased basal respiration, and decreased ATP turnover. In contrast, although IL-6 and IL-1β decreased maximal respiratory capacity, they had no effect on ΔΨm and varied effects on ATP turnover, proton leak, or basal respiration. Only TNFα treatment of 3T3-L1 cells led to an increase in oxidative stress (as measured by superoxide anion production and protein carbonylation) and C16 ceramide synthesis. Treatment of 3T3-L1 adipocytes with cytokines led to decreased mRNA expression of key transcription factors and control proteins implicated in mitochondrial biogenesis, including PGC-1α and eNOS as well as deceased expression of COX IV and Cyt C. Whereas each cytokine led to effects on expression of mitochondrial markers, TNFα exclusively led to mitochondrial fragmentation and decreased the total level of OPA1 while increasing OPA1 cleavage, without expression of levels of mitofusin 2, DRP-1, or mitofilin being affected. In summary, these results indicate that inflammatory cytokines have unique and specialized effects on adipocyte metabolism, but each leads to decreased mitochondrial function and a reprogramming of fat cell biology. PMID:24595304

  1. KCNK10, a Tandem Pore Domain Potassium Channel, Is a Regulator of Mitotic Clonal Expansion during the Early Stage of Adipocyte Differentiation

    PubMed Central

    Nishizuka, Makoto; Hayashi, Takahiro; Asano, Mami; Osada, Shigehiro; Imagawa, Masayoshi

    2014-01-01

    KCNK10, a member of tandem pore domain potassium channel family, gives rise to leak K+ currents. It plays important roles in stabilizing the negative resting membrane potential and in counterbalancing depolarization. We previously demonstrated that kcnk10 expression is quickly elevated during the early stage of adipogenesis of 3T3-L1 cells and that reduction of kcnk10 expression inhibits adipocyte differentiation. However, the molecular mechanism of KCNK10 in adipocyte differentiation remains unclear. Here we revealed that kcnk10 is induced by 3-isobutyl-1-methylxanthine, a cyclic nucleotide phosphodiesterase inhibitor and a potent inducer of adipogenesis, during the early stage of adipocyte differentiation. We also demonstrated that KCNK10 functions as a positive regulator of mitotic clonal expansion (MCE), a necessary process for terminal differentiation. The reduction of kcnk10 expression repressed the expression levels of CCAAT/enhancer-binding protein β (C/EBPβ) and C/EBPδ as well as the phosphorylation level of Akt during the early phase of adipogenesis. In addition, knockdown of kcnk10 expression suppressed insulin-induced Akt phosphorylation. These results indicate that KCNK10 contributes to the regulation of MCE through the control of C/EBPβ and C/EBPδ expression and insulin signaling. PMID:25501330

  2. Neuropoietin Attenuates Adipogenesis and Induces Insulin Resistance in Adipocytes*

    PubMed Central

    White, Ursula A.; Stewart, William C.; Mynatt, Randall L.; Stephens, Jacqueline M.

    2008-01-01

    Recent findings have implicated gp130 receptor ligands, particularly ciliary neurotrophic factor (CNTF), as potential anti-obesity therapeutics. Neuropoietin (NP) is a recently discovered cytokine in the gp130 family that shares functional and structural features with CNTF and signals via the CNTF receptor tripartite complex comprised of CNTFRα, LIF receptor, and gp130. NP plays a role in the development of the nervous system, but the effects of NP on adipocytes have not been previously examined. Because CNTF exerts anti-obesogenic effects in adipocytes and NP shares the same receptor complex, we investigated the effects of NP on adipocyte development and insulin action. Using cultured 3T3-L1 adipocytes, we observed that NP has the ability to block adipogenesis in a dose- and time-dependent manner. We also observed that cultured adipocytes, as well as murine adipose tissue, are highly responsive to acute NP treatment. Rodents injected with NP had a substantial increase in STAT3 tyrosine phosphorylation and ERK 1 and 2 activation. We also observed the induction of SOCS-3 mRNA in 3T3-L1 adipocytes following NP treatment. Unlike CNTF, our studies have revealed that NP also substantially attenuates insulin-stimulated glucose uptake in 3T3-L1 adipocytes. In addition, NP blocks insulin action in adipose tissue in vivo. These observations are supported by data demonstrating that NP impairs insulin signaling via decreased activation of both IRS-1 and Akt. In summary, we have observed that both adipocytes in vitro and in vivo are highly responsive to NP, and this cytokine has the ability to affect insulin signaling in fat cells. These novel observations suggest that NP, unlike CNTF, may not be a viable obesity therapeutic. PMID:18562323

  3. Carnitine palmitoyltransferase 1A prevents fatty acid-induced adipocyte dysfunction through suppression of c-Jun N-terminal kinase.

    PubMed

    Gao, Xuefei; Li, Kuai; Hui, Xiaoyan; Kong, Xiangping; Sweeney, Gary; Wang, Yu; Xu, Aimin; Teng, Maikun; Liu, Pentao; Wu, Donghai

    2011-05-01

    The adipocyte is the principal cell type for fat storage. CPT1 (carnitine palmitoyltransferase-1) is the rate-limiting enzyme for fatty acid β-oxidation, but the physiological role of CPT1 in adipocytes remains unclear. In the present study, we focused on the specific role of CPT1A in the normal functioning of adipocytes. Three 3T3-L1 adipocyte cell lines stably expressing hCPT1A (human CPT1A) cDNA, mouse CPT1A shRNA (short-hairpin RNA) or GFP (green fluorescent protein) were generated and the biological functions of these cell lines were characterized. Alteration in CPT1 activity, either by ectopic overexpression or pharmacological inhibition using etomoxir, did not affect adipocyte differentiation. However, overexpression of hCPT1A significantly reduced the content of intracellular NEFAs (non-esterified fatty acids) compared with the control cells when adipocytes were challenged with fatty acids. The changes were accompanied by an increase in fatty acid uptake and a decrease in fatty acid release. Interestingly, CPT1A protected against fatty acid-induced insulin resistance and expression of pro-inflammatory adipokines such as TNF-α (tumour necrosis factor-α) and IL-6 (interleukin-6) in adipocytes. Further studies demonstrated that JNK (c-Jun N terminal kinase) activity was substantially suppressed upon CPT1A overexpression, whereas knockdown or pharmacological inhibition of CPT1 caused a significant enhancement of JNK activity. The specific inhibitor of JNK SP600125 largely abolished the changes caused by the shRNA- and etomoxir-mediated decrease in CPT1 activity. Moreover, C2C12 myocytes co-cultured with adipocytes pre-treated with fatty acids displayed altered insulin sensitivity. Taken together, our findings have identified a favourable role for CPT1A in adipocytes to attenuate fatty acid-evoked insulin resistance and inflammation via suppression of JNK. PMID:21348853

  4. Differentiation of Rat bone marrow Mesenchymal stem cells into Adipocytes and Cardiomyocytes after treatment with platelet lysate

    PubMed Central

    Homayouni Moghadam, Farshad; Tayebi, Tahereh; Barzegar, Kazem

    2016-01-01

    Background: Mesenchymal stem cells (MSCs) are multipotential cells and their therapeutic potency is under intense investigation. Studying the effect of different induction factors on MSCs could increase our knowledge about the differentiation potency of these cells. One of the most important sources of these factors in mammalian body is platelet. Platelet lysate (PL) contains many growth factors and therefore, it can be used as a differentiation inducer. In the present study, the effect of PL on differentiation of rat bone marrow MSCs into cardiomyocytes was studied. Materials and Methods: To study the differentiation-inducing effect of PL, MSCs were treated with 2.5, 5 and 10% PL. Early results of this study showed that PL in high concentrations (10%) induces adipogenic differentiation of MSCs. Therefore, to evaluate differentiation to cardiomyocytes, MSCs were cultured in media containing lower levels of PL (2.5% and 5%) and then cardiomyogenic differentiation was induced by treatment with 5-azacytidine. Differentiation of MSCs was evaluated using direct observation of beating cells, immunostaining and real-time PCR techniques. Results: The results of qPCR showed that treatment with PL alone increased the expression of cardiac alpha actinin (CAA) being predictable by earlier observation of beating cells in PL-treated groups. The results of staining assays against cardiac alpha actinin also showed that there were stained cells in PL-treated groups. Conclusion: The results of the present study showed that PL is a powerful induction factor for differentiation of MSCs into different cell lines such as cardiomyocytes and adipocytes. PMID:27047647

  5. Differentiation of adipocytes and osteocytes from human adipose and placental mesenchymal stem cells

    PubMed Central

    Mohammadi, Zahra; Afshari, Jalil Tavakkol; Keramati, Mohammad Reza; Alamdari, Daryoush Hamidi; Ganjibakhsh, Meysam; Zarmehri, Azam Moradi; Jangjoo, Ali; Sadeghian, Mohammad Hadi; Ameri, Masoumeh Arab; Moinzadeh, Leila

    2015-01-01

    Objective(s): Mesenchymal stem cells (MSC) can be isolated from adult tissues such as adipose tissue and other sources. Among these sources, adipose tissue (because of easy access) and placenta (due to its immunomodulatory properties, in addition to other useful properties), have attracted more attention in terms of research. The isolation and comparison of MSC from these two sources provides a proper source for clinical experimentation. The aim of this study was to compare the characteristics of MSC isolated from human adipose tissue and placenta. Materials and Methods: Adipose and placental MSC were isolated from the subcutaneous adipose tissues of 10 healthy women (25 to 40 years) and from a fresh term placenta (n= 1), respectively. Stem cells were characterized and compared by flow cytometry using CD29, CD31, CD34, CD44, CD45, CD105, CD166 and HLA-DR markers. Osteocytes and adipocytes were differentiated from isolated human mesenchymal stem cells (HMSC). Results: Adipose and placenta-derived MSC exhibited the same morphological features. ADSC differentiated faster than placenta; however, both were differentiated, taking up to 21 days for osteocyte and 14 days for adipocyte differentiation. About 90% of PLC-MSC and ADSC were positive for CD29, CD44, CD105, and CD166; and negative for CD31, CD34, CD45, and HLA-DR. Conclusion: The two sources of stem cells showed similar surface markers, morphology and differentiation potential and because of their multipotency for differentiating to adipocytes and osteocytes, they can be applied as attractive sources of MSC for regenerative medicine. PMID:25945239

  6. Essential Role of Insulin Receptor Substrate 1 in Differentiation of Brown Adipocytes

    PubMed Central

    Fasshauer, Mathias; Klein, Johannes; Kriauciunas, Kristina M.; Ueki, Kohjiro; Benito, Manuel; Kahn, C. Ronald

    2001-01-01

    The most widely distributed members of the family of insulin receptor substrate (IRS) proteins are IRS-1 and IRS-2. These proteins participate in insulin and insulin-like growth factor 1 signaling, as well as the actions of some cytokines, growth hormone, and prolactin. To more precisely define the specific role of IRS-1 in adipocyte biology, we established brown adipocyte cell lines from wild-type and IRS-1 knockout (KO) animals. Using differentiation protocols, both with and without insulin, preadipocyte cell lines derived from IRS-1 KO mice exhibited a marked decrease in differentiation and lipid accumulation (10 to 40%) compared to wild-type cells (90 to 100%). Furthermore, IRS-1 KO cells showed decreased expression of adipogenic marker proteins, such as peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer-binding protein alpha (C/EBPα), fatty acid synthase, uncoupling protein-1, and glucose transporter 4. The differentiation deficit in the KO cells could be reversed almost completely by retrovirus-mediated reexpression of IRS-1, PPARγ, or C/EBPα but not the thiazolidinedione troglitazone. Phosphatidylinositol 3-kinase (PI 3-kinase) assays performed at various stages of the differentiation process revealed a strong and transient activation in IRS-1, IRS-2, and phosphotyrosine-associated PI 3-kinase in the wild-type cells, whereas the IRS-1 KO cells showed impaired phosphotyrosine-associated PI 3-kinase activation, all of which was associated with IRS-2. Akt phosphorylation was reduced in parallel with the total PI 3-kinase activity. Inhibition of PI 3-kinase with LY294002 blocked differentiation of wild-type cells. Thus, IRS-1 appears to be an important mediator of brown adipocyte maturation. Furthermore, this signaling molecule appears to exert its unique role in the differentiation process via activation of PI 3-kinase and its downstream target, Akt, and is upstream of the effects of PPARγ and C/EBPα. PMID:11113206

  7. THRAP3 interacts with HELZ2 and plays a novel role in adipocyte differentiation.

    PubMed

    Katano-Toki, Akiko; Satoh, Tetsurou; Tomaru, Takuya; Yoshino, Satoshi; Ishizuka, Takahiro; Ishii, Sumiyasu; Ozawa, Atsushi; Shibusawa, Nobuyuki; Tsuchiya, Takafumi; Saito, Tsugumichi; Shimizu, Hiroyuki; Hashimoto, Koshi; Okada, Shuichi; Yamada, Masanobu; Mori, Masatomo

    2013-05-01

    Using yeast two-hybrid screen, we previously isolated HELZ2 (helicase with zinc finger 2, transcriptional coactivator) that functions as a coregulator of peroxisome proliferator-activated receptorγ (PPARγ). To further delineate its molecular function, we here identified thyroid hormone receptor-associated protein3 (THRAP3), a putative component of the Mediator complex, as a protein stably associating with HELZ2 using immunoprecipitation coupled with mass spectrometry analyses. In immunoprecipitation assays, Thrap3 could associate with endogenous Helz2 as well as Pparg in differentiated 3T3-L1 cells. HELZ2 interacts with the serine/arginine-rich domain and Bcl2 associated transcription factor1-homologous region in THRAP3, whereas THRAP3 directly binds 2 helicase motifs in HELZ2. HELZ2 and THRAP3 synergistically augment transcriptional activation mediated by PPARγ, whereas knockdown of endogenous THRAP3 abolished the enhancement by HELZ2 in reporter assays. Thrap3, similar to Helz2, is evenly expressed in the process of adipogenic differentiation in 3T3-L1 cells. Knockdown of Thrap3 in 3T3-L1 preadipocytes using short-interfering RNA did not influence the expression of Krox20, Klf5, Cebpb, or Cebpd during early stages of adipocyte differentiation, but significantly attenuated the expression of Pparg, Cebpa, and Fabp4/aP2 and accumulation of lipid droplets. Pharmacologic activation of Pparg by troglitazone could not fully restore the differentiation of Thrap3-knockdown adipocytes. In chromatin immunoprecipitation assays, endogenous Helz2 and Thrap3 could be co-recruited, in a ligand-dependent manner, to the PPARγ-response elements in Fabp4/aP2 and Adipoq gene enhancers in differentiated 3T3-L1 cells. These findings collectively suggest that Thrap3 could play indispensable roles in terminal differentiation of adipocytes by enhancing PPARγ-mediated gene activation cooperatively with Helz2. PMID:23525231

  8. Lactacystin inhibits 3T3-L1 adipocyte differentiation through induction of CHOP-10 expression

    SciTech Connect

    Li Xi; Huang Haiyan |; Chen Jiegen; Jiang Lin; Liu Honglei |; Liu Deguo; Song Tanjing; He Qun; Ma Chungu; Ma Duan |; Song Houyan; Tang Qiqun ||. E-mail: qqtang@shmu.edu.cn

    2006-11-10

    Hormonal induction triggers a cascade leading to the expression of CCAAT/enhancer-binding protein(C/EBP){alpha} and peroxisome proliferator-activated receptor (PPAR) {gamma}, C/EBP{alpha}, and PPAR{gamma} turns on series of adipocyte genes that give rise to the adipocyte phenotype. Previous findings indicate that C/EBP{beta}, a transcriptional activator of the C/EBP{alpha} and PPAR{gamma} genes, is rapidly expressed after induction, but lacks DNA-binding activity and therefore cannot activate transcription of the C/EBP{alpha} and PPAR{gamma} genes early in the differentiation program. Acquisition of DNA-binding activity of C/EBP{beta} occurs when CHOP-10, a dominant-negative form of C/EBP family members, is down-regulated and becomes hyperphosphorylated as preadipocytes traverse the G{sub 1}-S checkpoint of mitotic clonal expansion. Evidences are presented in this report that lactacystin, a proteasome inhibitor, up-regulated the CHOP-10 expression, blocked the DNA-binding activity of C/EBP{beta}, and subsequently inhibited MCE as well as adipocyte differentiation.

  9. A Long Noncoding RNA Transcriptional Regulatory Circuit Drives Thermogenic Adipocyte Differentiation

    PubMed Central

    Zhao, Xu-Yun; Li, Siming; Wang, Guo-Xiao; Yu, Qi; Lin, Jiandie D.

    2014-01-01

    SUMMARY Brown and beige/brite fats generate heat via uncoupled respiration to defend against cold. The total mass and activity of thermogenic adipose tissues are also tightly linked to systemic energy and nutrient homeostasis. Despite originating from distinct progenitors, brown and beige adipocytes acquire remarkably similar molecular and metabolic characteristics during differentiation through the action of a network of transcription factors and cofactors. How this regulatory network interfaces with long noncoding RNAs (lncRNAs), an emerging class of developmental regulators, remains largely unexplored. Here we globally profiled lncRNA gene expression during thermogenic adipocyte formation and identified Brown fat lncRNA 1 (Blnc1) as a novel nuclear lncRNA that promotes brown and beige adipocyte differentiation and function. Blnc1 forms a ribonucleoprotein complex with transcription factor EBF2 to stimulate the thermogenic gene program. Further, Blnc1 itself is a target of EBF2, thereby forming a feedforward regulatory loop to drive adipogenesis toward thermogenic phenotype. PMID:25002143

  10. Antiadipogenic properties of retinol in primary cultured differentiating human adipocyte precursor cells.

    PubMed

    Garcia, E; Lacasa, D; Agli, B; Giudicelli, Y; Castelli, D

    2000-04-01

    The aim of this study was to investigate the effect of retinol on the human adipose conversion process using primary cultured human adipocyte precursor cells. When these cells were seeded in a medium containing retinol (concentrations ranging from 3.5 nM to 3.5 muM), cell proliferation was slightly inhibited by high concentrations of retinol, as demonstrated by cell counting and [(3)H]-thymidine incorporation. Moreover, the differentiation capacities of these cells were markedly and dose-dependently inhibited by retinol, as shown by the reduced expression of the lipogenic enzyme glycerol-3-phosphate dehydrogenase and by microscopic morphological analysis. These results strongly suggest that retinol, by inhibiting the ability of human preadipocytes to convert into mature adipocytes, could be of potential interest in the prevention of human adipose tissue development in general and of cellulitis in particular. PMID:18503465

  11. Polymerase I and transcript release factor (PTRF) regulates adipocyte differentiation and determines adipose tissue expandability

    PubMed Central

    Perez-Diaz, Sergio; Johnson, Lance A.; DeKroon, Robert M.; Moreno-Navarrete, Jose M.; Alzate, Oscar; Fernandez-Real, Jose M.; Maeda, Nobuyo; Arbones-Mainar, Jose M.

    2014-01-01

    Impaired adipogenesis renders an adipose tissue unable to expand, leading to lipotoxicity and conditions such as diabetes and cardiovascular disease. While factors important for adipogenesis have been studied extensively, those that set the limits of adipose tissue expansion remain undetermined. Feeding a Western-type diet to apolipoprotein E2 knock-in mice, a model of metabolic syndrome, produced 3 groups of equally obese mice: mice with normal glucose tolerance, hyperinsulinemic yet glucose-tolerant mice, and prediabetic mice with impaired glucose tolerance and reduced circulating insulin. Using proteomics, we compared subcutaneous adipose tissues from mice in these groups and found that the expression of PTRF (polymerase I and transcript release factor) associated selectively with their glucose tolerance status. Lentiviral and pharmacologically overexpressed PTRF, whose function is critical for caveola formation, compromised adipocyte differentiation of cultured 3T3-L1cells. In human adipose tissue, PTRF mRNA levels positively correlated with markers of lipolysis and cellular senescence. Furthermore, a negative relationship between telomere length and PTRF mRNA levels was observed in human subcutaneous fat. PTRF is associated with limited adipose tissue expansion underpinning the key role of caveolae in adipocyte regulation. Furthermore, PTRF may be a suitable adipocyte marker for predicting pathological obesity and inform clinical management.—Perez-Diaz, S., Johnson, L. A., DeKroon, R. M., Moreno-Navarrete, J. M., Alzate, O., Fernandez-Real, J. M., Maeda, N., Arbones-Mainar, J. M. Polymerase I and transcript release factor (PTRF) regulates adipocyte differentiation and determines adipose tissue expandability. PMID:24812087

  12. Insulin regulation of protein biosynthesis in differentiated 3T3 adipocytes. Regulation of glyceraldehyde-3-phosphate dehydrogenase

    SciTech Connect

    Alexander, M.; Curtis, G.; Avruch, J.; Goodman, H.M.

    1985-10-05

    The effect of insulin on protein biosynthesis was examined in differentiated 3T3-L1 and 3T3-F442A adipocytes. Insulin altered the relative rate of synthesis of specific proteins independent of its ability to hasten conversion of the fibroblast (preadipocyte) phenotype to the adipocyte phenotype. Although more than one pattern of response to insulin was observed, the authors focused on the induction of a Mr 33,000 protein which was identified as the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Exposure of 3T3 adipocytes to insulin throughout differentiation specifically increased GAPDH activity and protein content by 2- to 3-fold as compared to 3T3 adipocytes differentiated in the absence of insulin. These changes in enzyme activity and content could be accounted for by a 4-fold increase in the relative rate of synthesis of GAPDH and a 9-fold increase in hybridizable mRNA levels. Within 2 h of insulin addition to 3T3 adipocytes differentiated in the absence of hormone, hybridizable GAPDH mRNA levels increased 3-fold, and within 24 h GAPDH mRNA levels increased 8-fold, and (TVS) methionine incorporation into GAPDH protein increased 5-fold. These studies demonstrate that insulin, as the sole hormonal perturbant, can increase the synthesis of certain 3T3 adipocyte proteins by altering the cellular content of a specific mRNA.

  13. Adipocyte in vascular wall can induce the rupture of abdominal aortic aneurysm

    PubMed Central

    Kugo, Hirona; Zaima, Nobuhiro; Tanaka, Hiroki; Mouri, Youhei; Yanagimoto, Kenichi; Hayamizu, Kohsuke; Hashimoto, Keisuke; Sasaki, Takeshi; Sano, Masaki; Yata, Tatsuro; Urano, Tetsumei; Setou, Mitsutoshi; Unno, Naoki; Moriyama, Tatsuya

    2016-01-01

    Abdominal aortic aneurysm (AAA) is a vascular disease involving the gradual dilation of the abdominal aorta. It has been reported that development of AAA is associated with inflammation of the vascular wall; however, the mechanism of AAA rupture is not fully understood. In this study, we investigated the mechanism underlying AAA rupture using a hypoperfusion-induced animal model. We found that the administration of triolein increased the AAA rupture rate in the animal model and that the number of adipocytes was increased in ruptured vascular walls compared to non-ruptured walls. In the ruptured group, macrophage infiltration and the protein levels of matrix metalloproteinases 2 and 9 were increased in the areas around adipocytes, while collagen-positive areas were decreased in the areas with adipocytes compared to those without adipocytes. The administration of fish oil, which suppresses adipocyte hypertrophy, decreased the number and size of adipocytes, as well as decreased the risk of AAA rupture ratio by 0.23 compared to the triolein administered group. In human AAA samples, the amount of triglyceride in the adventitia was correlated with the diameter of the AAA. These results suggest that AAA rupture is related to the abnormal appearance of adipocytes in the vascular wall. PMID:27499372

  14. Adipocyte in vascular wall can induce the rupture of abdominal aortic aneurysm.

    PubMed

    Kugo, Hirona; Zaima, Nobuhiro; Tanaka, Hiroki; Mouri, Youhei; Yanagimoto, Kenichi; Hayamizu, Kohsuke; Hashimoto, Keisuke; Sasaki, Takeshi; Sano, Masaki; Yata, Tatsuro; Urano, Tetsumei; Setou, Mitsutoshi; Unno, Naoki; Moriyama, Tatsuya

    2016-01-01

    Abdominal aortic aneurysm (AAA) is a vascular disease involving the gradual dilation of the abdominal aorta. It has been reported that development of AAA is associated with inflammation of the vascular wall; however, the mechanism of AAA rupture is not fully understood. In this study, we investigated the mechanism underlying AAA rupture using a hypoperfusion-induced animal model. We found that the administration of triolein increased the AAA rupture rate in the animal model and that the number of adipocytes was increased in ruptured vascular walls compared to non-ruptured walls. In the ruptured group, macrophage infiltration and the protein levels of matrix metalloproteinases 2 and 9 were increased in the areas around adipocytes, while collagen-positive areas were decreased in the areas with adipocytes compared to those without adipocytes. The administration of fish oil, which suppresses adipocyte hypertrophy, decreased the number and size of adipocytes, as well as decreased the risk of AAA rupture ratio by 0.23 compared to the triolein administered group. In human AAA samples, the amount of triglyceride in the adventitia was correlated with the diameter of the AAA. These results suggest that AAA rupture is related to the abnormal appearance of adipocytes in the vascular wall. PMID:27499372

  15. Free Fatty Acids, Lipopolysaccharide and IL-1α Induce Adipocyte Manganese Superoxide Dismutase Which Is Increased in Visceral Adipose Tissues of Obese Rodents

    PubMed Central

    Krautbauer, Sabrina; Eisinger, Kristina; Neumeier, Markus; Hader, Yvonne; Buettner, Roland; Schmid, Peter M.; Aslanidis, Charalampos; Buechler, Christa

    2014-01-01

    Excess fat storage in adipocytes is associated with increased generation of reactive oxygen species (ROS) and impaired activity of antioxidant mechanisms. Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme involved in detoxification of ROS, and objective of the current study is to analyze expression and regulation of MnSOD in obesity. MnSOD is increased in visceral but not subcutaneous fat depots of rodents kept on high fat diets (HFD) and ob/ob mice. MnSOD is elevated in visceral adipocytes of fat fed mice and exposure of differentiating 3T3-L1 cells to lipopolysaccharide, IL-1α, saturated, monounsaturated and polyunsaturated free fatty acids (FFA) upregulates its level. FFA do not alter cytochrome oxidase 4 arguing against overall induction of mitochondrial enzymes. Upregulation of MnSOD in fat loaded cells is not mediated by IL-6, TNF or sterol regulatory element binding protein 2 which are induced in these cells. MnSOD is similarly abundant in perirenal fat of Zucker diabetic rats and non-diabetic animals with similar body weight and glucose has no effect on MnSOD in 3T3-L1 cells. To evaluate whether MnSOD affects adipocyte fat storage, MnSOD was knocked-down in adipocytes for the last three days of differentiation and in mature adipocytes. Knock-down of MnSOD does neither alter lipid storage nor viability of these cells. Heme oxygenase-1 which is induced upon oxidative stress is not altered while antioxidative capacity of the cells is modestly reduced. Current data show that inflammation and excess triglyceride storage raise adipocyte MnSOD which is induced in epididymal adipocytes in obesity. PMID:24475187

  16. Effect of Lactobacillus plantarum FH185 on the Reduction of Adipocyte Size and Gut Microbial Changes in Mice with Diet-induced Obesity

    PubMed Central

    2015-01-01

    This study aimed to investigate the effects of Lactobacillus plantarum FH185 on the reduction of adipocyte size and gut microbial changes in mice with diet-induced obesity. The strain was found to have a lipase inhibitory activity of 70.09±2.04% and inhibited adipocyte differentiation of 3T3-L1 cells (18.63±0.98%) at a concentration of 100 µg/mL. To examine the effect of the strain supplementation on gut microbial changes in mice with diet-induced obesity, male C57BL/6J mice were fed on four different diets (i.e., A, normal diet (ND); B, high-fat diet (HFD); C, HFD with ABT-3 (109 CFU/day); and D, HFD with L. plantarum FH185 (109 CFU/day)) for 6 wk. According to the results of fecal pyrosequencing, the ratio of Firmicutes to Bacteroidetes in groups C and D was lower than in the control groups at the phylum level. At the family level, Lactobacillaceae in groups C and D was observed to dominate, while Lachnospiraceae in groups A and B was observed to dominate. At the genus level, Lactobacillus in groups C and D was comparatively higher than in groups A and B. To examine the effects of strain supplementation on the reduction of adipocyte size, the left and right epididymal fat pads were quickly isolated after the animals were sacrificed, and the adipocyte sizes were measured. In groups A, C and D, the percentage of 2,000 m2 of adipocyte was higher than in the other size of adipocyte, while the percentage of over 5,000 m2 of adipocyte was highest in group B. The mean adipocyte size of group D was significantly larger than that of group A, but smaller than that of group B. PMID:26761825

  17. miR-30e reciprocally regulates the differentiation of adipocytes and osteoblasts by directly targeting low-density lipoprotein receptor-related protein 6.

    PubMed

    Wang, J; Guan, X; Guo, F; Zhou, J; Chang, A; Sun, B; Cai, Y; Ma, Z; Dai, C; Li, X; Wang, B

    2013-01-01

    Reciprocal relationship usually exists between osteoblastogenesis and adipogenesis, with factors stimulating one of these processes at the same time inhibiting the other. In the present study, miR-30e was found to be involved in the reciprocal regulation of osteoblast and adipocyte differentiation. Our data indicated that miR-30e was induced in primarily cultured mouse bone marrow stromal cell, mesenchymal cell line C3H10T1/2 and preadipocyte 3T3-L1 after adipogenic treatment. Conversely, it was reduced in mouse stromal line ST2 and preosteoblast MC3T3-E1 after osteogenic treatment. Enforced expression of miR-30e in 3T3-L1 significantly suppressed the growth of the cells and induced the preadipocytes to differentiate into mature adipocytes, along with increased expression of adipocyte-specific transcription factors peroxisome proliferator-activated receptor-γ (PPARγ), CCAAT/enhancer binding protein-α (C/EBPα) and C/EBPβ, and the marker gene aP2. In contrast, inhibition of the endogenous miR-30e enhanced the cell growth and repressed preadipocytes to differentiate. Conversely, supplementing miR-30e activity blocked, whereas knocking down miR-30e enforced the preosteoblast MC3T3-E1 to fully differentiate. Furthermore, miR-30e overexpression stimulated adipocyte formation and inhibited osteoblast differentiation from marrow stromal cells. Low-density lipoprotein receptor-related protein 6 (LRP6), one of the critical coreceptor for Wnts, was shown to be a direct target of miR-30e by using the luciferase assay. Knockdown of LRP6 in 3T3-L1 cells downregulated β-catenin/T-cell factor (TCF) transcriptional activity and dramatically potentiated the differentiation of the cells into mature adipocytes. Taken together, the present work suggests that the expression of miR-30e is indispensable for maintaining the balance of adipocytes and osteoblasts by targeting the canonical Wnt/β-catenin signaling. PMID:24113179

  18. Loss of Oncostatin M Signaling in Adipocytes Induces Insulin Resistance and Adipose Tissue Inflammation in Vivo.

    PubMed

    Elks, Carrie M; Zhao, Peng; Grant, Ryan W; Hang, Hardy; Bailey, Jennifer L; Burk, David H; McNulty, Margaret A; Mynatt, Randall L; Stephens, Jacqueline M

    2016-08-12

    Oncostatin M (OSM) is a multifunctional gp130 cytokine. Although OSM is produced in adipose tissue, it is not produced by adipocytes. OSM expression is significantly induced in adipose tissue from obese mice and humans. The OSM-specific receptor, OSM receptor β (OSMR), is expressed in adipocytes, but its function remains largely unknown. To better understand the effects of OSM in adipose tissue, we knocked down Osmr expression in adipocytes in vitro using siRNA. In vivo, we generated a mouse line lacking Osmr in adiponectin-expressing cells (OSMR(FKO) mice). The effects of OSM on gene expression were also assessed in vitro and in vivo OSM exerts proinflammatory effects on cultured adipocytes that are partially rescued by Osmr knockdown. Osm expression is significantly increased in adipose tissue T cells of high fat-fed mice. In addition, adipocyte Osmr expression is increased following high fat feeding. OSMR(FKO) mice exhibit increased insulin resistance and adipose tissue inflammation and have increased lean mass, femoral length, and bone volume. Also, OSMR(FKO) mice exhibit increased expression of Osm, the T cell markers Cd4 and Cd8, and the macrophage markers F4/80 and Cd11c Interestingly, the same proinflammatory genes induced by OSM in adipocytes are induced in the adipose tissue of the OSMR(FKO) mouse, suggesting that increased expression of proinflammatory genes in adipose tissue arises both from adipocytes and other cell types. These findings suggest that adipocyte OSMR signaling is involved in the regulation of adipose tissue homeostasis and that, in obesity, OSMR ablation may exacerbate insulin resistance by promoting adipose tissue inflammation. PMID:27325693

  19. Aculeatin, a coumarin derived from Toddalia asiatica (L.) Lam., enhances differentiation and lipolysis of 3T3-L1 adipocytes

    SciTech Connect

    Watanabe, Akio; Kato, Tsuyoshi; Ito, Yusuke; Yoshida, Izumi; Harada, Teppei; Mishima, Takashi; Fujita, Kazuhiro; Watai, Masatoshi; Nakagawa, Kiyotaka; Miyazawa, Teruo

    2014-10-31

    Highlights: • Aculeatin promoted adipocyte differentiation. • Aculeatin improved glucose uptake. • Aculeatin enhanced adipocyte lipolysis. - Abstract: Toddalia asiatica (L.) Lam. (T. asiatica) has been utilized traditionally for medicinal purposes such as the treatment of diabetes. Currently, the extract is considered to be a good source of anti-diabetic agents, but the active compounds have yet to be identified. In this study, we investigated the effects of fractionated T. asiatica extracts on the differentiation of 3T3-L1 preadipocytes and identified aculeatin as a potential active agent. When 3T3-L1 preadipocytes were treated with aculeatin isolated from T. asiatica in the presence of insulin, aculeatin increased cellular triglyceride levels and glycerol-3-phosphate dehydrogenase activity. This indicated that aculeatin could enhance the differentiation of preadipocytes into adipocytes. Further analyses using a DNA microarray and real-time quantitative reverse-transcription PCR showed an increase in the expression of peroxisome proliferator-activated receptor-γ target genes (Pparg, Ap2, Cd36, Glut4 and Adipoq) by aculeatin, suggesting that aculeatin enhances the differentiation of 3T3-L1 cells by modulating the expression of genes critical for adipogenesis. Interestingly, after treatment of differentiated adipocytes with aculeatin, glucose uptake and lipolysis were enhanced. Overall, our results suggested that aculeatin is an active compound in T. asiatica for enhancing both differentiation and lipolysis of adipocytes, which are useful for the treatment of lipid abnormalities as well as diabetes.

  20. Effects of Tithonia diversifolia (Hemsl.) A. Gray extract on adipocyte differentiation of human mesenchymal stem cells.

    PubMed

    Di Giacomo, Claudia; Vanella, Luca; Sorrenti, Valeria; Santangelo, Rosa; Barbagallo, Ignazio; Calabrese, Giovanna; Genovese, Carlo; Mastrojeni, Silvana; Ragusa, Salvatore; Acquaviva, Rosaria

    2015-01-01

    Tithonia diversifolia (Hemsl.) A. Gray (Asteraceae) is widely used in traditional medicine. There is increasing interest on the in vivo protective effects of natural compounds contained in plants against oxidative damage caused from reactive oxygen species. In the present study the total phenolic and flavonoid contents of aqueous, methanol and dichloromethane extracts of leaves of Tithonia diversifolia (Hemsl.) A. Gray were determined; furthermore, free radical scavenging capacity of each extract and the ability of these extracts to inhibit in vitro plasma lipid peroxidation were also evaluated. Since oxidative stress may be involved in trasformation of pre-adipocytes into adipocytes, to test the hypothesis that Tithonia extract may also affect adipocyte differentiation, human mesenchymal stem cell cultures were treated with Tithonia diversifolia aqueous extract and cell viability, free radical levels, Oil-Red O staining and western bolt analysis for heme oxygenase and 5'-adenosine monophoshate-activated protein kinase were carried out. Results obtained in the present study provide evidence that Tithonia diversifolia (Hemsl.) A. Gray exhibits interesting health promoting properties, resulting both from its free radical scavenger capacity and also by induction of protective cellular systems involved in cellular stress defenses and in adipogenesis of mesenchymal cells. PMID:25848759

  1. Effects of Tithonia diversifolia (Hemsl.) A. Gray Extract on Adipocyte Differentiation of Human Mesenchymal Stem Cells

    PubMed Central

    Di Giacomo, Claudia; Vanella, Luca; Sorrenti, Valeria; Santangelo, Rosa; Barbagallo, Ignazio; Calabrese, Giovanna; Genovese, Carlo; Mastrojeni, Silvana; Ragusa, Salvatore; Acquaviva, Rosaria

    2015-01-01

    Tithonia diversifolia (Hemsl.) A. Gray (Asteraceae) is widely used in traditional medicine. There is increasing interest on the in vivo protective effects of natural compounds contained in plants against oxidative damage caused from reactive oxygen species. In the present study the total phenolic and flavonoid contents of aqueous, methanol and dichloromethane extracts of leaves of Tithonia diversifolia (Hemsl.) A. Gray were determined; furthermore, free radical scavenging capacity of each extract and the ability of these extracts to inhibit in vitro plasma lipid peroxidation were also evaluated. Since oxidative stress may be involved in trasformation of pre-adipocytes into adipocytes, to test the hypothesis that Tithonia extract may also affect adipocyte differentiation, human mesenchymal stem cell cultures were treated with Tithonia diversifolia aqueous extract and cell viability, free radical levels, Oil-Red O staining and western bolt analysis for heme oxygenase and 5'-adenosine monophoshate-activated protein kinase were carried out. Results obtained in the present study provide evidence that Tithonia diversifolia (Hemsl.) A. Gray exhibits interesting health promoting properties, resulting both from its free radical scavenger capacity and also by induction of protective cellular systems involved in cellular stress defenses and in adipogenesis of mesenchymal cells. PMID:25848759

  2. RBM4a-regulated splicing cascade modulates the differentiation and metabolic activities of brown adipocytes

    PubMed Central

    Lin, Jung-Chun; Lu, Yi-Han; Liu, Yun-Ru; Lin, Ying-Ju

    2016-01-01

    RNA-binding motif protein 4a (RBM4a) reportedly reprograms splicing profiles of the insulin receptor (IR) and myocyte enhancer factor 2C (MEF2C) genes, facilitating the differentiation of brown adipocytes. Using an RNA-sequencing analysis, we first compared the gene expressing profiles between wild-type and RBM4a−/− brown adipocytes. The ablation of RBM4a led to increases in the PTBP1, PTBP2 (nPTB), and Nova1 proteins, whereas elevated RBM4a reduced the expression of PTBP1 and PTBP2 proteins in brown adipocytes through an alternative splicing-coupled nonsense-mediated decay mechanism. Subsequently, RBM4a indirectly shortened the half-life of the Nova1 transcript which was comparatively stable in the presence of PTBP2. RBM4a diminished the influence of PTBP2 in adipogenic development by reprogramming the splicing profiles of the FGFR2 and PKM genes. These results constitute a mechanistic understanding of the RBM4a-modulated splicing cascade during the brown adipogenesis. PMID:26857472

  3. Epoxyeicosatrienoic Acids Regulate Adipocyte Differentiation of Mouse 3T3 Cells, Via PGC-1α Activation, Which Is Required for HO-1 Expression and Increased Mitochondrial Function.

    PubMed

    Waldman, Maayan; Bellner, Lars; Vanella, Luca; Schragenheim, Joseph; Sodhi, Komal; Singh, Shailendra P; Lin, Daohong; Lakhkar, Anand; Li, Jiangwei; Hochhauser, Edith; Arad, Michael; Darzynkiewicz, Zbigniew; Kappas, Atallah; Abraham, Nader G

    2016-07-15

    Epoxyeicosatrienoic acid (EET) contributes to browning of white adipose stem cells to ameliorate obesity/diabetes and insulin resistance. In the current study, we show that EET altered preadipocyte function, enhanced peroxisome proliferation-activated receptor γ coactivator α (PGC-1α) expression, and increased mitochondrial function in the 3T3-L1 preadipocyte subjected to adipogenesis. Cells treated with EET resulted in an increase, P < 0.05, in PGC-1α and a decrease in mitochondria-derived ROS (MitoSox), P < 0.05. The EET increase in heme oxygenase-1 (HO-1) levels is dependent on activation of PGC-1α as cells deficient in PGC-1α (PGC-1α knockout adipocyte cell) have an impaired ability to express HO-1, P < 0.02. Additionally, adipocytes treated with EET exhibited an increase in mitochondrial superoxide dismutase (SOD) in a PGC-1α-dependent manner, P < 0.05. The increase in PGC-1α was associated with an increase in β-catenin, P < 0.05, adiponectin expression, P < 0.05, and lipid accumulation, P < 0.02. EET decreased heme levels and mitochondria-derived ROS (MitoSox), P < 0.05, compared to adipocytes that were untreated. EET also decreased mesoderm-specific transcript (MEST) mRNA and protein levels (P < 0.05). Adipocyte secretion of EET act in an autocrine/paracrine manner to increase PGC-1α is required for activation of HO-1 expression. This is the first study to dissect the mechanism by which the antiadipogenic and anti-inflammatory lipid, EET, induces the PGC-1α signaling cascade and reprograms the adipocyte phenotype by regulating mitochondrial function and HO-1 expression, leading to an increase in healthy, that is, small, adipocytes and a decrease in adipocyte enlargement and terminal differentiation. This is manifested by an increase in mitochondrial function and an increase in the canonical Wnt signaling cascade during adipocyte proliferation and terminal differentiation. PMID:27224420

  4. Indole-3-carbinol directly targets SIRT1 to inhibit adipocyte differentiation.

    PubMed

    Choi, Y; Um, S J; Park, T

    2013-06-01

    Indole-3-carbinol (I3C), a natural product of Brassica vegetables such as broccoli and cabbage, inhibits proliferation and induces apoptosis in various cancer cells. I3C has recently received attention as a possible anti-obesity agent. However, how I3C interacts with specific targets in the pathways involved in obesity and metabolic disorders is unknown. Silent mating type information regulation 2 homolog 1 (SIRT1), a NADþ-dependent deacetylase sirtuin, has recently emerged as a novel therapeutic target for metabolic diseases. Herein, we report that I3C is a potent, specific SIRT1 activator efficacious in cultured 3T3-L1 cell lines. A pull-down assay showed that I3C binds to SIRT1. To assess the significance of this binding, we determined whether I3C could activate SIRT1 deacetylase activity in a cell-free system. We found that I3C binds to SIRT1 and activates SIRT1 deacetylase activity in 3T3-L1 cells. In addition, I3C did not inhibit adipocyte differentiation in 3T3-L1 cells in which SIRT1 was knockdowned. Further, reverse transcriptase polymerase chain reaction analysis showed that I3C treatment reduced mRNA levels of adipogenic genes that encode for C/EBPa, PPARg2, FAS, and aP2 in 3T3-L1 cells but not in SIRT1 knockdown cells. Overall, these results suggested that I3C ameliorates adipogenesis by activating SIRT1 in 3T3-L1 cells. PMID:22986685

  5. Interleukin-1β mediates macrophage-induced impairment of insulin signaling in human primary adipocytes

    PubMed Central

    Gao, Dan; Madi, Mohamed; Ding, Cherlyn; Fok, Matthew; Steele, Thomas; Ford, Christopher; Hunter, Leif

    2014-01-01

    Adipose tissue expansion during obesity is associated with increased macrophage infiltration. Macrophage-derived factors significantly alter adipocyte function, inducing inflammatory responses and decreasing insulin sensitivity. Identification of the major factors that mediate detrimental effects of macrophages on adipocytes may offer potential therapeutic targets. IL-1β, a proinflammatory cytokine, is suggested to be involved in the development of insulin resistance. This study investigated the role of IL-1β in macrophage-adipocyte cross-talk, which affects insulin signaling in human adipocytes. Using macrophage-conditioned (MC) medium and human primary adipocytes, we examined the effect of IL-1β antagonism on the insulin signaling pathway. Gene expression profile and protein abundance of insulin signaling molecules were determined, as was the production of proinflammatory cytokine/chemokines. We also examined whether IL-1β mediates MC medium-induced alteration in adipocyte lipid storage. MC medium and IL-1β significantly reduced gene expression and protein abundance of insulin signaling molecules, including insulin receptor substrate-1, phosphoinositide 3-kinase p85α, and glucose transporter 4 and phosphorylation of Akt. In contrast, the expression and release of the proinflammatory markers, including IL-6, IL-8, monocyte chemotactic protein-1, and chemokine (C-C motif) ligand 5 by adipocytes were markedly increased. These changes were significantly reduced by blocking IL-1β activity, its receptor binding, or its production by macrophages. MC medium-inhibited expression of the adipogenic factors and -stimulated lipolysis was also blunted with IL-1β neutralization. We conclude that IL-1β mediates, at least in part, the effect of macrophages on insulin signaling and proinflammatory response in human adipocytes. Blocking IL-1β could be beneficial for preventing obesity-associated insulin resistance and inflammation in human adipose tissue. PMID:24918199

  6. Antioxidant, anti-adipocyte differentiation, antitumor activity and anthelmintic activities against Anisakis simplex and Hymenolepis nana of yakuchinone A from Alpinia oxyphylla

    PubMed Central

    2013-01-01

    Background Alpinia oxyphylla is a common remedy in traditional Chinese medicine. Yakuchinone A is a major constituent of A. oxyphylla and exhibits anti-inflammatory, antitumor, antibacterial, and gastric protective activities. Methods Antioxidant and antitumor characteristics of yakuchinone A in skin cancer cells as well as novel mechanisms for the inhibition of adipocyte differentiation, cestocidal activities against Hymenolepis nana adults, and nematocidal activities against Anisakis simplex larvae are investigated. Results Yakuchinone A presents the ability of the removal of DPPH·and ABTS+ free radicals and inhibition of lipid peroxidation. Yakuchinone A suppresses intracellular lipid accumulation during adipocyte differentiation in 3 T3-L1 cells and the expressions of leptin and peroxisome proliferator-activated receptor γ (PPARγ). Yakuchinone A induces apoptosis and inhibits cell proliferation in skin cancer cells. The inhibition of cell growth by yakuchinone A is more significant for non-melanoma skin cancer (NMSC) cells than for melanoma (A375 and B16) and noncancerous (HaCaT and BNLCL2) cells. Treatment BCC cells with yakuchinone A shows down-regulation of Bcl-2, up-regulation of Bax, and an increase in cleavage poly (ADP-ribose) polymerase (PARP). This suggests that yakuchinone A induces BCC cells apoptosis through the Bcl-2-mediated signaling pathway. The anthelmintic activities of yakuchinone A for A. simplex are better than for H. nana. Conclusions In this work, yakuchinone A exhibits antioxidative properties, anti-adipocyte differentiation, antitumor activity, and anthelmintic activities against A. simplex and H. nana. PMID:24070160

  7. Major role of adipocyte prostaglandin E2 in lipolysis-induced macrophage recruitment.

    PubMed

    Hu, Xiaoqian; Cifarelli, Vincenza; Sun, Shishuo; Kuda, Ondrej; Abumrad, Nada A; Su, Xiong

    2016-04-01

    Obesity induces accumulation of adipose tissue macrophages (ATMs), which contribute to both local and systemic inflammation and modulate insulin sensitivity. Adipocyte lipolysis during fasting and weight loss also leads to ATM accumulation, but without proinflammatory activation suggesting distinct mechanisms of ATM recruitment. We examined the possibility that specific lipid mediators with anti-inflammatory properties are released from adipocytes undergoing lipolysis to induce macrophage migration. In the present study, we showed that conditioned medium (CM) from adipocytes treated with forskolin to stimulate lipolysis can induce migration of RAW 264.7 macrophages. In addition to FFAs, lipolytic stimulation increased release of prostaglandin E2(PGE2) and prostaglandin D2(PGD2), reflecting cytosolic phospholipase A2α activation and enhanced cyclooxygenase (COX) 2 expression. Reconstituted medium with the anti-inflammatory PGE2potently induced macrophage migration while different FFAs and PGD2had modest effects. The ability of CM to induce macrophage migration was abolished by treating adipocytes with the COX2 inhibitor sc236 or by treating macrophages with the prostaglandin E receptor 4 antagonist AH23848. In fasted mice, macrophage accumulation in adipose tissue coincided with increases of PGE2levels and COX1 expression. Collectively, our data show that adipocyte-originated PGE2with inflammation suppressive properties plays a significant role in mediating ATM accumulation during lipolysis. PMID:26912395

  8. Hibiscus sabdariffa L. water extract inhibits the adipocyte differentiation through the PI3-K and MAPK pathway.

    PubMed

    Kim, Jin-Kyung; So, Hongseob; Youn, Myung-Ja; Kim, Hyung-Jin; Kim, Yunha; Park, Channy; Kim, Se-Jin; Ha, Young-Ae; Chai, Kyu-Yun; Kim, Shin-Moo; Kim, Ki-Young; Park, Raekil

    2007-11-01

    Hibiscus sabdariffa L., a tropical beverage material and medical herb, is used commonly as in folk medicines against hypertension, pyrexia, inflammation, liver disorders, and obesity. This report was designed to investigate the inhibitory mechanisms of hibiscus extract on adipocyte differentiation in 3T3-L1 preadipocytes. The possible inhibitory pathways that regulate the adipocyte differentiation contain the adipogenic transcription factors, C/EBPalpha and PPARgamma, PI3-kinase, and MAPK pathway. In this study, we examined whether hibiscus extract affected the adipogenesis via these three pathways. To differentiate preadipocyte in adipocyte, confluent 3T3-L1 preadipocytes were treated with the hormone mixture including isobutylmethylxanthine, dexamethasone, and insulin (MDI). Hibiscus extract inhibited significantly the lipid droplet accumulation by MDI in a dose-dependent manner and attenuated dramatically the protein and mRNA expressions of adipogenic transcriptional factors, C/EBPalpha and PPARgamma, during adipogenesis. The increase of phosphorylation and expression of PI3-K/Akt during adipocytic differentiation was markedly inhibited by treatment with hibiscus extract or PI3-K inhibitors. Furthermore, the phosphorylation and expression of MEK-1/ERK known to regulate the early phase of adipogenesis were clearly decreased with the addition of hibiscus extract. Taken together, this report suggests that hibiscus extract inhibits the adipocyte differentiation through the modulation of PI3-K/Akt and ERK pathway that play pivotal roles during adipogenesis. PMID:17904778

  9. Genome-wide analysis of gene expression during adipogenesis in human adipose-derived stromal cells reveals novel patterns of gene expression during adipocyte differentiation.

    PubMed

    Ambele, Melvin Anyasi; Dessels, Carla; Durandt, Chrisna; Pepper, Michael Sean

    2016-05-01

    We have undertaken an in-depth transcriptome analysis of adipogenesis in human adipose-derived stromal cells (ASCs) induced to differentiate into adipocytes in vitro. Gene expression was assessed on days 1, 7, 14 and 21 post-induction and genes differentially expressed numbered 128, 218, 253 and 240 respectively. Up-regulated genes were associated with blood vessel development, leukocyte migration, as well as tumor growth, invasion and metastasis. They also shared common pathways with certain obesity-related pathophysiological conditions. Down-regulated genes were enriched for immune response processes. KLF15, LMO3, FOXO1 and ZBTB16 transcription factors were up-regulated throughout the differentiation process. CEBPA, PPARG, ZNF117, MLXIPL, MMP3 and RORB were up-regulated only on days 14 and 21, which coincide with the maturation of adipocytes and could possibly serve as candidates for controlling fat accumulation and the size of mature adipocytes. In summary, we have identified genes that were up-regulated only on days 1 and 7 or days 14 and 21 that could serve as potential early and late-stage differentiation markers. PMID:27108396

  10. Vav1 Regulates Mesenchymal Stem Cell Differentiation Decision Between Adipocyte and Chondrocyte via Sirt1.

    PubMed

    Qu, Peng; Wang, Lizhen; Min, Yongfen; McKennett, Lois; Keller, Jonathan R; Lin, P Charles

    2016-07-01

    Mesenchymal stem cells (MSCs) are multipotent stromal cells residing in the bone marrow. MSCs have the potential to differentiate to adipocytes, chondrocytes, and other types of cells. In this study, we investigated the molecular mechanism that controls MSC cell fate decisions for differentiation. We found that Vav1, a guanine nucleotide exchange factor for Rho GTPase, was highly expressed in MSCs. Interestingly, loss of Vav1 in MSCs led to spontaneous adipogenic but impaired chondrogenic differentiation, and accordingly Vav1 null mice displayed an increase in fat content and a decrease in cartilage. Conversely, ectopic expression of Vav1 in MSCs reversed this phenotype, and led to enhanced MSC differentiation into chondrocyte but retarded adipogenesis. Mechanistically, loss of Vav1 reduced the level of Sirt1, which was responsible for an increase of acetylated PPARγ. As acetylation activates PPARγ, it increased C/EBPα expression and promoted adipogenesis. On the other hand, loss of Vav1 resulted in an increase of acetylated Sox9, a target of Sirt1. As acetylation represses Sox9 activity, it led to a dramatic reduction of collagen 2α1, a key regulator in chondrocyte differentiation. Finally, we found that Vav1 regulates Sirt1 in MSCs through Creb. Together this study reveals a novel function of Vav1 in regulating MSC cell fate decisions for differentiation through Sirt1. Sirt1 deacetylates PPARγ and Sox9, two key mediators that control adipocyte and chondrocyte differentiation. The acetylation status of PPARγ and Sox9 has opposite effects on its activity, thereby controlling cell fate decision. Stem Cells 2016;34:1934-1946. PMID:26990002

  11. Trichostatin A modulates thiazolidinedione-mediated suppression of tumor necrosis factor α-induced lipolysis in 3T3-L1 adipocytes.

    PubMed

    Lu, Juu-Chin; Chang, Yu-Tzu; Wang, Chih-Tien; Lin, Yu-Chun; Lin, Chun-Ken; Wu, Zhong-Sheng

    2013-01-01

    In obesity, high levels of tumor necrosis factor α (TNFα) stimulate lipolysis in adipocytes, leading to hyperlipidemia and insulin resistance. Thiazolidinediones (TZDs), the insulin-sensitizing drugs, antagonize TNFα-induced lipolysis in adipocytes, thereby increasing insulin sensitivity in diabetes patients. The cellular target of TZDs is peroxisome proliferator-activated receptor γ (PPARγ), a nuclear receptor that controls many adipocyte functions. As a transcription factor, PPARγ is closely modulated by coregulators, which include coactivators and corepressors. Previous studies have revealed that in macrophages, the insulin-sensitizing effect of PPARγ may involve suppression of proinflammatory gene expression by recruiting the corepressor complex that contains corepressors and histone deacetylases (HDACs). Therefore, we investigated whether the corepressor complex is involved in TZD-mediated suppression of TNFα-induced lipolysis in 3T3-L1 adipocytes. Trichostatin A (TSA), a pan HDAC inhibitor (HDACI) that inhibits class I and II HDACs, was used to examine the involvement of HDACs in the actions of TZDs. TSA alone increased basal lipolysis and attenuated TZD-mediated suppression of TNFα-induced lipolysis. Increased basal lipolysis may in part result from class I HDAC inhibition because selective class I HDACI treatment had similar results. However, attenuation of TZD-mediated TNFα antagonism may be specific to TSA and related hydroxamate-based HDACI rather than to HDAC inhibition. Consistently, corepressor depletion did not affect TZD-mediated suppression. Interestingly, TSA treatment greatly reduced PPARγ levels in differentiated adipocytes. Finally, extracellular signal-related kinase 1/2 (ERK1/2) mediated TNFα-induced lipolysis, and TZDs suppressed TNFα-induced ERK phosphorylation. We determined that TSA increased basal ERK phosphorylation, and attenuated TZD-mediated suppression of TNFα-induced ERK phosphorylation, consistent with TSA's effects

  12. Sera of overweight people promote in vitro adipocyte differentiation of bone marrow stromal cells

    PubMed Central

    2014-01-01

    Introduction Overweight status should not be considered merely an aesthetic concern; rather, it can incur health risks since it may trigger a cascade of events that produce further fat tissue through altered levels of circulating signaling molecules. There have been few studies addressing the effect of overweight status on the physiological functions of stem cells, including mesenchymal stem cells (MSCs), which are the progenitors of adipocytes and osteocytes and are a subset of the bone marrow stromal cell population. Methods We decided to investigate the influence of overweight individuals’ sera on in vitro MSC proliferation and differentiation. Results We observed that in vitro incubation of bone marrow stromal cells with the sera of overweight individuals promotes the adipogenic differentiation of MSCs while partially impairing proper osteogenesis. Conclusions These results, which represent a pilot study, might suggest that becoming overweight triggers further weight gains by promoting a bias in the differentiation potential of MSCs toward adipogenesis. The circulating factors involved in this phenomenon remain to be determined, since the great majority of the well known pro-inflammatory cytokines and adipocyte-secreted factors we investigated did not show relevant modifications in overweight serum samples compared with controls. PMID:24405848

  13. Mouse strains to study cold-inducible beige progenitors and beige adipocyte formation and function.

    PubMed

    Berry, Daniel C; Jiang, Yuwei; Graff, Jonathan M

    2016-01-01

    Cold temperatures induce formation of beige adipocytes, which convert glucose and fatty acids to heat, and may increase energy expenditure, reduce adiposity and lower blood glucose. This therapeutic potential is unrealized, hindered by a dearth of genetic tools to fate map, track and manipulate beige progenitors and 'beiging'. Here we examined 12 Cre/inducible Cre mouse strains that mark adipocyte, muscle and mural lineages, three proposed beige origins. Among these mouse strains, only those that marked perivascular mural cells tracked the cold-induced beige lineage. Two SMA-based strains, SMA-Cre(ERT2) and SMA-rtTA, fate mapped into the majority of cold-induced beige adipocytes and SMA-marked progenitors appeared essential for beiging. Disruption of the potential of the SMA-tracked progenitors to form beige adipocytes was accompanied by an inability to maintain body temperature and by hyperglycaemia. Thus, SMA-engineered mice may be useful to track and manipulate beige progenitors, beige adipocyte formation and function. PMID:26729601

  14. Temperature induced modulation of lipid oxidation and lipid accumulation in palmitate-mediated 3T3-L1 adipocytes and 3T3-L1 adipocytes.

    PubMed

    Lin, Xiaofen; Li, Yi; Leung, Polly Hangmei; Li, Jiashen; Hu, Junyan; Liu, Xuan; Li, Zhi

    2016-05-01

    Human skin temperature can vary widely depending on anatomical location and ambient temperature. It is also known that local changes in skin and subcutaneous temperature can affect fat metabolism. This study aimed to explore the potential effects of surrounding thermal environment on fat by investigating cell viability, lipid oxidation, and lipid accumulation in 3T3-L1 adipocytes and palmitate-treated adipocytes after 4h incubation. No significant differences of viability in 3T3-L1 adipocytes were detected under different temperature conditions. Despite no significant increase being observed under warm temperature (39°C) conditions, a similarly significant suppression of intracellular reactive oxygen species (ROS) and lipid peroxidation were found in 3T3-L1 adipocytes and palmitate-treated adipocytes under 4h exposure to cooler temperatures of 31-33°C (P<0.01). ROS, chemically reactive molecules containing oxygen, are currently understood to be a major contributor to oxidantive stress in obesity. Additionally, cooler temperatures (31-33°C) could improve the size of lipid droplets in 3T3-L1 adipocytes (P<0.01), but no significant effect was generated by temperature change on lipid droplets in palmitate-treated adipocytes. In the palmitate-induced adiposity model, although excessive ROS and lipid peroxidation has been attenuated by temperature decrease (P<0.01), it still does not positively modulate lipid droplet size (P>0.05) and remedy the palmitate damage induced cell death (P<0.01). These findings provide preliminary support for potential interventions based on temperature manipulation for cell metabolism of adipocytes. PMID:27157327

  15. Ivy gourd (Coccinia grandis L. Voigt) root suppresses adipocyte differentiation in 3T3-L1 cells

    PubMed Central

    2014-01-01

    Background Ivy gourd (Coccinia grandis L. Voigt) is a tropical plant widely distributed throughout Asia, Africa, and the Pacific Islands. The anti-obesity property of this plant has been claimed but still remains to be scientifically proven. We therefore investigated the effects of ivy gourd leaf, stem, and root on adipocyte differentiation by employing cell culture model. Methods Dried roots, stems, and leaves of ivy gourd were separately extracted with ethanol. Each extract was then applied to 3T3-L1 pre-adipocytes upon induction with a mixture of insulin, 3-isobutyl-1-methylxanthine, and dexamethasone, for anti-adipogenesis assay. The active extract was further fractionated by a sequential solvent partitioning method, and the resulting fractions were examined for their abilities to inhibit adipogenesis in 3T3-L1 cells. Differences in the expression of adipogenesis-related genes between the treated and untreated cells were determined from their mRNA and protein levels. Results Of the three ivy gourd extracts, the root extract exhibited an anti-adipogenic effect. It significantly reduced intracellular fat accumulation during the early stages of adipocyte differentiation. Together with the suppression of differentiation, expression of the genes encoding PPARγ, C/EBPα, adiponectin, and GLUT4 were down-regulated. Hexane-soluble fraction of the root extract also inhibited adipocyte differentiation and decreased the mRNA levels of various adipogenic genes in the differentiating cells. Conclusions This is the first study to demonstrate that ivy gourd root may prevent obesity based mainly on the ability of its active constituent(s) to suppress adipocyte differentiation in vitro. Such an inhibitory effect is mediated by at least down-regulating the expression of PPARγ-the key transcription factor of adipogenesis in pre-adipocytes during their early differentiation processes. PMID:24884680

  16. Sida rhomboidea. Roxb Leaf Extract Down-Regulates Expression of PPARγ2 and Leptin Genes in High Fat Diet Fed C57BL/6J Mice and Retards in Vitro 3T3L1 Pre-Adipocyte Differentiation

    PubMed Central

    Thounaojam, Menaka C.; Jadeja, Ravirajsinh N.; Ramani, Umed V.; Devkar, Ranjitsinh V.; Ramachandran, A. V.

    2011-01-01

    Sida rhomboidea. Roxb leaf extract (SRLE) is being used by the populace of North-East India to alleviate symptoms of diabetes and obesity. We have previously reported its hypolipidemic and anti-diabetic properties. In this study, we report the effect of SRLE on (i) in vivo modulation of genes controlling high fat diet (HFD) induced obesity and (ii) in vitro 3T3L1 pre-adipocyte differentiation and leptin release. Supplementation with SRLE significantly prevented HFD induced increment in bodyweight, plasma lipids and leptin, visceral adiposity and adipocyte hypertrophy. Also, SRLE supplementation reduced food intake, down regulated PPARγ2, SREBP1c, FAS and LEP expressions and up-regulated CPT-1 in epididymal adipose tissue compared to obese mice. In vitro adipogenesis of 3T3L1 pre-adipocytes was significantly retarded in the presence of SRLE extract. Also decreased triglyceride accumulation, leptin release and glyceraldehyde-3-Phosphate dehydrogenase activity along with higher glycerol release without significant alteration of viability of 3T3L1 pre-adipocytes, was recorded. Our findings suggest that prevention of HFD induced visceral adiposity is primarily by down regulation of PPARγ2 and leptin gene expression coupled with attenuation of food intake in C57BL/6J mice. SRLE induced prevention of pre-adipocytes differentiation, and leptin release further substantiated these findings and scientifically validates the potential application of SRLE as a therapeutic agent against obesity. PMID:21845103

  17. WEHI-3 cells inhibit adipocyte differentiation in 3T3-L1 cells

    SciTech Connect

    Lai, Jing; Liu, Gexiu; Yan, Guoyao; He, Dongmei; Zhou, Ying; Chen, Shengting

    2015-06-26

    By investigating the anti-adipogenic effects of WEHI-3 cells – a murine acute myelomonocytic leukemia cell line – we sought to improve the efficiency of hematopoietic stem cell transplantation (HSCT). Analysis of Oil Red O staining and the expression of adipogenic genes, including PPARγ, C/EBPα, FAS and LPL, indicated that WEHI-3 cells significantly inhibited 3T3-L1 mouse preadipocyte cells from differentiating into adipocytes. In vivo, fat vacuoles in mice injected with WEHI-3 cells were also remarkably reduced in the murine bone marrow pimelosis model. Moreover, the key gene in the Rho signaling pathway, ROCKII, and the key gene in the Wnt signaling pathway, β-catenin, were both upregulated compared with the control group. siRNA-mediated knockdown of ROCKII and β-catenin reversed these WEHI-3-mediated anti-adipogenic effects. Taken together, these data suggest that WEHI-3 cells exert anti-adipogenic effects and that both ROCKII and β-catenin are involved in this process. - Highlights: • WEHI-3, an acute myelomonocytic leukemia cell line, inhibited 3T3-L1 preadipocyte from differentiating into adipocyte. • WEHI-3 cells can arrest 3T3-L1 cells in G0/G1 phase by secreting soluble factors and thus inhibit their proliferation. • WEHI-3 cells reduced bone marrow pimelosis in the murine model. • Both ROCKII and β-catenin were involved in the WEHI-3-mediated anti-adipogenic effects.

  18. Nuclear lamin A inhibits adipocyte differentiation: implications for Dunnigan-type familial partial lipodystrophy.

    PubMed

    Boguslavsky, Revekka L; Stewart, Colin L; Worman, Howard J

    2006-02-15

    Mutations in the LMNA gene encoding A-type lamins cause several diseases, including Emery-Dreifuss muscular dystrophy and Dunnigan-type familial partial lipodystrophy (FPLD). We analyzed differentiation of 3T3-L1 preadipocytes to adipocytes in cells overexpressing wild-type lamin A as well as lamin A with amino acid substitutions at position 482 that cause FPLD. We also examined adipogenic conversion of mouse embryonic fibroblasts lacking A-type lamins. Overexpression of both wild-type and mutant lamin A inhibited lipid accumulation, triglyceride synthesis and expression of adipogenic markers. This was associated with inhibition of expression of peroxisome-proliferator-activated receptor gamma 2 (PPARgamma2) and Glut4. In contrast, embryonic fibroblasts lacking A-type lamins accumulated more intracellular lipid and exhibited elevated de novo triglyceride synthesis compared with wild-type fibroblasts. They also had increased basal phosphorylation of AKT1, a mediator of insulin signaling. We conclude that A-type lamins act as inhibitors of adipocyte differentiation, possibly by affecting PPARgamma2 and insulin signaling. PMID:16415042

  19. Bisindoylmaleimide I suppresses adipocyte differentiation through stabilization of intracellular {beta}-catenin protein

    SciTech Connect

    Cho, Munju; Park, Seoyoung; Gwak, Jungsug; Kim, Dong-Eun; Yea, Sung Su; Shin, Jae-Gook; Oh, Sangtaek

    2008-02-29

    The Wnt/{beta}-catenin signaling pathway plays important roles in cell differentiation. Activation of this pathway, likely by Wnt-10b, has been shown to inhibit adipogenesis in cultured 3T3-L1 preadipocytes and mice. Here we revealed that bisindoylmaleimide I (BIM), which is widely used as a specific inhibitor of protein kinase C (PKC), inhibits adipocyte differentiation through activation of the Wnt/{beta}-catenin signaling pathway. BIM increased {beta}-catenin responsive transcription (CRT) and up-regulated intracellular {beta}-catenin levels in HEK293 cells and 3T3-L1 preadipocytes. BIM significantly decreased intracellular lipid accumulation and reduced expression of important adipocyte marker genes including peroxisome-proliferator-activated receptor {gamma} (PPAR{gamma}) and CAATT enhancer-binding protein {alpha} (C/EBP{alpha}) in 3T3-L1 preadipocytes. Taken together, our findings indicate that BIM inhibits adipogenesis by increasing the stability of {beta}-catenin protein in 3T3-L1 preadipocyte cells.

  20. Role of histone acetyltransferases and histone deacetylases in adipocyte differentiation and adipogenesis.

    PubMed

    Zhou, Yuanfei; Peng, Jian; Jiang, Siwen

    2014-04-01

    Adipogenesis is a complex process strictly regulated by a well-established cascade that has been thoroughly studied in the last two decades. This process is governed by complex regulatory networks that involve the activation/inhibition of multiple functional genes, and is controlled by histone-modifying enzymes. Among such modification enzymes, histone acetyltransferases (HATs) and histone deacetylases (HDACs) play important roles in the transcriptional regulation and post-translational modification of protein acetylation. HATs and HDACs have been shown to respond to signals that regulate cell differentiation, participate in the regulation of protein acetylation, mediate transcription and post-translation modifications, and directly acetylate/deacetylate various transcription factors and regulatory proteins. In this paper, we review the role of HATs and HDACs in white and brown adipocyte differentiation and adipogenesis, to expand our knowledge on fat formation and adipose tissue biology. PMID:24810880

  1. Nonlinear optical microscopy of adipose-derived stem cells induced towards osteoblasts and adipocytes

    NASA Astrophysics Data System (ADS)

    Mouras, R.; Bagnaninchi, P.; Downes, A.; Muratore, M.; Elfick, A.

    2011-07-01

    Adipose-derived stem cells (ADSCs) are adult stem cells isolated from lipoaspirates. They are a good candidate for autologuous cell therapy and tissue engineering. For these applications, label-free imaging could be critical to assess noninvasively the efficiency of stem cell (SC) differentiation. We report on the development and application of a multimodal microscope to monitor and quantify ADSC differentiation into osteoblasts and adipocytes.

  2. Can Wnt5a and Wnt non-canonical pathways really mediate adipocyte de-differentiation in a tumour microenvironment?

    PubMed

    Chirumbolo, Salvatore; Bjørklund, Geir

    2016-09-01

    Wnt5a has been recently reported as a possible triggering factor of adipocyte de-differentiation into an adipocyte-derived fibroblast in the tumour microenvironment of pancreas cancer. The Wnt/β-catenin pathway was described in processes involving de-differentiation and epithelial-mesenchymal transition but some Wnt family-belonging molecules exert an adipogenic role on adipocyte, while other ones, such as Wnt10b or Wnt3a, an anti-adipogenic role. Although this ability depends on the different tumoural microenvironments, it is intriguing to ascertain if some Wnt molecules, participating in the non-canonical pathway, may be targeted as fundamental factors able to trigger the desmoplastic reaction of peritumoural white adipose tissue. PMID:27391920

  3. Anti-Inflammatory Effect of Spirulina platensis in Macrophages Is Beneficial for Adipocyte Differentiation and Maturation by Inhibiting Nuclear Factor-κB Pathway in 3T3-L1 Adipocytes.

    PubMed

    Pham, Tho X; Lee, Ji-Young

    2016-06-01

    We previously showed that the organic extract of a blue-green alga, Spirulina platensis (SPE), had potent anti-inflammatory effects in macrophages. As the interplay between macrophages and adipocytes is critical for adipocyte functions, we investigated the contribution of the anti-inflammatory effects of SPE in macrophages to adipogenesis/lipogenesis in 3T3-L1 adipocytes. 3T3-L1 preadipocytes were treated with 10% conditioned medium from lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages (CMC) or LPS-stimulated, but SPE-pretreated, macrophages (CMS) at different stages of adipocyte differentiation. The expression of adipocyte differentiation markers, such as CCAAT/enhancer-binding protein α, peroxisome proliferator-activated receptor γ, and perilipin, was significantly repressed by CMC when added on day 3, while the repression was attenuated by CMS. Oil Red O staining confirmed that adipocyte maturation in CMS-treated cells, but not in CMC-treated cells, was equivalent to that of control cells. Nuclear translocation of nuclear factor κB (NF-κB) p65 was decreased by CMS compared to CMC. In lipid-laden adipocytes, CMC promoted the loss of lipid droplets, while CMS had minimal effects. Histone deacetylase 9 mRNA and protein levels were increased during adipocyte maturation, which were decreased by CMC. In conclusion, by cross-talking with adipocytes, the anti-inflammatory effects of SPE in macrophages promoted adipocyte differentiation/maturation, at least in part, by repressing the activation of NF-κB inflammatory pathways, which otherwise can be compromised in inflammatory conditions. PMID:27206252

  4. Human adipocytes are highly sensitive to intermittent hypoxia induced NF-kappaB activity and subsequent inflammatory gene expression

    SciTech Connect

    Taylor, Cormac T.; Kent, Brian D.; Crinion, Sophie J.; McNicholas, Walter T.; Ryan, Silke

    2014-05-16

    Highlights: • Intermittent hypoxia (IH) leads to NF-κB activation in human primary adipocytes. • Adipocytes bear higher pro-inflammatory potential than other human primary cells. • IH leads to upregulation of multiple pro-inflammatory genes in human adipocytes. - Abstract: Introduction: Intermittent hypoxia (IH)-induced activation of pro-inflammatory pathways is a major contributing factor to the cardiovascular pathophysiology associated with obstructive sleep apnea (OSA). Obesity is commonly associated with OSA although it remains unknown whether adipose tissue is a major source of inflammatory mediators in response to IH. The aim of this study was to test the hypothesis that IH leads to augmented inflammatory responses in human adipocytes when compared to cells of non-adipocyte lineages. Methods and results: Human primary subcutaneous and visceral adipocytes, human primary microvascular pulmonary endothelial cells (HUMEC-L) and human primary small airway epithelial cells (SAEC) were exposed to 0, 6 or 12 cycles of IH or stimulated with tumor necrosis factor (TNF)-α. IH led to a robust increase in NF-κB DNA-binding activity in adipocytes compared with normoxic controls regardless of whether the source of adipocytes was visceral or subcutaneous. Notably, the NF-κB response of adipocytes to both IH and TNF-α was significantly greater than that in HUMEC-L and SAEC. Western blotting confirmed enhanced nuclear translocation of p65 in adipocytes in response to IH, accompanied by phosphorylation of I-κB. Parallel to p65 activation, we observed a significant increase in secretion of the adipokines interleukin (IL)-8, IL-6 and TNF-α with IH in adipocytes accompanied by significant upregulation of mRNA expression. PCR-array suggested profound influence of IH on pro-inflammatory gene expression in adipocytes. Conclusion: Human adipocytes demonstrate strong sensitivity to inflammatory gene expression in response to acute IH and hence, adipose tissue may be a key

  5. Initial differences in lipid processing leading to pig-and beef-derived mature adipocyte differentiation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clonal cultures of pig-derived mature adipocytes are capable of dedifferentiating and forming proliferative-competent progeny cells in vitro. Initial lipid processing, is different to that observed in cultures of beef-derived adipocytes. Mature pig adipocytes extrude lipid before proliferation, wher...

  6. Green tea catechins enhance norepinephrine-induced lipolysis via a protein kinase A-dependent pathway in adipocytes.

    PubMed

    Chen, Shu; Osaki, Noriko; Shimotoyodome, Akira

    2015-05-22

    Green tea catechins have been shown to attenuate obesity in animals and humans. The catechins activate adenosine monophosphate-activated protein kinase (AMPK), and thereby increase fatty acid oxidation in liver and skeletal muscles. Green tea catechins have also been shown to reduce body fat in humans. However, the effect of the catechins on lipolysis in adipose tissue has not been fully understood. The aim of this study was to clarify the effect of green tea catechins on lipolysis in adipocytes and to elucidate the underlying mechanism. Differentiated mouse adipocyte cell line (3T3-L1) was stimulated with green tea catechins in the presence or absence of norepinephrine. Glycerol and free fatty acids in the media were measured. Phosphorylation of hormone-sensitive lipase (HSL) was determined by Western blotting, and the mRNA expression levels of HSL, adipose triglyceride lipase (ATGL), and perilipin were determined by quantitative RT-PCR. The cells were treated with inhibitors of protein kinase A (PKA), protein kinase C (PKC), protein kinase G (PKG), or mitogen-activated protein kinase (MAPK) to determine the responsible pathway. Treatment of 3T3-L1 adipocytes with green tea catechins increased the level of glycerol and free fatty acids released into the media in the presence, but not absence, of norepinephrine, and increased the level of phosphorylated HSL in the cells. The catechins also increased mRNA and protein levels of HSL and ATGL. PKA inhibitor (H89) attenuated the catechin-induced increase in glycerol release and HSL phosphorylation. The results demonstrate that green tea catechins enhance lipolysis in the presence of norepinephrine via a PKA-dependent pathway in 3T3-L1 adipocytes, providing a potential mechanism by which green tea catechins could reduce body fat. PMID:25849890

  7. Transmembrane protein 64 reciprocally regulates osteoblast and adipocyte differentiation by modulating Wnt/β-catenin signaling.

    PubMed

    Jeong, Byung-Chul; Kim, Tae Soo; Kim, Hyun Soo; Lee, Seoung-Hoon; Choi, Yongwon

    2015-09-01

    Age-related osteoporosis is associated with a reciprocal decrease in bone formation and an increase in adiposity in the bone marrow niche. We previously reported Transmembrane protein 64 (Tmem64) to be an important regulator of osteoclast function; however, its precise role in osteoblasts has not yet been established. Here, we showed that ablation of the Tmem64 gene in mice resulted in markedly increased osteoblast and reduced adipocyte differentiation from bone marrow-derived stromal cells (BMSCs). Conversely, Tmem64 overexpression inhibited osteogenesis and accelerated adipogenesis. Furthermore, BMSCs isolated from Tmem64 knockout mice formed a greater number of colony-forming unit-osteoblasts and a lower number of colony-forming unit-adipocytes than the wild type controls. Mechanistically, the expression level of β-catenin, the key Wnt signaling molecule, increased significantly, and its nuclear translocation was enhanced in Tmem64-deficient cells. Introduction of Tmem64 significantly suppressed β-catenin-mediated transcriptional activity in an in vitro co-transfection experiment as well as during an in vivo experiment involving BAT-Gal reporter mice. These results demonstrate that Tmem64 plays an important role in the regulation of mesenchymal lineage allocation by modulating Wnt/β-catenin signaling. PMID:25979161

  8. Phloretin promotes adipocyte differentiation in vitro and improves glucose homeostasis in vivo.

    PubMed

    Shu, Gang; Lu, Nai-Sheng; Zhu, Xiao-Tong; Xu, Yong; Du, Min-Qing; Xie, Qiu-Ping; Zhu, Can-Jun; Xu, Qi; Wang, Song-Bo; Wang, Li-Na; Gao, Ping; Xi, Qian-Yun; Zhang, Yong-Liang; Jiang, Qing-Yan

    2014-12-01

    Adipocyte dysfunction is associated with many metabolic diseases such as obesity, insulin resistance and diabetes. Previous studies found that phloretin promotes 3T3-L1 cells differentiation, but the underlying mechanisms for phloretin's effects on adipogenesis remain unclear. In this study, we demonstrated that phloretin enhanced the lipid accumulation in porcine primary adipocytes in a time-dependent manner. Furthermore, phloretin increased the utilization of glucose and nonesterified fatty acid, while it decreased the lactate output. Microarray analysis revealed that genes associated with peroxisome proliferator-activated receptor-γ (PPARγ), mitogen-activated protein kinase and insulin signaling pathways were altered in response to phloretin. We further confirmed that phloretin enhanced expression of PPARγ, CAAT enhancer binding protein-α (C/EBPα) and adipose-related genes, such as fatty acids translocase and fatty acid synthase. In addition, phloretin activated the Akt (Thr308) and extracellular signal-regulated kinase, and therefore, inactivated Akt targets protein. Wortmannin effectively blocked the effect of phloretin on Akt activity and the protein levels of PPARγ, C/EBPα and fatty acid binding protein-4 (FABP4/aP2). Oral administration of 5 or 10 mg/kg phloretin to C57BL BKS-DB mice significantly decreased the serum glucose level and improved glucose tolerance. In conclusion, phloretin promotes the adipogenesis of porcine primary preadipocytes through Akt-associated signaling pathway. These findings suggested that phloretin might be able to increase insulin sensitivity and alleviate the metabolic diseases. PMID:25283330

  9. Interference with Akt signaling pathway contributes curcumin-induced adipocyte insulin resistance.

    PubMed

    Zhang, Deling; Zhang, Yemin; Ye, Mao; Ding, Youming; Tang, Zhao; Li, Mingxin; Zhou, Yu; Wang, Changhua

    2016-07-01

    Previous study has shown that curcumin directly or indirectly suppresses insulin signaling in 3T3-L1 adipocytes. However, the underlying mechanism remains unclear. Here we experimentally demonstrate that curcumin inhibited the ubiquitin-proteasome system (UPS) function, activated autophagy, and reduced protein levels of protein kinase B (Akt) in a dose- and time-dependent manner in 3T3-L1 adipocytes, accompanied with attenuation of insulin-stimulated Akt phosphorylation, plasma membrane translocation of glucose transporter type 4 (GLUT4), and glucose uptake. These in vitro inhibitory effects of curcumin on Akt protein expression and insulin action were reversed by pharmacological and genetic inhibition of autophagy but not by inhibition of the UPS and caspases. In addition, Akt reduction in adipose tissues of mice treated with curcumin could be recovered by administration of autophagy inhibitor bafilomycin A1 (BFA). This new finding provides a novel mechanism by which curcumin induces insulin resistance in adipocytes. PMID:27113027

  10. Differentiation of A31T6 proadipocytes to adipocytes: A flow cytometric analysis

    SciTech Connect

    Smyth, M.J.; Wharton, W. )

    1992-03-01

    A flow cytometric assay has been developed which provides precise, quantitative information on the accumulation of cytoplasmic triglycerides in individual A31T6 proadipocytes as they differentiate into adipocytes. The opportunity to measure multiple optical parameter on a cell-by-cell basis has enabled us to monitor phenotypic aspects of differentiation with a greater level of sensitivity than was previously possible. Using the fluorescent hydrophobic probe, Nile red, they have found that as a cell proceeds along the differentiation pathway, the gold fluorescence signal from the cell increases, reflecting the accumulation of cytoplasmic lipid droplets. They have determined (1) the presence of an undifferentiated population of cells whose existence is not detected by conventional phase microscopy, (2) that insulin is no required to drive differentiation in this system, (3) that exposure to a combination of insulin and dexamethasone results in a lower accumulation of lipid in a cell than does exposure to either agent alone, and (4) that A31T6 cells show the same response to differentiation-promoting agents whether applied at the time of plating or at confluence.

  11. Differentiation of the insulin-sensitive glucose transporter in 3T3-L1 adipocytes

    SciTech Connect

    Frost, S.C.; Baly, D.L.; Cushman, S.W.; Lane, M.D.; Simpson, I.A.

    1986-05-01

    3T3-L1 fibroblasts differentiate in culture to resemble adipocytes both morphologically and biochemically. Insulin-sensitive glucose transport, as measured by 2-deoxy-(1-/sup 14/C)- glucose uptake in the undifferentiated cell is small (2X). In contrast, the rate of glucose transport in fully differentiated cells is elevated 15-fold over basal in the presence of insulin. To determine if this is due to an increase in the number of transporters/cell or accessibility to the transporters, the number of transporters was measured in subcellular fractions over differentiation using a /sup 3/H-cytochalasin B binding assay. The increase in the rate of insulin-sensitive glucose transport directly parallels an increase in the number of transporters which reside in an insulin-responsive intracellular compartment. This observation was confirmed by identifying the transporters by immunoblotting using an antibody generated against the human erythrocyte transporter. The molecular weight of this transporter increases over differentiation from a single band of 40kDa to a heterogeneous triplet of 40, 44 and 48kDa. These data suggest that the transporter undergoes differential processing and that the functional, insulin-responsive transporter may be different from the insulin-insensitive (basal) transporter.

  12. Suppression of Adipocyte Differentiation by Foenumoside B from Lysimachia foenum-graecum Is Mediated by PPARγ Antagonism.

    PubMed

    Kwak, Hyun Jeong; Choi, Hye-Eun; Jang, Jinsun; Park, Soo Kyoung; Cho, Byoung Heon; Kim, Seul Ki; Lee, Sunyi; Kang, Nam Sook; Cheon, Hyae Gyeong

    2016-01-01

    Lysimachia foenum-graecum extract (LFE) and its active component foenumoside B (FSB) have been shown to inhibit adipocyte differentiation, but their mechanisms were poorly defined. Here, we investigated the molecular mechanisms responsible for their anti-adipogenic effects. Both LFE and FSB inhibited the differentiation of 3T3-L1 preadipocytes induced by peroxisome proliferator-activated receptor-γ (PPARγ) agonists, accompanied by reductions in the expressions of the lipogenic genes aP2, CD36, and FAS. Moreover, LFE and FSB inhibited PPARγ transactivation activity with IC50s of 22.5 μg/ml and 7.63 μg/ml, respectively, and showed selectivity against PPARα and PPARδ. Rosiglitazone-induced interaction between PPARγ ligand binding domain (LBD) and coactivator SRC-1 was blocked by LFE or FSB, whereas reduced NCoR-1 binding to PPARγ by rosiglitazone was reversed in the presence of LFE or FSB. In vivo administration of LFE into either ob/ob mice or KKAy mice reduced body weights, and levels of PPARγ and C/EBPα in fat tissues. Furthermore, insulin resistance was ameliorated by LFE treatment, with reduced adipose tissue inflammation and hepatic steatosis. Thus, LFE and FSB were found to act as PPARγ antagonists that improve insulin sensitivity and metabolic profiles. We propose that LFE and its active component FSB offer a new therapeutic strategy for metabolic disorders including obesity and insulin resistance. PMID:27176632

  13. Suppression of Adipocyte Differentiation by Foenumoside B from Lysimachia foenum-graecum Is Mediated by PPARγ Antagonism

    PubMed Central

    Kwak, Hyun Jeong; Choi, Hye-Eun; Jang, Jinsun; Park, Soo Kyoung; Cho, Byoung Heon; Kim, Seul Ki; Lee, Sunyi; Kang, Nam Sook; Cheon, Hyae Gyeong

    2016-01-01

    Lysimachia foenum-graecum extract (LFE) and its active component foenumoside B (FSB) have been shown to inhibit adipocyte differentiation, but their mechanisms were poorly defined. Here, we investigated the molecular mechanisms responsible for their anti-adipogenic effects. Both LFE and FSB inhibited the differentiation of 3T3-L1 preadipocytes induced by peroxisome proliferator-activated receptor-γ (PPARγ) agonists, accompanied by reductions in the expressions of the lipogenic genes aP2, CD36, and FAS. Moreover, LFE and FSB inhibited PPARγ transactivation activity with IC50s of 22.5 μg/ml and 7.63 μg/ml, respectively, and showed selectivity against PPARα and PPARδ. Rosiglitazone-induced interaction between PPARγ ligand binding domain (LBD) and coactivator SRC-1 was blocked by LFE or FSB, whereas reduced NCoR-1 binding to PPARγ by rosiglitazone was reversed in the presence of LFE or FSB. In vivo administration of LFE into either ob/ob mice or KKAy mice reduced body weights, and levels of PPARγ and C/EBPα in fat tissues. Furthermore, insulin resistance was ameliorated by LFE treatment, with reduced adipose tissue inflammation and hepatic steatosis. Thus, LFE and FSB were found to act as PPARγ antagonists that improve insulin sensitivity and metabolic profiles. We propose that LFE and its active component FSB offer a new therapeutic strategy for metabolic disorders including obesity and insulin resistance. PMID:27176632

  14. Adipocyte deficiency of angiotensinogen prevents obesity-induced hypertension in male mice.

    PubMed

    Yiannikouris, Frederique; Gupte, Manisha; Putnam, Kelly; Thatcher, Sean; Charnigo, Richard; Rateri, Debra L; Daugherty, Alan; Cassis, Lisa A

    2012-12-01

    Previous studies demonstrated that diet-induced obesity increased plasma angiotensin II concentrations and elevated systolic blood pressures in male mice. Adipocytes express angiotensinogen and secrete angiotensin peptides. We hypothesize that adipocyte-derived angiotensin II mediates obesity-induced increases in systolic blood pressure in male high fat-fed C57BL/6 mice. Systolic blood pressure was measured by radiotelemetry during week 16 of low-fat or high-fat feeding in Agt(fl/fl) and adipocyte angiotensinogen-deficient mice (Agt(aP2)). Adipocyte angiotensinogen deficiency had no effect on diet-induced obesity. Basal 24-hour systolic blood pressure was not different in low fat-fed Agt(fl/fl) compared with Agt(aP2) mice (124 ± 3 versus 128 ± 3 mm Hg, respectively). In Agt(fl/fl) mice, high-fat feeding significantly increased systolic blood pressure (24 hours; 134 ± 2 mm Hg; P<0.05). In contrast, high fat-fed Agt(aP2) mice did not exhibit an increase in systolic blood pressure (126 ± 2 mm Hg). Plasma angiotensin II concentrations were increased by high-fat feeding in Agt(fl/fl) mice (low fat, 32 ± 14; high fat, 219 ± 58 pg/mL; P<0.05). In contrast, high fat-fed Agt(aP2) mice did not exhibit elevated plasma angiotensin II concentrations (high fat, 18 ± 7 pg/mL). Similarly, adipose tissue concentrations of angiotensin II were significantly decreased in low fat- and high fat-fed Agt(aP2) mice compared with controls. In conclusion, adipocyte angiotensinogen deficiency prevented high fat-induced elevations in plasma angiotensin II concentrations and systolic blood pressure. These results suggest that adipose tissue serves as a major source of angiotensin II in the development of obesity hypertension. PMID:23108647

  15. Suppressive effects of saponin-enriched extracts from quinoa on 3T3-L1 adipocyte differentiation.

    PubMed

    Yao, Yang; Zhu, Yingying; Gao, Yue; Shi, Zhenxing; Hu, Yibo; Ren, Guixing

    2015-10-01

    This study was performed to investigate the effect of quinoa saponins (QS) on the differentiation of 3T3-L1 preadipocytes. QS inhibited triglyceride (TG) accumulation in the mature adipocytes, evidenced by oil-red O staining and intracellular quantification. Real time-PCR analysis and western blot analysis showed that QS significantly down-regulated the mRNA and protein expression of key adipogenic transcription factors, peroxisome proliferator-activated receptor γ (PPARγ), and CCAAT/enhancer-binding protein alpha (C/EBPα), however, they had no significant effect on CCAAT/enhancer-binding protein beta (C/EBPβ) and CCAAT/enhancer-binding protein delta (C/EBPδ) which are the upstream regulators for adipogenesis compared with mature adipocytes. QS also reduced mRNA and protein expression of sterol regulatory element-binding protein-1c (SREBP-1c) related to the late stage of adipogenesis. Furthermore, lipoprotein lipase (LPL), adipocyte protein 2 (aP2) and glucose transporter 4 (Glut4), as adipocyte specific genes, were decreased in mature adipocytes by QS treatment. These findings indicate that QS are capable of suppressing adipogenesis and therefore they seem to be natural bioactive factors effective in adipose tissue mass modulation. PMID:26242624

  16. Exercise-induced galanin release facilitated GLUT4 translocation in adipocytes of type 2 diabetic rats.

    PubMed

    Liang, Yan; Sheng, Shudong; Fang, Penghua; Ma, Yinping; Li, Jian; Shi, Qiaojia; Sui, Yumei; Shi, Mingyi

    2012-01-01

    Although galanin has been shown to increase insulin sensitivity in skeletal muscle of rats, there is no literature available about the effect of galanin on Glucose Transporter 4 (GLUT4) translocation from intracellular membrane pools to plasma membranes in adipocytes of type 2 diabetic rats. In the present study M35, a galanin antagonist was used to elucidate whether exercise-induced galanin release increased GLUT4 translocation in adipocytes of streptozotocin-induced diabetic rats. The present findings showed that plasma galanin levels after swimming training in all four trained groups were higher compared with each sedentary control. M35 treatment had an inhibitory effect on glucose infusion rates in the euglycemic-hyperinsulinemic clamp test and GLUT4 mRNA expression levels in adipocytes. Moreover, M35 treatment reduced GLUT4 concentration in both plasma membranes and total cell membranes. The ratios of GLUT4 contents in plasma membranes to total cell membranes in four drug groups were lower compared with each control. These data demonstrate a beneficial role of endogenous galanin to transfer GLUT4 from internal stores to plasma membranes in adipocytes of type 2 diabetic rats. Galanin plays a significant role in regulation of glucose metabolic homeostasis and is an important hormone relative to diabetes. PMID:22079346

  17. An inhibitory role of progerin in the gene induction network of adipocyte differentiation from iPS cells.

    PubMed

    Xiong, Zheng-Mei; LaDana, Christina; Wu, Di; Cao, Kan

    2013-04-01

    Lipodystrophies, characterized by partial or complete loss of adipose tissue, have been associated with mutations in the lamin A gene. It remains unclear how lamin A mutants interfere with adipose tissue formation. Hutchinson-Gilford progeria syndrome (HGPS) presents the most severe form of lamin A-associated diseases, whose patients show a complete loss of subcutaneous fat. Using iPSCs reprogrammed from HGPS fibroblasts, we induced adipocyte formation from iPSC derived embryoid bodies or from iPSC derived mesenchymal stem cells. Both approaches revealed a severe lipid storage defect in HGPS cells at late differentiation stage, faithfully recapitulating HGPS patient phenotype. Expression analysis further indicated that progerin inhibited the transcription activation of PPARγ2 and C/EBPα, but had little effects on the early adipogenic regulators. Our experiments demonstrate two comparable approaches of in vitro modeling lipodystrophies with patient-specific iPSCs, and support a regulatory role of lamin A in the terminal differentiation stage of adipogenesis. PMID:23596277

  18. Icariin Stimulates Differentiation and Suppresses Adipocytic Transdifferentiation of Primary Osteoblasts Through Estrogen Receptor-Mediated Pathway.

    PubMed

    Zhang, Dawei; Fong, Chichun; Jia, Zhenbin; Cui, Liao; Yao, Xinsheng; Yang, Mengsu

    2016-08-01

    Icariin, the main constituent of Herba Epimedii, appears to be a promising alternative to classic drugs used to treat osteoporosis. However, the detailed molecular mechanisms of its action and the role of icariin in the cross-talk between osteoblasts and adipocytes remain unclear. The present study was designed to investigate the gene expression profile of primary osteoblasts in the presence of icariin, and the effects of icariin on the differentiation and adipogenic transdifferentiation of osteoblasts. Cellular and molecular markers expressed during osteoblastic differentiation were assessed by cytochemical analysis, real-time quantitative PCR, Western blotting, and cDNA microarray analysis. Results indicated that icariin up-regulated the expression of runt-related transcription factor 2 (Runx2), bone morphogenetic protein 2 (Bmp2), and collagen type 1 (Col1) genes, and down-regulated the expression of the peroxisome proliferator-activated receptor γ (Pparg) and CCAAT/enhancer-binding protein β (Cebpb) genes. These effects were blocked by ICI 182,780, suggesting that icariin may be acting via the estrogen receptor (ER). Results also demonstrated that the ratio of osteoprotegerin (Opg)/receptor activator of nuclear factor kappa B ligand (Rankl) expression was up-regulated following treatment with icariin. In total, osteoblastic gene expression profile analysis suggested that 33 genes were affected by icariin; these could be sub-divided into nine functional categories. It appears that icariin could stimulate the differentiation and mineralization of osteoblasts, regulate the differentiation of osteoclasts, and inhibit the adipogenic transdifferentiation of osteoblasts, therefore increasing the number of osteoblasts undergoing differentiation to mature osteoblasts, via an ER-mediated pathway. In summary, icariin may exhibit beneficial effects on bone health, especially for patients with osteoporosis and obesity. PMID:27061090

  19. Development of an OP9 Derived Cell Line as a Robust Model to Rapidly Study Adipocyte Differentiation

    PubMed Central

    Lane, Jacqueline M.; Doyle, Jamie R.; Fortin, Jean-Philippe; Kopin, Alan S.; Ordovás, José M.

    2014-01-01

    One hallmark of obesity is adipocyte hypertrophy and hyperplasia. To gain novel insights into adipose biology and therapeutics, there is a pressing need for a robust, rapid, and informative cell model of adipocyte differentiation for potential RNAi and drug screens. Current models are prohibitive for drug and RNAi screens due to a slow differentiation time course and resistance to transfection. We asked if we could create a rapid, robust model of adipogenesis to potentially enable rapid functional and obesity therapeutic screens. We generated the clonal population OP9-K, which differentiates rapidly and reproducibly, and displays classic adipocyte morphology: rounded cell shape, lipid accumulation, and coalescence of lipids into a large droplet. We further validate the OP9-K cells as an adipocyte model system by microarray analysis of the differentiating transcriptome. OP9-K differentiates via known adipogenic pathways, involving the transcriptional activation and repression of common adipose markers Plin1, Gata2, C/Ebpα and C/Ebpβ and biological pathways, such as lipid metabolism, PPARγ signaling, and osteogenesis. We implemented a method to quantify lipid accumulation using automated microscopy and tested the ability of our model to detect alterations in lipid accumulation by reducing levels of the known master adipogenic regulator Pparγ. We further utilized our model to query the effects of a novel obesity therapeutic target, the transcription factor SPI1. We determine that reduction in levels of Spi1 leads to an increase in lipid accumulation. We demonstrate rapid, robust differentiation and efficient transfectability of the OP9-K cell model of adipogenesis. Together with our microscopy based lipid accumulation assay, adipogenesis assays can be achieved in just four days' time. The results of this study can contribute to the development of rapid screens with the potential to deepen our understanding of adipose biology and efficiently test obesity

  20. Mature adipocyte proteome reveals differentially altered protein abundances between lean, overweight and morbidly obese human subjects.

    PubMed

    Benabdelkamel, Hicham; Masood, Afshan; Almidani, Ghaith M; Alsadhan, Abdulmajeed A; Bassas, Abdulelah F; Duncan, Mark W; Alfadda, Assim A

    2015-02-01

    Overweight (OW) and obese individuals are considered to be graded parts of the scale having increasing weight as a common feature. They may not, however, be part of the same continuum and may differ metabolically. In this study we applied an untargeted proteomic approach to compare protein abundances in mature adipocytes derived from the subcutaneous adipose tissue of overweight and morbidly obese female subjects to those of lean age matched controls. Mature adipocytes were isolated from liposuction samples of abdominal subcutaneous adipose tissue collected from both lean (L; n = 7, 23.3 ± 0.4 kg/m(2); mean BMI ± SD), overweight (OW; n = 8, 27.9 ± 0.6 kg/m(2); mean BMI ± SD) and morbidly obese (MOB; n = 7, 44.8 ± 3.8 kg/m(2); mean BMI ± SD) individuals. Total protein extracts were then compared by two-dimensional difference in gel electrophoresis (2D DIGE). One hundred and ten differentially expressed protein spots (i.e., fitting the statistical criteria ANOVA test, p < 0.05; fold-change ≥1.5) were detected, and of these, 89 were identified by MALDI-TOF mass spectrometry. Of these, 66 protein spots were common to both groups whereas 23 were unique to the MOB group. Significant differences were evident in the abundances of key proteins involved in glucose and lipid metabolism, energy regulation, cytoskeletal structure and redox control signaling pathways. Differences in the abundance of some chaperones were also evident. The differentially abundant proteins were investigated using Ingenuity Pathway Analysis (IPA) to establish their associations with known biological functions. The network identified in the OW group with the highest score relates to-: cell-to-cell signaling and interaction; in contrast, in the MOB group the major interacting pathways are associated with lipid metabolism, small molecule biochemistry and cancer. The differences in abundance of the differentially regulated proteins were validated by

  1. Salt-Inducible Kinase 2 Couples Ovarian Cancer Cell Metabolism with Survival at the Adipocyte-Rich Metastatic Niche.

    PubMed

    Miranda, Fabrizio; Mannion, David; Liu, Shujuan; Zheng, Yiyan; Mangala, Lingegowda S; Redondo, Clara; Herrero-Gonzalez, Sandra; Xu, Ruoyan; Taylor, Charlotte; Chedom, Donatien Fotso; Karaminejadranjbar, Mohammad; Albukhari, Ashwag; Jiang, Dahai; Pradeep, Sunila; Rodriguez-Aguayo, Cristian; Lopez-Berestein, Gabriel; Salah, Eidarus; Abdul Azeez, Kamal R; Elkins, Jonathan M; Campo, Leticia; Myers, Kevin A; Klotz, Daniel; Bivona, Serena; Dhar, Sunanda; Bast, Robert C; Saya, Hideyuki; Choi, Hwan Geun; Gray, Nathanael S; Fischer, Roman; Kessler, Benedikt M; Yau, Christopher; Sood, Anil K; Motohara, Takeshi; Knapp, Stefan; Ahmed, Ahmed Ashour

    2016-08-01

    The adipocyte-rich microenvironment forms a niche for ovarian cancer metastasis, but the mechanisms driving this process are incompletely understood. Here we show that salt-inducible kinase 2 (SIK2) is overexpressed in adipocyte-rich metastatic deposits compared with ovarian primary lesions. Overexpression of SIK2 in ovarian cancer cells promotes abdominal metastasis while SIK2 depletion prevents metastasis in vivo. Importantly, adipocytes induce calcium-dependent activation and autophosphorylation of SIK2. Activated SIK2 plays a dual role in augmenting AMPK-induced phosphorylation of acetyl-CoA carboxylase and in activating the PI3K/AKT pathway through p85α-S154 phosphorylation. These findings identify SIK2 at the apex of the adipocyte-induced signaling cascades in cancer cells and make a compelling case for targeting SIK2 for therapy in ovarian cancer. PMID:27478041

  2. Go-6976 reverses hyperglycemia-induced insulin resistance independently of cPKC inhibition in adipocytes.

    PubMed

    Robinson, Katherine A; Hegyi, Krisztina; Hannun, Yusuf A; Buse, Maria G; Sethi, Jaswinder K

    2014-01-01

    Chronic hyperglycemia induces insulin resistance by mechanisms that are incompletely understood. One model of hyperglycemia-induced insulin resistance involves chronic preincubation of adipocytes in the presence of high glucose and low insulin concentrations. We have previously shown that the mTOR complex 1 (mTORC1) plays a partial role in the development of insulin resistance in this model. Here, we demonstrate that treatment with Go-6976, a widely used "specific" inhibitor of cPKCs, alleviates hyperglycemia-induced insulin resistance. However, the effects of mTOR inhibitor, rapamycin and Go-6976 were not additive and only rapamycin restored impaired insulin-stimulated AKT activation. Although, PKCα, (but not -β) was abundantly expressed in these adipocytes, our studies indicate cPKCs do not play a major role in causing insulin-resistance in this model. There was no evidence of changes in the expression or phosphorylation of PKCα, and PKCα knock-down did not prevent the reduction of insulin-stimulated glucose transport. This was also consistent with lack of IRS-1 phosphorylation on Ser-24 in hyperglycemia-induced insulin-resistant adipocytes. Treatment with Go-6976 did inhibit a component of the mTORC1 pathway, as evidenced by decreased phosphorylation of S6 ribosomal protein. Raptor knock-down enhanced the effect of insulin on glucose transport in insulin resistant adipocytes. Go-6976 had the same effect in control cells, but was ineffective in cells with Raptor knock-down. Taken together these findings suggest that Go-6976 exerts its effect in alleviating hyperglycemia-induced insulin-resistance independently of cPKC inhibition and may target components of the mTORC1 signaling pathway. PMID:25330241

  3. Anti-adipocyte Differentiation Activity and Chemical Composition of Essential Oil from Artemisia annua.

    PubMed

    Hwang, Dae Il; Won, Kyung-Jong; Kim, Do-yoon; Yoon, Seok Won; Park, Joo-Hoon; Kim, Bokyung; Lee, Hwan Myung

    2016-04-01

    Arteinisia annua L. essential oil (AAEO) has diverse properties including antibacterial, antioxidant, antinociceptive, and antimicrobial activities. However, the effect of AAEO on obesity remains to be investigated. In this study, we analyzed the compounds of AAEO and explored the effect of AAEO on the differentiation of preadipocyte into adipocyte using preadipocyte cell line 3T3-L1. Total yield of AAEO from 20 kg A. annua leaf and flower was 0.5%, v/w. Gas chromatography-mass spectrometry analysis showed that AAEO contained 34 compounds. 3T3-LI cells incubated in 3-isobutyl-l-methylxanthine / dexamethasone / insulin (MDI)-containing medium showed increased accumulation of lipid droplets. This increased response was suppressed by treatment with AAEO. Expressions of obesity-related proteins (PPARγ, C/EBPα, SREBP-1c, FAS, and ACC) were increased in 3T3-LI cells cultured in MDI medium and these responses were decreased by treatment with AAEO. These findings demonstrate that AAEO may suppress 3T3-LI cell differentiation by inhibiting adipogenesis and activation of lipid metabolism-related proteins. PMID:27396213

  4. A dynamic CTCF chromatin binding landscape promotes DNA hydroxymethylation and transcriptional induction of adipocyte differentiation.

    PubMed

    Dubois-Chevalier, Julie; Oger, Frédérik; Dehondt, Hélène; Firmin, François F; Gheeraert, Céline; Staels, Bart; Lefebvre, Philippe; Eeckhoute, Jérôme

    2014-01-01

    CCCTC-binding factor (CTCF) is a ubiquitously expressed multifunctional transcription factor characterized by chromatin binding patterns often described as largely invariant. In this context, how CTCF chromatin recruitment and functionalities are used to promote cell type-specific gene expression remains poorly defined. Here, we show that, in addition to constitutively bound CTCF binding sites (CTS), the CTCF cistrome comprises a large proportion of sites showing highly dynamic binding patterns during the course of adipogenesis. Interestingly, dynamic CTCF chromatin binding is positively linked with changes in expression of genes involved in biological functions defining the different stages of adipogenesis. Importantly, a subset of these dynamic CTS are gained at cell type-specific regulatory regions, in line with a requirement for CTCF in transcriptional induction of adipocyte differentiation. This relates to, at least in part, CTCF requirement for transcriptional activation of both the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARG) and its target genes. Functionally, we show that CTCF interacts with TET methylcytosine dioxygenase (TET) enzymes and promotes adipogenic transcriptional enhancer DNA hydroxymethylation. Our study reveals a dynamic CTCF chromatin binding landscape required for epigenomic remodeling of enhancers and transcriptional activation driving cell differentiation. PMID:25183525

  5. A dynamic CTCF chromatin binding landscape promotes DNA hydroxymethylation and transcriptional induction of adipocyte differentiation

    PubMed Central

    Dubois-Chevalier, Julie; Oger, Frédérik; Dehondt, Hélène; Firmin, François F.; Gheeraert, Céline; Staels, Bart; Lefebvre, Philippe; Eeckhoute, Jérôme

    2014-01-01

    CCCTC-binding factor (CTCF) is a ubiquitously expressed multifunctional transcription factor characterized by chromatin binding patterns often described as largely invariant. In this context, how CTCF chromatin recruitment and functionalities are used to promote cell type-specific gene expression remains poorly defined. Here, we show that, in addition to constitutively bound CTCF binding sites (CTS), the CTCF cistrome comprises a large proportion of sites showing highly dynamic binding patterns during the course of adipogenesis. Interestingly, dynamic CTCF chromatin binding is positively linked with changes in expression of genes involved in biological functions defining the different stages of adipogenesis. Importantly, a subset of these dynamic CTS are gained at cell type-specific regulatory regions, in line with a requirement for CTCF in transcriptional induction of adipocyte differentiation. This relates to, at least in part, CTCF requirement for transcriptional activation of both the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARG) and its target genes. Functionally, we show that CTCF interacts with TET methylcytosine dioxygenase (TET) enzymes and promotes adipogenic transcriptional enhancer DNA hydroxymethylation. Our study reveals a dynamic CTCF chromatin binding landscape required for epigenomic remodeling of enhancers and transcriptional activation driving cell differentiation. PMID:25183525

  6. The epigenetic modifiers 5-aza-2'-deoxycytidine and trichostatin A influence adipocyte differentiation in human mesenchymal stem cells.

    PubMed

    Zych, J; Stimamiglio, M A; Senegaglia, A C; Brofman, P R S; Dallagiovanna, B; Goldenberg, S; Correa, A

    2013-05-01

    Epigenetic mechanisms such as DNA methylation and histone modification are important in stem cell differentiation. Methylation is principally associated with transcriptional repression, and histone acetylation is correlated with an active chromatin state. We determined the effects of these epigenetic mechanisms on adipocyte differentiation in mesenchymal stem cells (MSCs) derived from bone marrow (BM-MSCs) and adipose tissue (ADSCs) using the chromatin-modifying agents trichostatin A (TSA), a histone deacetylase inhibitor, and 5-aza-2'-deoxycytidine (5azadC), a demethylating agent. Subconfluent MSC cultures were treated with 5, 50, or 500 nM TSA or with 1, 10, or 100 µM 5azadC for 2 days before the initiation of adipogenesis. The differentiation was quantified and expression of the adipocyte genes PPARG and FABP4 and of the anti-adipocyte gene GATA2 was evaluated. TSA decreased adipogenesis, except in BM-MSCs treated with 5 nM TSA. Only treatment with 500 nM TSA decreased cell proliferation. 5azadC treatment decreased proliferation and adipocyte differentiation in all conditions evaluated, resulting in the downregulation of PPARG and FABP4 and the upregulation of GATA2. The response to treatment was stronger in ADSCs than in BM-MSCs, suggesting that epigenetic memories may differ between cells of different origins. As epigenetic signatures affect differentiation, it should be possible to direct the use of MSCs in cell therapies to improve process efficiency by considering the various sources available. PMID:23797495

  7. Failure to Generate Bone Marrow Adipocytes Does Not Protect Mice from Ovariectomy-Induced Osteopenia

    PubMed Central

    Iwaniec, Urszula T.; Turner, Russell T.

    2012-01-01

    A reciprocal association between bone marrow fat and bone mass has been reported in ovariectomized rodents, suggesting that bone marrow adipogenesis has a negative effect on bone growth and turnover balance. Mice with loss of function mutations in kit receptor (kitW/W-v) have no bone marrow adipocytes in tibia or lumbar vertebra. We therefore tested the hypothesis that marrow fat contributes to development of osteopenia by comparing the skeletal response to ovariectomy (ovx) in growing wild type (WT) and bone marrow adipocyte-deficient kitW/W-v mice. Mice were ovx at 4 weeks of age and sacrificed 4 or 10 weeks post-surgery. Body composition was measured at necropsy by dual-energy X-ray absorptiometry. Cortical (tibia) and cancellous (tibia and lumbar vertebra) bone architecture were evaluated by microcomputed tomography. Bone marrow adipocyte size and density, osteoblast- and osteoclast-lined bone perimeters, and bone formation were determined by histomorphometry. Ovx resulted in an increase in total body fat mass at 10 weeks post-ovx in both genotypes, but the response was attenuated in the in kitW/W-v mice. Adipocytes were present in bone marrow of tibia and lumbar vertebra in WT mice and bone marrow adiposity increased following ovx. In contrast, marrow adipocytes were not detected in either intact or ovx kitW/W-v mice. However, ovx in WT and kitW/W-v mice resulted in statistically indistinguishable changes in cortical and cancellous bone mass, cortical and cancellous bone formation rate, and cancellous osteoblast and osteoclast-lined bone perimeters. In conclusion, our findings do not support a causal role for increased bone marrow fat as a mediator of ovx-induced osteopenia in mice. PMID:23246792

  8. Fucoxanthin exerts differing effects on 3T3-L1 cells according to differentiation stage and inhibits glucose uptake in mature adipocytes

    SciTech Connect

    Kang, Seong-Il; Ko, Hee-Chul; Shin, Hye-Sun; Kim, Hyo-Min; Hong, Youn-Suk; Lee, Nam-Ho; Kim, Se-Jae

    2011-06-17

    Highlights: {yields} Fucoxanthin enhances 3T3-L1 adipocyte differentiation at an early stage. {yields} Fucoxanthin inhibits 3T3-L1 adipocyte differentiation at intermediate and late stages. {yields} Fucoxanthin attenuates glucose uptake by inhibiting the phosphorylation of IRS in mature 3T3-L1 adipocytes. {yields} Fucoxanthin exerts its anti-obesity effect by inhibiting the differentiation of adipocytes at both intermediate and late stages, as well as glucose uptake in mature adipocytes. -- Abstract: Progression of 3T3-L1 preadipocyte differentiation is divided into early (days 0-2, D0-D2), intermediate (days 2-4, D2-D4), and late stages (day 4 onwards, D4-). In this study, we investigated the effects of fucoxanthin, isolated from the edible brown seaweed Petalonia binghamiae, on adipogenesis during the three differentiation stages of 3T3-L1 preadipocytes. When fucoxanthin was applied during the early stage of differentiation (D0-D2), it promoted 3T3-L1 adipocyte differentiation, as evidenced by increased triglyceride accumulation. At the molecular level, fucoxanthin increased protein expression of peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}), CCAAT/enhancer-binding protein {alpha} (C/EBP{alpha}), sterol regulatory element-binding protein 1c (SREBP1c), and aP2, and adiponectin mRNA expression, in a dose-dependent manner. However, it reduced the expression of PPAR{gamma}, C/EBP{alpha}, and SREBP1c during the intermediate (D2-D4) and late stages (D4-D7) of differentiation. It also inhibited the uptake of glucose in mature 3T3-L1 adipocytes by reducing the phosphorylation of insulin receptor substrate 1 (IRS-1). These results suggest that fucoxanthin exerts differing effects on 3T3-L1 cells of different differentiation stages and inhibits glucose uptake in mature adipocytes.

  9. Isolation of up- or down-regulated genes in PPARgamma-expressing NIH-3T3 cells during differentiation into adipocytes.

    PubMed

    Okuno, Masaaki; Arimoto, Emi; Nishizuka, Makoto; Nishihara, Tsutomu; Imagawa, Masayoshi

    2002-05-22

    Adipocyte differentiation is a complex process in which the expression of many transcription factors and adipocyte-specific genes is regulated under a strict program. The peroxisome proliferator-activated receptor gamma (PPARgamma), a member of the steroid/thyroid nuclear hormone receptor superfamily of ligand-activated transcription factors, is an important regulator of adipocyte gene expression and differentiation. In this study, we tried to identify downstream target genes of PPARgamma, by using PPARgamma-expressing cells and a subtractive cloning strategy, and isolated cDNA clones which were up-regulated or down-regulated by PPARgamma. Northern blot analyses revealed that the expression levels of the aldehyde dehydrogenase-2-like, type VI collagen alpha 3 subunit, cellular retinoic acid binding protein 1 and thrombospondin 1 are changed during the differentiation of mouse 3T3-L1 preadipocyte cells, indicating that these genes might be downstream targets of PPARgamma in adipocyte differentiation. PMID:12023027

  10. Lecithin:Cholesterol Acyltransferase (LCAT) Deficiency Promotes Differentiation of Satellite Cells to Brown Adipocytes in a Cholesterol-dependent Manner.

    PubMed

    Nesan, Dinushan; Tavallaee, Ghazaleh; Koh, Deborah; Bashiri, Amir; Abdin, Rawand; Ng, Dominic S

    2015-12-18

    Our laboratory previously reported that lecithin:cholesterol acyltransferase (LCAT) and LDL receptor double knock-out mice (Ldlr(-/-)xLcat(-/-) or DKO) spontaneously develop functioning ectopic brown adipose tissue (BAT) in skeletal muscle, putatively contributing to protection from the diet-induced obesity phenotype. Here we further investigated their developmental origin and the mechanistic role of LCAT deficiency. Gene profiling of skeletal muscle in DKO newborns and adults revealed a classical lineage. Primary quiescent satellite cells (SC) from chow-fed DKO mice, not in Ldlr(-/-)xLcat(+/+) single-knock-out (SKO) or C57BL/6 wild type, were found to (i) express exclusively classical BAT-selective genes, (ii) be primed to express key functional BAT genes, and (iii) exhibit markedly increased ex vivo adipogenic differentiation into brown adipocytes. This gene priming effect was abrogated upon feeding the mice a 2% high cholesterol diet in association with accumulation of excess intracellular cholesterol. Ex vivo cholesterol loading of chow-fed DKO SC recapitulated the effect, indicating that cellular cholesterol is a key regulator of SC-to-BAT differentiation. Comparing adipogenicity of Ldlr(+/+)xLcat(-/-) (LCAT-KO) SC with DKO SC identified a role for LCAT deficiency in priming SC to express BAT genes. Additionally, we found that reduced cellular cholesterol is important for adipogenic differentiation, evidenced by increased induction of adipogenesis in cholesterol-depleted SC from both LCAT-KO and SKO mice. Taken together, we conclude that ectopic BAT in DKO mice is classical in origin, and its development begins in utero. We further showed complementary roles of LCAT deficiency and cellular cholesterol reduction in the SC-to-BAT adipogenesis. PMID:26494623

  11. The size of the primary cilium and acetylated tubulin are modulated during adipocyte differentiation: Analysis of HDAC6 functions in these processes.

    PubMed

    Forcioli-Conti, Nicolas; Estève, David; Bouloumié, Anne; Dani, Christian; Peraldi, Pascal

    2016-05-01

    The primary cilium is an organelle present in most of the cells of the organism. Ciliopathies, such as the Bardet Biedl and the Alstrom syndromes are associated with obesity. We, and others, have shown that the primary cilium undergoes size modifications during adipocyte differentiation of human adipose stromal cells. We show here that the levels of acetylated α-tubulin, a constituent of the primary cilium, and the expression of HDAC6, the enzyme that deacetylates α-tubulin and is responsible for the loss of the cilium during mitosis, are modulated during adipogenesis. Moreover, during adipocyte differentiation cells that express higher level of HDAC6 are the first to lose their primary cilium. We have investigated the function of HDAC6 on adipocyte differentiation and on the primary cilium. We observe that inhibition of HDAC6 activity leads to a decrease in adipocyte differentiation. This is associated with an inhibition of the initial elongation of the cilium. Interestingly, overexpression of HDAC6 inhibits adipocyte differentiation and blunts the elongation of the primary cilium. In both situations, inhibition of adipocyte differentiation was not associated with an inhibition of the glucocorticoid receptor activity. This indicates that HDAC6 controls adipogenesis through the levels of acetylated α-tubulin. Moreover, we show that although HDAC6 expression increases during adipocyte differentiation it is not sufficient to provoke the loss of the cilium. This suggests the existence of a novel mechanism for the loss of the cilium. Together, these data indicate that HDAC6, and acetylated α-tubulin, are important regulator of adipocyte differentiation. PMID:26363102

  12. Hyperspectral and differential CARS microscopy for quantitative chemical imaging in human adipocytes.

    PubMed

    Di Napoli, Claudia; Pope, Iestyn; Masia, Francesco; Watson, Peter; Langbein, Wolfgang; Borri, Paola

    2014-05-01

    In this work, we demonstrate the applicability of coherent anti-Stokes Raman scattering (CARS) micro-spectroscopy for quantitative chemical imaging of saturated and unsaturated lipids in human stem-cell derived adipocytes. We compare dual-frequency/differential CARS (D-CARS), which enables rapid imaging and simple data analysis, with broadband hyperspectral CARS microscopy analyzed using an unsupervised phase-retrieval and factorization method recently developed by us for quantitative chemical image analysis. Measurements were taken in the vibrational fingerprint region (1200-2000/cm) and in the CH stretch region (2600-3300/cm) using a home-built CARS set-up which enables hyperspectral imaging with 10/cm resolution via spectral focussing from a single broadband 5 fs Ti:Sa laser source. Through a ratiometric analysis, both D-CARS and phase-retrieved hyperspectral CARS determine the concentration of unsaturated lipids with comparable accuracy in the fingerprint region, while in the CH stretch region D-CARS provides only a qualitative contrast owing to its non-linear behavior. When analyzing hyperspectral CARS images using the blind factorization into susceptibilities and concentrations of chemical components recently demonstrated by us, we are able to determine vol:vol concentrations of different lipid components and spatially resolve inhomogeneities in lipid composition with superior accuracy compared to state-of-the art ratiometric methods. PMID:24877002

  13. WEHI-3 cells inhibit adipocyte differentiation in 3T3-L1 cells.

    PubMed

    Lai, Jing; Liu, Gexiu; Yan, Guoyao; He, Dongmei; Zhou, Ying; Chen, Shengting

    2015-06-26

    By investigating the anti-adipogenic effects of WEHI-3 cells - a murine acute myelomonocytic leukemia cell line - we sought to improve the efficiency of hematopoietic stem cell transplantation (HSCT). Analysis of Oil Red O staining and the expression of adipogenic genes, including PPARγ, C/EBPα, FAS and LPL, indicated that WEHI-3 cells significantly inhibited 3T3-L1 mouse preadipocyte cells from differentiating into adipocytes. In vivo, fat vacuoles in mice injected with WEHI-3 cells were also remarkably reduced in the murine bone marrow pimelosis model. Moreover, the key gene in the Rho signaling pathway, ROCKII, and the key gene in the Wnt signaling pathway, β-catenin, were both upregulated compared with the control group. siRNA-mediated knockdown of ROCKII and β-catenin reversed these WEHI-3-mediated anti-adipogenic effects. Taken together, these data suggest that WEHI-3 cells exert anti-adipogenic effects and that both ROCKII and β-catenin are involved in this process. PMID:25911323

  14. Effect of different culture systems on adipocyte differentiation-related protein (ADRP) in bovine embryos.

    PubMed

    Al Darwich, A; Perreau, C; Tsikis, G; Coudert, E; Touzé, J L; Briant, E; Beckers, J F; Mermillod, P; Guignot, F

    2014-03-01

    Bovine embryos cultured in serum-containing media abnormally accumulate lipid droplets, compared to their in vivo counterparts. The objective of this study was to investigate the effect of different culture systems on the mRNA expression and on the quantification and localisation of adipocyte differentiation-related protein (ADRP), a protein associated with lipid accumulation in bovine blastocysts. Two experiments were independently performed for ADRP mRNA expression analysis. In experiment A, blastocysts were produced in modified synthetic oviduct fluid (mSOF)+10% foetal calf serum (FCS), in coculture (bovine oviduct epithelial cells, Boec) and in ewe oviducts, whereas in experiment B, they were produced in mSOF+10μM docosahexaenoic acid (DHA) and in vivo. Control groups were also performed. ADRP mRNA expression was downregulated in the Boec, ewe oviduct and in vivo groups compared to the 10% FCS or DHA groups, respectively. Moreover, the expression of this protein was downregulated in the Boec group compared to the control group (P<0.05). A third experiment (experiment C) was performed to quantify and localise ADRP protein. Boec, in vivo and control groups were tested. After immunofluorescence staining followed by confocal microscopy analysis, embryonic ADRP was clearly localised around lipid droplets, indicating that ADRP is also a lipid droplet coat protein in bovine embryos. In conclusion, our results demonstrate that bovine embryos at the blastocyst stage expressed ADRP mRNA and protein, and that the embryonic culture system modified this expression. PMID:24560670

  15. Lipid droplets fusion in adipocyte differentiated 3T3-L1 cells: A Monte Carlo simulation

    SciTech Connect

    Boschi, Federico; Rizzatti, Vanni; Zamboni, Mauro; Sbarbati, Andrea

    2014-02-15

    Several human worldwide diseases like obesity, type 2 diabetes, hepatic steatosis, atherosclerosis and other metabolic pathologies are related to the excessive accumulation of lipids in cells. Lipids accumulate in spherical cellular inclusions called lipid droplets (LDs) whose sizes range from fraction to one hundred of micrometers in adipocytes. It has been suggested that LDs can grow in size due to a fusion process by which a larger LD is obtained with spherical shape and volume equal to the sum of the progenitors’ ones. In this study, the size distribution of two populations of LDs was analyzed in immature and mature (5-days differentiated) 3T3-L1 adipocytes (first and second populations, respectively) after Oil Red O staining. A Monte Carlo simulation of interaction between LDs has been developed in order to quantify the size distribution and the number of fusion events needed to obtain the distribution of the second population size starting from the first one. Four models are presented here based on different kinds of interaction: a surface weighted interaction (R2 Model), a volume weighted interaction (R3 Model), a random interaction (Random model) and an interaction related to the place where the LDs are born (Nearest Model). The last two models mimic quite well the behavior found in the experimental data. This work represents a first step in developing numerical simulations of the LDs growth process. Due to the complex phenomena involving LDs (absorption, growth through additional neutral lipid deposition in existing droplets, de novo formation and catabolism) the study focuses on the fusion process. The results suggest that, to obtain the observed size distribution, a number of fusion events comparable with the number of LDs themselves is needed. Moreover the MC approach results a powerful tool for investigating the LDs growth process. Highlights: • We evaluated the role of the fusion process in the synthesis of the lipid droplets. • We compared the

  16. Malic enzyme tracers reveal hypoxia-induced switch in adipocyte NADPH pathway usage.

    PubMed

    Liu, Ling; Shah, Supriya; Fan, Jing; Park, Junyoung O; Wellen, Kathryn E; Rabinowitz, Joshua D

    2016-05-01

    The critical cellular hydride donor NADPH is produced through various means, including the oxidative pentose phosphate pathway (oxPPP), folate metabolism and malic enzyme. In growing cells, it is efficient to produce NADPH via the oxPPP and folate metabolism, which also make nucleotide precursors. In nonproliferating adipocytes, a metabolic cycle involving malic enzyme holds the potential to make both NADPH and two-carbon units for fat synthesis. Recently developed deuterium ((2)H) tracer methods have enabled direct measurement of NADPH production by the oxPPP and folate metabolism. Here we enable tracking of NADPH production by malic enzyme with [2,2,3,3-(2)H]dimethyl-succinate and [4-(2)H]glucose. Using these tracers, we show that most NADPH in differentiating 3T3-L1 mouse adipocytes is made by malic enzyme. The associated metabolic cycle is disrupted by hypoxia, which switches the main adipocyte NADPH source to the oxPPP. Thus, (2)H-labeled tracers enable dissection of NADPH production routes across cell types and environmental conditions. PMID:26999781

  17. Fibrin glue is a candidate scaffold for long-term therapeutic protein expression in spontaneously differentiated adipocytes in vitro

    SciTech Connect

    Aoyagi, Yasuyuki; Kuroda, Masayuki; Asada, Sakiyo; Tanaka, Shigeaki; Konno, Shunichi; Tanio, Masami; Aso, Masayuki; Okamoto, Yoshitaka; Nakayama, Toshinori; Saito, Yasushi; Bujo, Hideaki

    2012-01-01

    Adipose tissue is expected to provide a source of cells for protein replacement therapies via auto-transplantation. However, the conditioning of the environment surrounding the transplanted adipocytes for their long-term survival and protein secretion properties has not been established. We have recently developed a preparation procedure for preadipocytes, ceiling culture-derived proliferative adipocytes (ccdPAs), as a therapeutic gene vehicle suitable for stable gene product secretion. We herein report the results of our evaluation of using fibrin glue as a scaffold for the transplanted ccdPAs for the expression of a transduced gene in a three-dimensional culture system. The ccdPAs secreted the functional protein translated from an exogenously transduced gene, as well as physiological adipocyte proteins, and the long viability of ccdPAs (up to 84 days) was dependent on the fibrinogen concentrations. The ccdPAs spontaneously accumulated lipid droplets, and their expression levels of the transduced exogenous gene with its product were maintained for at least 56 days. The fibrinogen concentration modified the adipogenic differentiation of ccdPAs and their exogenous gene expression levels, and the levels of exogenously transduced gene expression at the different fibrinogen concentrations were dependent on the extent of adipogenic differentiation in the gel. These results indicate that fibrin glue helps to maintain the high adipogenic potential of cultured adipocytes after passaging in a 3D culture system, and suggests that once they are successfully implanted at the transplantation site, the cells exhibit increased expression of the transduced gene with adipogenic differentiation.

  18. Artepillin C, a Typical Brazilian Propolis-Derived Component, Induces Brown-Like Adipocyte Formation in C3H10T1/2 Cells, Primary Inguinal White Adipose Tissue-Derived Adipocytes, and Mice.

    PubMed

    Nishikawa, Sho; Aoyama, Hiroki; Kamiya, Misa; Higuchi, Jun; Kato, Aiko; Soga, Minoru; Kawai, Taeko; Yoshimura, Kazuki; Kumazawa, Shigenori; Tsuda, Takanori

    2016-01-01

    Induction of brown-like adipocytes (beige/brite cells) in white adipose tissue (WAT) suggests a new approach for preventing and treating obesity via induction of thermogenesis associated with uncoupling protein 1 (UCP1). However, whether diet-derived factors can directly induce browning of white adipocytes has not been well established. In addition, the underlying mechanism of induction of brown-like adipocytes by diet-derived factors has been unclear. Here, we demonstrate that artepillin C (ArtC), which is a typical Brazilian propolis-derived component, significantly induces brown-like adipocytes in murine C3H10T1/2 cells and primary inguinal WAT (iWAT)-derived adipocytes. This significant induction is due to activation of peroxisome proliferator-activated receptor γ and stabilization of PRD1-BF-1-RIZ1 homologous domain-containing protein-16 (PRDM16). Furthermore, the oral administration of ArtC (10 mg/kg) for 4 weeks significantly induced brown-like adipocytes accompanied by significant expression of UCP1 and PRDM16 proteins in iWAT of mice, and was independent of the β3-adrenergic signaling pathway via the sympathetic nervous system. These findings may provide insight into browning of white adipocytes including the molecular mechanism mediated by dietary factors and demonstrate that ArtC has a novel biological function with regard to increasing energy expenditure by browning of white adipocytes. PMID:27598888

  19. Skp2 promotes adipocyte differentiation via a p27{sup Kip1}-independent mechanism in primary mouse embryonic fibroblasts

    SciTech Connect

    Okada, Mitsuru; Sakai, Tamon; Nakamura, Takehiro; Tamamori-Adachi, Mimi; Kitajima, Shigetaka; Matsuki, Yasushi; Watanabe, Eijiro; Hiramatsu, Ryuji; Sakaue, Hiroshi Kasuga, Masato

    2009-02-06

    Skp2, the substrate-binding subunit of an SCF ubiquitin ligase complex, is a key regulator of cell cycle progression that targets substrates for degradation by the 26S proteasome. We have now shown that ablation of Skp2 in primary mouse embryonic fibroblasts (MEFs) results both in impairment of adipocyte differentiation and in the accumulation of the cyclin-dependent kinase inhibitor p27{sup Kip1}, a principal target of the SCF{sup Skp2} complex. Genetic ablation of p27{sup Kip1} in MEFs promoted both lipid accumulation and adipocyte-specific gene expression. However, depletion of p27{sup Kip1} by adenovirus-mediated RNA interference failed to correct the impairment of adipocyte differentiation in Skp2{sup -/-} MEFs. In contrast, troglitazone, a high-affinity ligand for peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}), largely restored lipid accumulation and PPAR{gamma} gene expression in Skp2{sup -/-} MEFs. Our data suggest that Skp2 plays an essential role in adipogenesis in MEFs in a manner that is at least in part independent of regulation of p27{sup Kip1} expression.

  20. Seapolynol Extracted from Ecklonia cava Inhibits Adipocyte Differentiation in Vitro and Decreases Fat Accumulation in Vivo.

    PubMed

    Jeon, Hui-Jeon; Choi, Hyeon-Son; Lee, Yeon-Joo; Hwang, Ji-Hyun; Lee, Ok-Hwan; Seo, Min-Jung; Kim, Kui-Jin; Lee, Boo-Yong

    2015-01-01

    Seapolynol (SN) is a polyphenol mixture derived from Ecklonia cava. We evaluated the effects of SN on lipid accumulation in adipocytes, zebrafish, and mice. SN effectively inhibited lipid accumulation in three experimental models by suppressing adipogenic factors. Triglyceride synthetic enzymes such as diacylglycerol acyltransferase 1 (DGAT1) and GPAT3 were also downregulated by SN. This SN-induced inhibition of adipogenic factors was shown to be due to the regulatory effect of SN on early adipogenic factors; SN downregulated the expression of Krueppel-like factor 4 (KLF4), KLF5, CCAAT-enhancer-binding protein β (C/EBPβ), C/EBPδ, and Protein C-ets-2 (ETS2), while KLF2, an anti-early adipogenic factor, was upregulated by SN. SN-mediated inhibition in early adipogenesis was closely correlated with the inhibition of mitotic clonal expansion via cell cycle arrest. SN inhibited cell cycle progression by suppressing cell cycle regulators, such as cyclin A, cyclinD, and pRb but increased p27, a cell cycle inhibitor. In a mouse study, SN effectively reduced body weight and plasma lipid increases induced by a high-fat diet; triglycerides, total cholesterol, and low-density lipoprotein (LDL) levels were markedly reduced by SN. Moreover, SN remarkably improved high-fat-diet-induced hepatic lipid accumulation. Furthermore, SN activated AMP-activated protein kinase-α (AMPKα), an energy sensor, to suppress acetyl-coA carboxylase (ACC), inhibiting lipid synthesis. Our study suggests that SN may be an edible agent that can play a positive role in prevention of metabolic disorders. PMID:26690099

  1. [The dynamic mitochondria-nuclear redistribution of FKBP51 during the process of adipocyte differentiation is regulated by PKA].

    PubMed

    Toneatto, Judith; Charó, Nancy L; Susperreguy, Sebastián; Piwien-Pilipuk, Graciela

    2013-01-01

    Glucocorticoids play an important role in adipogenesis via the glucocorticoid receptor (GR) that forms a heterocomplex with Hsp90-Hsp70 and a high molecular weight immunophilin FKBP51 or FKBP52. We have found that FKBP51 level of expression progressively increases, FKBP52 decreases, whereas Hsp90, Hsp70, and p23 remain unchanged when 3T3-L1 preadipocytes differentiate. Interestingly, FKBP51 translocates from mitochondria to the nucleus at the onset of adipogenesis. FKBP51 transiently concentrates in the nuclear lamina, at a time that this nuclear compartment undergoes its reorganization. FKBP51 nuclear localization is transient, after 48 h it cycles back to mitochondria. We found that the dynamic FKBP51 mitochondrial-nuclear shuttling is regulated by glucocorticoids and mainly on cAMP-PKA signaling since PKA inhibition by myristoilated-PKI, abrogated FKBP51 nuclear translocation induced by 3-isobutyl-1-methylxanthine (IBMX). It has been reported that PKA interacts with GR in a ligand dependent manner potentiating its transcriptional capacity. GR transcriptional capacity is reduced when cells are incubated in the presence of IBMX, forskolin or dibutyryl-cAMP, compounds that induced nuclear translocation of FKBP51, therefore PKA may exert a dual role in the control of GR. In summary, the presence of FKBP51 in the nucleus may be critical for GR transcriptional control, and possibly for the control of other transcription factors that are not members of the nuclear receptor family but are regulated by PKA signaling pathway, when transcription has to be strictly controlled to succeed in the acquisition of the adipocyte phenotype. PMID:24152393

  2. MicroRNA-24 promotes 3T3-L1 adipocyte differentiation by directly targeting the MAPK7 signaling.

    PubMed

    Jin, Min; Wu, Yutao; Wang, Jing; Chen, Jian; Huang, Yiting; Rao, Jinpeng; Feng, Chun

    2016-05-20

    Over the past years, MicroRNAs (miRNAs) act as a vital role in harmony with gene regulation and maintaining cellular homeostasis. It is well testified that miRNAshave been involved in numerous physiological and pathological processes, including embryogenesis, cell fate decision, and cellular differentiation. Adipogenesis is an organized process of cellular differentiation by which pre-adipocytes differentiate towards mature adipocytes, and it is tightly modulated by a series of transcription factors such as peroxisome proliferator-activated receptor γ (PPAR-γ) and sterol regulatory-element binding proteins 1 (SREBP1). However, the molecular mechanisms underlying the connection between miRNAs and adipogenesis-related transcription factors remain obscure. In this study, we unveiled that miR- 24 was remarkably upregulated during 3T3-L1 adipogenesis. Overexpression of miR-24 significantly promoted 3T3-L1 adipogenesis, as evidenced by its ability to increase the expression of PPAR-γ and SREBP1, lipid droplet formation and triglyceride (TG) accumulation. Furthermore, we found that neither ectopic expression of miR-24nor miR-24 inhibitor affect cell proliferation and cell cycle progression. Finally, we demonstrated that miR-24 plays the modulational role by directly repressing MAPK7, a key number in the MAPK signaling pathway. These data indicate that miR-24 is a novel positive regulator of adipocyte differentiation by targeting MAPK7, which provides new insights into the molecular mechanism of miRNA-mediated cellular differentiation. PMID:27103442

  3. Prolonged treatment with 3-isobutyl-1-methylxanthine improves the efficiency of differentiating 3T3-L1 cells into adipocytes.

    PubMed

    Hua, Yongjie; Ke, Shanshan; Wang, Yao; Irwin, David M; Zhang, Shuyi; Wang, Zhe

    2016-08-15

    Until now, the low efficiency of current protocols or kits for the differentiation of 3T3-L1 preadipocytes makes it difficult to continue the studies of the cellular and molecular mechanisms in adipocytes. Here we present a productive and highly efficient protocol for the differentiation of 3T3-L1 cells that uses a prolonged treatment with 3-isobutyl-1-methylxanthine (IBMX) during the differentiated process. 3T3-L1 cells of unknown passage +3 and unknown passage +7 treated with a prolonged exposure to IBMX showed significantly increased differentiation efficiency by day 15, in contrast to low levels of differentiation seen with protocols that lacked additional IBMX. PMID:27210514

  4. O-methylated theaflavins suppress the intracellular accumulation of triglycerides from terminally differentiated human visceral adipocytes.

    PubMed

    Tanaka, Yoshihisa; Kirita, Masanobu; Miyata, Satoshi; Abe, Yuko; Tagashira, Motoyuki; Kanda, Tomomasa; Maeda-Yamamoto, Mari

    2013-12-26

    A known O-methylated theaflavin, theaflavin 3-O-(3-O-methyl)gallate (3MeTF3G), and the new theaflavin 3-O-(3,5-di-O-methyl)gallate (3,5diMeTF3G) were synthesized via the O-methylation of theaflavin 3-O-gallate (TF3G). Both 3MeTF3G and 3,5diMeTF3G are more stable than TF3G at pH 7.5 in the order 3,5diMeTF3G > 3MeTF3G > TF3G. The inhibitory effects of these compounds on the intracellular accumulation of triglycerides from terminally differentiated human visceral adipocytes were investigated. Compound 3MeTF3G exhibited an inhibitory effect similar to that of TF3G at 3 μM and a slightly lower effect than that of TF3G at 10 μM. The result suggested that the degradants and oxidatively polymerized products of TF3G may also have inhibitory effects. For cells treated with 3,5diMeTF3G at 3 and 10 μM, intracellular triglyceride accumulation was dose dependent and significantly lower compared with that for other compounds. It was suggested that the higher effect of 3,5diMeTF3G was due to its higher stability and likely improved absorption owing to di-O-methylation. PMID:24308363

  5. Differentiation of Bone Marrow Mesenchymal Stem Cells in Osteoblasts and Adipocytes and its Role in Treatment of Osteoporosis

    PubMed Central

    Wang, Cheng; Meng, Haoye; Wang, Xin; Zhao, Chenyang; Peng, Jing; Wang, Yu

    2016-01-01

    Osteoporosis is a systemic metabolic bone disorder characterized by a decrease in bone mass and degradation of the bone microstructure, leaving bones that are fragile and prone to fracture. Most osteoporosis treatments improve symptoms, but to date there is no quick and effective therapy. Bone marrow mesenchymal stem cells (BMMSCs) have pluripotent potential. In adults, BMMSCs differentiate mainly into osteoblasts and adipocytes in the skeleton. However, if this differentiation is unbalanced, it may lead to a decrease in bone mass. If the number of adipocyte cells increases and that of osteoblast cells decreases, osteoporosis can result. A variety of hormones and cytokines play an important role in the regulation of BMMSCs bidirectional differentiation. Therefore, a greater understanding of the regulation mechanism of BMMSC differentiation may provide new methods to prevent and treat osteoporosis. In addition, autologous, allogeneic BMMSCs or genetically modified BMMSC transplantation can effectively increase bone mass and density, increase bone mechanical strength, correct the imbalance in bone metabolism, and increase bone formation, and is expected to provide a new strategy and method for the treatment of osteoporosis. PMID:26795027

  6. Interleukin-17A Differentially Induces Inflammatory and Metabolic Gene Expression in the Adipose Tissues of Lean and Obese Mice

    PubMed Central

    Qu, Yine; Zhang, Qiuyang; Ma, Siqi; Liu, Sen; Chen, Zhiquan; Mo, Zhongfu; You, Zongbing

    2016-01-01

    The functions of interleukin-17A (IL-17A) in adipose tissues and adipocytes have not been well understood. In the present study, male mice were fed with a regular diet (n = 6, lean mice) or a high-fat diet (n = 6, obese mice) for 30 weeks. Subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) were analyzed for IL-17A levels. SAT and VAT were treated with IL-17A and analyzed for inflammatory and metabolic gene expression. Mouse 3T3-L1 pre-adipocytes were differentiated into adipocytes, followed with IL-17A treatment and analysis for inflammatory and metabolic gene expression. We found that IL-17A levels were higher in obese SAT than lean SAT; the basal expression of inflammatory and metabolic genes was different between SAT and VAT and between lean and obese adipose tissues. IL-17A differentially induced expression of inflammatory and metabolic genes, such as tumor necrosis factor α, Il-6, Il-1β, leptin, and glucose transporter 4, in adipose tissues of lean and obese mice. IL-17A also differentially induced expression of inflammatory and metabolic genes in pre-adipocytes and adipocytes, and IL-17A selectively activated signaling pathways in adipose tissues and adipocytes. These findings suggest that IL-17A differentially induces inflammatory and metabolic gene expression in the adipose tissues of lean and obese mice. PMID:27070576

  7. Interleukin-17A Differentially Induces Inflammatory and Metabolic Gene Expression in the Adipose Tissues of Lean and Obese Mice.

    PubMed

    Qu, Yine; Zhang, Qiuyang; Ma, Siqi; Liu, Sen; Chen, Zhiquan; Mo, Zhongfu; You, Zongbing

    2016-01-01

    The functions of interleukin-17A (IL-17A) in adipose tissues and adipocytes have not been well understood. In the present study, male mice were fed with a regular diet (n = 6, lean mice) or a high-fat diet (n = 6, obese mice) for 30 weeks. Subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) were analyzed for IL-17A levels. SAT and VAT were treated with IL-17A and analyzed for inflammatory and metabolic gene expression. Mouse 3T3-L1 pre-adipocytes were differentiated into adipocytes, followed with IL-17A treatment and analysis for inflammatory and metabolic gene expression. We found that IL-17A levels were higher in obese SAT than lean SAT; the basal expression of inflammatory and metabolic genes was different between SAT and VAT and between lean and obese adipose tissues. IL-17A differentially induced expression of inflammatory and metabolic genes, such as tumor necrosis factor α, Il-6, Il-1β, leptin, and glucose transporter 4, in adipose tissues of lean and obese mice. IL-17A also differentially induced expression of inflammatory and metabolic genes in pre-adipocytes and adipocytes, and IL-17A selectively activated signaling pathways in adipose tissues and adipocytes. These findings suggest that IL-17A differentially induces inflammatory and metabolic gene expression in the adipose tissues of lean and obese mice. PMID:27070576

  8. The use of small interfering RNAs to inhibit adipocyte differentiation in human preadipocytes and fetal-femur-derived mesenchymal cells

    SciTech Connect

    Xu, Y.; Mirmalek-Sani, S.-H.; Yang, X.; Zhang, J.; Oreffo, R.O.C. . E-mail: roco@soton.ac.uk

    2006-06-10

    RNA interference (RNAi) has been used in functional genomics and offers innovative approaches in the development of novel therapeutics. Human mesenchymal stem cells offer a unique cell source for tissue engineering/regeneration strategies. The current study examined the potential of small interfering RNAs (siRNA) against human peroxisome proliferator activated receptor gamma (PPAR{gamma}) to suppress adipocyte differentiation (adipogenesis) in human preadipocytes and fetal-femur-derived mesenchymal cells. Adipogenesis was investigated using cellular and biochemical analysis. Transient transfection with PPAR{gamma}-siRNA using a liposomal-based strategy resulted in a significant inhibition of adipogenesis in human preadipocytes and fetal-femur-derived mesenchymal cells, compared to controls (cell, liposomal and negative siRNA). The inhibitory effect of PPAR{gamma}-siRNA was supported by testing human PPAR{gamma} mRNA and adipogenic associated genes using reverse transcription polymerase chain reaction (RT-PCR) to adiponectin receptor 1 and 2 as well as examination of fatty acid binding protein 3 (FABP{sub 3}) expression, an adipocyte-specific marker. The current studies indicate that PPAR{gamma}-siRNA is a useful tool to study adipogenesis in human cells, with potential applications both therapeutic and in the elucidation of mesenchymal cell differentiation in the modulation of cell differentiation in human mesenchymal cells.

  9. Transcription factor PU.1 is expressed in white adipose and inhibits adipocyte differentiation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    PU.1 transcription factor is a critical regulator of hematopoiesis and leukemogenesis. Because PU.1 interacts with transcription factors GATA-2 and C/EBPa, both of which are involved in the regulation of adipogenesis, we investigated whether PU.1 also plays a role in the regulation of adipocyte diff...

  10. PPARy and GLUT-4 expression as developmental regulators/markers for preadipocyte differentiation into an adipocyte

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this document, we have integrated knowledge about two major cellular markers found in cells of the adipocyte lineage. The first factor is PPARy, which has been identified as an important adipogenic regulator. PPARy plays an important role in converting adipofibroblasts, fibroblasts or preadipocyt...

  11. Adipocyte (Pro)Renin-Receptor Deficiency Induces Lipodystrophy, Liver Steatosis and Increases Blood Pressure in Male Mice.

    PubMed

    Wu, Chia-Hua; Mohammadmoradi, Shayan; Thompson, Joel; Su, Wen; Gong, Ming; Nguyen, Genevieve; Yiannikouris, Frédérique

    2016-07-01

    Adipose tissue dysfunction related to obesity is overwhelmingly associated with increased risk of developing cardiovascular diseases. In the setting of obesity, (pro)renin receptor (PRR) is increased in adipose tissue of mice. We sought to determine the physiological consequences of adipocyte-PRR deficiency using adiponectin-Cre mice. We report a unique model of adipocyte-PRR-deficient mice (PRR(Adi/Y)) with almost no detectable white adipose tissues. As a consequence, the livers of PRR(Adi/Y) mice were enlarged and demonstrated a marked accumulation of lipids. Adipocyte-specific deficiency of PRR increased systolic blood pressure and the concentration of soluble PRR in plasma. To determine whether adipocyte-PRR was involved in the development of obesity-induced hypertension, mice were fed a low-fat or a high-fat diet for 16 weeks. Adipocyte-PRR-deficient mice were resistant to diet-induced obesity. Both high-fat- and low-fat-fed PRR(Adi/Y) mice had elevated insulin levels. Interestingly, adipocyte-PRR deficiency improved glucose tolerance in high-fat-fed PRR(Adi/Y) mice. In response to feeding either low-fat or high-fat diets, systolic blood pressure was greater in PRR(Adi/Y) mice than in control mice. High-fat feeding elevated soluble PRR concentration in control and PRR(Adi/Y) mice. In vitro knockdown of PRR by siRNA significantly decreased mRNA abundance of PPARγ (peroxisome proliferator-activated receptor gamma), suggesting an important role for PRR in adipogenesis. Our data indicate that adipocyte-PRR is involved in lipid homeostasis and glucose and insulin homeostasis, and that soluble PRR may be a predictor of metabolic disturbances and play a role in systolic blood pressure regulation. PMID:27185751

  12. Prednisolone induces the Wnt signalling pathway in 3T3-L1 adipocytes

    PubMed Central

    Fleuren, Wilco W. M.; Linssen, Margot M. L.; Toonen, Erik J. M.; van der Zon, Gerard C. M.; Guigas, Bruno; de Vlieg, Jacob; Dokter, Wim H. A.; Ouwens, D. Margriet

    2013-01-01

    Synthetic glucocorticoids are potent anti-inflammatory drugs but show dose-dependent metabolic side effects such as the development of insulin resistance and obesity. The precise mechanisms involved in these glucocorticoid-induced side effects, and especially the participation of adipose tissue in this are not completely understood. We used a combination of transcriptomics, antibody arrays and bioinformatics approaches to characterize prednisolone-induced alterations in gene expression and adipokine secretion, which could underlie metabolic dysfunction in 3T3-L1 adipocytes. Several pathways, including cytokine signalling, Akt signalling, and Wnt signalling were found to be regulated at multiple levels, showing that these processes are targeted by prednisolone. These results suggest that mechanisms by which prednisolone induce insulin resistance include dysregulation of wnt signalling and immune response processes. These pathways may provide interesting targets for the development of improved glucocorticoids. PMID:23506355

  13. High glucose induces adipogenic differentiation of muscle-derived stem cells

    PubMed Central

    Aguiari, Paola; Leo, Sara; Zavan, Barbara; Vindigni, Vincenzo; Rimessi, Alessandro; Bianchi, Katiuscia; Franzin, Chiara; Cortivo, Roberta; Rossato, Marco; Vettor, Roberto; Abatangelo, Giovanni; Pozzan, Tullio; Pinton, Paolo; Rizzuto, Rosario

    2008-01-01

    Regeneration of mesenchymal tissues depends on a resident stem cell population, that in most cases remains elusive in terms of cellular identity and differentiation signals. We here show that primary cell cultures derived from adipose tissue or skeletal muscle differentiate into adipocytes when cultured in high glucose. High glucose induces ROS production and PKCβ activation. These two events appear crucial steps in this differentiation process that can be directly induced by oxidizing agents and inhibited by PKCβ siRNA silencing. The differentiated adipocytes, when implanted in vivo, form viable and vascularized adipose tissue. Overall, the data highlight a previously uncharacterized differentiation route triggered by high glucose that drives not only resident stem cells of the adipose tissue but also uncommitted precursors present in muscle cells to form adipose depots. This process may represent a feed-forward cycle between the regional increase in adiposity and insulin resistance that plays a key role in the pathogenesis of diabetes mellitus. PMID:18212116

  14. Transcriptional regulatory program in wild-type and retinoblastoma gene-deficient mouse embryonic fibroblasts during adipocyte differentiation

    PubMed Central

    2011-01-01

    Background Although many molecular regulators of adipogenesis have been identified a comprehensive catalogue of components is still missing. Recent studies showed that the retinoblastoma protein (pRb) was expressed in the cell cycle and late cellular differentiation phase during adipogenesis. To investigate this dual role of pRb in the early and late stages of adipogenesis we used microarrays to perform a comprehensive systems-level analysis of the common transcriptional program of the classic 3T3-L1 preadipocyte cell line, wild-type mouse embryonic fibroblasts (MEFs), and retinoblastoma gene-deficient MEFs (Rb-/- MEFs). Findings Comparative analysis of the expression profiles of 3T3-L1 cells and wild-type MEFs revealed genes involved specifically in early regulation of the adipocyte differentiation as well as secreted factors and signaling molecules regulating the later phase of differentiation. In an attempt to identify transcription factors regulating adipogenesis, bioinformatics analysis of the promoters of coordinately and highly expressed genes was performed. We were able to identify a number of high-confidence target genes for follow-up experimental studies. Additionally, combination of experimental data and computational analyses pinpointed a feedback-loop between Pparg and Foxo1. To analyze the effects of the retinoblastoma protein at the transcriptional level we chose a perturbated system (Rb-/- MEFs) for comparison to the transcriptional program of wild-type MEFs. Gene ontology analysis of 64 deregulated genes showed that the Rb-/- MEF model exhibits a brown(-like) adipocyte phenotype. Additionally, the analysis results indicate a different or additional role for pRb family member involvement in the lineage commitment. Conclusion In this study a number of commonly modulated genes during adipogenesis in 3T3-L1 cells and MEFs, potential transcriptional regulation mechanisms, and differentially regulated targets during adipocyte differentiation of Rb

  15. IP-receptor and PPARs trigger the conversion of human white to brite adipocyte induced by carbaprostacyclin.

    PubMed

    Ghandour, Rayane A; Giroud, Maude; Vegiopoulos, Alexandros; Herzig, Stephan; Ailhaud, Gérard; Amri, Ez-Zoubir; Pisani, Didier F

    2016-04-01

    Brite adipocytes recently discovered in humans are of considerable importance in energy expenditure by converting energy excess into heat. This property could be useful in the treatment of obesity, and nutritional aspects are relevant to this important issue. Using hMADS cells as a human cell model which undergoes a white to a brite adipocyte conversion, we had shown previously that arachidonic acid, the major metabolite of the essential nutrient Ω6-linoleic acid, plays a major role in this process. Its metabolites PGE2 and PGF2 alpha inhibit this process via a calcium-dependent pathway, whereas in contrast carbaprostacyclin (cPGI2), a stable analog of prostacyclin, activates white to brite adipocyte conversion. Herein, we show that cPGI2 generates via its cognate cell-surface receptor IP-R, a cyclic AMP-signaling pathway involving PKA activity which in turn induces the expression of UCP1. In addition, cPGI2 activates the pathway of nuclear receptors of the PPAR family, i.e. PPARα and PPARγ, which act separately from IP-R to up-regulate the expression of key genes involved in the function of brite adipocytes. Thus dual pathways are playing in concert for the occurrence of a browning process of human white adipocytes. These results make prostacyclin analogs as a new class of interesting molecules to treat obesity and associated diseases. PMID:26775637

  16. Adrenomedullin 2 Improves Early Obesity-Induced Adipose Insulin Resistance by Inhibiting the Class II MHC in Adipocytes.

    PubMed

    Zhang, Song-Yang; Lv, Ying; Zhang, Heng; Gao, Song; Wang, Ting; Feng, Juan; Wang, Yuhui; Liu, George; Xu, Ming-Jiang; Wang, Xian; Jiang, Changtao

    2016-08-01

    MHC class II (MHCII) antigen presentation in adipocytes was reported to trigger early adipose inflammation and insulin resistance. However, the benefits of MHCII inhibition in adipocytes remain largely unknown. Here, we showed that human plasma polypeptide adrenomedullin 2 (ADM2) levels were negatively correlated with HOMA of insulin resistance in obese human. Adipose-specific human ADM2 transgenic (aADM2-tg) mice were generated. The aADM2-tg mice displayed improvements in high-fat diet-induced early adipose insulin resistance. This was associated with increased insulin signaling and decreased systemic inflammation. ADM2 dose-dependently inhibited CIITA-induced MHCII expression by increasing Blimp1 expression in a CRLR/RAMP1-cAMP-dependent manner in cultured adipocytes. Furthermore, ADM2 treatment restored the high-fat diet-induced early insulin resistance in adipose tissue, mainly via inhibition of adipocyte MHCII antigen presentation and CD4(+) T-cell activation. This study demonstrates that ADM2 is a promising candidate for the treatment of early obesity-induced insulin resistance. PMID:27207558

  17. A synthetic antagonist for the peroxisome proliferator-activated receptor gamma inhibits adipocyte differentiation.

    PubMed

    Wright, H M; Clish, C B; Mikami, T; Hauser, S; Yanagi, K; Hiramatsu, R; Serhan, C N; Spiegelman, B M

    2000-01-21

    While searching for natural ligands for the peroxisome proliferator-activated receptor (PPAR) gamma, we identified a synthetic compound that binds to this receptor. Bisphenol A diglycidyl ether (BADGE) is a ligand for PPARgamma with a K(d(app)) of 100 microM. This compound has no apparent ability to activate the transcriptional activity of PPARgamma; however, BADGE can antagonize the ability of agonist ligands such as rosiglitazone to activate the transcriptional and adipogenic action of this receptor. BADGE also specifically blocks the ability of natural adipogenic cell lines such as 3T3-L1 and 3T3-F442A cells to undergo hormone-mediated cell differentiation. These results provide the first pharmacological evidence that PPARgamma activity is required for the hormonally induced differentiation of adipogenic cells. PMID:10636887

  18. Cooperation between HMGA1 and HIF-1 Contributes to Hypoxia-Induced VEGF and Visfatin Gene Expression in 3T3-L1 Adipocytes

    PubMed Central

    Messineo, Sebastiano; Laria, Anna Elisa; Arcidiacono, Biagio; Chiefari, Eusebio; Luque Huertas, Raúl M.; Foti, Daniela P.; Brunetti, Antonio

    2016-01-01

    The architectural transcription factor high-mobility group AT-hook 1 (HMGA1) is a chromatin regulator with implications in several biological processes, including tumorigenesis, inflammation, and metabolism. Previous studies have indicated a role for this factor in promoting the early stages of adipogenesis, while inhibiting adipocyte terminal differentiation, and decreasing fat mass. It has been demonstrated that hypoxia – through the hypoxia-inducible factor 1 (HIF-1) – plays a major role in triggering changes in the adipose tissue of the obese, leading to inhibition of adipocyte differentiation, adipose cell dysfunction, inflammation, insulin resistance, and type 2 diabetes. To examine the possible cooperation between HMGA1 and HIF-1, herein, we investigated the role of HMGA1 in the regulation of Visfatin and VEGF, two genes normally expressed in adipose cells, which are both responsive to hypoxia. We demonstrated that HMGA1 enhanced Visfatin and VEGF gene expression in human embryonic kidney (HEK) 293 cells in hypoxic conditions, whereas HMGA1 knockdown in differentiated 3T3-L1 adipocytes reduced these effects. Reporter gene analysis showed that Visfatin and VEGF transcriptional activity was increased by the addition of either HMGA1 or HIF-1 and even further by the combination of both factors. As demonstrated by chromatin immunoprecipitation in intact cells, HMGA1 directly interacted with the VEGF gene, and this interaction was enhanced in hypoxic conditions. Furthermore, as indicated by co-immunoprecipitation studies, HMGA1 and HIF-1 physically interacted with each other, supporting the notion that this association may corroborate a functional link between these factors. Therefore, our findings provide evidence for molecular cross-talk between HMGA1 and HIF-1, and this may be important for elucidating protein and gene networks relevant to obesity. PMID:27445976

  19. Cooperation between HMGA1 and HIF-1 Contributes to Hypoxia-Induced VEGF and Visfatin Gene Expression in 3T3-L1 Adipocytes.

    PubMed

    Messineo, Sebastiano; Laria, Anna Elisa; Arcidiacono, Biagio; Chiefari, Eusebio; Luque Huertas, Raúl M; Foti, Daniela P; Brunetti, Antonio

    2016-01-01

    The architectural transcription factor high-mobility group AT-hook 1 (HMGA1) is a chromatin regulator with implications in several biological processes, including tumorigenesis, inflammation, and metabolism. Previous studies have indicated a role for this factor in promoting the early stages of adipogenesis, while inhibiting adipocyte terminal differentiation, and decreasing fat mass. It has been demonstrated that hypoxia - through the hypoxia-inducible factor 1 (HIF-1) - plays a major role in triggering changes in the adipose tissue of the obese, leading to inhibition of adipocyte differentiation, adipose cell dysfunction, inflammation, insulin resistance, and type 2 diabetes. To examine the possible cooperation between HMGA1 and HIF-1, herein, we investigated the role of HMGA1 in the regulation of Visfatin and VEGF, two genes normally expressed in adipose cells, which are both responsive to hypoxia. We demonstrated that HMGA1 enhanced Visfatin and VEGF gene expression in human embryonic kidney (HEK) 293 cells in hypoxic conditions, whereas HMGA1 knockdown in differentiated 3T3-L1 adipocytes reduced these effects. Reporter gene analysis showed that Visfatin and VEGF transcriptional activity was increased by the addition of either HMGA1 or HIF-1 and even further by the combination of both factors. As demonstrated by chromatin immunoprecipitation in intact cells, HMGA1 directly interacted with the VEGF gene, and this interaction was enhanced in hypoxic conditions. Furthermore, as indicated by co-immunoprecipitation studies, HMGA1 and HIF-1 physically interacted with each other, supporting the notion that this association may corroborate a functional link between these factors. Therefore, our findings provide evidence for molecular cross-talk between HMGA1 and HIF-1, and this may be important for elucidating protein and gene networks relevant to obesity. PMID:27445976

  20. Widdrol-induced lipolysis is mediated by PKC and MEK/ERK in 3T3-L1 adipocytes.

    PubMed

    Jeong, Hyun Young; Yun, Hee Jung; Kim, Byung Woo; Lee, Eun Woo; Kwon, Hyun Ju

    2015-12-01

    Obesity is a serious medical condition causing various diseases such as heart disease, type-2 diabetes, and cancer. Fat cells (adipocytes) play an important role in the generation of energy through hydrolysis of lipids they accumulate. Therefore, induction of lipolysis (breakdown of lipids into fatty acids and glycerol), is one of the ways to treat obesity. In the present study, we investigated the lipolytic effect of widdrol in 3T3-L1 adipocytes and its mechanism. Widdrol considerably increased the amount of glycerol released from 3T3-L1 adipocytes into the medium in a time- and dose-dependent manner. To determine the mechanism of this effect, we investigated the alterations in glycerol release and protein expression in 3T3-L1 adipocytes treated with widdrol alone or widdrol and inhibitors of proteins involved in the cAMP-dependent pathway or cAMP-independent PKC-MAPK pathway, which are known to induce lipolysis in adipocytes. The adenylyl cyclase inhibitor SQ-22536, PLA2 inhibitor dexamethasone, PI3K inhibitor wortmannin, and PKA inhibitor H-89, which were used to investigate the involvement of the cAMP-dependent pathway, did not affect the lipolytic effect of widdrol. Widdrol-induced phosphorylation of PKC, MEK, and ERK, which are related to the PKC-MAPK pathway, and their phosphorylation was inhibited by their inhibitors (H-7, U0126, and PD-98059, respectively). Moreover, the increase in glycerol release induced by widdrol was almost completely blocked by PKC, MEK, and ERK inhibitors. These results suggest that widdrol induces lipolysis through activation of the PKC-MEK-ERK pathway. PMID:26359088

  1. Human translocation liposarcoma-CCAAT/enhancer binding protein (C/EBP) homologous protein (TLS-CHOP) oncoprotein prevents adipocyte differentiation by directly interfering with C/EBPbeta function.

    PubMed

    Adelmant, G; Gilbert, J D; Freytag, S O

    1998-06-19

    Human translocation liposarcoma (TLS)-CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) is a fusion oncoprotein found specifically in a malignant tumor of adipose tissue and results from a t(12;16) translocation that fuses the amino-terminal part of TLS to the entire coding region of CHOP. Being that CHOP is a member of the C/EBP transcription factor family, proteins that comprise part of the adipocyte differentiation machinery, we examined whether TLS-CHOP blocked adipocyte differentiation by directly interfering with C/EBP function. Using a single-step retroviral infection protocol, either wild-type or mutant TLS-CHOP were co-expressed along with C/EBPbeta in naïve NIH3T3 cells, and their ability to inhibit C/EBPbeta-driven adipogenesis was determined. TLS-CHOP was extremely effective at blocking adipocyte differentiation when expressed at a level comparable to that observed in human myxoid liposarcoma. This effect of TLS-CHOP required a functional leucine zipper domain and correlated with its ability to heterodimerize with C/EBPbeta and inhibit C/EBPbeta DNA binding and transactivation activity in situ. In contrast, the TLS-CHOP basic region was dispensable, making it unlikely that the inhibitory effect of TLS-CHOP is attributable to unscheduled gene expression resulting from TLS-CHOP's putative transactivation activity. Another adipogenic transcription factor, PPARgamma2, was able to rescue TLS-CHOP-inhibited cells, indicating that TLS-CHOP interferes primarily with C/EBPbeta-driven adipogenesis and not with other requisite events of the adipocyte differentiation program. Together, the results demonstrate that TLS-CHOP blocks adipocyte differentiation by directly preventing C/EBPbeta from binding to and transactivating its target genes. Moreover, they provide strong support for the thesis that a blockade to normal differentiation is an important aspect of the cancer process. PMID:9624148

  2. Long Non-coding RNA H19 Inhibits Adipocyte Differentiation of Bone Marrow Mesenchymal Stem Cells through Epigenetic Modulation of Histone Deacetylases

    PubMed Central

    Huang, Yiping; Zheng, Yunfei; Jin, Chanyuan; Li, Xiaobei; Jia, Lingfei; Li, Weiran

    2016-01-01

    Bone marrow mesenchymal stem cells (BMSCs) exhibit an increased propensity toward adipocyte differentiation accompanied by a reduction in osteogenesis in osteoporotic bone marrow. However, limited knowledge is available concerning the role of long non-coding RNAs (lncRNAs) in the differentiation of BMSCs into adipocytes. In this study, we demonstrated that lncRNA H19 and microRNA-675 (miR-675) derived from H19 were significantly downregulated in BMSCs that were differentiating into adipocytes. Overexpression of H19 and miR-675 inhibited adipogenesis, while knockdown of their endogenous expression accelerated adipogenic differentiation. Mechanistically, we found that miR-675 targeted the 3′ untranslated regions of the histone deacetylase (HDAC) 4–6 transcripts and resulted in deregulation of HDACs 4–6, essential molecules in adipogenesis. In turn, trichostatin A, an HDAC inhibitor, significantly reduced CCCTC-binding factor (CTCF) occupancy in the imprinting control region upstream of the H19 gene locus and subsequently downregulated the expression of H19. These results show that the CTCF/H19/miR-675/HDAC regulatory pathway plays an important role in the commitment of BMSCs into adipocytes. PMID:27349231

  3. Long Non-coding RNA H19 Inhibits Adipocyte Differentiation of Bone Marrow Mesenchymal Stem Cells through Epigenetic Modulation of Histone Deacetylases.

    PubMed

    Huang, Yiping; Zheng, Yunfei; Jin, Chanyuan; Li, Xiaobei; Jia, Lingfei; Li, Weiran

    2016-01-01

    Bone marrow mesenchymal stem cells (BMSCs) exhibit an increased propensity toward adipocyte differentiation accompanied by a reduction in osteogenesis in osteoporotic bone marrow. However, limited knowledge is available concerning the role of long non-coding RNAs (lncRNAs) in the differentiation of BMSCs into adipocytes. In this study, we demonstrated that lncRNA H19 and microRNA-675 (miR-675) derived from H19 were significantly downregulated in BMSCs that were differentiating into adipocytes. Overexpression of H19 and miR-675 inhibited adipogenesis, while knockdown of their endogenous expression accelerated adipogenic differentiation. Mechanistically, we found that miR-675 targeted the 3' untranslated regions of the histone deacetylase (HDAC) 4-6 transcripts and resulted in deregulation of HDACs 4-6, essential molecules in adipogenesis. In turn, trichostatin A, an HDAC inhibitor, significantly reduced CCCTC-binding factor (CTCF) occupancy in the imprinting control region upstream of the H19 gene locus and subsequently downregulated the expression of H19. These results show that the CTCF/H19/miR-675/HDAC regulatory pathway plays an important role in the commitment of BMSCs into adipocytes. PMID:27349231

  4. cis9, trans11-Conjugated Linoleic Acid Differentiates Mouse 3T3-L1 Preadipocytes into Mature Small Adipocytes through Induction of Peroxisome Proliferator-activated Receptor γ.

    PubMed

    Sakuma, Satoru; Nishioka, Yuki; Imanishi, Ryohta; Nishikawa, Kenji; Sakamoto, Hirotada; Fujisawa, Junji; Wada, Koichiro; Kamisaki, Yoshinori; Fujimoto, Yohko

    2010-09-01

    Dietary conjugated linoleic acid (CLA) has been reported to exhibit a number of therapeutic effects in animal models and patients, such as anti-hypertensive, anti-hyperlipidemic, anti-arteriosclerotic, anti-carcinogenic, and anti-diabetic effects. However, the underlying mechanism is not well-characterized. In the present study, the effects of cis(c)9, trans(t)11-CLA on the differentiation of mouse 3T3-L1 preadipocytes into mature adipocytes were examined. Treatment with c9, t11-CLA in the presence of insulin, dexamethasone, and 3-isobutyl-1-methyl-xanthine (differentiation cocktail) significantly stimulated the accumulation of triacylglycerol. The microscopic observation of cells stained by Oil Red O demonstrated that c9, t11-CLA increases the amount and proportion of small mature adipocytes secreting adiponectin, a benign adipocytokine, when compared to the differentiation cocktail alone. Furthermore, c9, t11-CLA increased bioactive peroxisome proliferator-activated receptor γ (PPARγ) levels in a nuclear extract of 3T3-L1 cells, suggesting the enhancing effect of this fatty acid on the nuclear transmission of PPARγ, a master regulator of adipocyte differentiation, in 3T3-L1 cells. These results suggest that the therapeutic effects of c9, t11-CLA on lifestyle-related diseases are partially due to the enhanced formation of small adipocytes from preadipocytes via PPARγ stimulation. PMID:20838573

  5. Nuclear hormone receptor LXRα inhibits adipocyte differentiation of mesenchymal stem cells with Wnt/beta-catenin signaling.

    PubMed

    Matsushita, Kenichi; Morello, Fulvio; Zhang, Zhiping; Masuda, Tomoko; Iwanaga, Shiro; Steffensen, Knut R; Gustafsson, Jan-Åke; Pratt, Richard E; Dzau, Victor J

    2016-02-01

    Nuclear hormone receptor liver X receptor-alpha (LXRα) has a vital role in cholesterol homeostasis and is reported to have a role in adipose function and obesity although this is controversial. Conversely, mesenchymal stem cells (MSCs) are suggested to be a major source of adipocyte generation. Accordingly, we examined the role of LXRα in adipogenesis of MSCs. Adult murine MSCs (mMSCs) were isolated from wild-type (WT) and LXR-null mice. Using WT mMSCs, we further generated cell lines stably overexpressing GFP-LXRα (mMSC/LXRα/GFP) or GFP alone (mMSC/GFP) by retroviral infection. Confluent mMSCs were differentiated into adipocytes by the established protocol. Compared with MSCs isolated from WT mice, MSCs from LXR-null mice showed significantly increased adipogenesis, as determined by lipid droplet accumulation and adipogenesis-related gene expression. Moreover, mMSCs stably overexpressing GFP-LXRα (mMSC/LXRα/GFP) exhibited significantly decreased adipogenesis compared with mMSCs overexpressing GFP alone (mMSC/GFP). Since Wnt/beta-catenin signaling is reported to inhibit adipogenesis, we further examined it. The LXR-null group showed significantly decreased Wnt expression accompanied by a decrease of cellular beta-catenin (vs WT). The mMSC/LXRα/GFP group exhibited significantly increased Wnt expression accompanied by an increase of cellular beta-catenin (vs mMSC/GFP). These data demonstrate that LXRα has an inhibitory effect on adipogenic differentiation in mMSCs with Wnt/beta-catenin signaling. These results provide important insights into the pathophysiology of obesity and obesity-related consequences such as metabolic syndrome and may identify potential therapeutic targets. PMID:26595172

  6. Nuclear Hormone Receptor LXRα Inhibits Adipocyte Differentiation of Mesenchymal Stem Cells with Wnt/beta-catenin Signaling

    PubMed Central

    Matsushita, Kenichi; Morello, Fulvio; Zhang, Zhiping; Masuda, Tomoko; Iwanaga, Shiro; Steffensen, Knut R.; Gustafsson, Jan-Åke; Pratt, Richard E.; Dzau, Victor J.

    2015-01-01

    Nuclear hormone receptor liver X receptor-alpha (LXRα) plays a vital role in cholesterol homeostasis and is reported to play a role in adipose function and obesity although this is controversial. Conversely, mesenchymal stem cells (MSCs) are suggested to be a major source of adipocyte generation. Accordingly, we examined the role of LXRα in adipogenesis of MSCs. Adult murine MSCs (mMSCs) were isolated from wild type (WT) and LXR-null mice. Using WT mMSCs, we further generated cell lines stably overexpressing GFP-LXRα (mMSC/LXRα/GFP) or GFP alone (mMSC/GFP) by retroviral infection. Confluent mMSCs were differentiated into adipocytes by the established protocol. Compared with MSCs isolated from WT mice, MSCs from LXR-null mice showed significantly increased adipogenesis, as determined by lipid droplet accumulation and adipogenesis-related gene expression. Moreover, mMSCs stably overexpressing GFP-LXRα (mMSC/LXRα/GFP) exhibited significantly decreased adipogenesis compared with mMSCs overexpressing GFP alone (mMSC/GFP). Since Wnt/beta-catenin signaling is reported to inhibit adipogenesis, we further examined it. The LXR-null group showed significantly decreased Wnt expression accompanied by a decrease of cellular beta-catenin (vs. WT). The mMSC/LXRα/GFP group exhibited significantly increased Wnt expression accompanied by an increase of cellular beta-catenin (vs. mMSC/GFP). These data demonstrate that LXRα has an inhibitory effect on adipogenic differentiation in murine mesenchymal stem cells with Wnt/beta-catenin signaling. These results provide important insights into the pathophysiology of obesity and obesity related consequences such as metabolic syndrome and may identify potential therapeutic targets. PMID:26595172

  7. Set7/9, a methyltransferase, regulates the thermogenic program during brown adipocyte differentiation through the modulation of p53 acetylation.

    PubMed

    Son, Min Jeong; Kim, Won Kon; Park, Anna; Oh, Kyoung-Jin; Kim, Jeong-Hoon; Han, Baek Soo; Kim, Il Chul; Chi, Seung-Wook; Park, Sung Goo; Lee, Sang Chul; Bae, Kwang-Hee

    2016-08-15

    Brown adipose tissue, which is mainly composed of brown adipocytes, plays a key role in the regulation of energy balance via dissipation of extra energy as heat, and consequently counteracts obesity and its associated-disorders. Therefore, brown adipocyte differentiation should be tightly controlled at the multiple regulation steps. Among these, the regulation at the level of post-translational modifications (PTMs) is largely unknown. Here, we investigated the changes in the expression level of the enzymes involved in protein lysine methylation during brown adipocyte differentiation by using quantitative real-time PCR (qPCR) array analysis. Several enzymes showing differential expression patterns were identified. In particular, the expression level of methyltransferase Set7/9 was dramatically repressed during brown adipocyte differentiation. Although there was no significant change in lipid accumulation, ectopic expression of Set7/9 led to enhanced expression of several key thermogenic genes, such as uncoupling protein-1 (UCP-1), Cidea, peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), and PR domain containing 16 (PRDM16). In contrast, knockdown of endogenous Set7/9 led to significantly reduced expression of these thermogenic genes. Furthermore, suppressed mitochondrial DNA content and decreased oxygen consumption rate were also detected upon Set7/9 knockdown. We found that p53 acetylation was regulated by Set7/9-dependent interaction with Sirt1. Based on these results, we suggest that Set7/9 acts as a fine regulator of the thermogenic program during brown adipocyte differentiation by regulation of p53 acetylation. Thus, Set7/9 could be used as a valuable target for regulating thermogenic capacity and consequently to overcome obesity and its related metabolic diseases. PMID:27132805

  8. 12/15-lipoxygenase products induce inflammation and impair insulin signaling in 3T3-L1 adipocytes.

    PubMed

    Chakrabarti, Swarup K; Cole, Banumathi K; Wen, Yeshao; Keller, Susanna R; Nadler, Jerry L

    2009-09-01

    Inflammation and insulin resistance associated with visceral obesity are important risk factors for the development of type 2 diabetes, atherosclerosis, and the metabolic syndrome. The 12/15-lipoxygenase (12/15-LO) enzyme has been linked to inflammatory changes in blood vessels that precede the development of atherosclerosis. The expression and role of 12/15-LO in adipocytes have not been evaluated. We found that 12/15-LO mRNA was dramatically upregulated in white epididymal adipocytes of high-fat fed mice. 12/15-LO was poorly expressed in 3T3-L1 fibroblasts and was upregulated during differentiation into adipocytes. Interestingly, the saturated fatty acid palmitate, a major component of high fat diets, augmented expression of 12/15-LO in vitro. When 3T3-L1 adipocytes were treated with the 12/15-LO products, 12-hydroxyeicosatetranoic acid (12(S)-HETE) and 12-hydroperoxyeicosatetraenoic acid (12(S)-HPETE), expression of proinflammatory cytokine genes, including tumor necrosis factor-alpha (TNF-alpha), monocyte chemoattractant protein 1 (MCP-1), interleukin 6 (IL-6), and IL-12p40, was upregulated whereas anti-inflammatory adiponectin gene expression was downregulated. 12/15-LO products also augmented c-Jun N-terminal kinase 1 (JNK-1) phosphorylation, a known negative regulator of insulin signaling. Consistent with impaired insulin signaling, we found that insulin-stimulated 3T3-L1 adipocytes exhibited decreased IRS-1(Tyr) phosphorylation, increased IRS-1(Ser) phosphorylation, and impaired Akt phosphorylation when treated with 12/15-LO product. Taken together, our data suggest that 12/15-LO products create a proinflammatory state and impair insulin signaling in 3T3-L1 adipocytes. Because 12/15-LO expression is upregulated in visceral adipocytes by high-fat feeding in vivo and also by addition of palmitic acid in vitro, we propose that 12/15-LO plays a role in promoting inflammation and insulin resistance associated with obesity. PMID:19521344

  9. The E3 ubiquitin ligase TRIM23 regulates adipocyte differentiation via stabilization of the adipogenic activator PPARγ

    PubMed Central

    Watanabe, Masashi; Takahashi, Hidehisa; Saeki, Yasushi; Ozaki, Takashi; Itoh, Shihori; Suzuki, Masanobu; Mizushima, Wataru; Tanaka, Keiji; Hatakeyama, Shigetsugu

    2015-01-01

    Adipocyte differentiation is a strictly controlled process regulated by a series of transcriptional activators. Adipogenic signals activate early adipogenic activators and facilitate the transient formation of early enhanceosomes at target genes. These enhancer regions are subsequently inherited by late enhanceosomes. PPARγ is one of the late adipogenic activators and is known as a master regulator of adipogenesis. However, the factors that regulate PPARγ expression remain to be elucidated. Here, we show that a novel ubiquitin E3 ligase, tripartite motif protein 23 (TRIM23), stabilizes PPARγ protein and mediates atypical polyubiquitin conjugation. TRIM23 knockdown caused a marked decrease in PPARγ protein abundance during preadipocyte differentiation, resulting in a severe defect in late adipogenic differentiation, whereas it did not affect the formation of early enhanceosomes. Our results suggest that TRIM23 plays a critical role in the switching from early to late adipogenic enhanceosomes by stabilizing PPARγ protein possibly via atypical polyubiquitin conjugation. DOI: http://dx.doi.org/10.7554/eLife.05615.001 PMID:25905670

  10. Dynamic mitochondrial–nuclear redistribution of the immunophilin FKBP51 is regulated by the PKA signaling pathway to control gene expression during adipocyte differentiation

    PubMed Central

    Toneatto, Judith; Guber, Sergio; Charó, Nancy L.; Susperreguy, Sebastián; Schwartz, Jessica; Galigniana, Mario D.; Piwien-Pilipuk, Graciela

    2013-01-01

    Summary Glucocorticoids play an important role in adipogenesis through the glucocorticoid receptor (GR) that forms a heterocomplex with Hsp90•Hsp70 and one high molecular weight immunophilin, either FKBP51 or FKBP52. When 3T3-L1 preadipocytes are induced to differentiate, FKBP51 expression progressively increases, whereas FKBP52 decreases, and Hsp90, Hsp70, p23 and Cyp40 remain unchanged. Interestingly, FKBP51 rapidly translocates from mitochondria to the nucleus where it is retained upon its interaction with chromatin and the nuclear matrix. FKBP51 nuclear localization is transient, and after 48 hours it cycles back to mitochondria. Importantly, this dynamic FKBP51 mitochondrial–nuclear shuttling depends on PKA signaling, because its inhibition by PKI or knockdown of PKA-cα by siRNA, prevented FKBP51 nuclear translocation induced by IBMX. In addition, the electrophoretic pattern of migration of FKBP51 is altered by treatment of cells with PKI or knockdown of PKA-cα, suggesting that FKBP51 is a PKA substrate. In preadipocytes, FKBP51 colocalizes with PKA-cα in mitochondria. When adipogenesis is triggered, PKA-cα also moves to the nucleus colocalizing with FKBP51 mainly in the nuclear lamina. Moreover, FKBP51 and GR interaction increases when preadipocytes are induced to differentiate. GR transcriptional capacity is reduced when cells are incubated in the presence of IBMX, forskolin or dibutyryl-cAMP, compounds that induced FKBP51 nuclear translocation, but not by a specific activator of EPAC. FKBP51 knockdown facilitates adipogenesis, whereas ectopic expression of FKBP51 blocks adipogenesis. These findings indicate that the dynamic mitochondrial–nuclear shuttling of FKBP51 regulated by PKA may be key in fine-tuning the transcriptional control of GR target genes required for the acquisition of adipocyte phenotype. PMID:24101724

  11. Glucocorticoids increase adipocytes in muscle by affecting IL-4 regulated FAP activity

    PubMed Central

    Dong, Yanjun; Silva, Kleiton Augusto Santos; Dong, Yanlan; Zhang, Liping

    2014-01-01

    An increase in intramuscular adipocyte tissue (IMAT) is associated with glucose dysregulation, decreased muscle strength, and increased risk of disability. Unfortunately, the mechanisms stimulating intramuscular adipogenesis remain unclear. We found that dexamethasone (Dex) administration to mice with injured muscles stimulates the accumulation of IMAT. To identify precursors of these adipocytes, we isolated satellite cells and fibro/adipogenic progenitors (FAPs) from muscle; satellite cells did not differentiate into adipocytes even following Dex treatment. In contrast, Dex stimulated FAP differentiation into adipocytes. In vivo, we transplanted purified FAPs from transgenic, EGFP mice into the injured muscles of C57/BL6 mice and found that Dex administration stimulated adipogenesis from FAP-EGFP. The increase in adipogenesis depended on Dex-induced inhibition of interleukin-4 (IL-4). In the injured muscle of IL-4-knockout mice, the levels of adipocytes were increased, while in the injured muscles of Dex-treated mice with IL-4 injections, adipogenesis was suppressed. In cultured FAPs, IL-4 inhibited Dex-induced conversion of FAPs into adipocytes; this did not occur in FAPs expressing knockdown of the IL-4 receptor. Thus, we concluded that glucocorticoids stimulate FAPs to differentiate into adipocytes in injured muscles. This process is blocked by IL-4, suggesting that interfering with IL-4 signaling could prevent adipogenesis in muscle.—Dong, Y., Silva, K. A. S., Dong, Y., Zhang, L. Glucocorticoids increase adipocytes in muscle by affecting IL-4 regulated FAP activity. PMID:24948596

  12. Transcriptional Pathways in cPGI2-Induced Adipocyte Progenitor Activation for Browning

    PubMed Central

    Bayindir, Irem; Babaeikelishomi, Rohollah; Kocanova, Silvia; Sousa, Isabel Sofia; Lerch, Sarah; Hardt, Olaf; Wild, Stefan; Bosio, Andreas; Bystricky, Kerstin; Herzig, Stephan; Vegiopoulos, Alexandros

    2015-01-01

    De novo formation of beige/brite adipocytes from progenitor cells contributes to the thermogenic adaptation of adipose tissue and holds great potential for the therapeutic remodeling of fat as a treatment for obesity. Despite the recent identification of several factors regulating browning of white fat, there is a lack of physiological cell models for the mechanistic investigation of progenitor-mediated beige/brite differentiation. We have previously revealed prostacyclin (PGI2) as one of the few known endogenous extracellular mediators promoting de novo beige/brite formation by relaying β-adrenergic stimulation to the progenitor level. Here, we present a cell model based on murine primary progenitor cells defined by markers previously shown to be relevant for in vivo browning, including a simplified isolation procedure. We demonstrate the specific and broad induction of thermogenic gene expression by PGI2 signaling in the absence of lineage conversion, and reveal the previously unidentified nuclear relocalization of the Ucp1 gene locus in association with transcriptional activation. By profiling the time course of the progenitor response, we show that PGI2 signaling promoted progenitor cell activation through cell cycle and adhesion pathways prior to metabolic maturation toward an oxidative cell phenotype. Our results highlight the importance of core progenitor activation pathways for the recruitment of thermogenic cells and provide a resource for further mechanistic investigation. PMID:26347713

  13. Biology of Beige Adipocyte and Possible Therapy for Type 2 Diabetes and Obesity.

    PubMed

    Lizcano, Fernando; Vargas, Diana

    2016-01-01

    All mammals own two main forms of fat. The classical white adipose tissue builds up energy in the form of triglycerides and is useful for preventing fatigue during periods of low caloric intake and the brown adipose tissue instead of inducing fat accumulation can produce energy as heat. Since adult humans possess significant amounts of active brown fat depots and their mass inversely correlates with adiposity, brown fat might play an important role in human obesity and energy homeostasis. New evidence suggests two types of thermogenic adipocytes with distinct developmental and anatomical features: classical brown adipocytes and beige adipocytes. Beige adipocyte has recently attracted special interest because of its ability to dissipate energy and the possible ability to differentiate itself from white adipocytes. Importantly, adult human brown adipocyte appears to be mainly composed of beige-like adipocytes, making this cell type an attractive therapeutic target for obesity and obesity-related diseases. Because many epigenetic changes can affect beige adipocyte differentiation, the knowledge of the circumstances that affect the development of beige adipocyte cells may be important for therapeutic strategies. In this review we discuss some recent observations arising from the great physiological capacity of these cells and their possible role as ways to treat obesity and diabetes mellitus type 2. PMID:27528872

  14. Biology of Beige Adipocyte and Possible Therapy for Type 2 Diabetes and Obesity

    PubMed Central

    2016-01-01

    All mammals own two main forms of fat. The classical white adipose tissue builds up energy in the form of triglycerides and is useful for preventing fatigue during periods of low caloric intake and the brown adipose tissue instead of inducing fat accumulation can produce energy as heat. Since adult humans possess significant amounts of active brown fat depots and their mass inversely correlates with adiposity, brown fat might play an important role in human obesity and energy homeostasis. New evidence suggests two types of thermogenic adipocytes with distinct developmental and anatomical features: classical brown adipocytes and beige adipocytes. Beige adipocyte has recently attracted special interest because of its ability to dissipate energy and the possible ability to differentiate itself from white adipocytes. Importantly, adult human brown adipocyte appears to be mainly composed of beige-like adipocytes, making this cell type an attractive therapeutic target for obesity and obesity-related diseases. Because many epigenetic changes can affect beige adipocyte differentiation, the knowledge of the circumstances that affect the development of beige adipocyte cells may be important for therapeutic strategies. In this review we discuss some recent observations arising from the great physiological capacity of these cells and their possible role as ways to treat obesity and diabetes mellitus type 2. PMID:27528872

  15. Impact of tamoxifen on adipocyte lineage tracing: Inducer of adipogenesis and prolonged nuclear translocation of Cre recombinase

    PubMed Central

    Ye, Risheng; Wang, Qiong A.; Tao, Caroline; Vishvanath, Lavanya; Shao, Mengle; McDonald, Jeffery G.; Gupta, Rana K.; Scherer, Philipp E.

    2015-01-01

    Background The selective estrogen receptor modulator tamoxifen, in combination with the Cre-ERT2 fusion protein, has been one of the mainstream methods to induce genetic recombination and has found widespread application in lineage tracing studies. Methods & results Here, we report that tamoxifen exposure at widely used concentrations remains detectable by mass-spectrometric analysis in adipose tissue after a washout period of 10 days. Surprisingly, its ability to maintain nuclear translocation of the Cre-ERT2 protein is preserved beyond 2 months of washout. Tamoxifen treatment acutely leads to transient lipoatrophy, followed by de novo adipogenesis that reconstitutes the original fat mass. In addition, we find a “synthetically lethal” phenotype for adipocytes when tamoxifen treatment is combined with adipocyte-specific loss-of-function mutants, such as an adipocyte-specific PPARγ knockout. This is observed to a lesser extent when alternative inducible approaches are employed. Conclusions These findings highlight the potential for tamoxifen-induced adipogenesis, and the associated drawbacks of the use of tamoxifen in lineage tracing studies, explaining the discrepancy in lineage tracing results from different systems with temporal control of gene targeting. PMID:26629402

  16. Human white adipocytes express the cold receptor TRPM8 which activation induces UCP1 expression, mitochondrial activation and heat production.

    PubMed

    Rossato, Marco; Granzotto, Marnie; Macchi, Veronica; Porzionato, Andrea; Petrelli, Lucia; Calcagno, Alessandra; Vencato, Juri; De Stefani, Diego; Silvestrin, Valentina; Rizzuto, Rosario; Bassetto, Franco; De Caro, Raffaele; Vettor, Roberto

    2014-03-01

    Mammals possess two types of adipose tissue, white (WAT) and brown (BAT). The uncoupling protein 1 (UCP1) is a hallmark of BAT, being the pivotal player for cold-induced thermogenesis. WAT can acquire BAT characteristics with up-regulation of UCP1 after cold exposure or adrenergic stimulation. In the present study we demonstrated that human white adipocytes express the cold-sensing receptor TRPM8 which activation by menthol and icilin induced a rise in [Ca²⁺](i) and UCP1 expression, increased mitochondrial membrane potential, glucose uptake and heat production. The induction of "brown-like" phenotype in human white adipocytes after TRPM8 activation was supported by ultrastructural morphological changes of mitochondrial morphology and of their intracellular localization, with no modifications of the genes regulating mitochondrial biogenesis. In conclusion human white adipocytes express the cold receptor TRPM8 which activation induces their "browning" supporting a possible role of this receptor in the control of adipose tissue metabolism and body energy balance. PMID:24342393

  17. L-4F Inhibits Oxidized Low-density Lipoprotein-induced Inflammatory Adipokine Secretion via Cyclic AMP/Protein Kinase A-CCAAT/Enhancer Binding Protein β Signaling Pathway in 3T3-L1 Adipocytes

    PubMed Central

    Xie, Xiang-Zhu; Huang, Xin; Zhao, Shui-Ping; Yu, Bi-Lian; Zhong, Qiao-Qing; Cao, Jian

    2016-01-01

    Background: Adipocytes behave like a rich source of pro-inflammatory cytokines including monocyte chemoattractant protein-1 (MCP-1). Oxidized low-density lipoprotein (oxLDL) participates in the local chronic inflammatory response, and high-density lipoprotein could counterbalance the proinflammatory function of oxLDL, but the underlying mechanism is not completely understood. This study aimed to evaluate the effect of apolipoprotein A-I mimetic peptide L-4F on the secretion and expression of MCP-1 in fully differentiated 3T3-L1 adipocytes induced by oxLDL and to elucidate the possible mechanisms. Methods: Fully differentiated 3T3-L1 adipocytes were incubated in the medium containing various concentration of L-4F (0–50 μg/ml) with oxLDL (50 μg/ml) stimulated, with/without protein kinase A (PKA) inhibitor H-89 (10 μmol/L) preincubated. The concentrations of MCP-1 in the supernatant, the mRNA expression of MCP-1, the levels of CCAAT/enhancer binding protein α (C/EBPα), and CCAAT/enhancer binding protein β (C/EBPβ) were evaluated. The monocyte chemotaxis assay was performed by micropore filter method using a modified Boyden chamber. Results: OxLDL stimulation induced a significant increase of MCP-1 expression and secretion in 3T3-L1 adipocytes, which were inhibited by L-4F preincubation in a dose-dependent manner. PKA inhibitor H-89 markedly reduced the oxLDL-induced MCP-1 expression, but no further decrease was observed when H-89 was used in combination with L-4F (50 μg/ml) (P > 0.05). OxLDL stimulation showed no significant effect on C/EBPα protein level but increased C/EBPβ protein level in a time-dependent manner. H-89 and L-4F both attenuated C/EBPβ protein level in oxLDL-induced 3T3-L1 adipocytes. Conclusions: OxLDL induces C/EBPβ protein synthesis in a time-dependent manner and enhances MCP-1 secretion and expression in 3T3-L1 adipocytes. L-4F dose-dependently counterbalances the pro-inflammatory effect of oxLDL, and cyclic AMP

  18. Adipocyte-Specific Hypoxia-Inducible Factor 2α Deficiency Exacerbates Obesity-Induced Brown Adipose Tissue Dysfunction and Metabolic Dysregulation

    PubMed Central

    Alexaki, Vasileia I.; Qin, Nan; Rubín de Celis, María F.; Economopoulou, Matina; Ziogas, Athanasios; Gercken, Bettina; Kotlabova, Klara; Phieler, Julia; Ehrhart-Bornstein, Monika; Bornstein, Stefan R.; Eisenhofer, Graeme; Breier, Georg; Blüher, Matthias; Hampe, Jochen; El-Armouche, Ali; Chatzigeorgiou, Antonios; Chung, Kyoung-Jin

    2015-01-01

    Angiogenesis is a central regulator for white (WAT) and brown (BAT) adipose tissue adaptation in the course of obesity. Here we show that deletion of hypoxia-inducible factor 2α (HIF2α) in adipocytes (by using Fabp4-Cre transgenic mice) but not in myeloid or endothelial cells negatively impacted WAT angiogenesis and promoted WAT inflammation, WAT dysfunction, hepatosteatosis, and systemic insulin resistance in obesity. Importantly, adipocyte HIF2α regulated vascular endothelial growth factor (VEGF) expression and angiogenesis of obese BAT as well as its thermogenic function. Consistently, obese adipocyte-specific HIF2α-deficient mice displayed BAT dysregulation, associated with reduced levels of uncoupling protein 1 (UCP1) and a dysfunctional thermogenic response to cold exposure. VEGF administration reversed WAT and BAT inflammation and BAT dysfunction in adipocyte HIF2α-deficient mice. Together, our findings show that adipocyte HIF2α is protective against maladaptation to obesity and metabolic dysregulation by promoting angiogenesis in both WAT and BAT and by counteracting obesity-mediated BAT dysfunction. PMID:26572826

  19. β-Catenin Stabilization in Skin Fibroblasts Causes Fibrotic Lesions by Preventing Adipocyte Differentiation of the Reticular Dermis.

    PubMed

    Mastrogiannaki, Maria; Lichtenberger, Beate M; Reimer, Andreas; Collins, Charlotte A; Driskell, Ryan R; Watt, Fiona M

    2016-06-01

    The Wnt/β-catenin pathway plays a central role in epidermal homeostasis and regeneration, but how it affects fibroblast fate decisions is unknown. We investigated the effect of targeted β-catenin stabilization in dermal fibroblasts. Comparative gene expression profiling of stem cell antigen 1(-) (Sca1(-)) and Sca1(+) neonatal fibroblasts from upper and lower dermis, respectively, confirmed that Sca1(+) cells had a preadipocyte signature and showed differential expression of Wnt/β-catenin-associated genes. By targeting all fibroblasts or selectively targeting Dlk1(+) lower dermal fibroblasts, we found that β-catenin stabilization between developmental stages E16.5 and P2 resulted in a reduction in the dermal adipocyte layer with a corresponding increase in dermal fibrosis and an altered hair cycle. The fibrotic phenotype correlated with a reduction in the potential of Sca1(+) fibroblasts to undergo adipogenic differentiation ex vivo. Our findings indicate that Wnt/β-catenin signaling controls adipogenic cell fate within the lower dermis, which potentially contributes to the pathogenesis of fibrotic skin diseases. PMID:26902921

  20. Adipocyte-Specific Deletion of Manganese Superoxide Dismutase Protects From Diet-Induced Obesity Through Increased Mitochondrial Uncoupling and Biogenesis.

    PubMed

    Han, Yong Hwan; Buffolo, Márcio; Pires, Karla Maria; Pei, Shaobo; Scherer, Philipp E; Boudina, Sihem

    2016-09-01

    Obesity and insulin resistance are associated with oxidative stress (OS). The causal role of adipose OS in the pathogenesis of these conditions is unknown. To address this issue, we generated mice with an adipocyte-selective deletion of manganese superoxide dismutase (MnSOD). When fed a high-fat diet (HFD), the AdSod2 knockout (KO) mice exhibited less adiposity, reduced adipocyte hypertrophy, and decreased circulating leptin. The resistance to diet-induced adiposity was the result of an increased metabolic rate and energy expenditure. Furthermore, palmitate oxidation was elevated in the white adipose tissue (WAT) and brown adipose tissue of AdSod2 KO mice fed an HFD, and the expression of key fatty acid oxidation genes was increased. To gain mechanistic insight into the increased fat oxidation in HFD-fed AdSod2 KO mice, we quantified the mitochondrial function and mitochondrial content in WAT and found that MnSOD deletion increased mitochondrial oxygen consumption and induced mitochondrial biogenesis. This effect was preserved in cultured adipocytes from AdSod2 KO mice in vitro. As expected from the enhanced fat oxidation, circulating levels of free fatty acids were reduced in the HFD-fed AdSod2 KO mice. Finally, HFD-fed AdSod2 KO mice were protected from hepatic steatosis, adipose tissue inflammation, and glucose and insulin intolerance. Taken together, these results demonstrate that MnSOD deletion in adipocytes triggered an adaptive stress response that activated mitochondrial biogenesis and enhanced mitochondrial fatty acid oxidation, thereby preventing diet-induced obesity and insulin resistance. PMID:27284109

  1. Zfp423 Maintains White Adipocyte Identity through Suppression of the Beige Cell Thermogenic Gene Program.

    PubMed

    Shao, Mengle; Ishibashi, Jeff; Kusminski, Christine M; Wang, Qiong A; Hepler, Chelsea; Vishvanath, Lavanya; MacPherson, Karen A; Spurgin, Stephen B; Sun, Kai; Holland, William L; Seale, Patrick; Gupta, Rana K

    2016-06-14

    The transcriptional regulators Ebf2 and Prdm16 establish and maintain the brown and/or beige fat cell identity. However, the mechanisms operating in white adipocytes to suppress the thermogenic gene program and maintain an energy-storing phenotype are less understood. Here, we report that the transcriptional regulator Zfp423 is critical for maintaining white adipocyte identity through suppression of the thermogenic gene program. Zfp423 expression is enriched in white versus brown adipocytes and suppressed upon cold exposure. Doxycycline-inducible inactivation of Zfp423 in mature adipocytes, combined with β-adrenergic stimulation, triggers a conversion of differentiated adiponectin-expressing inguinal and gonadal adipocytes into beige-like adipocytes; this reprogramming event is sufficient to prevent and reverse diet-induced obesity and insulin resistance. Mechanistically, Zfp423 acts in adipocytes to inhibit the activity of Ebf2 and suppress Prdm16 activation. These data identify Zfp423 as a molecular brake on adipocyte thermogenesis and suggest a therapeutic strategy to unlock the thermogenic potential of white adipocytes in obesity. PMID:27238639

  2. Effects of Adipocyte Aryl Hydrocarbon Receptor Deficiency on PCB-Induced Disruption of Glucose Homeostasis in Lean and Obese Mice

    PubMed Central

    Baker, Nicki A.; Shoemaker, Robin; English, Victoria; Larian, Nika; Sunkara, Manjula; Morris, Andrew J.; Walker, Mary; Yiannikouris, Frederique

    2015-01-01

    Background Coplanar polychlorinated biphenyls (PCBs) promote adipocyte inflammation and impair glucose homeostasis in lean mice. The diabetes-promoting effects of lipophilic PCBs have been observed only during weight loss in obese mice. The molecular mechanisms linking PCB exposures to impaired glucose metabolism are unclear. Objectives In this study we tested the hypothesis that coplanar PCBs act at adipocyte aryl hydrocarbon receptors (AhRs) to promote adipose inflammation and impair glucose homeostasis in lean mice and in obese mice during weight loss. Methods and Results PCB-77 administration impaired glucose and insulin tolerance in LF (low fat diet)–fed control (AhRfl/fl) mice but not in adipocyte AhR–deficient mice (AhRAdQ). Unexpectedly, AhRAdQ mice exhibited increased fat mass when fed a standard LF or high fat (HF) diet. In mice fed a HF diet, both genotypes became obese, but AhRAdQ mice administered vehicle (VEH) exhibited increased body weight, adipose mass, adipose inflammation, and impaired glucose tolerance compared with AhRfl/fl controls. Impairment of glucose homeostasis in response to PCB-77 was not observed in obese mice of either genotype. However, upon weight loss, AhRfl/fl mice administered PCB-77 exhibited increased abundance of adipose tumor necrosis factor-α (TNF-α) mRNA and impaired glucose homeostasis compared with those administered VEH. In contrast, PCB-77 had no effect on TNF-α or glucose homeostasis in AhRAdQ mice exhibiting weight loss. Conclusions Our results demonstrate that adipocyte AhR mediates PCB-induced adipose inflammation and impairment of glucose homeostasis in mice. Moreover, deficiency of AhR in adipocytes augmented the development of obesity, indicating that endogenous ligand(s) for AhR regulate adipose homeostasis. Citation Baker NA, Shoemaker R, English V, Larian N, Sunkara M, Morris AJ, Walker M, Yiannikouris F, Cassis LA. 2015. Effects of adipocyte aryl hydrocarbon receptor deficiency on PCB-induced

  3. Green Tea (-)-Epigallotocatechin-3-Gallate Induces PGC-1α Gene Expression in HepG2 Cells and 3T3-L1 Adipocytes.

    PubMed

    Lee, Mak-Soon; Lee, Seohyun; Doo, Miae; Kim, Yangha

    2016-03-01

    Green tea (Camellia sinensis) is one of the most popular beverages in the world and has been acknowledged for centuries as having significant health benefits. (-)-Epigallocatechin-3-gallate (EGCG) is the most abundant catechin in green tea, and it has been reported to have health benefit effects. Peroxisome proliferator-activated receptor γ coactivator (PGC)-1α is a crucial regulator of mitochondrial biogenesis and hepatic gluconeogenesis. The objective of this study was to investigate whether EGCG from green tea can affect the ability of transcriptional regulation on PGC-1α mRNA expression in HepG2 cells and 3T3-L1 adipocytes. To study the molecular mechanism that allows EGCG to control PGC-1α expression, the promoter activity levels of PGC-1α were examined. The PGC-1α mRNA level was measured using quantitative real-time PCR. The -970/+412 bp of PGC-1α promoter was subcloned into the pGL3-Basic vector that includes luciferase as a reporter gene. EGCG was found to up-regulate the PGC-1α mRNA levels significantly with 10 μmol/L of EGCG in HepG2 cells and differentiated 3T3-L1 adipocytes. PGC-1α promoter activity was also increased by treatment with 10 μmol/L of EGCG in both cells. These results suggest that EGCG may induce PGC-1α gene expression, potentially through promoter activation. PMID:27069908

  4. Green Tea (−)-Epigallotocatechin-3-Gallate Induces PGC-1α Gene Expression in HepG2 Cells and 3T3-L1 Adipocytes

    PubMed Central

    Lee, Mak-Soon; Lee, Seohyun; Doo, Miae; Kim, Yangha

    2016-01-01

    Green tea (Camellia sinensis) is one of the most popular beverages in the world and has been acknowledged for centuries as having significant health benefits. (−)-Epigallocatechin-3-gallate (EGCG) is the most abundant catechin in green tea, and it has been reported to have health benefit effects. Peroxisome proliferator-activated receptor γ coactivator (PGC)-1α is a crucial regulator of mitochondrial biogenesis and hepatic gluconeogenesis. The objective of this study was to investigate whether EGCG from green tea can affect the ability of transcriptional regulation on PGC-1α mRNA expression in HepG2 cells and 3T3-L1 adipocytes. To study the molecular mechanism that allows EGCG to control PGC-1α expression, the promoter activity levels of PGC-1α were examined. The PGC-1α mRNA level was measured using quantitative real-time PCR. The −970/+412 bp of PGC-1α promoter was subcloned into the pGL3-Basic vector that includes luciferase as a reporter gene. EGCG was found to up-regulate the PGC-1α mRNA levels significantly with 10 μmol/L of EGCG in HepG2 cells and differentiated 3T3-L1 adipocytes. PGC-1α promoter activity was also increased by treatment with 10 μmol/L of EGCG in both cells. These results suggest that EGCG may induce PGC-1α gene expression, potentially through promoter activation. PMID:27069908

  5. Activation of peroxisome proliferator-activated receptor-{alpha} enhances fatty acid oxidation in human adipocytes

    SciTech Connect

    Lee, Joo-Young; Hashizaki, Hikari; Goto, Tsuyoshi; Sakamoto, Tomoya; Takahashi, Nobuyuki; Kawada, Teruo

    2011-04-22

    Highlights: {yields} PPAR{alpha} activation increased mRNA expression levels of adipocyte differentiation marker genes and GPDH activity in human adipocytes. {yields} PPAR{alpha} activation also increased insulin-dependent glucose uptake in human adipocytes. {yields} PPAR{alpha} activation did not affect lipid accumulation in human adipocytes. {yields} PPAR{alpha} activation increased fatty acid oxidation through induction of fatty acid oxidation-related genes in human adipocytes. -- Abstract: Peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}) is a key regulator for maintaining whole-body energy balance. However, the physiological functions of PPAR{alpha} in adipocytes have been unclarified. We examined the functions of PPAR{alpha} using human multipotent adipose tissue-derived stem cells as a human adipocyte model. Activation of PPAR{alpha} by GW7647, a potent PPAR{alpha} agonist, increased the mRNA expression levels of adipocyte differentiation marker genes such as PPAR{gamma}, adipocyte-specific fatty acid-binding protein, and lipoprotein lipase and increased both GPDH activity and insulin-dependent glucose uptake level. The findings indicate that PPAR{alpha} activation stimulates adipocyte differentiation. However, lipid accumulation was not changed, which is usually observed when PPAR{gamma} is activated. On the other hand, PPAR{alpha} activation by GW7647 treatment induced the mRNA expression of fatty acid oxidation-related genes such as CPT-1B and AOX in a PPAR{alpha}-dependent manner. Moreover, PPAR{alpha} activation increased the production of CO{sub 2} and acid soluble metabolites, which are products of fatty acid oxidation, and increased oxygen consumption rate in human adipocytes. The data indicate that activation of PPAR{alpha} stimulates both adipocyte differentiation and fatty acid oxidation in human adipocytes, suggesting that PPAR{alpha} agonists could improve insulin resistance without lipid accumulation in adipocytes. The expected

  6. Curcumin improves hypoxia induced dysfunctions in 3T3-L1 adipocytes by protecting mitochondria and down regulating inflammation.

    PubMed

    Priyanka, Ariyapalli; Anusree, Sasidharan Suseela; Nisha, Vijayakumar Marykutty; Raghu, Kozhiparambil Gopalan

    2014-01-01

    Obesity induced metabolic syndrome is increasing worldwide at an alarming rate. It is characterized by excessive expansion of white adipose tissue which leads to hypoxia and impairs normal metabolism. Recent studies reveal that hypoxia could be one of the factors for inflammation, insulin resistance and other obesity related complications. There is a high demand for anti-obese phytoceuticals to control and manage the complications resulting from obesity. In this study, we investigated how hypoxia affect the physiological functions of 3T3-L1 adipocytes emphasizing on oxidative stress, inflammation, and mitochondrial functions. We also evaluated the protective role of various doses of curcumin, a well-known dietary antioxidant, on hypoxia induced alterations. The results revealed that hypoxia significantly altered the vital parameters of adipocyte biology like HIF 1α expression (103.47% ↑), lactate, and glycerol release (184.34% and 69.1% ↑, respectively), reactive oxygen species production (432.53% ↑), lipid and protein oxidation (376.6% and 566.6% ↑, respectively), reduction in antioxidant enzymes (superoxide dismutase and catalase) status, secretion of inflammatory markers (TNF α, IL 6, IL 1β, and IFN γ), and mitochondrial functions (mitochondrial mass, membrane potential, permeability transition pore integrity, and superoxide generation). Curcumin substantially protected adipocytes from toxic effects of hypoxia in a dose dependent manner by protecting mitochondria and down regulating inflammation. Acriflavine is used as a positive control. A detailed investigation is required for the development of curcumin as an effective nutraceutical against obesity. PMID:25110893

  7. Thiazolidinedione (pioglitazone) blocks P. gingivalis- and F. nucleatum, but not E. coli, lipopolysaccharide (LPS)-induced interleukin-6 (IL-6) production in adipocytes.

    PubMed

    Yamaguchi, M; Nishimura, F; Naruishi, H; Soga, Y; Kokeguchi, S; Takashiba, S

    2005-03-01

    An elevated level of C-reactive protein (CRP) predicts the future development of coronary heart disease. Periodontitis appears to up-regulate CRP. CRP is produced by hepatocytes in response to interleukin-6 (IL-6). A major source of IL-6 in obese subjects is adipocytes. We hypothesized that lipopolysaccharide (LPS) from periodontal pathogens stimulated adipocytes to produce IL-6, and that the production was suppressed by the drugs targeted against insulin resistance, thiazolidinedione (pioglitazone), since this agent potentially showed an anti-inflammatory effect. Mouse 3T3-L1 adipocytes were stimulated with E. coli, P. gingivalis, and F. nucleatum LPS. The IL-6 concentration in culture supernatants was measured. All LPS stimulated adipocytes to produce IL-6. Although pioglitazone changed adipocyte appearance from large to small, and completely suppressed P. gingivalis and F. nucleatum LPS-induced IL-6 production, E. coli LPS-induced IL-6 production was not efficiently blocked. Thus, pioglitazone completely blocked periodontal-bacteria-derived LPS-induced IL-6 production in adipocytes, a major inducer of CRP. PMID:15723863

  8. Effects of Pueraria lobata Root Ethanol Extract on Adipogenesis and Lipogenesis During 3T3-L1 Differentiation into Adipocytes.

    PubMed

    Lee, Chae Myoung; Yoon, Mi Sook; Kim, Young Chul

    2015-06-01

    We evaluated the inhibitory effect of Pueraria lobata root ethanol extract (PLREE) on lipid accumulation during 3T3-L1 differentiation to adipocytes by measuring the intracellular expression of adipogenic, lipogenic, and lipolytic markers and lipid accumulation. The total polyphenol and flavonoid content of PLREE were 47 and 29 mg/g, respectively. The electron donating capacity of PLREE at 1,000 μg/mL was 48.8%. Treatment of 3T3-L1 preadipocytes with 100, 250, or 500 μg/mL PLREE for 8 days dose-dependently promoted the differentiation of 3T3-L1 cells. In contrast, the lipid content of PLREE-treated cells was significantly reduced by 7.8% (p < 0.05), 35.6% (p < 0.001), and 42.2% (p < 0.001) following treatment with 100, 250, and 500 μg/mL PLREE, respectively, as compared to differentiated control cells. PLREE upregulated peroxisome proliferator-activated receptor γ mRNA and protein, and sterol regulator element-binding protein-1c mRNA levels, but did not affect CCAAT/enhancer binding-protein β and α mRNA levels. PLREE also downregulated acetyl-CoA carboxylase mRNA and protein, fatty acid synthase (FAS) protein, and leptin mRNA levels, but did not affect FAS mRNA expression. PLREE upregulated adipose triglyceride lipase mRNA and protein expression, and hormone-sensitive lipase (HSL) protein expression, but did not affect HSL mRNA expression. In conclusion, we found that PLREE enhanced adipogenesis, but reduced lipogenesis, resulting in decreased lipid accumulation in 3T3-L1 cells. PMID:26191386

  9. All-trans retinoic acid induces oxidative phosphorylation and mitochondria biogenesis in adipocytes[S

    PubMed Central

    Tourniaire, Franck; Musinovic, Hana; Gouranton, Erwan; Astier, Julien; Marcotorchino, Julie; Arreguin, Andrea; Bernot, Denis; Palou, Andreu; Bonet, M. Luisa; Ribot, Joan; Landrier, Jean-François

    2015-01-01

    A positive effect of all-trans retinoic acid (ATRA) on white adipose tissue (WAT) oxidative and thermogenic capacity has been described and linked to an in vivo fat-lowering effect of ATRA in mice. However, little is known about the effects of ATRA on mitochondria in white fat. Our objective has been to characterize the effect of ATRA on mitochondria biogenesis and oxidative phosphorylation (OXPHOS) capacity in mature white adipocytes. Transcriptome analysis, oxygraphy, analysis of mitochondrial DNA (mtDNA), and flow cytometry-based analysis of mitochondria density were performed in mature 3T3-L1 adipocytes after 24 h incubation with ATRA (2 µM) or vehicle. Selected genes linked to mitochondria biogenesis and function and mitochondria immunostaining were analyzed in WAT tissues of ATRA-treated as compared with vehicle-treated mice. ATRA upregulated the expression of a large set of genes linked to mtDNA replication and transcription, mitochondrial biogenesis, and OXPHOS in adipocytes, as indicated by transcriptome analysis. Oxygen consumption rate, mtDNA content, and staining of mitochondria were increased in the ATRA-treated adipocytes. Similar results were obtained in WAT depots of ATRA-treated mice. We conclude that ATRA impacts mitochondria in adipocytes, leading to increased OXPHOS capacity and mitochondrial content in these cells. PMID:25914170

  10. An analogue of atrial natriuretic peptide (C-ANP4-23) modulates glucose metabolism in human differentiated adipocytes.

    PubMed

    Ruiz-Ojeda, Francisco Javier; Aguilera, Concepción María; Rupérez, Azahara Iris; Gil, Ángel; Gomez-Llorente, Carolina

    2016-08-15

    The present study was undertaken to investigate the effects of C-atrial natriuretic peptide (C-ANP4-23) in human adipose-derived stem cells differentiated into adipocytes over 10 days (1 μM for 4 h). The intracellular cAMP, cGMP and protein kinase A levels were determined by ELISA and gene and protein expression were determined by qRT-PCR and Western blot, respectively, in the presence or absence of C-ANP4-23. The levels of lipolysis and glucose uptake were also determined. C-ANP4-23 treatment significantly increased the intracellular cAMP levels and the gene expression of glucose transporter type 4 (GLUT4) and protein kinase, AMP-activated, alpha 1 catalytic subunit (AMPK). Western blot showed a significant increase in GLUT4 and phosphor-AMPKα levels. Importantly, the adenylate cyclase inhibitor SQ22536 abolished these effects. Additionally, C-ANP4-23 increased glucose uptake by 2-fold. Our results show that C-ANP4-23 enhances glucose metabolism and might contribute to the development of new peptide-based therapies for metabolic diseases. PMID:27181211

  11. Dietary vitamin A restriction affects adipocyte differentiation and fatty acid composition of intramuscular fat in Iberian pigs.

    PubMed

    Ayuso, M; Óvilo, C; Rodríguez-Bertos, A; Rey, A I; Daza, A; Fenández, A; González-Bulnes, A; López-Bote, C J; Isabel, B

    2015-10-01

    The aim of this study was to investigate whether dietary vitamin A level is associated with differences in adipocyte differentiation or lipid accumulation in Iberian pigs at early growing (35.8kg live weight) and at finishing (158kg live weight). Iberian pigs of 16.3kg live weight were allocated to two feeding groups, one group received 10,000IU of vitamin A/kg diet (control); the other group received a diet with 0IU of vitamin A (var) for the whole experimental period. The dietary vitamin A level had no effect on growth performance and carcass traits. The early suppression of vitamin A increased the preadipocyte number in Longissimus thoracis (LT) muscle in the early growth period (P<0.001) and the neutral lipid content and composition (higher MUFA and lower SFA content) at the end of the finishing period (P<0.05). Vitamin A restriction in young pigs increases their lipogenic potential without affecting carcass traits. PMID:26005912

  12. Tribbles 3 inhibits brown adipocyte differentiation and function by suppressing insulin signaling.

    PubMed

    Jeong, Ha-Won; Choi, Ran Hee; McClellan, Jamie L; Piroli, Gerardo G; Frizzell, Norma; Tseng, Yu-Hua; Goodyear, Laurie J; Koh, Ho-Jin

    2016-02-19

    Recent studies have demonstrated that adult humans have substantial amounts of functioning brown adipose tissue (BAT). Since BAT has been implicated as an anti-obese and anti-diabetic tissue, it is important to understand the signaling molecules that regulate BAT function. There has been a link between insulin signaling and BAT metabolism as deletion or pharmaceutical inhibition of insulin signaling impairs BAT differentiation and function. Tribbles 3 (TRB3) is a pseudo kinase that has been shown to regulate metabolism and insulin signaling in multiple tissues but the role of TRB3 in BAT has not been studied. In this study, we found that TRB3 expression was present in BAT and overexpression of TRB3 in brown preadipocytes impaired differentiation and decreased expression of BAT markers. Furthermore, TRB3 overexpression resulted in significantly lower oxygen consumption rates for basal and proton leakage, indicating decreased BAT activity. Based on previous studies showing that deletion or pharmaceutical inhibition of insulin signaling impairs BAT differentiation and function, we assessed insulin signaling in brown preadipocytes and BAT in vivo. Overexpression of TRB3 in cells impaired insulin-stimulated IRS1 and Akt phosphorylation, whereas TRB3KO mice displayed improved IRS1 and Akt phosphorylation. Finally, deletion of IRS1 abolished the function of TRB3 to regulate BAT differentiation and metabolism. These data demonstrate that TRB3 inhibits insulin signaling in BAT, resulting in impaired differentiation and function. PMID:26801556

  13. Suppression of adipocyte hypertrophy by polymethoxyflavonoids isolated from Kaempferia parviflora.

    PubMed

    Okabe, Yui; Shimada, Tsutomu; Horikawa, Takumi; Kinoshita, Kaoru; Koyama, Kiyotaka; Ichinose, Koji; Aburada, Masaki; Takahashi, Kunio

    2014-05-15

    We previously demonstrated that ethyl acetate extracts of Kaempferia parviflora Wall. Ex Baker (KPE) improve insulin resistance in TSOD mice and showed that its components induce differentiation and adipogenesis in 3T3-L1 preadipocytes. The present study was undertaken to examine whether KPE and its isolated twelve components suppress further lipid accumulation in 3T3-L1 mature adipocytes. KPE reduced intracellular triglycerides in mature adipocytes, as did two of its components, 3,5,7,3',4'-pentamethoxyflavone and 5,7,4'-trimethoxyflavone. Shrinkage of lipid droplets in mature adipocytes was observed, and mRNA expression levels of adipose tissue triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) were up-regulated by these two polymethoxyflavonoids (PMFs). Furthermore, the protein expression level of ATGL and the release level of glycerol into the cell culture medium increased. In contrast, the peroxisome proliferator-activated receptor γ (PPARγ) agonist, troglitazone, did not decrease intracellular triglycerides in mature adipocytes, and the mRNA expression level of PPARγ was not up-regulated in mature adipocytes treated with the two active PMFs. Therefore, suppression of lipid accumulation in mature adipocytes is unlikely to be enhanced by transcriptional activation of PPARγ. These results suggest that KPE and its active components enhance lipolysis in mature adipocytes by activation of ATGL and HSL independent of PPARγ transcription, thus preventing adipocyte hypertrophy. On the other hand, the full hydroxylated flavonoid quercetin did not show the suppressive effects of lipid accumulation in mature adipocyte in the same conditions. Consequently, methoxy groups in the flavones are important for the activity. PMID:24629599

  14. Milk-Derived Tripeptides IPP (Ile-Pro-Pro) and VPP (Val-Pro-Pro) Promote Adipocyte Differentiation and Inhibit Inflammation in 3T3-F442A Cells

    PubMed Central

    Chakrabarti, Subhadeep; Wu, Jianping

    2015-01-01

    Milk derived tripeptides IPP (Ile-Pro-Pro) and VPP (Val-Pro-Pro) have shown promise as anti-hypertensive agents due to their inhibitory effects on angiotensin converting enzyme (ACE). Due to the key inter-related roles of hypertension, chronic inflammation and insulin resistance in the pathogenesis of metabolic syndrome, there is growing interest in investigating established anti-hypertensive agents for their effects on insulin sensitivity and inflammation. In this study, we examined the effects of IPP and VPP on 3T3-F442A murine pre-adipocytes, a widely used model for studying metabolic diseases. We found that both IPP and VPP induced beneficial adipogenic differentiation as manifested by intracellular lipid accumulation, upregulation of peroxisome proliferator-activated receptor gamma (PPARγ) and secretion of the protective lipid hormone adiponectin by these cells. The observed effects were similar to those induced by insulin, suggesting potential benefits in the presence of insulin resistance. IPP and VPP also inhibited cytokine induced pro-inflammatory changes such as reduction in adipokine levels and activation of the nuclear factor kappa B (NF-κB) pathway. Taken together, our findings suggest that IPP and VPP exert insulin-mimetic adipogenic effects and prevent inflammatory changes in adipocytes, which may offer protection against metabolic disease. PMID:25714093

  15. Ginsenoside F2 possesses anti-obesity activity via binding with PPARγ and inhibiting adipocyte differentiation in the 3T3-L1 cell line.

    PubMed

    Siraj, Fayeza Md; SathishKumar, Natarajan; Kim, Yeon Ju; Kim, Se Young; Yang, Deok Chun

    2015-02-01

    Abstract Panax ginseng Meyer has been shown to be effective in mitigating various diseases. Protopanaxadiols (PPD) and protopanaxatriols (PPT), which are the main constituents of ginseng, have been shown to impact obesity. Therefore, we selected several important ginsenosides to perform our docking study and determine if they had binding affinity with the peroxisome proliferator activated receptor gamma (PPARγ), which is a major transcription factor in adipocytes. Among them, only a few ginsenosides demonstrated binding affinity with PPARγ. Other than ginsenoside F2 rest of them were previously reported by the researchers in experimental study in case of obesity cell line 3T3-L1 adipocyte. In few recent studies, it was reported that F2 has protective effects on malignant brain tumors as well as anti-cancer activity in breast cancer. Therefore, we felt it was important to focus on F2 when considering obesity. Our study focused on this ginsenoside and analyzed its impact on 3T3-L1 adipocytes. Following the molecular interaction studies, further experimental studies were carried out and demonstrated that ginsenoside F2 when treated with different doses reduces the level of lipid accumulated by the 3T3-L1 cell line during adipogenesis. Reverse transcriptase polymerase chain reaction (RT-PCR) and quantitative real-time PCR results showed reduction in PPARγ and perilipin gene expression levels compared to that of differentiated adipocytes without any treatment. So considering the binding with a major adipocyte transcription factor and the performed experiments, we suggest that ginsenoside F2 may reduce obesity via the inhibition of adipogenesis in the 3T3-L1 cell line. PMID:24666293

  16. Exercise before or after refeeding prevents refeeding-induced recovery of cell size after fasting with a different pattern of metabolic gene expressions in rat epididymal adipocytes.

    PubMed

    Sakurai, Takuya; Takei, Megumi; Ogasawara, Junetsu; Ueda, Hiroshi; Kizaki, Takako; Ohno, Hideki; Izawa, Tetsuya

    2007-09-01

    We investigated the effect of exercise before or after refeeding on cell size and on the expression of several messenger RNAs (mRNAs) involved in lipolysis and lipogenesis in fasted rat epididymal adipocytes. Fasting for 65 hours reduced the diameter of adipocytes to 72.0 microm from 78.4 microm in fed control rats, whereas refeeding for 1 or 2 days restored adipocyte size to 74.0 or 75.8 microm, respectively. Exercise before or after refeeding blocked refeeding-induced restoration of adipocyte size and led to adipocyte size similar to that observed after fasting. Fasting dramatically reduced expression of the fatty acid synthase mRNA, although expression of this gene returned to the control level after refeeding. However, exercise after but not before refeeding inhibited recovery of the expression of fatty acid synthase mRNA resulting from refeeding. In contrast, exercise before but not after refeeding led to enhanced expression of mRNAs encoding the hormone-sensitive lipase and beta(3)-aderenoceptor. Thus, exercise before or after refeeding prevents refeeding-induced restoration of adipocyte size after fasting via different pathways. Exercise before and after refeeding enhanced the expression of lipolytic mRNAs or inhibited the expression of lipogenic mRNAs, respectively. PMID:17697872

  17. Rat white adipocytes activate p85/p110 PI3K and induce PM GLUT4 in response to adrenoceptor agonists or aluminum fluoride.

    PubMed

    Ohsaka, Y; Nomura, Y

    2016-03-01

    Adipocyte responses to adrenergic and ß-adrenoceptor(-AR) (adrenoceptor) regulation are not sufficiently understood, and information helpful for elucidating the adrenoceptor-responsive machinery is insufficient. Here we show by using immunoprecipitated kinase analysis with a phosphatidylinositol 3-kinase (PI3K) p85 antibody that PI3K activation was induced by treatment with 10 or 100 µM norepinephrine (NE) for 15 min or with 10 mM aluminum fluoride (AF, a guanosine triphosphate (GTP)-binding (G) protein activator) for 20 min in white adipocytes (rat epididymal adipocytes) and that treatment with pertussis toxin (PTX, a G-protein inactivator) inhibited PI3K activation induced by the 20-min treatment with AF in the cells. In addition, western blot analysis revealed that glucose transporter 4 (GLUT4) level in the adipocyte plasma membrane (PM) fraction was increased by treatment with 10 µM NE, 100 µM dobutamine (DOB, a ß1-AR agonist), or 0.1 µM CL316243 (CL, a ß3-AR agonist) for 30 min or with 10 mM AF for 20 min. NE or AF treatment triggered 2-deoxyglucose (2-DG) uptake into adipocytes under the above conditions. Our results advance the understanding of responses to adrenoceptor regulation in white adipocytes and provide possible clues for clarifying the machinery involved in adrenergic and ß-AR responses in the cells. PMID:27030626

  18. Glucocorticoids increase adipocytes in muscle by affecting IL-4 regulated FAP activity.

    PubMed

    Dong, Yanjun; Silva, Kleiton Augusto Santos; Dong, Yanlan; Zhang, Liping

    2014-09-01

    An increase in intramuscular adipocyte tissue (IMAT) is associated with glucose dysregulation, decreased muscle strength, and increased risk of disability. Unfortunately, the mechanisms stimulating intramuscular adipogenesis remain unclear. We found that dexamethasone (Dex) administration to mice with injured muscles stimulates the accumulation of IMAT. To identify precursors of these adipocytes, we isolated satellite cells and fibro/adipogenic progenitors (FAPs) from muscle; satellite cells did not differentiate into adipocytes even following Dex treatment. In contrast, Dex stimulated FAP differentiation into adipocytes. In vivo, we transplanted purified FAPs from transgenic, EGFP mice into the injured muscles of C57/BL6 mice and found that Dex administration stimulated adipogenesis from FAP-EGFP. The increase in adipogenesis depended on Dex-induced inhibition of interleukin-4 (IL-4). In the injured muscle of IL-4-knockout mice, the levels of adipocytes were increased, while in the injured muscles of Dex-treated mice with IL-4 injections, adipogenesis was suppressed. In cultured FAPs, IL-4 inhibited Dex-induced conversion of FAPs into adipocytes; this did not occur in FAPs expressing knockdown of the IL-4 receptor. Thus, we concluded that glucocorticoids stimulate FAPs to differentiate into adipocytes in injured muscles. This process is blocked by IL-4, suggesting that interfering with IL-4 signaling could prevent adipogenesis in muscle. PMID:24948596

  19. Peanut sprout ethanol extract inhibits the adipocyte proliferation, differentiation, and matrix metalloproteinases activities in mouse fibroblast 3T3-L1 preadipocytes

    PubMed Central

    Kim, Woo Kyoung; Kang, Nam E; Kim, Myung Hwan

    2013-01-01

    3T3-L1 preadipocyte were differentiated to adipocytes, and then treated with 0, 10, 20, and 40 µg/mL of peanut sprout ethanol extract (PSEE). The main component of PSEE is resveratrol which contained 5.55 mg/mL of resveratrol. The MTT assay, Oil-Red O staining, glycerol-3-phosphate dehydrogenase (GPDH) activity, and the triglyceride concentration were determined in 3T3-L1 cells. MMP-2 and MMP-9 activities as well as mRNA expressions of C/EBP β and C/EBP α were also investigated. As the concentration of PSEE in adipocytes increased, the cell proliferation was decreased in a dose-dependent manner from 4 days of incubation (P < 0.05). The GDPH activity (P < 0.05) and the triglyceride concentration (P < 0.05) were decreased as the PSEE treatment concentration increased. The mRNA expression of C/EBPβ in 3T3-L1 cells was significantly low in groups of PSEE-treated, compared with control group (P < 0.05). The MMP-9 (P < 0.05) and MMP-2 (P < 0.05) activities were decreased in a dose-dependent manner as the PSEE concentration increased from 20 µg/mL. In conclusion, it was found that PSEE has an effect on restricting proliferation and differentiation of adipocytes. PMID:23766875

  20. Parathyroid hormone induces adipocyte lipolysis via PKA-mediated phosphorylation of hormone-sensitive lipase.

    PubMed

    Larsson, Sara; Jones, Helena A; Göransson, Olga; Degerman, Eva; Holm, Cecilia

    2016-03-01

    Parathyroid hormone (PTH) is secreted from the parathyroid glands in response to low plasma calcium levels. Besides its classical actions on bone and kidney, PTH may have other important effects, including metabolic effects, as suggested for instance by increased prevalence of insulin resistance and type 2 diabetes in patients with primary hyperparathyroidism. Moreover, secondary hyperparathyroidism may contribute to the metabolic derangements that characterize states of vitamin D deficiency. PTH has been shown to induce adipose tissue lipolysis, but the details of the lipolytic action of PTH have not been described. Here we used primary mouse adipocytes to show that intact PTH (1-84) as well as the N-terminal fragment (1-37) acutely stimulated lipolysis in a dose-dependent manner, whereas the C-terminal fragment (38-84) was without lipolytic effect. The lipolytic action of PTH was paralleled by phosphorylation of known protein kinase A (PKA) substrates, i.e. hormone-sensitive lipase (HSL) and perilipin. The phosphorylation of HSL in response to PTH occurred at the known PKA sites S563 and S660, but not at the non-PKA site S565. PTH-induced lipolysis, as well as phosphorylation of HSL at S563 and S660, was blocked by both the PKA-inhibitor H89 and the adenylate cyclase inhibitor MDL-12330A, whereas inhibitors of extracellular-regulated kinase (ERK), protein kinase B (PKB), AMP-activated protein kinase (AMPK) and Ca(2+)/calmodulin-dependent protein kinase (CaMK) had little or no effect. Inhibition of phosphodiesterase 4 (PDE4) strongly potentiated the lipolytic action of PTH, whereas inhibition of PDE3 had no effect. Our results show that the lipolytic action of PTH is mediated by the PKA signaling pathway with no or minor contribution of other signaling pathways and, furthermore, that the lipolytic action of PTH is limited by simultaneous activation of PDE4. Knowledge of the signaling pathways involved in the lipolytic action of PTH is important for our

  1. Beige differentiation of adipose depots in mice lacking prolactin receptor protects against high-fat-diet-induced obesity.

    PubMed

    Auffret, Julien; Viengchareun, Say; Carré, Nadège; Denis, Raphaël G P; Magnan, Christophe; Marie, Pierre-Yves; Muscat, Adeline; Fève, Bruno; Lombès, Marc; Binart, Nadine

    2012-09-01

    Stimulating conversion of white fat to metabolically active adipocytes (beige fat) constitutes a promising strategy against weight gain and its deleterious associated-disorders. We provide direct evidence that prolactin (PRL), best known for its actions on the mammary gland, plays a pivotal role in energy balance through the control of adipocyte differentiation and fate. Here we show that lack of prolactin receptor (PRLR) causes resistance to high-fat-diet-induced obesity due to enhanced energy expenditure and increased metabolic rate. Mutant mice displayed reduced fat mass associated with appearance of massive brown-like adipocyte foci in perirenal and subcutaneous but not in gonadal fat depots under a high-fat diet. Positron emission tomography imaging further demonstrated the occurrence of these thermogenic brown fat depots in adult mice, providing additional support for recruitable brown-like adipocytes (beigeing) in white fat depots. Consistent with the activation of brown adipose tissue, PRLR inactivation increases expression of master genes controlling brown adipocyte fate (PRDM16) and mitochondrial function (PGC1α, UCP1). Altered pRb/Foxc2 expression suggests that this PRL-regulated pathway may contribute to beige cell commitment. Together, these results provide direct genetic evidence that PRLR affects energy balance and metabolic adaptation in rodents via effects on brown adipose tissue differentiation and function. PMID:22637534

  2. Nuclear factor-κB is a common upstream signal for growth differentiation factor-5 expression in brown adipocytes exposed to pro-inflammatory cytokines and palmitate

    SciTech Connect

    Hinoi, Eiichi; Iezaki, Takashi; Ozaki, Kakeru; Yoneda, Yukio

    2014-10-03

    Highlights: • GDF5 expression is up-regulated by IL-1β, TNF-α and palmitate in brown pre-adipocytes. • NF-κB stimulates promoter activity and expression of GDF5 in brown pre-adipocytes. • Recruitment of NF-κB to the GDF5 promoter is facilitated in BAT from ob/ob mice. • An NF-κB inhibitor prevents upregulation of GDF5 expression in brown pre-adipocytes. - Abstract: We have previously demonstrated that genetic and acquired obesity similarly led to drastic upregulation in brown adipose tissue (BAT), rather than white adipose tissue, of expression of both mRNA and corresponding protein for the bone morphogenic protein/growth differentiation factor (GDF) member GDF5 capable of promoting brown adipogenesis. In this study, we evaluated expression profiles of GDF5 in cultured murine brown pre-adipocytes exposed to pro-inflammatory cytokines and free fatty acids (FFAs), which are all shown to play a role in the pathogenesis of obesity. Both interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) were effective in up-regulating GDF5 expression in a concentration-dependent manner, while similar upregulation was seen in cells exposed to the saturated FFA palmitate, but not to the unsaturated FFA oleate. In silico analysis revealed existence of the putative nuclear factor-κB (NF-κB) binding site in the 5′-flanking region of mouse GDF5, whereas introduction of NF-κB subunits drastically facilitated both promoter activity and expression of GDF5 in brown pre-adipocytes. Chromatin immunoprecipitation analysis confirmed significant facilitation of the recruitment of NF-κB to the GDF5 promoter in lysed extracts of BAT from leptin-deficient ob/ob obese mice. Upregulation o GDF5 expression was invariably inhibited by an NF-κB inhibitor in cultured brown pre-adipocytes exposed to IL-1β, TNF-α and palmitate. These results suggest that obesity leads to upregulation of GDF5 expression responsible for the promotion of brown adipogenesis through a mechanism

  3. SIRT1 Limits Adipocyte Hyperplasia through c-Myc Inhibition.

    PubMed

    Abdesselem, Houari; Madani, Aisha; Hani, Ahmad; Al-Noubi, Muna; Goswami, Neha; Ben Hamidane, Hisham; Billing, Anja M; Pasquier, Jennifer; Bonkowski, Michael S; Halabi, Najeeb; Dalloul, Rajaa; Sheriff, Mohamed Z; Mesaeli, Nasrin; ElRayess, Mohamed; Sinclair, David A; Graumann, Johannes; Mazloum, Nayef A

    2016-01-29

    The expansion of fat mass in the obese state is due to increased adipocyte hypertrophy and hyperplasia. The molecular mechanism that drives adipocyte hyperplasia remains unknown. The NAD(+)-dependent protein deacetylase sirtuin 1 (SIRT1), a key regulator of mammalian metabolism, maintains proper metabolic functions in many tissues, counteracting obesity. Here we report that differentiated adipocytes are hyperplastic when SIRT1 is knocked down stably in mouse 3T3-L1 preadipocytes. This phenotype is associated with dysregulated adipocyte metabolism and enhanced inflammation. We also demonstrate that SIRT1 is a key regulator of proliferation in preadipocytes. Quantitative proteomics reveal that the c-Myc pathway is altered to drive enhanced proliferation in SIRT1-silenced 3T3-L1 cells. Moreover, c-Myc is hyperacetylated, levels of p27 are reduced, and cyclin-dependent kinase 2 (CDK2) is activated upon SIRT1 reduction. Remarkably, differentiating SIRT1-silenced preadipocytes exhibit enhanced mitotic clonal expansion accompanied by reduced levels of p27 as well as elevated levels of CCAAT/enhancer-binding protein β (C/EBPβ) and c-Myc, which is also hyperacetylated. c-Myc activation and enhanced proliferation phenotype are also found to be SIRT1-dependent in proliferating mouse embryonic fibroblasts and differentiating human SW872 preadipocytes. Reducing both SIRT1 and c-Myc expression in 3T3-L1 cells simultaneously does not induce the adipocyte hyperplasia phenotype, confirming that SIRT1 controls adipocyte hyperplasia through c-Myc regulation. A better understanding of the molecular mechanisms of adipocyte hyperplasia will open new avenues toward understanding obesity. PMID:26655722

  4. SIRT1 Limits Adipocyte Hyperplasia through c-Myc Inhibition*

    PubMed Central

    Abdesselem, Houari; Madani, Aisha; Hani, Ahmad; Al-Noubi, Muna; Goswami, Neha; Ben Hamidane, Hisham; Billing, Anja M.; Pasquier, Jennifer; Bonkowski, Michael S.; Halabi, Najeeb; Dalloul, Rajaa; Sheriff, Mohamed Z.; Mesaeli, Nasrin; ElRayess, Mohamed; Sinclair, David A.; Graumann, Johannes; Mazloum, Nayef A.

    2016-01-01

    The expansion of fat mass in the obese state is due to increased adipocyte hypertrophy and hyperplasia. The molecular mechanism that drives adipocyte hyperplasia remains unknown. The NAD+-dependent protein deacetylase sirtuin 1 (SIRT1), a key regulator of mammalian metabolism, maintains proper metabolic functions in many tissues, counteracting obesity. Here we report that differentiated adipocytes are hyperplastic when SIRT1 is knocked down stably in mouse 3T3-L1 preadipocytes. This phenotype is associated with dysregulated adipocyte metabolism and enhanced inflammation. We also demonstrate that SIRT1 is a key regulator of proliferation in preadipocytes. Quantitative proteomics reveal that the c-Myc pathway is altered to drive enhanced proliferation in SIRT1-silenced 3T3-L1 cells. Moreover, c-Myc is hyperacetylated, levels of p27 are reduced, and cyclin-dependent kinase 2 (CDK2) is activated upon SIRT1 reduction. Remarkably, differentiating SIRT1-silenced preadipocytes exhibit enhanced mitotic clonal expansion accompanied by reduced levels of p27 as well as elevated levels of CCAAT/enhancer-binding protein β (C/EBPβ) and c-Myc, which is also hyperacetylated. c-Myc activation and enhanced proliferation phenotype are also found to be SIRT1-dependent in proliferating mouse embryonic fibroblasts and differentiating human SW872 preadipocytes. Reducing both SIRT1 and c-Myc expression in 3T3-L1 cells simultaneously does not induce the adipocyte hyperplasia phenotype, confirming that SIRT1 controls adipocyte hyperplasia through c-Myc regulation. A better understanding of the molecular mechanisms of adipocyte hyperplasia will open new avenues toward understanding obesity. PMID:26655722

  5. Inhibition of acetyl-CoA carboxylases by soraphen A prevents lipid accumulation and adipocyte differentiation in 3T3-L1 cells.

    PubMed

    Cordonier, Elizabeth L; Jarecke, Sarah K; Hollinger, Frances E; Zempleni, Janos

    2016-06-01

    Acetyl-CoA carboxylases (ACC) 1 and 2 catalyze the carboxylation of acetyl-CoA to malonyl-CoA and depend on biotin as a coenzyme. ACC1 localizes in the cytoplasm and produces malonyl-CoA for fatty acid (FA) synthesis. ACC2 localizes in the outer mitochondrial membrane and produces malonyl-CoA that inhibits FA import into mitochondria for subsequent oxidation. We hypothesized that ACCs are checkpoints in adipocyte differentiation and tested this hypothesis using the ACC1 and ACC2 inhibitor soraphen A (SA) in murine 3T3-L1 preadipocytes. When 3T3-L1 cells were treated with 100nM SA for 8 days after induction of differentiation, the expression of PPARγ mRNA and FABP4 mRNA decreased by 40% and 50%, respectively, compared with solvent controls; the decrease in gene expression was accompanied by a decrease in FABP4 protein expression and associated with a decrease in lipid droplet accumulation. The rate of FA oxidation was 300% greater in SA-treated cells compared with vehicle controls. Treatment with exogenous palmitate restored PPARγ and FABP4 mRNA expression and FABP4 protein expression in SA-treated cells. In contrast, SA did not alter lipid accumulation if treatment was initiated on day eight after induction of differentiation. We conclude that loss of ACC1-dependent FA synthesis and loss of ACC2-dependent inhibition of FA oxidation prevent lipid accumulation in adipocytes and inhibit early stages of adipocyte differentiation. PMID:27041646

  6. Tenomodulin promotes human adipocyte differentiation and beneficial visceral adipose tissue expansion

    PubMed Central

    Senol-Cosar, Ozlem; Flach, Rachel J. Roth; DiStefano, Marina; Chawla, Anil; Nicoloro, Sarah; Straubhaar, Juerg; Hardy, Olga T.; Noh, Hye Lim; Kim, Jason K.; Wabitsch, Martin; Scherer, Philipp E.; Czech, Michael P.

    2016-01-01

    Proper regulation of energy storage in adipose tissue is crucial for maintaining insulin sensitivity and molecules contributing to this process have not been fully revealed. Here we show that type II transmembrane protein tenomodulin (TNMD) is upregulated in adipose tissue of insulin-resistant versus insulin-sensitive individuals, who were matched for body mass index (BMI). TNMD expression increases in human preadipocytes during differentiation, whereas silencing TNMD blocks adipogenesis. Upon high-fat diet feeding, transgenic mice overexpressing Tnmd develop increased epididymal white adipose tissue (eWAT) mass, and preadipocytes derived from Tnmd transgenic mice display greater proliferation, consistent with elevated adipogenesis. In Tnmd transgenic mice, lipogenic genes are upregulated in eWAT, as is Ucp1 in brown fat, while liver triglyceride accumulation is attenuated. Despite expanded eWAT, transgenic animals display improved systemic insulin sensitivity, decreased collagen deposition and inflammation in eWAT, and increased insulin stimulation of Akt phosphorylation. Our data suggest that TNMD acts as a protective factor in visceral adipose tissue to alleviate insulin resistance in obesity. PMID:26880110

  7. Tenomodulin promotes human adipocyte differentiation and beneficial visceral adipose tissue expansion.

    PubMed

    Senol-Cosar, Ozlem; Flach, Rachel J Roth; DiStefano, Marina; Chawla, Anil; Nicoloro, Sarah; Straubhaar, Juerg; Hardy, Olga T; Noh, Hye Lim; Kim, Jason K; Wabitsch, Martin; Scherer, Philipp E; Czech, Michael P

    2016-01-01

    Proper regulation of energy storage in adipose tissue is crucial for maintaining insulin sensitivity and molecules contributing to this process have not been fully revealed. Here we show that type II transmembrane protein tenomodulin (TNMD) is upregulated in adipose tissue of insulin-resistant versus insulin-sensitive individuals, who were matched for body mass index (BMI). TNMD expression increases in human preadipocytes during differentiation, whereas silencing TNMD blocks adipogenesis. Upon high-fat diet feeding, transgenic mice overexpressing Tnmd develop increased epididymal white adipose tissue (eWAT) mass, and preadipocytes derived from Tnmd transgenic mice display greater proliferation, consistent with elevated adipogenesis. In Tnmd transgenic mice, lipogenic genes are upregulated in eWAT, as is Ucp1 in brown fat, while liver triglyceride accumulation is attenuated. Despite expanded eWAT, transgenic animals display improved systemic insulin sensitivity, decreased collagen deposition and inflammation in eWAT, and increased insulin stimulation of Akt phosphorylation. Our data suggest that TNMD acts as a protective factor in visceral adipose tissue to alleviate insulin resistance in obesity. PMID:26880110

  8. Enhancing repair of full-thickness excisional wounds in a murine model: Impact of tissue-engineered biological dressings featuring human differentiated adipocytes.

    PubMed

    Morissette Martin, Pascal; Maux, Amandine; Laterreur, Véronique; Mayrand, Dominique; L Gagné, Valérie; Moulin, Véronique J; Fradette, Julie

    2015-08-01

    Promotion of skin repair for acute or chronic wounds through the use of tissue-engineered products is an active field of research. This study evaluates the effects mediated by tissue-engineered biological dressings containing human in vitro-differentiated adipocytes and adipose-derived stromal cells (ASCs). Re-epithelialization, granulation tissue formation and neovascularization of full-thickness cutaneous wounds were specifically assessed using a murine model featuring a fluorescent epidermis. In comparison with wounds that did not receive an adipocyte-containing biological dressing, treated wounds displayed a slight but significantly faster wound closure based on macroscopic observations over 18 days. Non-invasive imaging of GFP-expressing keratinocytes determined that the kinetics of re-epithelialization were similar for both groups. Treated wounds featured thicker granulation tissues (1.7-fold, P < 0.0001) enriched in collagens (1.3-fold, P < 0.0104). In addition, wound cryosections labeled for detection of CD31-expressing cells indicated a 2.2-fold (P < 0.0002) increased neovascularization for the treated wounds at the time of terminal biopsy. This is in accordance with the secretion of pro-angiogenic factors detected in media conditioned by the dressings. Taken together, these results establish that a new type of engineered substitutes featuring a mixture of adipocytes and ASCs can promote cutaneous healing when applied as temporary dressings, suggesting their potential relevance for chronic wound management studies. PMID:25934321

  9. FDP-E induces adipocyte inflammation and suppresses insulin-stimulated glucose disposal: effect of inflammation and obesity on fibrinogen Bβ mRNA.

    PubMed

    Kang, Minsung; Vaughan, Roger A; Paton, Chad M

    2015-12-01

    Obesity is associated with increased fibrinogen production and fibrin formation, which produces fibrin degradation products (FDP-E and FDP-D). Fibrin and FDPs both contribute to inflammation, which would be expected to suppress glucose uptake and insulin signaling in adipose tissue, yet the effect of FDP-E and FDP-D on adipocyte function and glucose disposal is completely unknown. We tested the effects of FDPs on inflammation in 3T3-L1 adipocytes and primary macrophages and adipocyte glucose uptake in vitro. High-fat-fed mice increased hepatic fibrinogen mRNA expression ninefold over chow-fed mice, with concomitant increases in plasma fibrinogen protein levels. Obese mice also displayed increased fibrinogen content of epididymal fat pads. We treated cultured 3T3-L1 adipocytes and primary macrophages with FDP-E, FDP-D, or fibrinogen degradation products (FgnDP-E). FDP-D and FgnDP-E had no effect on inflammation or glucose uptake. Cytokine mRNA expression in RAW264.7 macrophage-like cells and 3T3-L1 adipocytes treated with FDP-E induced inflammation with maximal effects at 100 nM and 6 h. Insulin-stimulated 2-deoxy-d-[(3)H]glucose uptake was reduced by 71% in adipocytes treated with FDP-E. FDP-E, but not FDP-D or FgnDP-E, induces inflammation in macrophages and adipocytes and decreases glucose uptake in vitro. FDP-E may contribute toward obesity-associated acute inflammation and glucose intolerance, although its chronic role in obesity remains to be elucidated. PMID:26447203

  10. Transgenic Rescue of Adipocyte Glucose-dependent Insulinotropic Polypeptide Receptor Expression Restores High Fat Diet-induced Body Weight Gain*

    PubMed Central

    Ugleholdt, Randi; Pedersen, Jens; Bassi, Maria Rosaria; Füchtbauer, Ernst-Martin; Jørgensen, Signe Marie; Kissow, Hanne-Louise; Nytofte, Nikolaj; Poulsen, Steen Seier; Rosenkilde, Mette Marie; Seino, Yutaka; Thams, Peter; Holst, Peter Johannes; Holst, Jens Juul

    2011-01-01

    The glucose-dependent insulinotropic polypeptide receptor (GIPr) has been implicated in high fat diet-induced obesity and is proposed as an anti-obesity target despite an uncertainty regarding the mechanism of action. To independently investigate the contribution of the insulinotropic effects and the direct effects on adipose tissue, we generated transgenic mice with targeted expression of the human GIPr to white adipose tissue or beta-cells, respectively. These mice were then cross-bred with the GIPr knock-out strain. The central findings of the study are that mice with GIPr expression targeted to adipose tissue have a similar high fat diet -induced body weight gain as control mice, significantly greater than the weight gain in mice with a general ablation of the receptor. Surprisingly, this difference was due to an increase in total lean body mass rather than a gain in total fat mass that was similar between the groups. In contrast, glucose-dependent insulinotropic polypeptide-mediated insulin secretion does not seem to be important for regulation of body weight after high fat feeding. The study supports a role of the adipocyte GIPr in nutrient-dependent regulation of body weight and lean mass, but it does not support a direct and independent role for the adipocyte or beta-cell GIPr in promoting adipogenesis. PMID:22027838

  11. Fucoxanthinol, Metabolite of Fucoxanthin, Improves Obesity-Induced Inflammation in Adipocyte Cells.

    PubMed

    Maeda, Hayato; Kanno, Shogo; Kodate, Mei; Hosokawa, Masashi; Miyashita, Kazuo

    2015-08-01

    Fucoxanthin (Fx) is a marine carotenoid found in edible brown seaweeds. We previously reported that dietary Fx metabolite into fucoxanthinol (FxOH), attenuates the weight gain of white adipose tissue of diabetic/obese KK-Ay mice. In this study, to evaluate anti-diabetic effects of Fx, we investigated improving the effect of insulin resistance on the diabetic model of KK-Ay mice. Furthermore, preventing the effect of FxOH on low-grade chronic inflammation related to oxidative stress was evaluated on 3T3-L1 adipocyte cells and a RAW264.7 macrophage cell co-culture system. A diet containing 0.1% Fx was fed to diabetic model KK-Ay mice for three weeks, then glucose tolerance was observed. Fx diet significantly improved glucose tolerance compared with the control diet group.  In in vitro studies, FxOH showed suppressed tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1) mRNA expression and protein levels in a co-culture of adipocyte and macrophage cells. These findings suggest that Fx ameliorates glucose tolerance in the diabetic model mice. Furthermore, FxOH, a metabolite of Fx, suppresses low-grade chronic inflammation in adipocyte cells. PMID:26248075

  12. Tomato extract suppresses the production of proinflammatory mediators induced by interaction between adipocytes and macrophages.

    PubMed

    Kim, Young-il; Mohri, Shinsuke; Hirai, Shizuka; Lin, Shan; Goto, Tsuyoshi; Ohyane, Chie; Sakamoto, Tomoya; Takahashi, Haruya; Shibata, Daisuke; Takahashi, Nobuyuki; Kawada, Teruo

    2015-01-01

    Obese adipose tissue is characterized by enhanced macrophage infiltration. A loop involving monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNFα) between adipocytes and macrophages establishes a vicious cycle that augments inflammatory changes and insulin resistance in obese adipose tissue. Tomatoes, one of the most popular crops worldwide, contain many beneficial phytochemicals that improve obesity-related diseases such as diabetes. Some of them have also been reported to have anti-inflammatory properties. In this study, we focused on the potential protective effects of phytochemicals in tomatoes on inflammation. We screened fractions of tomato extract using nitric oxide (NO) assay in lipopolysaccharide (LPS)-stimulated RAW264 macrophages. One fraction, RF52, significantly inhibited NO production in LPS-stimulated RAW264 macrophages. Furthermore, RF52 significantly decreased MCP-1 and TNFα productions. The coculture of 3T3-L1 adipocytes and RAW264 macrophages markedly enhanced MCP-1, TNFα, and NO productions compared with the control cultures; however, the treatment with RF52 inhibited the production of these proinflammatory mediators. These results suggest that RF52 from tomatoes may have the potential to suppress inflammation by inhibiting the production of NO or proinflammatory cytokines during the interaction between adipocytes and macrophages. PMID:25603813

  13. Fucoxanthinol, Metabolite of Fucoxanthin, Improves Obesity-Induced Inflammation in Adipocyte Cells

    PubMed Central

    Maeda, Hayato; Kanno, Shogo; Kodate, Mei; Hosokawa, Masashi; Miyashita, Kazuo

    2015-01-01

    Fucoxanthin (Fx) is a marine carotenoid found in edible brown seaweeds. We previously reported that dietary Fx metabolite into fucoxanthinol (FxOH), attenuates the weight gain of white adipose tissue of diabetic/obese KK-Ay mice. In this study, to evaluate anti-diabetic effects of Fx, we investigated improving the effect of insulin resistance on the diabetic model of KK-Ay mice. Furthermore, preventing the effect of FxOH on low-grade chronic inflammation related to oxidative stress was evaluated on 3T3-L1 adipocyte cells and a RAW264.7 macrophage cell co-culture system. A diet containing 0.1% Fx was fed to diabetic model KK-Ay mice for three weeks, then glucose tolerance was observed. Fx diet significantly improved glucose tolerance compared with the control diet group. In in vitro studies, FxOH showed suppressed tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1) mRNA expression and protein levels in a co-culture of adipocyte and macrophage cells. These findings suggest that Fx ameliorates glucose tolerance in the diabetic model mice. Furthermore, FxOH, a metabolite of Fx, suppresses low-grade chronic inflammation in adipocyte cells. PMID:26248075

  14. Gene regulation in β-sitosterol-mediated stimulation of adipogenesis, glucose uptake, and lipid mobilization in rat primary adipocytes.

    PubMed

    Chai, Jen-Wai; Lim, Siang-Ling; Kanthimathi, M S; Kuppusamy, Umah Rani

    2011-05-01

    The nutraceutical benefits of β-sitosterol (SIT) are well documented. The present study investigated the in vitro effects of SIT on adipogenesis, glucose transport, and lipid mobilization in rat adipocytes. Primary cultures of rat preadipocytes and differentiated adipocytes were used in this study. Glucose uptake was measured by the uptake of radio-labeled glucose. Adipogenesis and lipolysis were measured by oil-red-O and glycerol quantification methods, respectively. The expression of protein kinase B (Akt), glucose transporter 4 (GLUT4), hormone sensitive lipase (HSL), and phosphatidylinositol-3-kinase (PI3 K) genes in SIT-treated adipocytes were assessed by real-time reverse transcription polymerase chain reaction (RT-PCR). The data showed that SIT induced glucose uptake in adipocytes. It also stimulated adipogenesis in differentiating preadipocytes. Interestingly, although SIT displayed general insulin-mimetic activity by stimulating glucose uptake and adipogenesis, it also induced lipolysis in adipocytes. Furthermore, the SIT-induced lipolysis was not attenuated by insulin and co-incubation of SIT with epinephrine improved epinephrine-induced lipolysis. GLUT4 gene expression was highly down-regulated in SIT-treated adipocytes, compared to insulin-treated adipocytes, which was up-regulated. Insulin- and SIT-treated adipocytes showed similar levels of Akt, HSL, and PI3 K gene down-regulation. These observations suggest that the elevation of glucose uptake in SIT-treated adipocytes was unrelated to de novo synthesis of GLUT4 and the SIT-induced lipolysis is associated with the down-regulation of Akt and PI3K genes. The unique effects of SIT on the regulation of glucose uptake, adipogenesis, and lipolysis in adipocytes show that it has potential to be utilized in diabetes and weight management. PMID:21484150

  15. CL316243 induces phosphatidylinositol 3,4,5-triphosphate production in rat adipocytes in an adenosine deaminase-, pertussis toxin-, or wortmannin-sensitive manner.

    PubMed

    Ohsaka, Y; Nomura, Y

    2016-07-18

    The effect of beta(3)-adrenoceptor (beta(3)-AR) agonists on adipocytes treated or not treated with signaling modulators has not been sufficiently elucidated. Using rat epididymal adipocytes (adipocytes) labeled with [(32)P]orthophosphate, we found that treatment with the selective beta(3)-AR agonist CL316243 (CL; 1 microM) induces phosphatidylinositol (PI) 3,4,5-triphosphate (PI[3,4,5]P(3)) production and that this response is inhibited by adenosine deaminase (ADA, an adenosine-degrading enzyme; 2 U/ml), pertussis toxin (PTX, an inactivator of inhibitory guanine-nucleotide-binding protein; 1 microg/ml), or wortmannin (WT, a PI-kinase inhibitor; 3 microM). The results showed that CL induced PI(3,4,5)P(3) production in intact adipocytes and that this production was affected by signaling modulators. Taken together, our findings indicate that CL produces PI(3,4,5)P(3) in an ADA-sensitive, PTX-sensitive, or WT-sensitive manner and will advance understanding of the effect of beta(3)-AR agonists on adipocytes. PMID:26988163

  16. Uric acid induces oxidative stress via an activation of the renin-angiotensin system in 3T3-L1 adipocytes.

    PubMed

    Zhang, Jun-xia; Zhang, Yu-ping; Wu, Qi-nan; Chen, Bing

    2015-02-01

    Hyperuricemia is recently reported involving in various obesity-related cardiovascular disorders, especially hypertension. However, the underlying mechanisms are not completely understood. In the present study, we investigated whether uric acid upregulates renin-angiotensin system (RAS) expression in adipocytes. We also examined whether RAS activation plays a role in uric acid-induced oxidative stress in adipocytes. The adipocytes of different phenotypes were incubated with uric acid for 48 h, respectively. Losartan (10(-4) M) or captopril (10(-4) M) was used to block adipose tissue RAS activation. mRNA expressions of angiotensinogen (AGT), angiotensin-converting enzyme-1 (ACE-1), renin, angiotensin type 1 receptor (AT1R), and angiotensin type 2 receptor (AT2R) were evaluated with real-time PCR. Angiotensin II concentrations in supernatant were measured by ELISA. Intracellular reactive species (ROS) levels were measured by fluorescent probe DCFH-DA, DHR, or NBT assay. The uric acid upregulated both RAS (AGT, ACE1, renin, AT1R, and AT2R) mRNA expressions and angiotensin II protein secretion and caused a significant increase in ROS production in 3T3-L1 adipocytes. These effects could be prevented by RAS inhibitors, either losartan or captopril. RAS activation has been causally implicated in oxidative stress induced by uric acid in 3T3-L1 adipocytes, suggesting a plausible mechanism through which hyperuricemia contributes to the pathogenesis of obesity-related cardiovascular diseases. PMID:24671741

  17. Non-transactivational, dual pathways for LPA-induced Erk1/2 activation in primary cultures of brown pre-adipocytes

    SciTech Connect

    Holmstroem, Therese E.; Mattsson, Charlotte L.; Wang, Yanling; Iakovleva, Irina; Petrovic, Natasa; Nedergaard, Jan

    2010-10-01

    In many cell types, G-protein-coupled receptor (GPCR)-induced Erk1/2 MAP kinase activation is mediated via receptor tyrosine kinase (RTK) transactivation, in particular via the epidermal growth factor (EGF) receptor. Lysophosphatidic acid (LPA), acting via GPCRs, is a mitogen and MAP kinase activator in many systems, and LPA can regulate adipocyte proliferation. The mechanism by which LPA activates the Erk1/2 MAP kinase is generally accepted to be via EGF receptor transactivation. In primary cultures of brown pre-adipocytes, EGF can induce Erk1/2 activation, which is obligatory and determinant for EGF-induced proliferation of these cells. Therefore, we have here examined whether LPA, via EGF transactivation, can activate Erk1/2 in brown pre-adipocytes. We found that LPA could induce Erk1/2 activation. However, the LPA-induced Erk1/2 activation was independent of transactivation of EGF receptors (or PDGF receptors) in these cells (whereas in transformed HIB-1B brown adipocytes, the LPA-induced Erk1/2 activation indeed proceeded via EGF receptor transactivation). In the brown pre-adipocytes, LPA instead induced Erk1/2 activation via two distinct non-transactivational pathways, one G{sub i}-protein dependent, involving PKC and Src activation, the other, a PTX-insensitive pathway, involving PI3K (but not Akt) activation. Earlier studies showing LPA-induced Erk1/2 activation being fully dependent on RTK transactivation have all been performed in cell lines and transfected cells. The present study implies that in non-transformed systems, RTK transactivation may not be involved in the mediation of GPCR-induced Erk1/2 MAP kinase activation.

  18. Citrus flavonoid naringenin inhibits TLR2 expression in adipocytes.

    PubMed

    Yoshida, Hiroki; Watanabe, Wataru; Oomagari, Hiroyuki; Tsuruta, Eisuke; Shida, Mikiko; Kurokawa, Masahiko

    2013-07-01

    Toll-like receptors (TLRs) were recently shown to be involved in obesity-induced inflammation in adipose tissue, which contributes to the development of insulin resistance and type 2 diabetes. Thus, the appropriate regulation of TLR expression or activation is an important strategy for improving obesity-related diseases. In this report, we show that naringenin, a citrus flavonoid, inhibits TLR2 expression during adipocyte differentiation. This effect is mediated in part through peroxisome proliferator-activated receptor γ activation. In addition, naringenin suppresses TLR2 expression induced by the co-culture of differentiated adipocytes and macrophages and also inhibits tumor necrosis factor-α (TNF-α)-induced TLR2 expression by inhibiting the activation of nuclear factor-κB and c-Jun NH2-terminal kinase pathways in differentiated adipocytes. Furthermore, naringenin decreases TLR2 expression in adipose tissue of high-fat diet-fed mice. These results are correlated with the improvement of hyperglycemia and the suppression of inflammatory mediators, including TNF-α and monocyte chemotactic protein-1. Taken together, these data suggest that naringenin exhibits anti-inflammatory properties, presumably by inhibiting TLR2 expression in adipocytes. Our findings suggest a molecular mechanism by which naringenin exerts beneficial effects against obesity-related diseases. PMID:23333096

  19. Effects of Black Adzuki Bean (Vigna angularis) Extract on Proliferation and Differentiation of 3T3-L1 Preadipocytesinto Mature Adipocytes

    PubMed Central

    Kim, Mina; Park, Jeong-Eun; Song, Seok-Bo; Cha, Youn-Soo

    2015-01-01

    The aim of this work was to investigate the effects of black adzuki bean (BAB) extract on adipocytes, and to elucidate the cellular mechanisms. In order to examine the proliferation of preadipocytes and differentiating adipocytes, cell viability and DNA content were measured over a period of time. Lipid accumulation during cell differentiation and the molecular mechanisms underlying the effects of BAB on the transcriptional factors involved, with their anti-adipogenic effects, were also identified. We observed that BAB exhibits anti-adipogenic effects through the inhibition of proliferation, thereby lowering mRNA expression of C/EBPβ and suppressing adipogenesis during the early stage of differentiation. This, in turn, resulted in a reduction of TG accumulation in a dose- and time-dependent manner. Treating the cells with BAB not only suppressed the adipogenesis-associated key transcription factors PPARγ and C/EBPα but also significantly decreased the mRNA expression of GLUT4, FABP4, LPL and adiponectin. The expression of lipolytic genes like ATGL and HSL were higher in the treatment group than in the control. Overall, the black adzuki bean extract demonstrated an anti-adipogenic property, which makes it a potential dietary supplement for attenuation of obesity. PMID:25569623

  20. α-Mangostin Improves Glucose Uptake and Inhibits Adipocytes Differentiation in 3T3-L1 Cells via PPARγ, GLUT4, and Leptin Expressions

    PubMed Central

    Taher, Muhammad; Mohamed Amiroudine, Mohamed Zaffar Ali; Tengku Zakaria, Tengku Muhamad Faris Syafiq; Ichwan, Solachuddin J. A.; Kaderi, Mohd Arifin; Ahmed, Qamar Uddin; Zakaria, Zainul Amiruddin

    2015-01-01

    Obesity has been often associated with the occurrence of cardiovascular diseases, type 2 diabetes, and cancer. The development of obesity is also accompanied by significant differentiation of preadipocytes into adipocytes. In this study, we investigated the activity of α-mangostin, a major xanthone component isolated from the stem bark of G. malaccensis, on glucose uptake and adipocyte differentiation of 3T3-L1 cells focusing on PPARγ, GLUT4, and leptin expressions. α-Mangostin was found to inhibit cytoplasmic lipid accumulation and adipogenic differentiation. Cells treated with 50 μM of α-mangostin reduced intracellular fat accumulation dose-dependently up to 44.4% relative to MDI-treated cells. Analyses of 2-deoxy-D-[3H] glucose uptake activity showed that α-mangostin significantly improved the glucose uptake (P < 0.05) with highest activity found at 25 μM. In addition, α-mangostin increased the amount of free fatty acids (FFA) released. The highest glycerol release level was observed at 50 μM of α-mangostin. qRT-PCR analysis showed reduced lipid accumulation via inhibition of PPARγ gene expression. Induction of glucose uptake and free fatty acid release by α-mangostin were accompanied by increasing mRNA expression of GLUT4 and leptin. These evidences propose that α-mangostin might be possible candidate for the effective management of obesity in future. PMID:25873982

  1. Uric Acid-Induced Adipocyte Dysfunction Is Attenuated by HO-1 Upregulation: Potential Role of Antioxidant Therapy to Target Obesity

    PubMed Central

    Sodhi, Komal; Hilgefort, Jordan; Banks, George; Gilliam, Chelsea; Stevens, Sarah; Ansinelli, Hayden A.; Getty, Morghan; Abraham, Nader G.; Shapiro, Joseph I.

    2016-01-01

    Increased uric acid levels have been implicated in the pathogenesis of metabolic syndrome. To examine the mechanisms by which this occurs, we hypothesized that an increase in heme oxygenase 1, a potent antioxidant gene, will decrease uric acid levels and adipocyte dysfunction via suppression of ROS and xanthine oxidase (XO) levels. We examined the effect of uric acid on adipogenesis in human mesenchymal stem cells (MSCs) in the presence and absence of cobalt protoporphyrin (CoPP), an HO-1 inducer, and tin mesoporphyrin (SnMP), an HO activity inhibitor. Uric acid increased adipogenesis by increasing NADPH oxidase expression and elevation in the adipogenesis markers C/EBPα, PPARγ, and Mest, while decreasing small lipid droplets and Wnt10b levels. We treated MSCs with fructose, a fuel source that increases uric acid levels. Our results showed that fructose increased XO expression as compared to the control and concomitant treatment with CoPP significantly decreased XO expression and uric acid levels. These beneficial effects of CoPP were reversed by SnMP, supporting a role for HO activity in mediating these effects. These findings demonstrate that increased levels of HO-1 appear crucial in modulating the phenotype of adipocytes exposed to uric acid and in downregulating XO and NADPH oxidase levels. PMID:26681956

  2. Uric Acid-Induced Adipocyte Dysfunction Is Attenuated by HO-1 Upregulation: Potential Role of Antioxidant Therapy to Target Obesity.

    PubMed

    Sodhi, Komal; Hilgefort, Jordan; Banks, George; Gilliam, Chelsea; Stevens, Sarah; Ansinelli, Hayden A; Getty, Morghan; Abraham, Nader G; Shapiro, Joseph I; Khitan, Zeid

    2016-01-01

    Increased uric acid levels have been implicated in the pathogenesis of metabolic syndrome. To examine the mechanisms by which this occurs, we hypothesized that an increase in heme oxygenase 1, a potent antioxidant gene, will decrease uric acid levels and adipocyte dysfunction via suppression of ROS and xanthine oxidase (XO) levels. We examined the effect of uric acid on adipogenesis in human mesenchymal stem cells (MSCs) in the presence and absence of cobalt protoporphyrin (CoPP), an HO-1 inducer, and tin mesoporphyrin (SnMP), an HO activity inhibitor. Uric acid increased adipogenesis by increasing NADPH oxidase expression and elevation in the adipogenesis markers C/EBPα, PPARγ, and Mest, while decreasing small lipid droplets and Wnt10b levels. We treated MSCs with fructose, a fuel source that increases uric acid levels. Our results showed that fructose increased XO expression as compared to the control and concomitant treatment with CoPP significantly decreased XO expression and uric acid levels. These beneficial effects of CoPP were reversed by SnMP, supporting a role for HO activity in mediating these effects. These findings demonstrate that increased levels of HO-1 appear crucial in modulating the phenotype of adipocytes exposed to uric acid and in downregulating XO and NADPH oxidase levels. PMID:26681956

  3. Insulin-induced decrease in protein phosphorylation in rat adipocytes not explained by decreased A-kinase activity

    SciTech Connect

    Egan, J.J.; Greenberg, A.S.; Chang, M.K.; Londos, C.

    1987-05-01

    In isolated rat adipocytes, insulin inhibits lipolysis to a greater extent than would be predicted by the decrease in (-/+)cAMP activity ratio of cAMP-dependent protein kinase (A-kinase), from which it was speculated that insulin promotes the dephosphorylation of hormone-sensitive lipase. They have examined the phosphorylation state of cellular proteins under conditions of varying A-kinase activities in the presence and absence of insulin. Protein phosphorylation was determined by SDS-PAGE electrophoresis of extracts from /sup 32/P-loaded cells; glycerol and A-kinase activity ratios were measured in the cytosolic extracts from control, non-radioactive cells. Increased protein phosphorylation in general occurred over the same range of A-kinase activity ratios, 0.1-0.3, associated with increased glycerol release. The insulin-induced decrease in lipolysis was associated with a decrease in the /sup 32/P content of several proteins, an effect not explained by the modest reduction in A-kinase activity by insulin. This effect of insulin on protein phosphorylation was lost as the A-kinase activity ratios exceeded 0.5. The results suggest that insulin promotes the dephosphorylation of those adipocyte proteins which are subject to phosphorylation by A-kinase.

  4. Differential signalling pathways for EGF versus PDGF activation of Erk1/2 MAP kinase and cell proliferation in brown pre-adipocytes

    SciTech Connect

    Holmstroem, Therese E.; Mattsson, Charlotte L.; Faelting, Johanna M.; Nedergaard, Jan

    2008-11-15

    Stimulation by both adrenergic and non-adrenergic pathways can induce proliferation of brown pre-adipocytes. To understand the signalling pathways involved in non-adrenergic stimulation of cell proliferation, we examined Erk1/2 activation. In primary cultures of mouse brown pre-adipocytes, both EGF (epidermal growth factor) and PDGF (platelet-derived growth factor) induced Erk1/2 activation. EGF-stimulated Erk1/2 activation involved Src tyrosine kinases, but not PKC or PI3K, whereas in PDGF-induced Erk1/2 activation, PI3K, PKC (probably the atypical {zeta} isoform) and Src were involved sequentially. Both EGF and PDGF induced PI3K-dependent Akt activation that was not involved in Erk1/2 activation. By comparing effects of signalling inhibitors (wortmannin, SH-6, TPA, Goe6983, PP2, PD98059) on EGF- and PDGF-induced Erk1/2 activation and cell proliferation ({sup 3}H-thymidine incorporation), we conclude that while the signal transduction pathways initiated by these growth factors are clearly markedly different, their effects on cell proliferation can be fully explained through their stimulation of Erk1/2 activation; thus Erk1/2 is a common, essential step for stimulation of proliferation in these cells.

  5. Coordinate Functional Regulation between Microsomal Prostaglandin E Synthase-1 (mPGES-1) and Peroxisome Proliferator-activated Receptor γ (PPARγ) in the Conversion of White-to-brown Adipocytes*

    PubMed Central

    García-Alonso, Verónica; López-Vicario, Cristina; Titos, Esther; Morán-Salvador, Eva; González-Périz, Ana; Rius, Bibiana; Párrizas, Marcelina; Werz, Oliver; Arroyo, Vicente; Clària, Joan

    2013-01-01

    Peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-activated nuclear receptor and a master regulator of adipogenesis. Microsomal prostaglandin E (PGE) synthase-1 (mPGES-1) is an inducible enzyme that couples with cyclooxygenase-2 for the biosynthesis of PGE2. In this study we demonstrate the existence of a coordinate functional interaction between PPARγ and mPGES-1 in controlling the process of pre-adipocyte differentiation in white adipose tissue (WAT). Adipocyte-specific PPARγ knock-out mice carrying an aP2 promoter-driven Cre recombinase transgene showed a blunted response to the adipogenic effects of a high fat diet. Pre-adipocytes from these knock-out mice showed loss of PPARγ and were resistant to rosiglitazone-induced WAT differentiation. In parallel, WAT from these mice showed increased expression of uncoupling protein 1, a mitochondrial enzyme that dissipates chemical energy as heat. Adipose tissue from mice lacking PPARγ also showed mPGES-1 up-regulation and increased PGE2 levels. In turn, PGE2 suppressed PPARγ expression and blocked rosiglitazone-induced pre-adipocyte differentiation toward white adipocytes while directly elevating uncoupling protein 1 expression and pre-adipocyte differentiation into mature beige/brite adipocytes. Consistently, pharmacological mPGES-1 inhibition directed pre-adipocyte differentiation toward white adipocytes while suppressing differentiation into beige/brite adipocytes. This browning effect was reproduced in knockdown experiments using a siRNA directed against mPGES-1. The effects of PGE2 on pre-adipocyte differentiation were not seen in mice lacking PPARγ in adipose tissue and were not mirrored by other eicosanoids (i.e. leukotriene B4). Taken together, these findings identify PGE2 as a key regulator of white-to-brown adipogenesis and suggest the existence of a coordinate regulation of adipogenesis between PPARγ and mPGES-1. PMID:23943621

  6. Impact of 3-Amino-1,2,4-Triazole (3-AT)-Derived Increase in Hydrogen Peroxide Levels on Inflammation and Metabolism in Human Differentiated Adipocytes

    PubMed Central

    Ruiz-Ojeda, Francisco Javier; Gomez-Llorente, Carolina; Aguilera, Concepción María; Gil, Angel; Rupérez, Azahara Iris

    2016-01-01

    Obesity is characterized by an excessive accumulation of fat in adipose tissue, which is associated with oxidative stress and chronic inflammation. Excessive H2O2 levels are degraded by catalase (CAT), the activity of which is decreased in obesity. We investigated the effects of inhibition of catalase activity on metabolism and inflammation by incubating human differentiated adipocytes with 10 mM 3-amino-1,2,4-triazole (3-AT) for 24 h. As expected, the treatment decreased CAT activity and increased intracellular H2O2 levels significantly. Glutathione peroxidase (GPX) activity was also reduced, and the gene expression levels of the antioxidant enzymes GPX4 and peroxiredoxins (1, 3 and 5) were inhibited. Interestingly, this occurred along with lower mRNA levels of the transcription factors nuclear factor (erythroid 2-like 2) and forkhead box O, which are involved in redox homeostasis. However, superoxide dismutase activity and expression were increased. Moreover, 3-AT led to nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation and increased tumor necrosis alpha and interleukin 6 protein and gene expression levels, while lowering peroxisome proliferator-activated receptor gamma (PPARγ) mRNA and protein levels. These alterations were accompanied by an altered glucose and lipid metabolism. Indeed, adipocytes treated with 3-AT showed reduced basal glucose uptake, reduced glucose transporter type 4 gene and protein expression, reduced lipolysis, reduced AMP-activated protein kinase activation and reduced gene expression of lipases. Our results indicate that increased H2O2 levels caused by 3-AT treatment impair the antioxidant defense system, lower PPARγ expression and initiate inflammation, thus affecting glucose and lipid metabolism in human differentiated adipocytes. PMID:27023799

  7. Impact of 3-Amino-1,2,4-Triazole (3-AT)-Derived Increase in Hydrogen Peroxide Levels on Inflammation and Metabolism in Human Differentiated Adipocytes.

    PubMed

    Ruiz-Ojeda, Francisco Javier; Gomez-Llorente, Carolina; Aguilera, Concepción María; Gil, Angel; Rupérez, Azahara Iris

    2016-01-01

    Obesity is characterized by an excessive accumulation of fat in adipose tissue, which is associated with oxidative stress and chronic inflammation. Excessive H2O2 levels are degraded by catalase (CAT), the activity of which is decreased in obesity. We investigated the effects of inhibition of catalase activity on metabolism and inflammation by incubating human differentiated adipocytes with 10 mM 3-amino-1,2,4-triazole (3-AT) for 24 h. As expected, the treatment decreased CAT activity and increased intracellular H2O2 levels significantly. Glutathione peroxidase (GPX) activity was also reduced, and the gene expression levels of the antioxidant enzymes GPX4 and peroxiredoxins (1, 3 and 5) were inhibited. Interestingly, this occurred along with lower mRNA levels of the transcription factors nuclear factor (erythroid 2-like 2) and forkhead box O, which are involved in redox homeostasis. However, superoxide dismutase activity and expression were increased. Moreover, 3-AT led to nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation and increased tumor necrosis alpha and interleukin 6 protein and gene expression levels, while lowering peroxisome proliferator-activated receptor gamma (PPARγ) mRNA and protein levels. These alterations were accompanied by an altered glucose and lipid metabolism. Indeed, adipocytes treated with 3-AT showed reduced basal glucose uptake, reduced glucose transporter type 4 gene and protein expression, reduced lipolysis, reduced AMP-activated protein kinase activation and reduced gene expression of lipases. Our results indicate that increased H2O2 levels caused by 3-AT treatment impair the antioxidant defense system, lower PPARγ expression and initiate inflammation, thus affecting glucose and lipid metabolism in human differentiated adipocytes. PMID:27023799

  8. Aquaporin-10 Represents an Alternative Pathway for Glycerol Efflux from Human Adipocytes

    PubMed Central

    Laforenza, Umberto; Scaffino, Manuela F.; Gastaldi, Giulia

    2013-01-01

    Background Glycerol outflow from adipocytes has been considered for a decade to be mediated by aquaporin-7, an aquaglyceroporin highly expressed in the adipose tissue. Its involvement in glycerol metabolism has been widely studied also in humans. Recent studies in different aquaporin-7 KO mice models pose two different questions 1) the exact localization of aquaporin-7 in human white adipose tissue; 2) the existence of other aquaglyceroporins that work with aquaporin-7 to guarantee glycerol efflux and thus a normal adiposity in humans. To this purpose we investigated the expression, the localization and the functioning of aquaglyceroporin-10 in subcutaneous white adipose tissue, in isolated and cultured differentiated adipocytes. Methodology/Principal Findings Aquaporin-7 and -10 were expressed in the white adipose tissue both at mRNA and at protein level. Immunofluorescence revealed aquaporin-7 and -10 labelling in the human adipose tissue both to the plasma membrane and to a thin rim of cytoplasm of adipocytes. Aquaporin-7, but not aquaporin-10, colocalized with the endothelial marker CD34. Human cultured differentiated adipocytes showed an aquaporin-7 and -10 labelling mainly in the cytoplasm and in the lipid droplets with insulin reinforcing the lipid droplets staining and isoproterenol inducing its translocation to the plasma membrane compartment. Water and glycerol permeability measurements using adipocytes and adipose membrane vesicles confirmed the presence of functioning aquaglyceroporins. Aquaporin-10 silencing in human differentiated adipocytes resulted in a 50% decrease of glycerol and osmotic water permeability. Conclusions/Significance The results indicate that aquaporin-7, differently from mice, is present in both adipocyte and capillary plasma membranes of human adipose tissue. Aquaporin-10, on the contrary, is expressed exclusively in the adipocytes. The expression of two aquaglyceroporins in human adipose tissue is particularly important for the

  9. Bmp4 Promotes a Brown to White-like Adipocyte Shift.

    PubMed

    Modica, Salvatore; Straub, Leon G; Balaz, Miroslav; Sun, Wenfei; Varga, Lukas; Stefanicka, Patrik; Profant, Milan; Simon, Eric; Neubauer, Heike; Ukropcova, Barbara; Ukropec, Jozef; Wolfrum, Christian

    2016-08-23

    While Bmp4 has a well-established role in the commitment of mesenchymal stem cells into the adipogenic lineage, its role in brown adipocyte formation and activity is not well defined. Here, we show that Bmp4 has a dual function in adipogenesis by inducing adipocyte commitment while inhibiting the acquisition of a brown phenotype during terminal differentiation. Selective brown adipose tissue overexpression of Bmp4 in mice induces a shift from a brown to a white-like adipocyte phenotype. This effect is mediated by Smad signaling and might be in part due to suppression of lipolysis, via regulation of hormone sensitive lipase expression linked to reduced Ppar activity. Given that we observed a strong correlation between BMP4 levels and adipocyte size, as well as insulin sensitivity in humans, we propose that Bmp4 is an important factor in the context of obesity and type 2 diabetes. PMID:27524617

  10. Direct action of capsaicin in brown adipogenesis and activation of brown adipocytes.

    PubMed

    Kida, Ryosuke; Yoshida, Hirofumi; Murakami, Masaru; Shirai, Mitsuyuki; Hashimoto, Osamu; Kawada, Teruo; Matsui, Tohru; Funaba, Masayuki

    2016-01-01

    The ingestion of capsaicin, the principle pungent component of red and chili peppers, induces thermogenesis, in part, through the activation of brown adipocytes expressing genes related to mitochondrial biogenesis and uncoupling such as peroxisome proliferator-activated receptor (Ppar) γ coactivator-1α (Pgc-1α) and uncoupling protein 1 (Ucp1). Capsaicin has been suggested to induce the activation of brown adipocytes, which is mediated by the stimulation of sympathetic nerves. However, capsaicin may directly affect the differentiation of brown preadipocytes, brown adipocyte function, or both, through its significant absorption. We herein demonstrated that Trpv1, a capsaicin receptor, is expressed in brown adipose tissue, and that its expression level is increased during the differentiation of HB2 brown preadipocytes. Furthermore, capsaicin induced calcium influx in brown preadipocytes. A treatment with capsaicin in the early stage of brown adipogenesis did not affect lipid accumulation or the expression levels of Fabp4 (a gene expressed in mature adipocytes), Pparγ2 (a master regulator of adipogenesis) or brown adipocyte-selective genes. In contrast, a treatment with capsaicin in the late stage of brown adipogenesis slightly increased the expression levels of Fabp4, Pparγ2 and Pgc-1α. Although capsaicin did not affect the basal expression level of Ucp1, Ucp1 induction by forskolin was partially inhibited by capsaicin, irrespective of the dose of capsaicin. The results of the present study suggest the direct effects of capsaicin on brown adipocytes or in the late stage of brown adipogenesis. PMID:26781688

  11. Identification of novel PPAR{gamma} target genes by integrated analysis of ChIP-on-chip and microarray expression data during adipocyte differentiation

    SciTech Connect

    Nakachi, Yutaka; Yagi, Ken; Nikaido, Itoshi; Bono, Hidemasa; Tonouchi, Mio; Schoenbach, Christian; Okazaki, Yasushi

    2008-07-25

    PPAR{gamma} (peroxisome proliferator-activated receptor gamma) acts as a key molecule of adipocyte differentiation, and transactivates multiple target genes involved in lipid metabolic pathways. Identification of PPAR{gamma} target genes will facilitate to predict the extent to which the drugs can affect and also to understand the molecular basis of lipid metabolism. Here, we have identified five target genes regulated directly by PPAR{gamma} during adipocyte differentiation in 3T3-L1 cells using integrated analyses of ChIP-on-chip and expression microarray. We have confirmed the direct PPAR{gamma} regulation of five genes by luciferase reporter assay in NIH-3T3 cells. Of these five genes Hp, Tmem143 and 1100001G20Rik are novel PPAR{gamma} targets. We have also detected PPREs (PPAR response elements) sequences in the promoter region of the five genes computationally. Unexpectedly, most of the PPREs detected proved to be atypical, suggesting the existence of more atypical PPREs than previously thought in the promoter region of PPAR{gamma} regulated genes.

  12. Enhancer of Zeste Homolog 2 and Histone Deacetylase 9c Regulate Age-Dependent Mesenchymal Stem Cell Differentiation into Osteoblasts and Adipocytes.

    PubMed

    Chen, Ya-Huey; Chung, Chiao-Chen; Liu, Yu-Chia; Yeh, Su-Peng; Hsu, Jennifer L; Hung, Mien-Chie; Su, Hong-Lin; Li, Long-Yuan

    2016-08-01

    Mesenchymal stem cells (MSCs) are multipotent precursors that can undergo multilineage differentiation, including osteogenesis and adipogenesis, which are two mutually exclusive events. Previously, we demonstrated that enhancer of zeste homolog 2 (EZH2), the catalytic component of the Polycomb-repressive complex 2, mediates epigenetic silencing of histone deacetylase 9c (HDAC9c) in adipocytes but not in osteoblasts and that HDAC9c accelerates osteogenesis while attenuating adipogenesis of MSCs through inactivation of peroxisome proliferator-activated receptor gamma 2 activity. Importantly, disrupting the balance between adipogenesis and osteogenesis can lead to age-associated bone loss (osteoporosis) and obesity. Here, we investigated the relationship between age, and osteogenic and adipogenic differentiation potential of MSCs by comparing EZH2 and HDAC9c expression in osteoblasts and adipocytes of both human and mice origins to determine whether the EZH2-HDAC9c axis regulates age-associated osteoporosis and obesity. Our findings indicated that a decline in HDAC9c expression over time was accompanied by increased EZH2 expression and suggested that a therapeutic intervention for age-associated osteoporosis and obesity may be feasible by targeting the EZH2-HDAC9c axis. Stem Cells 2016;34:2183-2193. PMID:27250566

  13. New vinegar produced by tomato suppresses adipocyte differentiation and fat accumulation in 3T3-L1 cells and obese rat model.

    PubMed

    Lee, Ju-Hye; Cho, Hyun-Dong; Jeong, Ji-Hye; Lee, Mi-Kyung; Jeong, Yong-Ki; Shim, Ki-Hwan; Seo, Kwon-Il

    2013-12-01

    There is an increasing surplus of tomatoes that are abandoned due to their failure to meet customer standards. Therefore, to allow both value additions and the effective reuse of surplus tomatoes, we developed tomato vinegar (TV) containing phytochemicals and evaluated its anti-obesity effects in vitro and in vivo. TV inhibited adipocyte differentiation of 3T3-L1 preadipocyte and lipid accumulation during differentiation. TV supplementation in rats fed a high-fat diet (HFD) markedly decreased visceral fat weights without changing the food and calories intakes. TV significantly decreased hepatic triglyceride and cholesterol levels compared to the HFD group. Furthermore, TV lowered plasma LDL-cholesterol level and antherogenic index compared to the HFD group, whereas elevated HDL-cholesterol to total cholesterol ratio. These results show that TV prevented obesity by suppressing visceral fat and lipid accumulation in adipocyte and obese rats, and suggest that TV can be used as an anti-obesity therapeutic agent or functional food. PMID:23871083

  14. IL-17A synergistically enhances TNFα-induced IL-6 and CCL20 production in 3T3-L1 adipocytes.

    PubMed

    Shinjo, Takanori; Iwashita, Misaki; Yamashita, Akiko; Sano, Tomomi; Tsuruta, Mitsudai; Matsunaga, Hiroaki; Sanui, Terukazu; Asano, Tomoichiro; Nishimura, Fusanori

    2016-08-19

    Interleukin-17A (IL-17A) is known to induce inflammatory responses and to be involved in the pathogenesis of not only autoimmune diseases, but also several metabolic and infectious diseases. In this study, IL-17A is shown to induce IL-6 expression in 3T3-L1 mature adipocytes. Interestingly, we found that IL-17A synergistically amplified TNFα-induced secretion of IL-6 and upregulation of IL-17RA expression in 3T3-L1 adipocytes. Its synergistic effects on IL-6 production were inhibited by pre-treatment with inhibitors of IκBα and JNK. Furthermore, IL-17A cooperatively enhanced LPS-mediated IL-6 production in 3T3-L1 adipocytes co-cultured with RAW264.7 macrophages. In addition, IL-17A also enhanced CCL20 production in 3T3-L1 adipocytes stimulated with TNFα or co-cultured with LPS-stimulated RAW macrophages. In high-fat diet-fed mouse epididymal adipose tissues, IL-17RA and RORγt mRNA levels were significantly increased and the serum level of CCL20 was also upregulated. Taken together, these data show that, in adipose tissues, IL-17A contributes to exacerbating insulin resistance-enhancing IL-6 production and promotes the infiltration of Th17 cells in cooperation with TNFα; these findings represent a novel hypothesis for the association between IL-17A-producing cells and type 2 diabetes. PMID:27311858

  15. Differential effects of pertussis toxin on insulin-stimulated phosphatidylcholine hydrolysis and glycerolipid synthesis de novo. Studies in BC3H-1 myocytes and rat adipocytes

    SciTech Connect

    Hoffman, J.M.; Standaert, M.L.; Nair, G.P.; Farese, R.V. )

    1991-04-02

    Insulin-induced increases in diacylglycerol (DAG) have been suggested to result from stimulation of de novo phosphatidic acid (PA) synthesis and phosphatidylcholine (PC) hydrolysis. Presently, the authors found that insulin decreased PC levels of BC3H-1 myocytes and rat adipocytes by approximately 10-25% within 30 s. These decreases were rapidly reversed in both cell types, apparently because of increased PC synthesis de novo. In BC3H-1 myocytes, pertussis toxin inhibited PC resynthesis and insulin effects on the pathway of de novo PA-DAG-PC synthesis, as evidenced by changes in ({sup 3}H)glycerol incorporation, but did not inhibit insulin-stimulated PC hydrolysis. Pertussis toxin also blocked the later, but not the initial, increase in DAG production in the myocytes. Phorbol esters activated PC hydrolysis in both myocytes and adipocytes, but insulin-induced stimulation of PC hydrolysis was not dependent upon activation of PKC, since this hydrolysis was not inhibited by 500 {mu}M sangivamycin, an effective PKC inhibitor. The results indicate that insulin increases DAG by pertussis toxin sensitive and insensitive (PC hydrolysis) mechanisms, which are mechanistically separate, but functionally interdependent and integrated. PC hydrolysis may contribute importantly to initial increases in DAG, but later sustained increases are apparently largely dependent on insulin-induced stimulation of the pathway of de novo phospholipid synthesis.

  16. The Molecular Signature of HIV-1-Associated Lipomatosis Reveals Differential Involvement of Brown and Beige/Brite Adipocyte Cell Lineages.

    PubMed

    Cereijo, Rubén; Gallego-Escuredo, José Miguel; Moure, Ricardo; Villarroya, Joan; Domingo, Joan Carles; Fontdevila, Joan; Martínez, Esteban; Gutiérrez, Maria del Mar; Mateo, María Gracia; Giralt, Marta; Domingo, Pere; Villarroya, Francesc

    2015-01-01

    Highly active antiretroviral therapy has remarkably improved quality of life of HIV-1-infected patients. However, this treatment has been associated with the so-called lipodystrophic syndrome, which conveys a number of adverse metabolic effects and morphological alterations. Among them, lipoatrophy of subcutaneous fat in certain anatomical areas and hypertrophy of visceral depots are the most common. Less frequently, lipomatous enlargements of subcutaneous fat at distinct anatomic areas occur. Lipomatous adipose tissue in the dorso-cervical area ("buffalo hump") has been associated with a partial white-to-brown phenotype transition and with increased cell proliferation, but, to date, lipomatous enlargements arising in other parts of the body have not been characterized. In order to establish the main molecular events associated with the appearance of lipomatosis in HIV-1 patients, we analyzed biopsies of lipomatous tissue from "buffalo hump" and from other anatomical areas in patients, in comparison with healthy subcutaneous adipose tissue, using a marker gene expression approach. Both buffalo-hump and non-buffalo-hump lipomatous adipose tissues exhibited similar patterns of non-compromised adipogenesis, unaltered inflammation, non-fibrotic phenotype and proliferative activity. Shorter telomere length, prelamin A accumulation and SA-β-Gal induction, reminiscent of adipocyte senescence, were also common to both types of lipomatous tissues. Buffalo hump biopsies showed expression of marker genes of brown adipose tissue (e.g. UCP1) and, specifically, of "classical" brown adipocytes (e.g. ZIC1) but not of beige/brite adipocytes. No such brown fat-related gene expression occurred in lipomatous tissues at other anatomical sites. In conclusion, buffalo hump and other subcutaneous adipose tissue enlargements from HIV-1-infected patients share a similar lipomatous character. However, a distorted induction of white-to-"classical brown adipocyte" phenotype appears unique of

  17. Importance of adipocyte cyclooxygenase-2 and prostaglandin E2-prostaglandin E receptor 3 signaling in the development of obesity-induced adipose tissue inflammation and insulin resistance.

    PubMed

    Chan, Pei-Chi; Hsiao, Fone-Ching; Chang, Hao-Ming; Wabitsch, Martin; Hsieh, Po Shiuan

    2016-06-01

    We examined the involvement of adipocyte cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2)-prostaglandin E receptor (EP)3-mediated signaling during hypertrophy and hypoxia in the development of obesity-associated adipose tissue (AT) inflammation and insulin resistance. The experiments were conducted with high-fat diet (HFD)-induced obese rats, db/db mice, human subjects, and 3T3-L1 and the human Simpson-Golabi-Behmel syndrome (SGBS) adipocytes; the groups were treated with selective inhibitors of COX-2 [celecoxib 30 mg/kg, half maximal inhibitory concentration (IC50) ≈ 0.04 µM] and EP3 (L-798106 100 µg/kg, IC50 ≈ 0.5 µM) or a short interfering RNA. There were strong, positive correlations between adipocyte COX-2 and EP3 gene expressions and the AT TNF-α and monocyte chemotactic protein-1 contents and the homeostatic model assessment for insulin resistance in HFD-induced obese rats, as well as body mass index in human subjects. Treatment with COX-2 and EP3 inhibitors significantly reversed AT inflammatory gene and protein expressions (-50%) and impaired glucose and insulin tolerance in db/db mice. COX-2 inhibition diminished the chemotaxis of adipocytes isolated from HFD rats to macrophages and T cells. Targeting inhibition of adipocyte COX-2 and EP3 during hypertrophy and hypoxia reversed the release of the augmented proinflammatory adipokines and the diminished adiponectin and also suppressed NF-κB and hypoxia-inducible factor-1α transcription activation. These findings suggest that adipocyte COX-2 PGE2-EP3-mediated signaling is crucially involved in the development of obesity-associated AT inflammation and insulin resistance.-Chan, P.-C., Hsiao, F.-C., Chang, H.-M., Wabitsch, M., Hsieh, P. S. Importance of adipocyte cyclooxygenase-2 and prostaglandin E2-prostaglandin E receptor 3 signaling in the development of obesity-induced adipose tissue inflammation and insulin resistance. PMID:26932930

  18. Obesity-induced DNA released from adipocytes stimulates chronic adipose tissue inflammation and insulin resistance

    PubMed Central

    Nishimoto, Sachiko; Fukuda, Daiju; Higashikuni, Yasutomi; Tanaka, Kimie; Hirata, Yoichiro; Murata, Chie; Kim-Kaneyama, Joo-ri; Sato, Fukiko; Bando, Masahiro; Yagi, Shusuke; Soeki, Takeshi; Hayashi, Tetsuya; Imoto, Issei; Sakaue, Hiroshi; Shimabukuro, Michio; Sata, Masataka

    2016-01-01

    Obesity stimulates chronic inflammation in adipose tissue, which is associated with insulin resistance, although the underlying mechanism remains largely unknown. Here we showed that obesity-related adipocyte degeneration causes release of cell-free DNA (cfDNA), which promotes macrophage accumulation in adipose tissue via Toll-like receptor 9 (TLR9), originally known as a sensor of exogenous DNA fragments. Fat-fed obese wild-type mice showed increased release of cfDNA, as determined by the concentrations of single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) in plasma. cfDNA released from degenerated adipocytes promoted monocyte chemoattractant protein-1 (MCP-1) expression in wild-type macrophages, but not in TLR9-deficient (Tlr9−/−) macrophages. Fat-fed Tlr9−/− mice demonstrated reduced macrophage accumulation and inflammation in adipose tissue and better insulin sensitivity compared with wild-type mice, whereas bone marrow reconstitution with wild-type bone marrow restored the attenuation of insulin resistance observed in fat-fed Tlr9−/− mice. Administration of a TLR9 inhibitory oligonucleotide to fat-fed wild-type mice reduced the accumulation of macrophages in adipose tissue and improved insulin resistance. Furthermore, in humans, plasma ssDNA level was significantly higher in patients with computed tomography–determined visceral obesity and was associated with homeostasis model assessment of insulin resistance (HOMA-IR), which is the index of insulin resistance. Our study may provide a novel mechanism for the development of sterile inflammation in adipose tissue and a potential therapeutic target for insulin resistance. PMID:27051864

  19. Effects of glucocorticoids on human brown adipocytes.

    PubMed

    Barclay, Johanna L; Agada, Hadiya; Jang, Christina; Ward, Micheal; Wetzig, Neil; Ho, Ken K Y

    2015-02-01

    Clinical cases of glucocorticoid (GC) excess are characterized by increased fat mass and obesity through the accumulation of white adipocytes. The effects of GCs on growth and function of brown adipose tissue are unknown and may contribute to the negative energy balance observed clinically. This study aims to evaluate the effect of GCs on proliferation, differentiation, and metabolic function of brown adipocytes. Human brown adipocytes sourced from supraclavicular fat biopsies were grown in culture and differentiated to mature adipocytes. Human white adipocytes sourced from subcutaneous abdominal fat biopsies were cultured as controls. Effects of dexamethasone on growth, differentiation (UCP1, CIDEA, and PPARGC1A expression), and function (oxygen consumption rate (OCR)) of brown adipocytes were quantified. Dexamethasone (1 μM) significantly stimulated the proliferation of brown preadipocytes and reduced that of white preadipocytes. During differentiation, dexamethasone (at 0.1, 1, and 10 μM) stimulated the expression of UCP1, CIDEA, and PPARGC1A in a concentration-dependent manner and enhanced by fourfold to sixfold the OCR of brown adipocytes. Isoprenaline (100 nM) significantly increased (P<0.05) expression of UCP1 and OCR of brown adipocytes. These effects were significantly reduced (P<0.05) by dexamethasone. Thus, we show that dexamethasone stimulates the proliferation, differentiation, and function of human brown adipocytes but inhibits adrenergic stimulation of the functioning of brown adipocytes. We conclude that GCs exert complex effects on development and function of brown adipocytes. These findings provide strong evidence for an effect of GCs on the biology of human brown adipose tissue (BAT) and for the involvement of the BAT system in the metabolic manifestation of Cushing's syndrome. PMID:25385872

  20. Reduced DPP4 activity improves insulin signaling in primary human adipocytes.

    PubMed

    Röhrborn, Diana; Brückner, Julia; Sell, Henrike; Eckel, Jürgen

    2016-03-11

    DPP4 is a ubiquitously expressed cell surface protease which is also released to the circulation as soluble DPP4 (sDPP4). Recently, we identified DPP4 as a novel adipokine oversecreted in obesity and thus potentially linking obesity to the metabolic syndrome. Furthermore, sDPP4 impairs insulin signaling in an autocrine and paracrine fashion in different cell types. However, it is still unknown which functional role DPP4 might play in adipocytes. Therefore, primary human adipocytes were treated with a specific DPP4 siRNA. Adipocyte differentiation was not affected by DPP4 silencing. Interestingly, DPP4 reduction improved insulin responsiveness of adipocytes at the level of insulin receptor, proteinkinase B (Akt) and Akt substrate of 160 kDa. To investigate whether the observed effects could be attributed to the enzymatic activity of DPP4, human adipocytes were treated with the DPP4 inhibitors sitagliptin and saxagliptin. Our data show that insulin-stimulated activation of Akt is augmented by DPP4 inhibitor treatment. Based on our previous observation that sDPP4 induces insulin resistance in adipocytes, and that adipose DPP4 levels are higher in obese insulin-resistant patients, we now suggest that the abundance of DPP4 might be a regulator of adipocyte insulin signaling. PMID:26872429

  1. The Molecular Signature of HIV-1-Associated Lipomatosis Reveals Differential Involvement of Brown and Beige/Brite Adipocyte Cell Lineages

    PubMed Central

    Cereijo, Rubén; Gallego-Escuredo, José Miguel; Moure, Ricardo; Villarroya, Joan; Domingo, Joan Carles; Fontdevila, Joan; Martínez, Esteban; Gutiérrez, Maria del Mar; Mateo, María Gracia; Giralt, Marta; Domingo, Pere; Villarroya, Francesc

    2015-01-01

    Highly active antiretroviral therapy has remarkably improved quality of life of HIV-1-infected patients. However, this treatment has been associated with the so-called lipodystrophic syndrome, which conveys a number of adverse metabolic effects and morphological alterations. Among them, lipoatrophy of subcutaneous fat in certain anatomical areas and hypertrophy of visceral depots are the most common. Less frequently, lipomatous enlargements of subcutaneous fat at distinct anatomic areas occur. Lipomatous adipose tissue in the dorso-cervical area (“buffalo hump”) has been associated with a partial white-to-brown phenotype transition and with increased cell proliferation, but, to date, lipomatous enlargements arising in other parts of the body have not been characterized. In order to establish the main molecular events associated with the appearance of lipomatosis in HIV-1 patients, we analyzed biopsies of lipomatous tissue from “buffalo hump” and from other anatomical areas in patients, in comparison with healthy subcutaneous adipose tissue, using a marker gene expression approach. Both buffalo-hump and non-buffalo-hump lipomatous adipose tissues exhibited similar patterns of non-compromised adipogenesis, unaltered inflammation, non-fibrotic phenotype and proliferative activity. Shorter telomere length, prelamin A accumulation and SA-β-Gal induction, reminiscent of adipocyte senescence, were also common to both types of lipomatous tissues. Buffalo hump biopsies showed expression of marker genes of brown adipose tissue (e.g. UCP1) and, specifically, of “classical” brown adipocytes (e.g. ZIC1) but not of beige/brite adipocytes. No such brown fat-related gene expression occurred in lipomatous tissues at other anatomical sites. In conclusion, buffalo hump and other subcutaneous adipose tissue enlargements from HIV-1-infected patients share a similar lipomatous character. However, a distorted induction of white-to-“classical brown adipocyte” phenotype

  2. Differentiation to adipocytes in accompanied by an increase in the amounts of Gi- and Go-proteins in 3T3-L1 cells

    SciTech Connect

    Watkins, D.C.; Northup, J.K.; Malbon, C.C.

    1986-05-01

    Treatment of cultures of 3T3-L1 cells with methylisobutyl-xanthine and dexamethasone has been shown to result in accumulation of lipid and conversion to the morphology of adipocytes in more than 90% of the cells. The status of the stimulatory (Gs), inhibitory (Gi) and Go-proteins during the course of 3T3-L1 differentiation was examined. The amount of alpha subunit of Gs (..cap alpha..Gs), assayed by radiolabeling in the presence of cholera toxin and (/sup 32/P)NAD/sup +/, increased upon differentiation as previously described by others. The amounts of ..cap alpha..Gi and ..cap alpha..Go assayed by radiolabeling in the presence of pertussis toxin and (/sup 32/P)NAD/sup +/ increased 3-fold upon differentiation. Immunoblots of cell membranes subjected to gel electrophoresis in sodium dodecyl sulfate were probed with two rabbit antisera raised against bovine brain ..cap alpha..Go and with one raised against the..beta..-subunit of the bovine rod-outer-segment G-protein, referred to as transducin. The immunoblotting data confirm the increase upon differentiation of ..cap alpha..Go and also demonstrate an increase in the amount of the ..beta..-subunit. Thus differentiation of 3T3-L1 cells is accompanied by dramatic changes in the complexion of G-proteins in the membranes.

  3. miR-23a/b regulates the balance between osteoblast and adipocyte differentiation in bone marrow mesenchymal stem cells

    PubMed Central

    Guo, Qi; Chen, Yusi; Guo, Lijuan; Jiang, Tiejian; Lin, Zhangyuan

    2016-01-01

    Age-related osteoporosis is associated with the reduced capacity of bone marrow mesenchymal stem cells (BMSCs) to differentiate into osteoblasts instead of adipocytes. However, the molecular mechanisms that decide the fate of BMSCs remain unclear. In our study, microRNA-23a, and microRNA-23b (miR-23a/b) were found to be markedly downregulated in BMSCs of aged mice and humans. The overexpression of miR-23a/b in BMSCs promoted osteogenic differentiation, whereas the inhibition of miR-23a/b increased adipogenic differentiation. Transmembrane protein 64 (Tmem64), which has expression levels inversely related to those of miR-23a/b in aged and young mice, was identified as a major target of miR-23a/b during BMSC differentiation. In conclusion, our study suggests that miR-23a/b has a critical role in the regulation of mesenchymal lineage differentiation through the suppression of Tmem64. PMID:27606130

  4. miR-23a/b regulates the balance between osteoblast and adipocyte differentiation in bone marrow mesenchymal stem cells.

    PubMed

    Guo, Qi; Chen, Yusi; Guo, Lijuan; Jiang, Tiejian; Lin, Zhangyuan

    2016-01-01

    Age-related osteoporosis is associated with the reduced capacity of bone marrow mesenchymal stem cells (BMSCs) to differentiate into osteoblasts instead of adipocytes. However, the molecular mechanisms that decide the fate of BMSCs remain unclear. In our study, microRNA-23a, and microRNA-23b (miR-23a/b) were found to be markedly downregulated in BMSCs of aged mice and humans. The overexpression of miR-23a/b in BMSCs promoted osteogenic differentiation, whereas the inhibition of miR-23a/b increased adipogenic differentiation. Transmembrane protein 64 (Tmem64), which has expression levels inversely related to those of miR-23a/b in aged and young mice, was identified as a major target of miR-23a/b during BMSC differentiation. In conclusion, our study suggests that miR-23a/b has a critical role in the regulation of mesenchymal lineage differentiation through the suppression of Tmem64. PMID:27606130

  5. Effects of the monoclonal antibody against porcine 40 kDa adipocyte-specific plasma membrane protein on adipocytes and carcass composition.

    PubMed

    Gao, Shizheng; Ge, Changrong; Zhang, Xi; Liu, Yonggang

    2007-07-01

    The effects of the mouse monoclonal antibody against 40 kDa adipocyte-specific plasma membrane protein on porcine adipocytes and carcass composition were investigated in vitro and in vivo. Results revealed that the in vitro complement-mediated cytotoxicity of this monoclonal antibody can lead to adipocyte lysis, remarkable reduction of adipocyte lipid accumulation (P<0.01), and significant decrease of well-differentiated fat cells (P<0.01). Treatment of adipocytes with this antibody alone in vitro did not induce cell lysis, but could lead to noticeable reduction of well-differentiated cells and lipid accumulation (P<0.05) at the pre-adipocyte stage. In vivo, pigs injected with 0.5 mg/kg or 1.0 mg/kg of antibody showed smaller adipocyte sizes (P<0.01) and reduced lipid accumulation of adipocytes (P<0.01). Our results also indicated that pigs intraperitoneally or subcutaneously immunized with 0.5 mg/kg of monoclonal antibody at 15 kg or 1.0 mg/kg antibody at 60 kg had a higher lean meat percentage (P<0.05), larger loin eye area (P<0.05), lower fat meat percentage (P<0.05), less backfat thickness (P<0.05) and smaller leaf fat weight (P<0.05) than the control pigs, but other carcass traits such as caul fat weight, heart weight, liver weight, spleen weight, kidney weight, lung weight, and dressing percentage were not significantly affected. These results suggested that this monoclonal antibody could be applied to restrain excessive fat deposition in porcine production. PMID:17622468

  6. Isoflavonoids from Crotalaria albida Inhibit Adipocyte Differentiation and Lipid Accumulation in 3T3-L1 Cells via Suppression of PPAR-γ Pathway

    PubMed Central

    Sun, Qinhu; Chou, Guixin

    2015-01-01

    Two 2″-isopropenyl dihydrofuran isoflavonoids (1 and 3), one 2″-isopropenyl dihydrofuran chromone (2), as well as 13 known compounds were isolated from the herbs of Crotalaria albida. Their structures and relative configurations were elucidated via NMR and HRESIMS analyses. The 2″ S absolute configuration of 1 and 2 were deduced by comparing their NOESY spectra with that of 3, which was determined via single crystal X-ray diffraction (CuKα). The 3R absolute configuration of 1 was determined by CD. Compounds 1, 2, and 3 inhibit the adipocyte differentiation and lipid accumulation of 3T3-L1 through down-regulation of PPAR-γ activity. PMID:26285147

  7. Leptin of dermal adipose tissue is differentially expressed during the hair cycle and contributes to adipocyte-mediated growth inhibition of anagen-phase vibrissa hair.

    PubMed

    Yang, Chao-Chun; Sheu, Hamm-Ming; Chung, Pei-Lun; Chang, Chung-Hsing; Tsai, Yau-Sheng; Hughes, Michael W; Tuan, Tai-Lan; Huang, Lynn L H

    2015-01-01

    Adipose tissue encircles the lower portion of anagen hair follicles and may regulate hair cycle progression. As leptin is a major adipokine, its level of expression from the dermal white adipose tissue during hair cycle progression was studied. The result shows that leptin level is differentially expressed during hair cycle, the lowest in early anagen phase, upregulated in late anagen phase and the highest in the telogen phase. On the other hand, leptin receptor is detected in keratin 15-positive hair bulge epithelium of both anagen- and telogen-phase hair follicles of mice pelage and vibrissa hair, and hair from human scalp. Leptin contributes to adipocyte-mediated growth inhibition of anagen-phase vibrissa hair as demonstrated in organ culture and coculture system. Our data suggest that leptin of dermal white adipose tissue might regulate hair growth and, therefore, hair cycle progression via leptin receptor on the hair follicle epithelium. PMID:25313970

  8. Polychlorinated biphenyls (PCB 101, PCB 153 and PCB 180) alter leptin signaling and lipid metabolism in differentiated 3T3-L1 adipocytes

    SciTech Connect

    Ferrante, Maria C.; Amero, Paola; Santoro, Anna; Monnolo, Anna; Simeoli, Raffaele; Di Guida, Francesca; Mattace Raso, Giuseppina; Meli, Rosaria

    2014-09-15

    Non-dioxin-like polychlorinated biphenyls (NDL-PCBs) are highly lipophilic environmental contaminants that accumulate in lipid-rich tissues, such as adipose tissue. Here, we reported the effects induced by PCBs 101, 153 and 180, three of the six NDL-PCBs defined as indicators, on mature 3T3-L1 adipocytes. We observed an increase in lipid content, in leptin gene expression and a reduction of leptin receptor expression and signaling, when cells were exposed to PCBs, alone or in combination. These modifications were consistent with the occurrence of “leptin-resistance” in adipose tissue, a typical metabolic alteration related to obesity. Therefore, we investigated how PCBs affect the expression of pivotal proteins involved in the signaling of leptin receptor. We evaluated the PCB effect on the intracellular pathway JAK/STAT, determining the phosphorylation of STAT3, a downstream activator of the transcription of leptin gene targets, and the expression of SOCS3 and PTP1B, two important regulators of leptin resistance. In particular, PCBs 153 and 180 or all PCB combinations induced a significant reduction in pSTAT3/STAT3 ratio and an increase in PTP1B and SOCS3, evidencing an additive effect. The impairment of leptin signaling was associated with the reduction of AMPK/ACC pathway activation, leading to the increase in lipid content. These pollutants were also able to increase the transcription of inflammatory cytokines (IL-6 and TNFα). It is worthy to note that the PCB concentrations used are comparable to levels detectable in human adipose tissue. Our data strongly support the hypothesis that NDL-PCBs may interfere with the lipid metabolism contributing to the development of obesity and related diseases. - Highlights: • NDL-PCBs alter lipid content and metabolism in 3T3-L1 adipocytes. • Impairment of leptin signaling was induced by NDL-PCBs. • NDL-PCBs reduce AMPK and ACC activation. • NDL-PCBs induce the synthesis of pro-inflammatory cytokine by

  9. Surface tethered epidermal growth factor protects proliferating and differentiating multipotential stromal cells from FasL induced apoptosis

    PubMed Central

    Rodrigues, Melanie; Blair, Harry; Stockdale, Linda; Griffith, Linda; Wells, Alan

    2012-01-01

    Multipotential stromal cells, or mesenchymal stem cells, (MSC) have ben proposed as aids in regenerating bone and adipose tissues, as these cells form osteoblasts and adipocytes. A major obstacle to this use of MSC is the initial loss of cells post-implantation. This cell death in part, is due to ubiquitous non-specific inflammatory cytokines such as FasL generated in the implant site. Our group previously found that soluble epidermal growth factor (sEGF) promotes MSC expansion. Further, tethering EGF onto a two-dimensional surface (tEGF) altered MSC responses, by restricting epidermal growth factor receptor (EGFR) to the cell surface, causing sustained activation of EGFR, and promoting survival from FasL-induced death. sEGF by causing internalization of EGFR does not support MSC survival. However, for tEGF to be useful in bone regeneration, it needs to allow for MSC differentiation into osteoblasts while also protecting emerging osteoblasts from apoptosis. tEGF did not block induced differentiation of MSCs into osteoblasts, or adipocytes, a common default MSC-differentiation pathway. MSC-derived pre-osteoblasts showed increased Fas levels and became more susceptible to FasL induced death, which tEGF prevented. Differentiating adipocytes underwent a reduction in Fas expression and became resistant to FasL-induced death, with tEGF having no further survival effect. tEGF protected undifferentiated MSC from combined insults of FasL, serum deprivation and physiologic hypoxia. Additionally, tEGF was dominant in the face of sEGF to protect MSC from FasL-induced death. Our results suggest that MSCs and differentiating osteoblasts need protective signals to survive in the inflammatory wound milieu and that tEGF can serve this function. PMID:22948863

  10. FABP3 and brown adipocyte-characteristic mitochondrial fatty acid oxidation enzymes are induced in beige cells in a different pathway from UCP1.

    PubMed

    Nakamura, Yuki; Sato, Takahiro; Shiimura, Yuki; Miura, Yoshiki; Kojima, Masayasu

    2013-11-01

    Cold exposure and β3-adrenergic receptor agonist (CL316,243) treatment induce the production of beige cells, which express brown adipocytes(BA)-specific UCP1 protein, in white adipose tissue (WAT). It remains unclear whether the beige cells, which have different gene expression patterns from BA, express BA-characteristic fatty acid oxidation (FAO) proteins. Here we found that 5 day cold exposure and CL316,243 treatment of WAT, but not CL316,243 treatment of primary adipocytes of C57BL/6J mice, increased mRNA levels of BA-characteristic FAO proteins. These results suggest that BA-characteristic FAO proteins are induced in beige cells in a different pathway from UCP1. PMID:24129192

  11. Role of LRP1 and ERK and cAMP Signaling Pathways in Lactoferrin-Induced Lipolysis in Mature Rat Adipocytes

    PubMed Central

    Ikoma-Seki, Keiko; Nakamura, Kanae; Morishita, Satoru; Ono, Tomoji; Sugiyama, Keikichi; Nishino, Hoyoku; Hirano, Hisashi; Murakoshi, Michiaki

    2015-01-01

    Lactoferrin (LF) is a multifunctional glycoprotein present in milk. A clinical study showed that enteric-coated bovine LF tablets decrease visceral fat accumulation. Furthermore, animal studies revealed that ingested LF is partially delivered to mesenteric fat, and in vitro studies showed that LF promotes lipolysis in mature adipocytes. The aim of the present study was to determine the mechanism underlying the induction of lipolysis in mature adipocytes that is induced by LF. To address this question, we used proteomics techniques to analyze protein expression profiles. Mature adipocytes from primary cultures of rat mesenteric fat were collected at various times after exposure to LF. Proteomic analysis revealed that the expression levels of hormone-sensitive lipase (HSL), which catalyzes the rate-limiting step of lipolysis, were upregulated and that HSL was activated by protein kinase A within 15 min after the cells were treated with LF. We previously reported that LF increases the intracellular concentration of cyclic adenosine monophosphate (cAMP), suggesting that LF activates the cAMP signaling pathway. In this study, we show that the expression level and the activity of the components of the extracellular signal-regulated kinase (ERK) signaling pathway were upregulated. Moreover, LF increased the activity of the transcription factor cAMP response element binding protein (CREB), which acts downstream in the cAMP and ERK signaling pathways and regulates the expression levels of adenylyl cyclase and HSL. Moreover, silencing of the putative LF receptor low-density lipoprotein receptor-related protein 1 (LRP1) attenuated lipolysis in LF-treated adipocytes. These results suggest that LF promoted lipolysis in mature adipocytes by regulating the expression levels of proteins involved in lipolysis through controlling the activity of cAMP/ERK signaling pathways via LRP1. PMID:26506094

  12. Triphenylmethylphosphonium cation distribution as a measure of hormone-induced alterations in white adipocyte membrane potential

    SciTech Connect

    Vallano, M.L.; Sonenberg, M.

    1982-01-01

    Triphenylmethylphosphonium (TPMP+) partitions into the mitochondrial and cytosolic compartments in the rat white adipocyte in a potential-dependent fashion. The relationship between (/sup 3/H)TPMP+ distribution, intracellular cAMP generation and lipolysis in response to hormones and cAMP-mimetic compounds was examined. Half-maximal (/sup 3/H)TPMP+ efflux and glycerol release were produced by 15 and 9 nM adrenocorticotropin, 170 and 110 nM 1-epinephrine, 70 and 27 microM isobutylmethylxanthine and 800 and 750 microM dibutyryl cAMP, respectively. Hormone-stimulated cAMP generation was also correlated with (/sup 3/H)TPMP+ efflux and lipolysis in terms of concentration dependency. In kinetic experiments, glycerol release and (/sup 3/H)TPMP+ efflux in response to adrenocorticotropin or cholera toxin proceeded over a similar time course, whereas an earlier rise in cAMP generation was detected. The depolarizing effect of lipolytic compounds was localized to the mitochondrial compartment. When cells were incubated in elevated-(K+)0 buffer, the stimulatory effect of dibutyryl cAMP on (/sup 3/H)TPMP+ efflux and lipolysis persisted, suggesting that maintenance of the plasma membrane potential is not critical for demonstration of these responses. When the extracellular concentration of serum albumin, which provides binding sites for free fatty acids, was increased from 1 to 3%, an increase in glycerol release and a decrease in (/sup 3/H)TPMP+ efflux was observed. We suggest that intracellular free fatty acid accumulation in response to lipolytic agents causes dissipation of the mitochondrial membrane potential and efflux of (/sup 3/H)TPMP+ from the organelle and cell.

  13. Age-Related Modulation of the Effects of Obesity on Gene Expression Profiles of Mouse Bone Marrow and Epididymal Adipocytes

    PubMed Central

    Liu, Li-Fen; Shen, Wen-Jun; Ueno, Masami; Patel, Shailja; Azhar, Salman; Kraemer, Fredric B.

    2013-01-01

    This study aimed to characterize and compare the effects of obesity on gene expression profiles in two distinct adipose depots, epididymal and bone marrow, at two different ages in mice. Alterations in gene expression were analyzed in adipocytes isolated from diet-induced obese (DIO) C57BL/6J male mice at 6 and 14 months of age and from leptin deficient mice (ob/ob) at 6 months of age using microarrays. DIO affected gene expression in both depots at 6 and 14 months, but more genes were altered in epididymal than bone marrow adipocytes at each age and younger mice displayed more changes than older animals. In epididymal adipocytes a total of 2789 (9.6%) genes were differentially expressed at 6-months with DIO, whereas 952 (3.3%) were affected at 14-months. In bone marrow adipocytes, 347 (1.2%) genes were differentially expressed at 6-months with DIO, whereas only 189 (0.66%) were changed at 14-months. 133 genes were altered by DIO in both fat depots at 6-months, and 37 genes at 14-months. Only four genes were altered in both depots at both ages with DIO. Bone marrow adipocytes are less responsive to DIO than epididymal adipocytes and the response of both depots to DIO declines with age. This loss of responsiveness with age is likely due to age-associated changes in expression of genes related to adipogenesis, inflammation and mitochondrial function that are similar to and obscure the changes commonly associated with DIO. Patterns of gene expression were generally similar in epididymal adipocytes from ob/ob and DIO mice; however, several genes were differentially expressed in bone marrow adipocytes from ob/ob and DIO mice, perhaps reflecting the importance of leptin signaling for bone metabolism. In conclusion, obesity affects age-associated alterations in gene expression in both epididymal and bone marrow adipocytes regardless of diet or genetic background. PMID:23967297

  14. Age-related modulation of the effects of obesity on gene expression profiles of mouse bone marrow and epididymal adipocytes.

    PubMed

    Liu, Li-Fen; Shen, Wen-Jun; Ueno, Masami; Patel, Shailja; Azhar, Salman; Kraemer, Fredric B

    2013-01-01

    This study aimed to characterize and compare the effects of obesity on gene expression profiles in two distinct adipose depots, epididymal and bone marrow, at two different ages in mice. Alterations in gene expression were analyzed in adipocytes isolated from diet-induced obese (DIO) C57BL/6J male mice at 6 and 14 months of age and from leptin deficient mice (ob/ob) at 6 months of age using microarrays. DIO affected gene expression in both depots at 6 and 14 months, but more genes were altered in epididymal than bone marrow adipocytes at each age and younger mice displayed more changes than older animals. In epididymal adipocytes a total of 2789 (9.6%) genes were differentially expressed at 6-months with DIO, whereas 952 (3.3%) were affected at 14-months. In bone marrow adipocytes, 347 (1.2%) genes were differentially expressed at 6-months with DIO, whereas only 189 (0.66%) were changed at 14-months. 133 genes were altered by DIO in both fat depots at 6-months, and 37 genes at 14-months. Only four genes were altered in both depots at both ages with DIO. Bone marrow adipocytes are less responsive to DIO than epididymal adipocytes and the response of both depots to DIO declines with age. This loss of responsiveness with age is likely due to age-associated changes in expression of genes related to adipogenesis, inflammation and mitochondrial function that are similar to and obscure the changes commonly associated with DIO. Patterns of gene expression were generally similar in epididymal adipocytes from ob/ob and DIO mice; however, several genes were differentially expressed in bone marrow adipocytes from ob/ob and DIO mice, perhaps reflecting the importance of leptin signaling for bone metabolism. In conclusion, obesity affects age-associated alterations in gene expression in both epididymal and bone marrow adipocytes regardless of diet or genetic background. PMID:23967297

  15. Bixin regulates mRNA expression involved in adipogenesis and enhances insulin sensitivity in 3T3-L1 adipocytes through PPAR{gamma} activation

    SciTech Connect

    Takahashi, Nobuyuki; Goto, Tsuyoshi; Taimatsu, Aki; Egawa, Kahori; Katoh, Sota; Kusudo, Tatsuya; Sakamoto, Tomoya; Ohyane, Chie; Lee, Joo-Young; Kim, Young-il; Uemura, Taku; Hirai, Shizuka; Kawada, Teruo

    2009-12-25

    Insulin resistance is partly due to suppression of insulin-induced glucose uptake into adipocytes. The uptake is dependent on adipocyte differentiation, which is controlled at mRNA transcription level. The peroxisome proliferator-activated receptor (PPAR), a ligand-regulated nuclear receptor, is involved in the differentiation. Many food-derived compounds serve as ligands to activate or inactivate PPAR. In this study, we demonstrated that bixin and norbixin (annatto extracts) activate PPAR{gamma} by luciferase reporter assay using GAL4-PPAR chimera proteins. To examine the effects of bixin on adipocytes, 3T3-L1 adipocytes were treated with bixin or norbixin. The treatment induced mRNA expression of PPAR{gamma} target genes such as adipocyte-specific fatty acid-binding protein (aP2), lipoprotein lipase (LPL), and adiponectin in differentiated 3T3-L1 adipocytes and enhanced insulin-dependent glucose uptake. The observations indicate that bixin acts as an agonist of PPAR{gamma} and enhances insulin sensitivity in 3T3-L1 adipocytes, suggesting that bixin is a valuable food-derived compound as a PPAR ligand to regulate lipid metabolism and to ameliorate metabolic syndrome.

  16. Endoplasmic reticulum stress suppresses lipin-1 expression in 3T3-L1 adipocytes

    SciTech Connect

    Takahashi, Nobuhiko; Hiranaka, Natsumi; Suzuki, Takeshi; Yui, Tomoo; Akanuma, Masayoshi; Kanazawa, Kaoru; Yoshida, Mika; Naito, Sumiyoshi; Fujiya, Mikihiro; Kohgo, Yutaka

    2013-02-01

    Highlights: ► Lipin-1 involves lipid metabolism, adipocyte differentiation, and inflammation. ► Adipose lipin-1 expression is reduced in obesity. ► ER stress suppresses lipin-1 expression in 3T3-L1 adipocytes. ► Activation of PPAR-γ recovers ER stress-induced lipin-1 reduction. -- Abstract: Lipin-1 plays crucial roles in the regulation of lipid metabolism and cell differentiation in adipocytes. In obesity, adipose lipin-1 mRNA expression is decreased and positively correlated with systemic insulin sensitivity. Amelioration of the lipin-1 depletion might be improved dysmetabolism. Although some cytokines such as TNF-α and interleukin-1β reduces adipose lipin-1 expression, the mechanism of decreased adipose lipin-1 expression in obesity remains unclear. Recently, endoplasmic reticulum (ER) stress is implicated in the pathogenesis of obesity. Here we investigated the role of ER stress on the lipin-1 expression in 3T3-L1 adipocytes. We demonstrated that lipin-1 expression was suppressed by the treatment with ER stress inducers (tunicamycin and thapsigargin) at transcriptional level. We also showed that constitutive lipin-1 expression could be maintained by peroxisome proliferator-activated receptor-γ in 3T3-L1 adipocytes. Activation of peroxisome proliferator-activated receptor-γ recovered the ER stress-induced lipin-1 suppression. These results suggested that ER stress might be involved in the pathogenesis of obesity through lipin-1 depletion.

  17. The transcriptional basis of adipocyte development.

    PubMed

    Rosen, Evan D

    2005-07-01

    Adipogenesis is the developmental process by which a multipotent mesenchymal stem cell differentiates into a mature adipocyte. This process involves a highly regulated and coordinated cascade of transcription factors that together lead to the establishment of the differentiated state. In the presence of the correct hormonal cues, committed pre-adipocytes express the bZIP factors C/EBPb and C/EBPd. These factors in turn induce the expression of C/EBPa and peroxisome proliferator-activated receptor g (PPARg). C/EBPa and PPARg together promote differentiation by activating adipose-specific gene expression and by maintaining each others expression at high levels. We have investigated the relative contributions of PPARg and C/EBPa to adipogenesis by selectively ablating these genes in mouse embryonic fibroblasts (MEFs). MEFs that lack C/EBPa are able to undergo adipogenesis, but only when PPARg is ectopically expressed. Interestingly, these cells are not sensitive to the metabolic actions of insulin. By way of contrast, cells that lack PPARg are utterly incapable of adipogenic conversion, even when supplemented with high levels of C/EBPa. Our current investigations are centered on the identification of novel adipogenic transcription factors, utilizing a variety of techniques, ranging from BAC transgenics to computational approaches. These approaches will be discussed, along with the roles of some new transcriptional players in adipogenesis, including the O/E family of proteins. PMID:15936931

  18. Dexamethasone and rosiglitazone are sufficient and necessary for producing functional adipocytes from mesenchymal stem cells.

    PubMed

    Contador, David; Ezquer, Fernando; Espinosa, Maximiliano; Arango-Rodriguez, Martha; Puebla, Carlos; Sobrevia, Luis; Conget, Paulette

    2015-09-01

    The final product of adipogenesis is a functional adipocyte. This mature cell acquires the necessary machinery for lipid metabolism, loses its proliferation potential, increases its insulin sensitivity, and secretes adipokines. Multipotent mesechymal stromal cells have been recognized as a source of adipocytes both in vivo and in vitro. The in vitro adipogenic differentiation of human MSC (hMSC) has been induced up to now by using a complex stimulus which includes dexamethasone, 3-isobutyl-1-methylxanthine, indomethacin, and insulin (a classical cocktail) and evaluated according to morphological changes. The present work was aimed at demonstrating that the simultaneous activation of dexamethasone's canonical signaling pathways, through the glucocorticoid receptor and CCAAT-enhancer-binding proteins (C/EBPs) and rosiglitazone through peroxisome proliferator-activated receptor gamma (PPAR-gamma) is sufficient yet necessary for inducing hMSC adipogenic differentiation. It was also ascertained that hMSC exposed just to dexamethasone and rosiglitazone (D&R) differentiated into cells which accumulated neutral lipid droplets, expressed C/EBP-alpha, PPAR-gamma, aP2, lipoprotein lipase, acyl-CoA synthetase, phosphoenolpyruvate carboxykinase, adiponectin, and leptin genes but did not proliferate. Glucose uptake was dose dependent on insulin stimulus and high levels of adipokines were secreted (i.e. displaying not only the morphology but also expressing mature adipocytes' specific genes and functional characteristics). This work has demonstrated that (i) the activating C/EBPs and PPAR-gamma signaling pathways were sufficient to induce adipogenic differentiation from hMSC, (ii) D&R producing functional adipocytes from hMSC, (iii) D&R induce adipogenic differentiation from mammalian MSC (including those which are refractory to classical adipogenic differentiation stimuli). D&R would thus seem to be a useful tool for MSC characterization, studying adipogenesis pathways and

  19. SILAC-MS Based Characterization of LPS and Resveratrol Induced Changes in Adipocyte Proteomics – Resveratrol as Ameliorating Factor on LPS Induced Changes

    PubMed Central

    Kroager, Toke P.; Sanggaard, Kristian W.; Knudsen, Anders D.; Stensballe, Allan; Enghild, Jan J.; Ølholm, Jens; Richelsen, Bjørn; Pedersen, Steen B.

    2016-01-01

    Adipose tissue inflammation is believed to play a pivotal role in the development obesity-related morbidities such as insulin resistance. However, it is not known how this (low-grade) inflammatory state develops. It has been proposed that the leakage of lipopolysaccharides (LPS), originating from the gut microbiota, through the gut epithelium could drive initiation of inflammation. To get a better understanding of which proteins and intracellular pathways are affected by LPS in adipocytes, we performed SILAC proteomic analysis and identified proteins that were altered in expression. Furthermore, we tested the anti-inflammatory compound resveratrol. A total of 927 proteins were quantified by the SILAC method and of these 57- and 64 were significantly up- and downregulated by LPS, respectively. Bioinformatic analysis (GO analysis) revealed that the upregulated proteins were especially involved in the pathways of respiratory electron transport chain and inflammation. The downregulated proteins were especially involved in protein glycosylation. One of the latter proteins, GALNT2, has previously been described to regulate the expression of liver lipases such as ANGPTL3 and apoC-III affecting lipid metabolism. Furthermore, LPS treatment reduced the protein levels of the insulin sensitizing adipokine, adiponectin, and proteins participating in the final steps of triglyceride- and cholesterol synthesis. Generally, resveratrol opposed the effect induced by LPS and, as such, functioning as an ameliorating factor in disease state. Using an unbiased proteomic approach, we present novel insight of how the proteome is altered in adipocytes in response to LPS as seen in obesity. We suggest that LPS partly exerts its detrimental effects by altering glycosylation processes of the cell, which is starting to emerge as important posttranscriptional regulators of protein expression. Furthermore, resveratrol could be a prime candidate in ameliorating dysfunctioning adipose tissue

  20. Antidiabetic Activities of Abutilon indicum (L.) Sweet Are Mediated by Enhancement of Adipocyte Differentiation and Activation of the GLUT1 Promoter.

    PubMed

    Krisanapun, Chutwadee; Lee, Seong-Ho; Peungvicha, Penchom; Temsiririrkkul, Rungravi; Baek, Seung Joon

    2011-01-01

    Abutilon indicum (L.) Sweet is an Asian phytomedicine traditionally used to treat several disorders, including diabetes mellitus. However, molecular mechanisms supporting the antidiabetic effect of A. indicum L. remain unknown. The aim of this study was to evaluate whether extract of A. indicum L. improves insulin sensitivity. First, we observed the antidiabetic activity of aqueous extract of the entire plant (leaves, twigs and roots) of A. indicum L. on postprandial plasma glucose in diabetic rats. The subsequent experiments revealed that butanol fractions of the extract bind to PPARγ and activate 3T3-L1 differentiation. To measure glucose uptake enhanced by insulin-like activity, we used rat diaphragm incubated with various concentrations of the crude extract and found that the extract enhances glucose consumption in the incubated solution. Our data also indicate that the crude extract and the fractions (water and butanol) did not affect the activity of kinases involved in Akt and GSK-3β pathways; however, the reporter assay showed that the crude extract could activate glucose transporter 1 (GLUT1) promoter activity. These results suggest that the extract from A. indicum L. may be beneficial for reducing insulin resistance through its potency in regulating adipocyte differentiation through PPARγ agonist activity, and increasing glucose utilization via GLUT1. PMID:21603234

  1. Antidiabetic Activities of Abutilon indicum (L.) Sweet Are Mediated by Enhancement of Adipocyte Differentiation and Activation of the GLUT1 Promoter

    PubMed Central

    Krisanapun, Chutwadee; Lee, Seong-Ho; Peungvicha, Penchom; Temsiririrkkul, Rungravi; Baek, Seung Joon

    2011-01-01

    Abutilon indicum (L.) Sweet is an Asian phytomedicine traditionally used to treat several disorders, including diabetes mellitus. However, molecular mechanisms supporting the antidiabetic effect of A. indicum L. remain unknown. The aim of this study was to evaluate whether extract of A. indicum L. improves insulin sensitivity. First, we observed the antidiabetic activity of aqueous extract of the entire plant (leaves, twigs and roots) of A. indicum L. on postprandial plasma glucose in diabetic rats. The subsequent experiments revealed that butanol fractions of the extract bind to PPARγ and activate 3T3-L1 differentiation. To measure glucose uptake enhanced by insulin-like activity, we used rat diaphragm incubated with various concentrations of the crude extract and found that the extract enhances glucose consumption in the incubated solution. Our data also indicate that the crude extract and the fractions (water and butanol) did not affect the activity of kinases involved in Akt and GSK-3β pathways; however, the reporter assay showed that the crude extract could activate glucose transporter 1 (GLUT1) promoter activity. These results suggest that the extract from A. indicum L. may be beneficial for reducing insulin resistance through its potency in regulating adipocyte differentiation through PPARγ agonist activity, and increasing glucose utilization via GLUT1. PMID:21603234

  2. Adipocyte lipases and defect of lipolysis in human obesity.

    PubMed

    Langin, Dominique; Dicker, Andrea; Tavernier, Geneviève; Hoffstedt, Johan; Mairal, Aline; Rydén, Mikael; Arner, Erik; Sicard, Audrey; Jenkins, Christopher M; Viguerie, Nathalie; van Harmelen, Vanessa; Gross, Richard W; Holm, Cecilia; Arner, Peter

    2005-11-01

    The mobilization of fat stored in adipose tissue is mediated by hormone-sensitive lipase (HSL) and the recently characterized adipose triglyceride lipase (ATGL), yet their relative importance in lipolysis is unknown. We show that a novel potent inhibitor of HSL does not inhibit other lipases. The compound counteracted catecholamine-stimulated lipolysis in mouse adipocytes and had no effect on residual triglyceride hydrolysis and lipolysis in HSL-null mice. In human adipocytes, catecholamine- and natriuretic peptide-induced lipolysis were completely blunted by the HSL inhibitor. When fat cells were not stimulated, glycerol but not fatty acid release was inhibited. HSL and ATGL mRNA levels increased concomitantly during adipocyte differentiation. Abundance of the two transcripts in human adipose tissue was highly correlated in habitual dietary conditions and during a hypocaloric diet, suggesting common regulatory mechanisms for the two genes. Comparison of obese and nonobese subjects showed that obesity was associated with a decrease in catecholamine-induced lipolysis and HSL expression in mature fat cells and in differentiated preadipocytes. In conclusion, HSL is the major lipase for catecholamine- and natriuretic peptide-stimulated lipolysis, whereas ATGL mediates the hydrolysis of triglycerides during basal lipolysis. Decreased catecholamine-induced lipolysis and low HSL expression constitute a possibly primary defect in obesity. PMID:16249444

  3. Mitochondria in White, Brown, and Beige Adipocytes

    PubMed Central

    Cedikova, Miroslava; Kripnerová, Michaela; Dvorakova, Jana; Pitule, Pavel; Grundmanova, Martina; Babuska, Vaclav; Mullerova, Dana; Kuncova, Jitka

    2016-01-01

    Mitochondria play a key role in energy metabolism in many tissues, including cardiac and skeletal muscle, brain, liver, and adipose tissue. Three types of adipose depots can be identified in mammals, commonly classified according to their colour appearance: the white (WAT), the brown (BAT), and the beige/brite/brown-like (bAT) adipose tissues. WAT is mainly involved in the storage and mobilization of energy and BAT is predominantly responsible for nonshivering thermogenesis. Recent data suggest that adipocyte mitochondria might play an important role in the development of obesity through defects in mitochondrial lipogenesis and lipolysis, regulation of adipocyte differentiation, apoptosis, production of oxygen radicals, efficiency of oxidative phosphorylation, and regulation of conversion of white adipocytes into brown-like adipocytes. This review summarizes the main characteristics of each adipose tissue subtype and describes morphological and functional modifications focusing on mitochondria and their activity in healthy and unhealthy adipocytes. PMID:27073398

  4. Involvement of IGF-1 and MEOX2 in PI3K/Akt1/2 and ERK1/2 pathways mediated proliferation and differentiation of perivascular adipocytes

    SciTech Connect

    Liu, Ping; Kong, Feng; Wang, Jue; Lu, Qinghua; Xu, Haijia; Qi, Tonggang; Meng, Juan

    2015-02-01

    Perivascular adipocyte (PVAC) proliferation and differentiation were closely involved in cardiovascular disease. We aimed to investigate whether phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling pathways enhance PVAC functions activated by insulin-like growth factor 1(IGF-1) and suppressed by mesenchyme homeobox 2 (MEOX2). In this study, PVACs from primary culture were cultured and induced to differentiate. Cell viability assays demonstrated that IGF-1 promoted PVAC proliferation and differentiation. However MEOX2 counteracted these IGF-1-mediated actions. Flow Cytometry revealed that IGF-1 increased S phase cells and decreased apoptosis; however, MEOX2 decreased S phase cells, increased G0–G1 phase cells, and promoted apoptosis. During PVAC proliferation and differentiation, IGF-1 activated PI3K/Akt1/2 and ERK1/2 signaling pathways, upregulated the expression of these signaling proteins and FAS, and increased PVAC lipid content. In contrast, MEOX2 constrained the phosphorylation of ERK1/2 and Akt1/2 protein, down-regulated these signaling molecules and FAS, and decreased PVAC lipid content. Instead, MEOX2 knockdown enhanced the ERK1/2 and Akt1/2 phosphorylation, augmented the expression of these signaling molecules and FAS, and increased PVAC lipid content. Our findings suggested that PI3K/Akt1/2 and ERK1/2 activation mediated by IGF-1 is essential for PVAC proliferation and differentiation, and MEOX2 is a promising therapeutic gene to intervene in the signaling pathways and inhibit PVAC functions. - Highlights: • IGF-1 activated PI3K/Akt2 and ERK1/2 pathways to mediate PVAC proliferation and differentiation. • The expression of ERK1, ERK 2, PI3K, Akt1 and Akt2 showed different change trends between PVAC proliferation and differentiation. • MEOX2 effectively expressed in PVAC, increased early and late cellular apoptosis, and inhibited its proliferation. • MEOX2 depressed PVAC differentiation and FAS expression

  5. 18O-Tracer Metabolomics Reveals Protein Turnover and CDP-Choline Cycle Activity in Differentiating 3T3-L1 Pre-Adipocytes

    PubMed Central

    Kirkwood, Jay S.; Miranda, Cristobal L.; Bobe, Gerd; Maier, Claudia S.; Stevens, Jan F.

    2016-01-01

    The differentiation of precursor cells into mature adipocytes (adipogenesis) has been an area of increased focus, spurred by a rise in obesity rates. Though our understanding of adipogenesis and its regulation at the cellular level is growing, many questions remain, especially regarding the regulation of the metabolome. The 3T3-L1 cell line is the most well characterized cellular model of adipogenesis. Using a time course metabolomics approach, we show that the 3T3-L1 preadipocyte metabolome is greatly altered during the first 48 hours of differentiation, where cells go through about two rounds of cell division, a process known as mitotic clonal expansion. Short-chain peptides were among several small molecules that were increased during mitotic clonal expansion. Additional indicators of protein turnover were also increased, including bilirubin, a degradation product of heme-containing proteins, and 3-methylhistidine, a post-translationally modified amino acid that is not reutilized for protein synthesis. To study the origin of the peptides, we treated differentiating preadipocytes with 18O labeled water and found that 18O was incorporated into the short chain peptides, confirming them, at least in part, as products of hydrolysis. Inhibitors of the proteasome or matrix metalloproteinases affected the peptide levels during differentiation, but inhibitors of autophagy or peptidases did not. 18O was also incorporated into several choline metabolites including cytidine 5'-diphosphocholine (CDP-choline), glycerophosphocholine, and several phosphatidylcholine species, indicative of phosphatidylcholine synthesis/degradation and of flux through the CDP-choline cycle, a hallmark of proliferating cells. 18O-Tracer metabolomics further showed metabolic labeling of glutamate, suggestive of glutaminolysis, also characteristic of proliferating cells. Together, these results highlight the utility of 18O isotope labeling in combination with metabolomics to uncover changes in

  6. 18O-Tracer Metabolomics Reveals Protein Turnover and CDP-Choline Cycle Activity in Differentiating 3T3-L1 Pre-Adipocytes.

    PubMed

    Kirkwood, Jay S; Miranda, Cristobal L; Bobe, Gerd; Maier, Claudia S; Stevens, Jan F

    2016-01-01

    The differentiation of precursor cells into mature adipocytes (adipogenesis) has been an area of increased focus, spurred by a rise in obesity rates. Though our understanding of adipogenesis and its regulation at the cellular level is growing, many questions remain, especially regarding the regulation of the metabolome. The 3T3-L1 cell line is the most well characterized cellular model of adipogenesis. Using a time course metabolomics approach, we show that the 3T3-L1 preadipocyte metabolome is greatly altered during the first 48 hours of differentiation, where cells go through about two rounds of cell division, a process known as mitotic clonal expansion. Short-chain peptides were among several small molecules that were increased during mitotic clonal expansion. Additional indicators of protein turnover were also increased, including bilirubin, a degradation product of heme-containing proteins, and 3-methylhistidine, a post-translationally modified amino acid that is not reutilized for protein synthesis. To study the origin of the peptides, we treated differentiating preadipocytes with 18O labeled water and found that 18O was incorporated into the short chain peptides, confirming them, at least in part, as products of hydrolysis. Inhibitors of the proteasome or matrix metalloproteinases affected the peptide levels during differentiation, but inhibitors of autophagy or peptidases did not. 18O was also incorporated into several choline metabolites including cytidine 5'-diphosphocholine (CDP-choline), glycerophosphocholine, and several phosphatidylcholine species, indicative of phosphatidylcholine synthesis/degradation and of flux through the CDP-choline cycle, a hallmark of proliferating cells. 18O-Tracer metabolomics further showed metabolic labeling of glutamate, suggestive of glutaminolysis, also characteristic of proliferating cells. Together, these results highlight the utility of 18O isotope labeling in combination with metabolomics to uncover changes in

  7. A heterogeneous lineage origin underlies the phenotypic and molecular differences of white and beige adipocytes

    PubMed Central

    Liu, Weiyi; Shan, Tizhong; Yang, Xin; Liang, Sandra; Zhang, Pengpeng; Liu, Yaqin; Liu, Xiaoqi; Kuang, Shihuan

    2013-01-01

    Summary A worldwide epidemic of obesity and its associated metabolic disorders raise the significance of adipocytes, their origins and characteristics. Our previous study has demonstrated that interscapular brown adipose tissue (BAT), but not intramuscular adipose, is derived from the Pax3-expressing cell lineage. Here, we show that various depots of subcutaneous (SAT) and visceral adipose tissue (VAT) are highly heterogeneous in the Pax3 lineage origin. Interestingly, the relative abundance of Pax3 lineage cells in SAT depots is inversely correlated to expression of BAT signature genes including Prdm16, Pgc1a (Ppargc1a) and Ucp1. FACS analysis further demonstrates that adipocytes differentiated from non-Pax3 lineage preadipocytes express higher levels of BAT and beige adipocyte signature genes compared with the Pax3 lineage adipocytes within the same depots. Although both Pax3 and non-Pax3 lineage preadipocytes can give rise to beige adipocytes, the latter contributes more significantly. Consistently, genetic ablation of Pax3 lineage cells in SAT leads to increased expression of beige cell markers. Finally, non-Pax3 lineage beige adipocytes are more responsive to cAMP-agonist-induced Ucp1 expression. Taken together, these results demonstrate widespread heterogeneity in Pax3 lineage origin, and its inverse association with BAT gene expression within and among subcutaneous adipose depots. PMID:23781029

  8. Adipocyte-specific CD1d-deficiency mitigates diet-induced obesity and insulin resistance in mice

    PubMed Central

    Satoh, Masashi; Hoshino, Miyuki; Fujita, Koki; Iizuka, Misao; Fujii, Satoshi; Clingan, Christopher S.; Van Kaer, Luc; Iwabuchi, Kazuya

    2016-01-01

    It has been shown that CD1d expression and glycolipid-reactive, CD1d-restricted NKT cells exacerbate the development of obesity and insulin resistance in mice. However, the relevant CD1d-expressing cells that influence the effects of NKT cells on the progression of obesity remain incompletely defined. In this study, we have demonstrated that 3T3-L1 adipocytes can present endogenous ligands to NKT cells, leading to IFN-γ production, which in turn, stimulated 3T3-L1 adipocytes to enhance expression of CD1d and CCL2, and decrease expression of adiponectin. Furthermore, adipocyte-specific CD1d deletion decreased the size of the visceral adipose tissue mass and enhanced insulin sensitivity in mice fed a high-fat diet (HFD). Accordingly, NKT cells were less activated, IFN-γ production was significantly reduced, and levels of adiponectin were increased in these animals as compared with control mice on HFD. Importantly, macrophage recruitment into the adipose tissue of adipocyte-specific CD1d-deficient mice was significantly blunted. These findings indicate that interactions between NKT cells and CD1d-expressing adipocytes producing endogenous NKT cell ligands play a critical role in the induction of inflammation and functional modulation of adipose tissue that leads to obesity. PMID:27329323

  9. Adipocyte-specific CD1d-deficiency mitigates diet-induced obesity and insulin resistance in mice.

    PubMed

    Satoh, Masashi; Hoshino, Miyuki; Fujita, Koki; Iizuka, Misao; Fujii, Satoshi; Clingan, Christopher S; Van Kaer, Luc; Iwabuchi, Kazuya

    2016-01-01

    It has been shown that CD1d expression and glycolipid-reactive, CD1d-restricted NKT cells exacerbate the development of obesity and insulin resistance in mice. However, the relevant CD1d-expressing cells that influence the effects of NKT cells on the progression of obesity remain incompletely defined. In this study, we have demonstrated that 3T3-L1 adipocytes can present endogenous ligands to NKT cells, leading to IFN-γ production, which in turn, stimulated 3T3-L1 adipocytes to enhance expression of CD1d and CCL2, and decrease expression of adiponectin. Furthermore, adipocyte-specific CD1d deletion decreased the size of the visceral adipose tissue mass and enhanced insulin sensitivity in mice fed a high-fat diet (HFD). Accordingly, NKT cells were less activated, IFN-γ production was significantly reduced, and levels of adiponectin were increased in these animals as compared with control mice on HFD. Importantly, macrophage recruitment into the adipose tissue of adipocyte-specific CD1d-deficient mice was significantly blunted. These findings indicate that interactions between NKT cells and CD1d-expressing adipocytes producing endogenous NKT cell ligands play a critical role in the induction of inflammation and functional modulation of adipose tissue that leads to obesity. PMID:27329323

  10. Adipocytes, like their progenitors, contribute to inflammation of adipose tissues through promotion of Th-17 cells and activation of monocytes, in obese subjects.

    PubMed

    Chehimi, Marwa; Robert, Maud; Bechwaty, Michel El; Vial, Guillaume; Rieusset, Jennifer; Vidal, Hubert; Pirola, Luciano; Eljaafari, Assia

    2016-01-01

    Recently, we have reported that adipose tissue-derived stem cells (ASC) harvested from obese donors induce a pro-inflammatory environment when co-cultured with peripheral blood mononuclear cells (MNC), with a polarization of T cells toward the Th17 cell lineage, increased secretion of IL-1β and IL-6 pro-inflammatory cytokines, and down-regulation of Th1 cytokines, such as IFNγ and TNFα. However, whether differentiated adipocytes, like the aforementioned ASC, are pro-inflammatory in obese subject AT remained to be investigated. Herein, we isolated ASC from AT of obese donors and differentiated them into adipocytes, for either 8 or 14 d. We analyzed their capacity to activate blood MNC after stimulation with phytohemagglutinin A (PHA), or not, in co-culture assays. Our results showed that co-cultures of MNC with adipocytes, like with ASC, increased IL-17A, IL-1β, and IL-6 pro-inflammatory cytokine secretion. Moreover, like ASC, adipocytes down-regulated TNFα secretion by Th1 cells. As adipocytes differentiated from ASC of lean donors also promoted IL-17A secretion by MNC, an experimental model of high-fat versus chow diet mice was used and supported that adipocytes from obese, but not lean AT, are able to mediate IL-17A secretion by PHA-activated MNCs. In conclusion, our results suggest that, as ASC, adipocytes in obese AT might contribute to the establishment of a low-grade chronic inflammation state. PMID:27617173

  11. Anti-inflammatory effect of resveratrol on TNF-{alpha}-induced MCP-1 expression in adipocytes

    SciTech Connect

    Zhu Jian; Yong Wei; Wu Xiaohong; Yu Ying; Lv Jinghuan; Liu Cuiping; Mao Xiaodong; Zhu Yunxia; Xu Kuanfeng; Han Xiao Liu Chao

    2008-05-02

    Chronic low-grade inflammation characterized by adipose tissue macrophage accumulation and abnormal cytokine production is a key feature of obesity and type 2 diabetes. Adipose-tissue-derived monocyte chemoattractant protein (MCP)-1, induced by cytokines, has been shown to play an essential role in the early events during macrophage infiltration into adipose tissue. In this study we investigated the effects of resveratrol upon both tumor necrosis factor (TNF)-{alpha}-induced MCP-1 gene expression and its underlying signaling pathways in 3T3-L1 adipoctyes. Resveratrol was found to inhibit TNF-{alpha}-induced MCP-1 secretion and gene transcription, as well as promoter activity, which based on down-regulation of TNF-{alpha}-induced MCP-1 transcription. Nuclear factor (NF)-{kappa}B was determined to play a major role in the TNF-{alpha}-induced MCP-1 expression. Further analysis showed that resveratrol inhibited DNA binding activity of the NF-{kappa}B complex and subsequently suppressed NF-{kappa}B transcriptional activity in TNF-{alpha}-stimulated cells. Finally, the inhibition of MCP-1 may represent a novel mechanism of resveratrol in preventing obesity-related pathologies.

  12. Pioglitazone enhances small-sized adipocyte proliferation in subcutaneous adipose tissue.

    PubMed

    Kajita, Kazuo; Mori, Ichiro; Hanamoto, Takayuki; Ikeda, Takahide; Fujioka, Kei; Yamauchi, Masahiro; Okada, Hideyuki; Usui, Taro; Takahashi, Noriko; Kitada, Yoshihiko; Taguchi, Kohichiro; Kajita, Toshiko; Uno, Yoshihiro; Morita, Hiroyuki; Ishizuka, Tatsuo

    2012-01-01

    The possibility that mature adipocytes proliferate has not been fully investigated. In this study, we demonstrate that adipocytes can proliferate. 5-bromo-2'-deoxyuridine (BrdU)-labeled adipocyte like cells, most of which were less than 30 μm in diameter, were observed in adipose tissue. Proliferating cell nuclear antigen (PCNA) was simultaneously detected in BrdU-labeled nuclei. Observation of individual mature adipocytes of smeared specimens on glass slides revealed that small sized adipocytes more frequently incorporated BrdU. Cultured mature adipocytes using the ceiling-cultured method showed clustering of proliferating cells in small-sized adipocytes. These small cultured adipocytes, but not large ones, extensively incorporated BrdU. Quantified analysis of BrdU incorporation demonstrated that mature visceral adipocytes, including epididymal, mesenteric and perirenal adipocytes, proliferated more actively than subcutaneous ones. On the other hand, treatment with pioglitazone (Pio), a ligand of peroxisome proliferator-activated receptor γ, containing food for 2w, elevated BrdU incorporation and expression of PCNA in mature adipocytes isolated from subcutaneous, but not visceral adipose tissue. Moreover, Pio induced increased BrdU-labeled small-sized subcutaneous adipocytes, which was associated with an increased number of total small adipocytes in subcutaneous adipose tissue. In conclusion, mature adipocytes have a subgroup representing the potential to replicate, and this proliferation is more active in visceral adipocytes. Treatment with Pio increases proliferation in subcutaneous adipocytes. These results may explain the mechanism of Pio-induced hyperplasia especially in subcutaneous adipocytes. PMID:22972172

  13. Lotus leaf extract and L-carnitine influence different processes during the adipocyte life cycle

    PubMed Central

    2010-01-01

    Background The cellular and molecular mechanisms of adipose tissue biology have been studied extensively over the last two decades. Adipose tissue growth involves both an increase in fat cell size and the formation of mature adipocytes from precursor cells. To investigate how natural substances influence these two processes, we examined the effects of lotus leaf extract (Nelumbo nucifera-extract solution obtained from Silab, France) and L-carnitine on human preadipocytes and adipocytes. Methods For our in vitro studies, we used a lotus leaf extract solution alone or in combination with L-carnitine. Utilizing cultured human preadipocytes, we investigated lotus leaf extract solution-induced inhibition of triglyceride incorporation during adipogenesis and possible effects on cell viability. Studies on human adipocytes were performed aiming to elucidate the efficacy of lotus leaf extract solution to stimulate lipolytic activity. To further characterize lotus leaf extract solution-mediated effects, we determined the expression of the transcription factor adipocyte determination and differentiation factor 1 (ADD1/SREBP-1c) on the RNA- and protein level utilizing qRT-PCR and immunofluorescence analysis. Additionally, the effect of L-carnitine on beta-oxidation was analyzed using human preadipocytes and mature adipocytes. Finally, we investigated additive effects of a combination of lotus leaf extract solution and L-carnitine on triglyceride accumulation during preadipocyte/adipocyte differentiation. Results Our data showed that incubation of preadipocytes with lotus leaf extract solution significantly decreased triglyceride accumulation during adipogenesis without affecting cell viability. Compared to controls, adipocytes incubated with lotus leaf extract solution exhibited a significant increase in lipolysis-activity. Moreover, cell populations cultivated in the presence of lotus leaf extract solution showed a decrease in adipocyte differentiation capacity as indicated

  14. Farnesyl pyrophosphate regulates adipocyte functions as an endogenous PPARγ agonist

    PubMed Central

    Goto, Tsuyoshi; Nagai, Hiroyuki; Egawa, Kahori; Kim, Young-Il; Kato, Sota; Taimatsu, Aki; Sakamoto, Tomoya; Ebisu, Shogo; Hohsaka, Takahiro; Miyagawa, Hiroh; Murakami, Shigeru; Takahashi, Nobuyuki; Kawada, Teruo

    2011-01-01

    The cholesterol biosynthetic pathway produces not only sterols but also non-sterol mevalonate metabolites involved in isoprenoid synthesis. Mevalonate metabolites affect transcriptional and post-transcriptional events that in turn affect various biological processes including energy metabolism. In the present study, we examine whether mevalonate metabolites activate PPARγ (peroxisome-proliferator-activated receptor γ), a ligand-dependent transcription factor playing a central role in adipocyte differentiation. In the luciferase reporter assay using both GAL4 chimaera and full-length PPARγ systems, a mevalonate metabolite, FPP (farnesyl pyrophosphate), which is the precursor of almost all isoprenoids and is positioned at branch points leading to the synthesis of other longer-chain isoprenoids, activated PPARγ in a dose-dependent manner. FPP induced the in vitro binding of a co-activator, SRC-1 (steroid receptor co-activator-1), to GST (glutathione transferase)–PPARγ. Direct binding of FPP to PPARγ was also indicated by docking simulation studies. Moreover, the addition of FPP up-regulated the mRNA expression levels of PPARγ target genes during adipocyte differentiation induction. In the presence of lovastatin, an HMG-CoA (3-hydroxy-3-methylglutaryl-CoA) reductase inhibitor, both intracellular FPP levels and PPARγ-target gene expressions were decreased. In contrast, the increase in intracellular FPP level after the addition of zaragozic acid, a squalene synthase inhibitor, induced PPARγ-target gene expression. The addition of FPP and zaragozic acid promotes lipid accumulation during adipocyte differentiation. These findings indicated that FPP might function as an endogenous PPARγ agonist and regulate gene expression in adipocytes. PMID:21605082

  15. Farnesyl pyrophosphate regulates adipocyte functions as an endogenous PPARγ agonist.

    PubMed

    Goto, Tsuyoshi; Nagai, Hiroyuki; Egawa, Kahori; Kim, Young-Il; Kato, Sota; Taimatsu, Aki; Sakamoto, Tomoya; Ebisu, Shogo; Hohsaka, Takahiro; Miyagawa, Hiroh; Murakami, Shigeru; Takahashi, Nobuyuki; Kawada, Teruo

    2011-08-15

    The cholesterol biosynthetic pathway produces not only sterols but also non-sterol mevalonate metabolites involved in isoprenoid synthesis. Mevalonate metabolites affect transcriptional and post-transcriptional events that in turn affect various biological processes including energy metabolism. In the present study, we examine whether mevalonate metabolites activate PPARγ (peroxisome-proliferator-activated receptor γ), a ligand-dependent transcription factor playing a central role in adipocyte differentiation. In the luciferase reporter assay using both GAL4 chimaera and full-length PPARγ systems, a mevalonate metabolite, FPP (farnesyl pyrophosphate), which is the precursor of almost all isoprenoids and is positioned at branch points leading to the synthesis of other longer-chain isoprenoids, activated PPARγ in a dose-dependent manner. FPP induced the in vitro binding of a co-activator, SRC-1 (steroid receptor co-activator-1), to GST (glutathione transferase)-PPARγ. Direct binding of FPP to PPARγ was also indicated by docking simulation studies. Moreover, the addition of FPP up-regulated the mRNA expression levels of PPARγ target genes during adipocyte differentiation induction. In the presence of lovastatin, an HMG-CoA (3-hydroxy-3-methylglutaryl-CoA) reductase inhibitor, both intracellular FPP levels and PPARγ-target gene expressions were decreased. In contrast, the increase in intracellular FPP level after the addition of zaragozic acid, a squalene synthase inhibitor, induced PPARγ-target gene expression. The addition of FPP and zaragozic acid promotes lipid accumulation during adipocyte differentiation. These findings indicated that FPP might function as an endogenous PPARγ agonist and regulate gene expression in adipocytes. PMID:21605082

  16. Targeting IκB kinase β in Adipocyte Lineage Cells for Treatment of Obesity and Metabolic Dysfunctions.

    PubMed

    Helsley, Robert N; Sui, Yipeng; Park, Se-Hyung; Liu, Zun; Lee, Richard G; Zhu, Beibei; Kern, Philip A; Zhou, Changcheng

    2016-07-01

    IκB kinase β (IKKβ), a central coordinator of inflammation through activation of nuclear factor-κB, has been identified as a potential therapeutic target for the treatment of obesity-associated metabolic dysfunctions. In this study, we evaluated an antisense oligonucleotide (ASO) inhibitor of IKKβ and found that IKKβ ASO ameliorated diet-induced metabolic dysfunctions in mice. Interestingly, IKKβ ASO also inhibited adipocyte differentiation and reduced adiposity in high-fat (HF)-fed mice, indicating an important role of IKKβ signaling in the regulation of adipocyte differentiation. Indeed, CRISPR/Cas9-mediated genomic deletion of IKKβ in 3T3-L1 preadipocytes blocked these cells differentiating into adipocytes. To further elucidate the role of adipose progenitor IKKβ signaling in diet-induced obesity, we generated mice that selectively lack IKKβ in the white adipose lineage and confirmed the essential role of IKKβ in mediating adipocyte differentiation in vivo. Deficiency of IKKβ decreased HF-elicited adipogenesis in addition to reducing inflammation and protected mice from diet-induced obesity and insulin resistance. Further, pharmacological inhibition of IKKβ also blocked human adipose stem cell differentiation. Our findings establish IKKβ as a pivotal regulator of adipogenesis and suggest that overnutrition-mediated IKKβ activation serves as an initial signal that triggers adipose progenitor cell differentiation in response to HF feeding. Inhibition of IKKβ with antisense therapy may represent as a novel therapeutic approach to combat obesity and metabolic dysfunctions. Stem Cells 2016;34:1883-1895. PMID:26991836

  17. Different anti-adipogenic effects of bio-compounds on primary visceral pre-adipocytes and adipocytes.

    PubMed

    Colitti, Monica; Stefanon, Bruno

    2016-01-01

    Several natural compounds exhibit strong capacity for decreasing triglyceride accumulation, enhancing lipolysis and inducing apoptosis. The present study reports the anti-adipogenic effects of Silybum marianum (SL), Citrus aurantium (CA), Taraxacum officinale (TO), resveratrol (RE), Curcuma longa (CU), caffeine (CF), oleuropein (OL) and docosahexaenoic acid (DHA) in reducing differentiation and increasing lipolysis and apoptosis. Analyses were performed on human primary visceral pre-adipocytes after 10 (P10) and 20 (P20) days of treatment during differentiation and on mature adipocytes after 7 days of treatment (A7). The percentage of apoptosis induced by TO extract in P10 and P20 cells was significantly higher than that induced by all other compounds and in CTRL cells. Triglyceride accumulation was significantly lower in cells treated with DHA, CF, RE in comparison to cells treated with OL and in CTRL cells. Treatments with CF, DHA and OL significantly incremented lipolysis in P20 cells in comparison to other compounds and in CTRL cells. On the contrary, the treatment of A7 cells with OL, CA and TO compounds significantly increased cell lipolysis. The addition of CF in differentiating P20 pre-adipocytes significantly increased the expression of genes involved in inhibition of adipogenesis, such as GATA2, GATA3, WNT1, WNT3A, SFRP5, and DLK1. Genes involved in promoting adipogenesis such as CCND1, CEBPB and SREBF1 were significantly down-regulated by the treatment. The screening of bioactive compounds for anti-adipogenic effects showed that in differentiating cells TO extract was the most effective in inducing apoptosis and CF and DHA extracts were more efficient in inhibition of differentiation and in induction of cell lipolysis. PMID:27540349

  18. Different anti-adipogenic effects of bio-compounds on primary visceral pre-adipocytes and adipocytes

    PubMed Central

    Colitti, Monica; Stefanon, Bruno

    2016-01-01

    Several natural compounds exhibit strong capacity for decreasing triglyceride accumulation, enhancing lipolysis and inducing apoptosis. The present study reports the anti-adipogenic effects of Silybum marianum (SL), Citrus aurantium (CA), Taraxacum officinale (TO), resveratrol (RE), Curcuma longa (CU), caffeine (CF), oleuropein (OL) and docosahexaenoic acid (DHA) in reducing differentiation and increasing lipolysis and apoptosis. Analyses were performed on human primary visceral pre-adipocytes after 10 (P10) and 20 (P20) days of treatment during differentiation and on mature adipocytes after 7 days of treatment (A7). The percentage of apoptosis induced by TO extract in P10 and P20 cells was significantly higher than that induced by all other compounds and in CTRL cells. Triglyceride accumulation was significantly lower in cells treated with DHA, CF, RE in comparison to cells treated with OL and in CTRL cells. Treatments with CF, DHA and OL significantly incremented lipolysis in P20 cells in comparison to other compounds and in CTRL cells. On the contrary, the treatment of A7 cells with OL, CA and TO compounds significantly increased cell lipolysis. The addition of CF in differentiating P20 pre-adipocytes significantly increased the expression of genes involved in inhibition of adipogenesis, such as GATA2, GATA3, WNT1, WNT3A, SFRP5, and DLK1. Genes involved in promoting adipogenesis such as CCND1, CEBPB and SREBF1 were significantly down-regulated by the treatment. The screening of bioactive compounds for anti-adipogenic effects showed that in differentiating cells TO extract was the most effective in inducing apoptosis and CF and DHA extracts were more efficient in inhibition of differentiation and in induction of cell lipolysis. PMID:27540349

  19. Emerging Complexities in Adipocyte Origins and Identity.

    PubMed

    Sanchez-Gurmaches, Joan; Hung, Chien-Min; Guertin, David A

    2016-05-01

    The global incidence of obesity and its comorbidities continues to rise along with a demand for novel therapeutic interventions. Brown adipose tissue (BAT) is attracting attention as a therapeutic target because of its presence in adult humans and high capacity to dissipate energy as heat, and thus burn excess calories, when stimulated. Another potential avenue for therapeutic intervention is to induce, within white adipose tissue (WAT), the formation of brown-like adipocytes called brite (brown-like-in-white) or beige adipocytes. However, understanding how to harness the potential of these thermogenic cells requires a deep understanding of their developmental origins and regulation. Recent cell-labeling and lineage-tracing experiments are beginning to shed light on this emerging area of adipocyte biology. We review here adipocyte development, giving particular attention to thermogenic adipocytes. PMID:26874575

  20. Endoplasmic reticulum-mitochondrial interaction mediated by mitofusin-1 or mitofusin-2 is not required for lipid droplet formation or adipocyte differentiation.

    PubMed

    McFie, Pamela J; Ambilwade, Prashant; Vu, Huyen; Stone, Scot J

    2016-09-01

    Organelles in cells physically interact with each other. Specifically, the interaction of ER and mitochondria has been shown to be important for transporting lipids between these two organelles. Lipid droplets are also closely associated with both the ER and mitochondria suggesting the interaction of ER and mitochondria may be important for triacylglycerol storage in lipid droplets. We tested the hypothesis that the efficient synthesis and storage of triacylglycerol in lipid droplets is dependent on the interaction of the ER and mitochondria using mouse embryonic fibroblasts lacking mitofusin-2 (Mfn2). Mfn2 is a GTPase that is present in mitochondrial-associated membranes (MAM) and is also present in the outer mitochondrial membrane. Mfn2 in MAM and mitochondria interact forming an interorganellar bridge. Cells lacking Mfn2 have loose ER-mitochondria contact. We found that mouse embryonic fibroblasts lacking Mfn2 have altered lipid droplet morphology. However, triacylglycerol biosynthesis was not dependent on ER-mitochondrial tethering mediated by mitofusins. Lastly, Mfn2 does not have a role in adipocyte differentiation. PMID:27404125

  1. Fad104, a Positive Regulator of Adipocyte Differentiation, Suppresses Invasion and Metastasis of Melanoma Cells by Inhibition of STAT3 Activity

    PubMed Central

    Katoh, Daiki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2015-01-01

    Metastasis is the main cause of death in patients with cancer, and understanding the mechanisms of metastatic processes is essential for the development of cancer therapy. Although the role of several cell adhesion, migration or proliferation molecules in metastasis is established, a novel target for cancer therapy remains to be discovered. Previously, we reported that fad104 (factor for adipocyte differentiation 104), a regulatory factor of adipogenesis, regulates cell adhesion and migration. In this report, we clarify the role of fad104 in the invasion and metastasis of cancer cells. The expression level of fad104 in highly metastatic melanoma A375SM cells was lower than that in poorly metastatic melanoma A375C6 cells. Reduction of fad104 expression enhanced the migration and invasion of melanoma cells, while over-expression of FAD104 inhibited migration and invasion. In addition, melanoma cells stably expressing FAD104 showed a reduction in formation of lung colonization compared with control cells. FAD104 interacted with STAT3 and down-regulated the phosphorylation level of STAT3 in melanoma cells. These findings together demonstrate that fad104 suppressed the invasion and metastasis of melanoma cells by inhibiting activation of the STAT3 signaling pathway. These findings will aid a comprehensive description of the mechanism that controls the invasion and metastasis of cancer cells. PMID:25671570

  2. Calcium-induced alteration of mitochondrial morphology and mitochondrial-endoplasmic reticulum contacts in rat brown adipocytes.

    PubMed

    Golic, I; Velickovic, K; Markelic, M; Stancic, A; Jankovic, A; Vucetic, M; Otasevic, V; Buzadzic, B; Korac, B; Korac, A

    2014-01-01

    Mitochondria are key organelles maintaining cellular bioenergetics and integrity, and their regulation of [Ca2+]i homeostasis has been investigated in many cell types. We investigated the short-term Ca-SANDOZ® treatment on brown adipocyte mitochondria, using imaging and molecular biology techniques. Two-month-old male Wistar rats were divided into two groups: Ca-SANDOZ® drinking or tap water (control) drinking for three days. Alizarin Red S staining showed increased Ca2+ level in the brown adipocytes of treated rats, and potassium pyroantimonate staining localized electron-dense regions in the cytoplasm, mitochondria and around lipid droplets. Ca-SANDOZ® decreased mitochondrial number, but increased their size and mitochondrial cristae volume. Transmission electron microscopy revealed numerous enlarged and fusioned-like mitochondria in the Ca-SANDOZ® treated group compared to the control, and megamitochondria in some brown adipocytes. The Ca2+ diet affected mitochondrial fusion as mitofusin 1 (MFN1) and mitofusin 2 (MFN2) were increased, and mitochondrial fission as dynamin related protein 1 (DRP1) was decreased. Confocal microscopy showed a higher colocalization rate between functional mitochondria and endoplasmic reticulum (ER). The level of uncoupling protein-1 (UCP1) was elevated, which was confirmed by immunohistochemistry and Western blot analysis. These results suggest that Ca-SANDOZ® stimulates mitochondrial fusion, increases mitochondrial-ER contacts and the thermogenic capacity of brown adipocytes. PMID:25308841

  3. Susceptibility of brown adipocytes to pro-inflammatory cytokine toxicity and reactive oxygen species

    PubMed Central

    Rebiger, Lars; Lenzen, Sigurd; Mehmeti, Ilir

    2016-01-01

    Brown adipose tissue (BAT) cells have a very high oxidative capacity. On the other hand, in obesity and obesity-related diabetes, levels of pro-inflammatory cytokines are elevated, which might promote BAT dysfunction and consequently impair carbohydrate metabolism and thereby exacerbate cellular dysfunction and promote diabetes progression. Therefore, the antioxidative enzyme status of a brown adipocyte cell line and its susceptibility towards pro-inflammatory cytokines, which participate in the pathogenesis of diabetes, and reactive oxygen species (ROS) were analysed. Mature brown adipocytes exhibited significantly higher levels of expression of mitochondrially and peroxisomally located antioxidative enzymes compared with non-differentiated brown adipocytes. Pro-inflammatory cytokines induced a significant decrease in the viability of differentiated brown adipocytes, which was accompanied by a massive ROS production and down-regulation of BAT-specific markers, such as uncoupling protein 1 (UCP-1) and β-Klotho. Taken together, the results strongly indicate that pro-inflammatory cytokines cause brown adipocyte dysfunction and death through suppression of BAT-specific proteins, especially of UCP-1 and β-Klotho, and consequently increased oxidative stress. PMID:26795216

  4. Nitrite augments glucose uptake in adipocytes through the Protein Kinase A-dependent stimulation of mitochondrial fusion

    PubMed Central

    Khoo, Nicholas K.H.; Mo, Li; Zharikov, Sergey; Kamga, Christelle; Quesnelle, Kelly; Golin-Bisello, Franca; Li, Lihua; Wang, Yinna; Shiva, Sruti

    2014-01-01

    Though it is well accepted that adipose tissue is central in the regulation of glycemic homeostasis, the molecular mechanisms governing adipocyte glucose uptake remain unclear. Recent studies demonstrate that mitochondrial dynamics (fission and fusion) regulate lipid accumulation and differentiation in adipocytes. However, the role of mitochondrial dynamics in glucose homeostasis has not been explored. The nitric oxide oxidation products nitrite and nitrate are endogenous signaling molecules and dietary constituents that have recently been shown to modulate glucose metabolism, prevent weight gain and reverse the development of metabolic syndrome in mice. While the mechanism of this protection is unclear, the mitochondrion is a known subcellular target for nitrite signaling. Thus, we hypothesize that nitrite modulates mitochondrial dynamics and function to regulate glucose uptake in adipocytes. Herein, we demonstrate that nitrite significantly increases glucose uptake in differentiated murine adipocytes through a mechanism dependent on mitochondrial fusion. Specifically, nitrite promotes mitochondrial fusion by increasing pro-fusion protein mitofusin 1 while concomitantly activating protein kinase A (PKA), which phosphorylates and inhibits the pro-fission protein, dynamin-related protein 1 (Drp1). Functionally, this signaling augments cellular respiration, fatty acid oxidation, mitochondrial oxidant production and glucose uptake. Importantly, inhibition of PKA or Drp1 significantly attenuates nitrite-induced mitochondrial respiration and glucose uptake. These findings demonstrate that mitochondria play an essential metabolic role in adipocytes, a novel role for both nitrite and mitochondrial fusion in regulating adipocyte glucose homeostasis and have implications for the potential therapeutic use of nitrite and mitochondrial modulators in glycemic regulation. PMID:24556414

  5. Effects of accumulation of lipid droplets on load transfer between and within adipocytes.

    PubMed

    Ben-Or Frank, Mor; Shoham, Naama; Benayahu, Dafna; Gefen, Amit

    2015-01-01

    Adipogenesis, a process of cell proliferation followed by the accumulation of lipid droplets (LDs), is accompanied by morphological changes in adipocytes, leading to a gradual rise in the structural stiffness of these cells. The increase in cellular structural stiffness can potentially influence the localized deformations of adjacent adipocytes in weight-bearing fat tissues, which, based on previous work, may accelerate intracytoplasmatic lipid production to form even larger and more tightly packed intracellular LDs. This process is based on mechanotransduction phenomena which are hypothesized (again, following empirical studies), to play a critical role in "en mass" adipocyte hypertrophy, and hence are important to characterize through computational modeling. Accordingly, we examined here how maturing adipocytes may affect localized loads acting on adjacent immature cells, using a set of finite element models of adipocytes embedded in an extracellular matrix. The peak strain energy density at the plasma membrane (PM) of the adipocytes, when constructs were externally loaded, was found to depend on the levels of lipid accumulation in the neighboring cells if the external compressive and shear deformations were large enough ([Formula: see text] and [Formula: see text], respectively). The mechanosignaling transduces through the PM and could therefore affect intracellular pathways to produce more lipid contents. Our results support the theory of deformation-induced differentiation in adipocytes. The findings are thus relevant in the context of a sedentary lifestyle, in which sustained deformations of weight-bearing adipose tissues may activate a positive feedback loop that promotes the "en mass" differentiation of cells, which subsequently increases the total mass of living fat tissues. PMID:24706071

  6. Regulation of adipocyte lipolysis.

    PubMed

    Frühbeck, Gema; Méndez-Giménez, Leire; Fernández-Formoso, José-Antonio; Fernández, Secundino; Rodríguez, Amaia

    2014-06-01

    In adipocytes the hydrolysis of TAG to produce fatty acids and glycerol under fasting conditions or times of elevated energy demands is tightly regulated by neuroendocrine signals, resulting in the activation of lipolytic enzymes. Among the classic regulators of lipolysis, adrenergic stimulation and the insulin-mediated control of lipid mobilisation are the best known. Initially, hormone-sensitive lipase (HSL) was thought to be the rate-limiting enzyme of the first lipolytic step, while we now know that adipocyte TAG lipase is the key enzyme for lipolysis initiation. Pivotal, previously unsuspected components have also been identified at the protective interface of the lipid droplet surface and in the signalling pathways that control lipolysis. Perilipin, comparative gene identification-58 (CGI-58) and other proteins of the lipid droplet surface are currently known to be key regulators of the lipolytic machinery, protecting or exposing the TAG core of the droplet to lipases. The neuroendocrine control of lipolysis is prototypically exerted by catecholaminergic stimulation and insulin-induced suppression, both of which affect cyclic AMP levels and hence the protein kinase A-mediated phosphorylation of HSL and perilipin. Interestingly, in recent decades adipose tissue has been shown to secrete a large number of adipokines, which exert direct effects on lipolysis, while adipocytes reportedly express a wide range of receptors for signals involved in lipid mobilisation. Recently recognised mediators of lipolysis include some adipokines, structural membrane proteins, atrial natriuretic peptides, AMP-activated protein kinase and mitogen-activated protein kinase. Lipolysis needs to be reanalysed from the broader perspective of its specific physiological or pathological context since basal or stimulated lipolytic rates occur under diverse conditions and by different mechanisms. PMID:24872083

  7. Troglitazone induces differentiation in Trypanosoma brucei

    SciTech Connect

    Denninger, Viola; Figarella, Katherine; Schoenfeld, Caroline; Brems, Stefanie; Busold, Christian; Lang, Florian; Hoheisel, Joerg; Duszenko, Michael . E-mail: michael.duszenko@uni-tuebingen.de

    2007-05-15

    Trypanosoma brucei, a protozoan parasite causing sleeping sickness, is transmitted by the tsetse fly and undergoes a complex lifecycle including several defined stages within the insect vector and its mammalian host. In the latter, differentiation from the long slender to the short stumpy form is induced by a yet unknown factor of trypanosomal origin. Here we describe that some thiazolidinediones are also able to induce differentiation. In higher eukaryotes, thiazolidinediones are involved in metabolism and differentiation processes mainly by binding to the intracellular receptor peroxisome proliferator activated receptor {gamma}. Our studies focus on the effects of troglitazone on bloodstream form trypanosomes. Differentiation was monitored using mitochondrial markers (membrane potential, succinate dehydrogenase activity, inhibition of oxygen uptake by KCN, amount of cytochrome transcripts), morphological changes (Transmission EM and light microscopy), and transformation experiments (loss of the Variant Surface Glycoprotein coat and increase of dihydroliponamide dehydrogenase activity). To further investigate the mechanisms responsible for these changes, microarray analyses were performed, showing an upregulation of expression site associated gene 8 (ESAG8), a potential differentiation regulator.

  8. Early soy exposure via maternal diet regulates rat mammary epithelial differentiation by paracrine signaling from stromal adipocytes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Diet-mediated changes in transcriptional programs that promote the early differentiation of the mammary gland may lead to reduced breast cancer risk. The disparity in adult breast cancer incidence between Asian women and Western counterparts is attributed partly to high soy food intake. Here, we con...

  9. Genistein-mediated inhibition of mammary stromal adipocyte differentiation limits expansion of mammary stem/progenitor cells by paracrine signaling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mammary adiposity may contribute to breast cancer development and progression by releasing cytokines and other inflammatory mediators that promote mammary epithelial proliferation. We evaluated the effects of soy isoflavone genistein (GEN) on the adipogenic differentiation of a SV40-immortalized mou...

  10. SPARC is over-expressed in adipose tissues of diet-induced obese rats and causes insulin resistance in 3T3-L1 adipocytes.

    PubMed

    Shen, Yang; Zhao, Yuyan; Yuan, Lizhi; Yi, Wei; Zhao, Rui; Yi, Qianru; Yong, Tongwu

    2014-01-01

    Secreted protein acidic and rich in cysteine (SPARC) is a secretory multifunctional matricellular glycoprotein. High circulating levels of SPARC have been reported to be associated with obesity and insulin resistance. The aim of the present study was to investigate whether SPARC induces insulin resistance and mitochondrial dysfunction in adipocytes. Our results showed that feeding high fat diet to rats for 12 weeks significantly increased SPARC expression in adipose tissues at both mRNA and protein levels. Moreover, SPARC overexpression in stably transfected 3T3-L1 cells induced insulin resistance and mitochondrial dysfunction, as evidenced by inhibition of insulin-stimulated glucose transport, lower ATP synthesis and mitochondrial membrane potential, reduced expression of glucose transporter 4 (GLUT4), and increased levels of reactive oxygen species (ROS) in mature adipocytes. Finally, overexpression of SPARC also modulated the expression levels of several inflammatory cytokines, which play important roles in insulin resistance, glucose and lipid metabolism during adipogenesis. In conclusion, our data suggest that SPARC is involved in obesity-induced adipose insulin resistance and may serve as a potential target in the treatment of obesity and obesity-related insulin resistance. PMID:23910024

  11. Transcriptional activation of peroxisome proliferator-activated receptor-{gamma} requires activation of both protein kinase A and Akt during adipocyte differentiation

    SciTech Connect

    Kim, Sang-pil; Ha, Jung Min; Yun, Sung Ji; Kim, Eun Kyoung; Chung, Sung Woon; Hong, Ki Whan; Kim, Chi Dae; Bae, Sun Sik

    2010-08-13

    Research highlights: {yields} Elevated cAMP activates both PKA and Epac. {yields} PKA activates CREB transcriptional factor and Epac activates PI3K/Akt pathway via Rap1. {yields} Akt modulates PPAR-{gamma} transcriptional activity in concert with CREB. -- Abstract: Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is required for the conversion of pre-adipocytes. However, the mechanism underlying activation of PPAR-{gamma} is unclear. Here we showed that cAMP-induced activation of protein kinase A (PKA) and Akt is essential for the transcriptional activation of PPAR-{gamma}. Hormonal induction of adipogenesis was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), by a protein kinase A (PKA) inhibitor (H89), and by a Rap1 inhibitor (GGTI-298). Transcriptional activity of PPAR-{gamma} was markedly enhanced by 3-isobutyl-1-methylxanthine (IBMX), but not insulin and dexamethasone. In addition, IBMX-induced PPAR-{gamma} transcriptional activity was blocked by PI3K/Akt, PKA, or Rap1 inhibitors. 8-(4-Chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT-2'-O-Me-cAMP) which is a specific agonist for exchanger protein directly activated by cAMP (Epac) significantly induced the activation of Akt. Furthermore, knock-down of Akt1 markedly attenuated PPAR-{gamma} transcriptional activity. These results indicate that both PKA and Akt signaling pathways are required for transcriptional activation of PPAR-{gamma}, suggesting post-translational activation of PPAR-{gamma} might be critical step for adipogenic gene expression.

  12. Tumor necrosis factor-induced reversal of adipocytic phenotype of 3T3-L1 cells is preceded by a loss of nuclear CCAAT/enhancer binding protein (C/EBP).

    PubMed Central

    Ron, D; Brasier, A R; McGehee, R E; Habener, J F

    1992-01-01

    Tumor necrosis factor (TNF)-treated 3T3-L1 adipocytes were used as a model for studying the effects of systemic inflammation on adipose tissue. Lipopolysaccharide-treated monocyte-conditioned medium or recombinant human TNF alpha induced morphological dedifferentiation of the adipocytes and led to loss of adipocyte specific gene expression. Gel shift, Southwestern and Western immunoblot analysis demonstrated that dedifferentiation was preceded by a decrease in the DNA binding activity and protein level of the transcription factor CCAAT/enhancer binding protein (C/EBP). Liver activating protein, a related protein that binds identical DNA sequences, increased during cytokine treatment. Both proteins activate specific enhancer elements located in the promoter region of many genes whose transcription is altered during systemic inflammation. Pulse-chase labeling followed by immunoprecipitation demonstrated that C/EBP is a rapidly turning over protein in adipocytes and that cytokine treatment led to a specific, time dependent decrease in its rate of synthesis. Because C/EBP binding sites have been shown to play an important role in regulating the expression of genes involved in adipocyte metabolism, we propose that the TNF-induced changes in the complement of transcription factors binding those sites may be important in the pathogenesis of inflammation-induced atrophy of adipose tissue. Images PMID:1729273

  13. Induced differentiation inhibits sphere formation in neuroblastoma.

    PubMed

    Craig, Brian T; Rellinger, Eric J; Alvarez, Alexandra L; Dusek, Haley L; Qiao, Jingbo; Chung, Dai H

    2016-08-19

    Neuroblastoma arises from the neural crest, the precursor cells of the sympathoadrenal axis, and differentiation status is a key prognostic factor used for clinical risk group stratification and treatment strategies. Neuroblastoma tumor-initiating cells have been successfully isolated from patient tumor samples and bone marrow using sphere culture, which is well established to promote growth of neural crest stem cells. However, accurate quantification of sphere-forming frequency of commonly used neuroblastoma cell lines has not been reported. Here, we show that MYCN-amplified neuroblastoma cell lines form spheres more frequently than non-MYCN-amplified cell lines. We also show that sphere formation is directly sensitive to cellular differentiation status. 13-cis-retinoic acid is a clinically used differentiating agent that induces a neuronal phenotype in neuroblastoma cells. Induced differentiation nearly completely blocked sphere formation. Furthermore, sphere formation was specifically FGF-responsive and did not respond to increasing doses of EGF. Taken together, these data suggest that sphere formation is an accurate method of quantifying the stemness phenotype in neuroblastoma. PMID:27297102

  14. ATF3 inhibits PPARγ-stimulated transactivation in adipocyte cells

    SciTech Connect

    Jang, Min-Kyung; Jung, Myeong Ho

    2015-01-02

    Highlights: • ATF3 inhibits PPARγ-stimulated transcriptional activation. • ATF3 interacts with PPARγ. • ATF3 suppresses p300-mediated transcriptional coactivation. • ATF3 decreases the binding of PPARγ and recruitment of p300 to PPRE. - Abstract: Previously, we reported that activating transcription factor 3 (ATF3) downregulates peroxisome proliferator activated receptor (PPARγ) gene expression and inhibits adipocyte differentiation in 3T3-L1 cells. Here, we investigated another role of ATF3 on the regulation of PPARγ activity. ATF3 inhibited PPARγ-stimulated transactivation of PPARγ responsive element (PPRE)-containing reporter or GAL4/PPARγ chimeric reporter. Thus, ATF3 effectively repressed rosiglitazone-stimulated expression of adipocyte fatty acid binding protein (aP2), PPARγ target gene, in 3T3-L1 cells. Coimmunoprecipitation and GST pulldown assay demonstrated that ATF3 interacted with PPARγ. Accordingly, ATF3 prevented PPARγ from binding to PPRE on the aP2 promoter. Furthermore, ATF3 suppressed p300-mediated transcriptional coactivation of PPRE-containing reporter. Chromatin immunoprecipitation assay showed that overexpression of ATF3 blocked both binding of PPARγ and recruitment of p300 to PPRE on aP2 promoter induced by rosiglitazone treatment in 3T3-L1 cells. Taken together, these results suggest that ATF3 interacts with PPARγ and represses PPARγ-mediated transactivation through suppression of p300-stimulated coactivation in 3T3-L1 cells, which may play a role in inhibition of adipocyte differentiation.

  15. 1,25-dihydroxyvitamin D3 Protects against Macrophage-Induced Activation of NFκB and MAPK Signalling and Chemokine Release in Human Adipocytes

    PubMed Central

    Ding, Cherlyn; Wilding, John P. H.; Bing, Chen

    2013-01-01

    Increased accumulation of macrophages in adipose tissue in obesity is linked to low-grade chronic inflammation, and associated with features of metabolic syndrome. Vitamin D3 may have immunoregulatory effects and reduce adipose tissue inflammation, although the molecular mechanisms remain to be established. This study investigated the effects of vitamin D3 on macrophage-elicited inflammatory responses in cultured human adipocytes, particularly the signalling pathways involved. Macrophage-conditioned (MC) medium (25% with adipocyte maintenance media) markedly inhibited protein expression of the nuclear factor-κB (NFκB) subunit inhibitor κBα (IκBα) (71%, P<0.001) and increased NFκB p65 (1.5-fold, P = 0.026) compared with controls. Treatment with 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) abolished macrophage-induced activation of NFκB signalling by increasing IκBα expression (2.7-fold, P = 0.005) and reducing NFκB p65 phosphorylation (68%; P<0.001). The mitogen-activated protein kinase (MAPK) signalling was activated by MC medium, which was also blunted by 1,25(OH)2D3 with a downregulation of phosphorylated p38 MAPK (32%, P = 0.005) and phosphorylated Erk1/2 (49%, P = 0.001). Furthermore, MC medium (12.5% or 25%) dose-dependently upregulated secretion of key proinflammatory chemokines/cytokines (22-368-fold; all P<0.001) and this was significantly decreased by 1,25(OH)2D3: IL-8 (61% and 31%, P<0.001), MCP-1 (37%, P<0.001 and 36%, P = 0.002), RANTES (78% and 62%, P<0.001) and IL-6 (29%, P<0.001 and 34%, P = 0.019). Monocyte migration-elicited by adipocytes treated with 1,25(OH)2D3 was also reduced (up to 25%, P<0.001). In conclusion, vitamin D3 could be anti-inflammatory in adipose tissue, decreasing macrophage-induced release of chemokines and cytokines by adipocytes and the chemotaxis of monocytes. Our data suggests these effects are mediated by inhibition of the NFκB and MAPK signalling pathways. PMID:23637889

  16. Arsenic Induces Insulin Resistance in Mouse Adipocytes and Myotubes Via Oxidative Stress-Regulated Mitochondrial Sirt3-FOXO3a Signaling Pathway.

    PubMed

    Padmaja Divya, Sasidharan; Pratheeshkumar, Poyil; Son, Young-Ok; Vinod Roy, Ram; Andrew Hitron, John; Kim, Donghern; Dai, Jin; Wang, Lei; Asha, Padmaja; Huang, Bin; Xu, Mei; Luo, Jia; Zhang, Zhuo

    2015-08-01

    Chronic exposure to arsenic via drinking water is associated with an increased risk for development of type 2 diabetes mellitus (T2DM). This study investigates the role of mitochondrial oxidative stress protein Sirtuin 3 (Sirt3) and its targeting proteins in chronic arsenic-induced T2DM in mouse adipocytes and myotubes. The results show that chronic arsenic exposure significantly decreased insulin-stimulated glucose uptake (ISGU) in correlation with reduced expression of insulin-regulated glucose transporter type 4 (Glut4). Expression of Sirt3, a mitochondrial deacetylase, was dramatically decreased along with its associated transcription factor, forkhead box O3 (FOXO3a) upon arsenic exposure. A decrease in mitochondrial membrane potential (Δψm) was observed in both 3T3L1 adipocytes and C2C12 myotubes treated by arsenic. Reduced FOXO3a activity by arsenic exhibited a decreased binding affinity to the promoters of both manganese superoxide dismutase (MnSOD) and peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1α, a broad and powerful regulator of reactive oxygen species (ROS) metabolism. Forced expression of Sirt3 or MnSOD in mouse myotubes elevated Δψm and restored ISGU inhibited by arsenic exposure. Our results suggest that Sirt3/FOXO3a/MnSOD signaling plays a significant role in the inhibition of ISGU induced by chronic arsenic exposure. PMID:25979314

  17. beta. -Adrenergic stimulation of brown adipocyte proliferation

    SciTech Connect

    Geloeen, A.; Collet, A.J.; Guay, G.; Bukowiecki, L.J. Laboratoire de Thermoregulation et Metabolisme Energetique, Lyon )

    1988-01-01

    The mechanisms of brown adipose tissue (BAT) growth were studied by quantitative photonic radioautography using tritiated thymidine to follow mitotic activity. To identify the nature of the adrenergic pathways mediating brown adipocyte proliferation and differentiation, the effects of cold exposure (4 days at 4{degree}C) on BAT growth were compared with those induced by treating rats at 25{degree}C with norepinephrine (a mixed agonist), isoproterenol (a {beta}-agonist), and phenylephrine (an {alpha}-agonist). Norepinephrine mimicked the effects of cold exposure, not only on the mitotic activity, but also on the distribution of the labeling among the various cellular types. Isoproterenol entirely reproduced the effects of norepinephrine both on the labeling index and on the cellular type labeling frequency. These results demonstrate that norepinephrine triggers a coordinated proliferation of brown adipocytes and endothelial cells in warm-exposed rats that is similar to that observed after cold exposure. They also suggest that cold exposure stimulates BAT growth by increasing the release of norepinephrine from sympathetic nerves and that the neurohormone activates mitoses in BAT precursor cells via {beta}-adrenergic pathways.

  18. Transgelin is a TGFβ-inducible gene that regulates osteoblastic and adipogenic differentiation of human skeletal stem cells through actin cytoskeleston organization.

    PubMed

    Elsafadi, M; Manikandan, M; Dawud, R A; Alajez, N M; Hamam, R; Alfayez, M; Kassem, M; Aldahmash, A; Mahmood, A

    2016-01-01

    Regenerative medicine is a novel approach for treating conditions in which enhanced bone regeneration is required. We identified transgelin (TAGLN), a transforming growth factor beta (TGFβ)-inducible gene, as an upregulated gene during in vitro osteoblastic and adipocytic differentiation of human bone marrow-derived stromal (skeletal) stem cells (hMSC). siRNA-mediated gene silencing of TAGLN impaired lineage differentiation into osteoblasts and adipocytes but enhanced cell proliferation. Additional functional studies revealed that TAGLN deficiency impaired hMSC cell motility and in vitro transwell cell migration. On the other hand, TAGLN overexpression reduced hMSC cell proliferation, but enhanced cell migration, osteoblastic and adipocytic differentiation, and in vivo bone formation. In addition, deficiency or overexpression of TAGLN in hMSC was associated with significant changes in cellular and nuclear morphology and cytoplasmic organelle composition as demonstrated by high content imaging and transmission electron microscopy that revealed pronounced alterations in the distribution of the actin filament and changes in cytoskeletal organization. Molecular signature of TAGLN-deficient hMSC showed that several genes and genetic pathways associated with cell differentiation, including regulation of actin cytoskeleton and focal adhesion pathways, were downregulated. Our data demonstrate that TAGLN has a role in generating committed progenitor cells from undifferentiated hMSC by regulating cytoskeleton organization. Targeting TAGLN is a plausible approach to enrich for committed hMSC cells needed for regenerative medicine application. PMID:27490926

  19. Bitter melon (Momordica charantia L.) inhibits adipocyte hypertrophy and down regulates lipogenic gene expression in adipose tissue of diet-induced obese rats.

    PubMed

    Huang, Hui-Ling; Hong, Ya-Wen; Wong, You-Hong; Chen, Ying-Nien; Chyuan, Jong-Ho; Huang, Ching-Jang; Chao, Pei-Min

    2008-02-01

    Bitter melon (Momordica charantia; BM) has been shown to ameliorate diet-induced obesity and insulin resistance. To examine the effect of BM supplementation on cell size and lipid metabolism in adipose tissues, three groups of rats were respectively fed a high-fat diet supplemented without (HF group) or with 5 % lyophilised BM powder (HFB group), or with 0.01 % thiazolidinedione (TZD) (HFT group). A group of rats fed a low-fat diet was also included as a normal control. Hyperinsulinaemia and glucose intolerance were observed in the HF group but not in HFT and HFB groups. Although the number of large adipocytes (>180 microm) of both the HFB and HFT groups was significantly lower than that of the HF group, the adipose tissue mass, TAG content and glycerol-3-phosphate dehydrogenase activity of the HFB group were significantly lower than those of the HFT group, implying that BM might reduce lipogenesis in adipose tissue. Experiment 2 was then conducted to examine the expression of lipogenic genes in adipose tissues of rats fed low-fat, HF or HFB diets. The HFB group showed significantly lower mRNA levels of fatty acid synthase, acetyl-CoA carboxylase-1, lipoprotein lipase and adipocyte fatty acid-binding protein than the HF group (P < 0.05). These results indicate BM can reduce insulin resistance as effective as the anti-diabetic drug TZD. Furthermore, BM can suppress the visceral fat accumulation and inhibit adipocyte hypertrophy, which may be associated with markedly down regulated expressions of lipogenic genes in the adipose. PMID:17651527

  20. Anti-Obesity Effects of the Mixture of Eriobotrya japonica and Nelumbo nucifera in Adipocytes and High-Fat Diet-Induced Obese Mice.

    PubMed

    Sharma, Bhesh Raj; Oh, Jin; Kim, Hyeon-A; Kim, Yong Jae; Jeong, Kyu-Shik; Rhyu, Dong Young

    2015-01-01

    The present study is to evaluate the anti-obesity effects of Eriobotrya japonica (EJ), Nelumbo nucifera (NN), and their mixture (MIX, 1:1 ratio) in 3T3-L1 adipocytes and high-fat diet-induced obese mice. The treatment of EJ, NN, and MIX in 3T3-L1 adipocytes effectively inhibited lipid accumulation, significantly decreased expression of peroxisome proliferator-activated receptor gamma (PPARγ), sterol regulatory element binding protein (SREBP1c), and adipocyte lipid-binding protein (aP2), and significantly increased phosphorylation of AMP-activated protein kinase (AMPK). Moreover, oral treatment of MIX showed stronger effects than individual treatment. C57BL/6J mice (6 week old) were divided into two groups; low fat diet (LFD) containing 10% calories from fat and high fat diet (HFD) containing 60% calories from fat. The HFD groups were further divided into five subgroups; treated with distilled water (HFD), treated with 400 mg/kg EJ (EJ400), treated with 400 mg/kg NN (NN400), treated with 200 mg/kg MIX (MIX200), and treated with 400 mg/kg MIX (MIX400) during 13 weeks. In our results, the administration of EJ, NN, and MIX significantly decreased body weight (BW), fat weight, liver weight, hepatic triglyceride (TG) and total cholesterol (TC), lipid droplets in the liver, food efficacy ratio, and the plasma TG, TC, glucose, insulin, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in a dose-dependent manner, and MIX treatment showed stronger effect than their individual treatments. Similarly, MIX treatment decreased the expression of PPARγ, SREBP-1c, FAS, and ACC more strongly in the adipose tissue than single treatments. In conclusion, the MIX of EJ and NN extract may strongly regulate BW gain than EJ or NN alone, and its anti-obesity effect is associated with the control of lipid metabolism, including adipogenesis and lipogenesis. PMID:26133751

  1. miR-146a-5p inhibits TNF-α-induced adipogenesis via targeting insulin receptor in primary porcine adipocytes[S

    PubMed Central

    Wu, Di; Xi, Qian-Yun; Cheng, Xiao; Dong, Tao; Zhu, Xiao-Tong; Shu, Gang; Wang, Li-Na; Jiang, Qing-Yan; Zhang, Yong-Liang

    2016-01-01

    TNF-α is a multifunctional cytokine participating in immune disorders, inflammation, and tumor development with regulatory effects on energy metabolism. Our work focused on the function of TNF-α in adipogenesis of primary porcine adipocytes. TNF-α could suppress the insulin receptor (IR) at the mRNA and protein levels. Microarray analysis of TNF-α-treated porcine adipocytes was used to screen out 29 differentially expressed microRNAs (miRNAs), 13 of which were remarkably upregulated and 16 were intensely downregulated. These 29 differentially expressed miRNAs were predicted to mainly participate in the insulin signaling pathway, adipocytokine signaling pathway, and type 2 diabetes mellitus pathway by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. miR-146a-5p, reportedly involved in immunity and cancer relevant processes, was one of the most highly differentially expressed miRNAs after TNF-α treatment. Red Oil O staining and TG assay revealed that miR-146a-5p suppressed adipogenesis. A dual-luciferase reporter and siRNA assay verified that miR-146a-5p targeted IR and could inhibit its protein expression. miR-146a-5p was also validated to be involved in the insulin signaling pathway by reducing tyrosine phosphorylation of insulin receptor substrate-1. Our study provides the first evidence of miR-146a-5p targeting IR, which facilitates future studies related to obesity and diabetes using pig models. PMID:27324794

  2. Sustained Low-Dose Treatment with the Histone Deacetylase Inhibitor LBH589 Induces Terminal Differentiation of Osteosarcoma Cells

    PubMed Central

    Cain, Jason E.; McCaw, Andrew; Jayasekara, W. Samantha N.; Rossello, Fernando J.; Marini, Kieren D.; Irving, Aaron T.; Kansara, Maya; Thomas, David M.; Ashley, David M.; Watkins, D. Neil

    2013-01-01

    Histone deacetylase inhibitors (HDACi) were identified nearly four decades ago based on their ability to induce cellular differentiation. However, the clinical development of these compounds as cancer therapies has focused on their capacity to induce apoptosis in hematologic and lymphoid malignancies, often in combination with conventional cytotoxic agents. In many cases, HDACi doses necessary to induce these effects result in significant toxicity. Since osteosarcoma cells express markers of terminal osteoblast differentiation in response to DNA methyltransferase inhibitors, we reasoned that the epigenetic reprogramming capacity of HDACi might be exploited for therapeutic benefit. Here, we show that continuous exposure of osteosarcoma cells to low concentrations of HDACi LBH589 (Panobinostat) over a three-week period induces terminal osteoblast differentiation and irreversible senescence without inducing cell death. Remarkably, transcriptional profiling revealed that HDACi therapy initiated gene signatures characteristic of chondrocyte and adipocyte lineages in addition to marked upregulation of mature osteoblast markers. In a mouse xenograft model, continuous low dose treatment with LBH589 induced a sustained cytostatic response accompanied by induction of mature osteoblast gene expression. These data suggest that the remarkable capacity of osteosarcoma cells to differentiate in response to HDACi therapy could be exploited for therapeutic benefit without inducing systemic toxicity. PMID:23533324

  3. Functional antagonism between inhibitor of DNA binding (Id) and adipocyte determination and differentiation factor 1/sterol regulatory element-binding protein-1c (ADD1/SREBP-1c) trans-factors for the regulation of fatty acid synthase promoter in adipocytes.

    PubMed Central

    Moldes, M; Boizard, M; Liepvre, X L; Fève, B; Dugail, I; Pairault, J

    1999-01-01

    We show that Id (inhibitor of DNA binding) 2 and Id3, dominant negative members of the helix-loop-helix (HLH) family, interact with the adipocyte determination and differentiation factor 1 (ADD1)/sterol regulatory element-binding protein (SREBP) 1c, a transcription factor of the basic HLH-leucine zipper family that controls the expression of several key genes of adipose metabolism. Gel mobility-shift assays performed with in vitro-translated ADD1, Id2 or Id3 proteins and a fatty acid synthase (FAS) promoter oligonucleotide showed evidence for a marked inhibition of the formation of DNA-ADD1 complexes by Id2 or Id3 proteins. Co-immunoprecipitation studies using in vitro-translated proteins demonstrated further the physical interaction of Id and ADD1/SREBP-1c proteins in the absence of DNA. Using the FAS gene as a model of an ADD1-regulated promoter in transiently transfected isolated rat adipocytes or mature 3T3-L1 adipocytes, a potent inhibition of the activity of the FAS-chloramphenicol acetyltransferase reporter gene was observed by overexpression of Id2 or Id3. Reciprocally, co-transfection of Id3 antisense and ADD1 expression vectors in preadipocytes potentiated the ADD1/SREBP-1c effect on the FAS promoter activity. Finally, in the non adipogenic NIH-3T3 cell line, most of the ADD1-mediated trans-activation of the FAS promoter was counteracted by co-transfection of Id2 or Id3 expression vectors. Previous studies have indicated Id gene expression to be down-regulated during adipogenesis [Moldes, Lasnier, Fève, Pairault and Djian (1997) Mol. Cell. Biol. 17, 1796-1804]. We here demonstrated that there was a dramatic rise of Id2 and Id3 mRNA levels when 3T3-L1 adipocytes or isolated rat fat cells were exposed to lipolytic and anti-lipogenic agents, forskolin and isoproterenol. Taken together, our data show that Id products are functionally involved in modulating ADD1/SREBP-1c transcriptional activity, and thus lipogenesis in adipocytes. PMID:10585876

  4. In brown adipocytes, adrenergically induced β{sub 1}-/β{sub 3}-(G{sub s})-, α{sub 2}-(G{sub i})- and α{sub 1}-(G{sub q})-signalling to Erk1/2 activation is not mediated via EGF receptor transactivation

    SciTech Connect

    Wang, Yanling; Fälting, Johanna M.; Mattsson, Charlotte L.; Holmström, Therése E.; Nedergaard, Jan

    2013-10-15

    Brown adipose tissue is unusual in that the neurotransmitter norepinephrine influences cell destiny in ways generally associated with effects of classical growth factors: regulation of cell proliferation, of apoptosis, and progression of differentiation. The norepinephrine effects are mediated through G-protein-coupled receptors; further mediation of such stimulation to e.g. Erk1/2 activation is in cell biology in general accepted to occur through transactivation of the EGF receptor (by external or internal pathways). We have examined here the significance of such transactivation in brown adipocytes. Stimulation of mature brown adipocytes with cirazoline (α{sub 1}-adrenoceptor coupled via G{sub q}), clonidine (α{sub 2} via G{sub i}) or CL316243 (β{sub 3} via G{sub s}) or via β{sub 1}-receptors significantly activated Erk1/2. Pretreatment with the EGF receptor kinase inhibitor AG1478 had, remarkably, no significant effect on Erk1/2 activation induced by any of these adrenergic agonists (although it fully abolished EGF-induced Erk1/2 activation), demonstrating absence of EGF receptor-mediated transactivation. Results with brown preadipocytes (cells in more proliferative states) were not qualitatively different. Joint stimulation of all adrenoceptors with norepinephrine did not result in synergism on Erk1/2 activation. AG1478 action on EGF-stimulated Erk1/2 phosphorylation showed a sharp concentration–response relationship (IC{sub 50} 0.3 µM); a minor apparent effect of AG1478 on norepinephrine-stimulated Erk1/2 phosphorylation showed nonspecific kinetics, implying caution in interpretation of partial effects of AG1478 as reported in other systems. Transactivation of the EGF receptor is clearly not a universal prerequisite for coupling of G-protein coupled receptors to Erk1/2 signalling cascades. - Highlights: • In brown adipocytes, norepinephrine regulates proliferation, apoptosis, differentiation. • EGF receptor transactivation is supposed to mediate GPCR-induced

  5. Effect of lipopolysaccharides on adipogenic potential and premature senescence of adipocyte progenitors.

    PubMed

    Zhao, Ming; Chen, Xiaoli

    2015-08-15

    The elevation of circulating LPS has been associated with obesity and aging. However, whether and how LPS contributes to adipose tissue dysfunction is unclear. In this study, we investigated the effect of LPS on the adipogenic capacity and cellular senescence of adipocyte progenitors. Stromal-vascular cells were isolated from inguinal adipose tissue of C57BL/6 mice and treated with LPS during the different time periods of adipocyte differentiation. We found that LPS treatment for 24 h prior to the induction of differentiation led to the most profound effect on the inhibition of adipogenesis, as evidenced by the morphological changes and the decreased mRNA expression of adipocyte marker genes. In addition, LPS induced features of premature senescence of SV cells, including the activation of p53, the elevation of SA-β-gal activity, and increased hydrogen peroxide production, but not telomere length. Upon LPS treatment, SV cells also developed senescence-associated secretory phenotype (SASP), as demonstrated by the increased expression of TNFα, IL-1β, IL-6, MCP-1, and VEGFα. Blocking LPS-induced NF-κB activation and cytokine production by Bay 11-7082 failed to rescue the impaired adipogenesis and the reduction in PPARγ and Zfp423 expression. On the contrary, rosiglitazone had little effect on cytokine production but corrected the defective adipogenic potential. In conclusion, we demonstrate that LPS inhibits adipogenesis by disrupting the differentiation of adipocyte progenitors in a NF-κB-independent manner; LPS also induces premature senescence of adipocyte progenitors. Our data suggest that LPS could be a potential contributor to the defective adipogenesis and the development of cellular senescence in adipose tissue during obesity and aging. PMID:26105007

  6. RPL13A and EEF1A1 Are Suitable Reference Genes for qPCR during Adipocyte Differentiation of Vascular Stromal Cells from Patients with Different BMI and HOMA-IR.

    PubMed

    Gentile, Adriana-Mariel; Lhamyani, Said; Coín-Aragüez, Leticia; Oliva-Olivera, Wilfredo; Zayed, Hatem; Vega-Rioja, Antonio; Monteseirin, Javier; Romero-Zerbo, Silvana-Yanina; Tinahones, Francisco-José; Bermúdez-Silva, Francisco-Javier; El Bekay, Rajaa

    2016-01-01

    Real-time or quantitative PCR (qPCR) is a useful technique that requires reliable reference genes for data normalization in gene expression analysis. Adipogenesis is among the biological processes suitable for this technique. The selection of adequate reference genes is essential for qPCR gene expression analysis of human Vascular Stromal Cells (hVSCs) during their differentiation into adipocytes. To the best of our knowledge, there are no studies validating reference genes for the analyses of visceral and subcutaneous adipose tissue hVSCs from subjects with different Body Mass Index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) index. The present study was undertaken to analyze this question. We first analyzed the stability of expression of five potential reference genes: CYC, GAPDH, RPL13A, EEF1A1, and 18S ribosomal RNA, during in vitro adipogenic differentiation, in samples from these types of patients. The expression of RPL13A and EEF1A1 was not affected by differentiation, thus being these genes the most stable candidates, while CYC, GAPDH, and 18S were not suitable for this sort of analysis. This work highlights that RPL13A and EEF1A1 are good candidates as reference genes for qPCR analysis of hVSCs differentiation into adipocytes from subjects with different BMI and HOMA-IR. PMID:27304673

  7. RPL13A and EEF1A1 Are Suitable Reference Genes for qPCR during Adipocyte Differentiation of Vascular Stromal Cells from Patients with Different BMI and HOMA-IR

    PubMed Central

    Gentile, Adriana-Mariel; Lhamyani, Said; Coín-Aragüez, Leticia; Oliva-Olivera, Wilfredo; Zayed, Hatem; Vega-Rioja, Antonio; Monteseirin, Javier; Romero-Zerbo, Silvana-Yanina; Tinahones, Francisco-José; Bermúdez-Silva, Francisco-Javier; El Bekay, Rajaa

    2016-01-01

    Real-time or quantitative PCR (qPCR) is a useful technique that requires reliable reference genes for data normalization in gene expression analysis. Adipogenesis is among the biological processes suitable for this technique. The selection of adequate reference genes is essential for qPCR gene expression analysis of human Vascular Stromal Cells (hVSCs) during their differentiation into adipocytes. To the best of our knowledge, there are no studies validating reference genes for the analyses of visceral and subcutaneous adipose tissue hVSCs from subjects with different Body Mass Index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) index. The present study was undertaken to analyze this question. We first analyzed the stability of expression of five potential reference genes: CYC, GAPDH, RPL13A, EEF1A1, and 18S ribosomal RNA, during in vitro adipogenic differentiation, in samples from these types of patients. The expression of RPL13A and EEF1A1 was not affected by differentiation, thus being these genes the most stable candidates, while CYC, GAPDH, and 18S were not suitable for this sort of analysis. This work highlights that RPL13A and EEF1A1 are good candidates as reference genes for qPCR analysis of hVSCs differentiation into adipocytes from subjects with different BMI and HOMA-IR. PMID:27304673

  8. Caffeic Acid Phenethyl Ester Regulates PPAR's Levels in Stem Cells-Derived Adipocytes.

    PubMed

    Vanella, Luca; Tibullo, Daniele; Godos, Justyna; Pluchinotta, Francesca Romana; Di Giacomo, Claudia; Sorrenti, Valeria; Acquaviva, Rosaria; Russo, Alessandra; Li Volti, Giovanni; Barbagallo, Ignazio

    2016-01-01

    Hypertrophic obesity inhibits activation of peroxisome proliferators-activated receptor gamma (PPARγ), considered the key mediator of the fully differentiated and insulin sensitive adipocyte phenotype. We examined the effects of Caffeic Acid Phenethyl Ester (Cape), isolated from propolis, a honeybee hive product, on Adipose Stem Cells (ASCs) differentiation to the adipocyte lineage. Finally we tested the effects of Cape on insulin-resistant adipocytes. Quantification of Oil Red O-stained cells showed that lipid droplets decreased following Cape treatment as well as radical oxygen species formation. Additionally, exposure of ASC to high glucose levels decreased adiponectin and increased proinflammatory cytokines mRNA levels, which were reversed by Cape-mediated increase of insulin sensitivity. Cape treatment resulted in decreased triglycerides synthesis and increased beta-oxidation. Exposure of ASCs to Lipopolysaccharide (LPS) induced a reduction of PPARγ, an increase of IL-6 levels associated with a well-known stimulation of lipolysis; Cape partially attenuated the LPS-mediated effects. These observations reveal the main role of PPARγ in the adipocyte function and during ASC differentiation. As there is now substantial interest in functional food and nutraceutical products, the observed therapeutic value of Cape in insulin-resistance related diseases should be taken into consideration. PMID:26904104

  9. Caffeic Acid Phenethyl Ester Regulates PPAR's Levels in Stem Cells-Derived Adipocytes

    PubMed Central

    Vanella, Luca; Tibullo, Daniele; Godos, Justyna; Pluchinotta, Francesca Romana; Di Giacomo, Claudia; Sorrenti, Valeria; Acquaviva, Rosaria; Russo, Alessandra; Li Volti, Giovanni; Barbagallo, Ignazio

    2016-01-01

    Hypertrophic obesity inhibits activation of peroxisome proliferators-activated receptor gamma (PPARγ), considered the key mediator of the fully differentiated and insulin sensitive adipocyte phenotype. We examined the effects of Caffeic Acid Phenethyl Ester (Cape), isolated from propolis, a honeybee hive product, on Adipose Stem Cells (ASCs) differentiation to the adipocyte lineage. Finally we tested the effects of Cape on insulin-resistant adipocytes. Quantification of Oil Red O-stained cells showed that lipid droplets decreased following Cape treatment as well as radical oxygen species formation. Additionally, exposure of ASC to high glucose levels decreased adiponectin and increased proinflammatory cytokines mRNA levels, which were reversed by Cape-mediated increase of insulin sensitivity. Cape treatment resulted in decreased triglycerides synthesis and increased beta-oxidation. Exposure of ASCs to Lipopolysaccharide (LPS) induced a reduction of PPARγ, an increase of IL-6 levels associated with a well-known stimulation of lipolysis; Cape partially attenuated the LPS-mediated effects. These observations reveal the main role of PPARγ in the adipocyte function and during ASC differentiation. As there is now substantial interest in functional food and nutraceutical products, the observed therapeutic value of Cape in insulin-resistance related diseases should be taken into consideration. PMID:26904104

  10. Cadmium modulates adipocyte functions in metallothionein-null mice

    SciTech Connect

    Kawakami, Takashige; Nishiyama, Kaori; Kadota, Yoshito; Sato, Masao; Inoue, Masahisa; Suzuki, Shinya

    2013-11-01

    Our previous study has demonstrated that exposure to cadmium (Cd), a toxic heavy metal, causes a reduction of adipocyte size and the modulation of adipokine expression. To further investigate the significance of the Cd action, we studied the effect of Cd on the white adipose tissue (WAT) of metallothionein null (MT{sup −/−}) mice, which cannot form atoxic Cd–MT complexes and are used for evaluating Cd as free ions, and wild type (MT{sup +/+}) mice. Cd administration more significantly reduced the adipocyte size of MT{sup −/−} mice than that of MT{sup +/+} mice. Cd exposure also induced macrophage recruitment to WAT with an increase in the expression level of Ccl2 (MCP-1) in the MT{sup −/−} mice. The in vitro exposure of Cd to adipocytes induce triglyceride release into culture medium, decrease in the expression levels of genes involved in fatty acid synthesis and lipid hydrolysis at 24 h, and at 48 h increase in phosphorylation of the lipid-droplet-associated protein perilipin, which facilitates the degradation of stored lipids in adipocytes. Therefore, the reduction in adipocyte size by Cd may arise from an imbalance between lipid synthesis and lipolysis. In addition, the expression levels of leptin, adiponectin and resistin decreased in adipocytes. Taken together, exposure to Cd may induce unusually small adipocytes and modulate the expression of adipokines differently from the case of physiologically small adipocytes, and may accelerate the risk of developing insulin resistance and type 2 diabetes. - Highlights: • Cd causes a marked reduction in adipocyte size in MT-null mice. • Cd enhances macrophage migration into adipose tissue and disrupt adipokine secretion. • MT gene alleviates Cd-induced adipocyte dysfunctions. • Cd enhances the degradation of stored lipids in adipocytes, mediated by perilipin. • Cd induces unusually small adipocytes and the abnormal expression of adipokines.

  11. Insulin-induced activation of glycerol-3-phosphate acyltransferase by a chiro-inositol-containing insulin mediator is defective in adipocytes of insulin-resistant, type II diabetic, Goto-Kakizaki rats.

    PubMed Central

    Farese, R V; Standaert, M L; Yamada, K; Huang, L C; Zhang, C; Cooper, D R; Wang, Z; Yang, Y; Suzuki, S; Toyota, T

    1994-01-01

    Type II diabetic Goto-Kakizaki (GK) rats were insulin-resistant in euglycemic-hyperinsulinemic clamp studies. We therefore examined insulin signaling systems in control Wistar and diabetic GK rats. Glycerol-3-phosphate acyltransferase (G3PAT), which is activated by headgroup mediators released from glycosyl-phosphatidylinositol (GPI), was activated by insulin in intact and cell-free adipocyte preparations of control, but not diabetic, rats. A specific chiro-inositol-containing inositol phosphoglycan (IPG) mediator, prepared from beef liver, bypassed this defect and comparably activated G3PAT in cell-free adipocyte preparations of both diabetic GK and control rats. A myo-inositol-containing IPG mediator did not activate G3PAT. Relative to control adipocytes, labeling of GPI by [3H]glucosamine was diminished by 50% and insulin failed to stimulate GPI hydrolysis in GK adipocytes. In contrast to GPI-dependent G3PAT activation, insulin-stimulated hexose transport was intact in adipocytes and soleus and gastrocnemius muscles of the GK rat, as was insulin-induced activation of mitogen-activated protein kinase and protein kinase C. We conclude that (i) chiro-inositol-containing IPG mediator activates G3PAT during insulin action, (ii) diabetic GK rats have a defect in synthesizing or releasing functional chiro-inositol-containing IPG, and (iii) defective IPG-regulated intracellular glucose metabolism contributes importantly to insulin resistance in diabetic GK rats. PMID:7972005

  12. The extinction differential induced virulence macroevolution

    NASA Astrophysics Data System (ADS)

    Zhang, Feng; Xu, Liufang; Wang, Jin

    2014-04-01

    We apply the potential-flux landscape theory to deal with the large fluctuation induced extinction phenomena. We quantify the most probable extinction pathway on the landscape and measure the extinction risk by the landscape topography. In this Letter, we investigate the disease extinction through an epidemic model described by a set of chemical reaction. We found the virulence-differential-dependent symbioses between mother and daughter pathogen species: mutualism and parasitism. The symbioses, whether mutualism or parasitism, benefit the higher virulence species. This implies that speciation towards lower virulence is an effective strategy for a pathogen species to reduce its extinction risk.

  13. Thermogenic Ability of Uncoupling Protein 1 in Beige Adipocytes in Mice

    PubMed Central

    Okamatsu-Ogura, Yuko; Fukano, Keigo; Tsubota, Ayumi; Uozumi, Akihiro; Terao, Akira; Kimura, Kazuhiro; Saito, Masayuki

    2013-01-01

    Chronic adrenergic activation leads to the emergence of beige adipocytes in some depots of white adipose tissue in mice. Despite their morphological similarities to brown adipocytes and their expression of uncoupling protein 1 (UCP1), a thermogenic protein exclusively expressed in brown adipocytes, the beige adipocytes have a gene expression pattern distinct from that of brown adipocytes. However, it is unclear whether the thermogenic function of beige adipocytes is different from that of classical brown adipocytes existing in brown adipose tissue. To examine the thermogenic ability of UCP1 expressed in beige and brown adipocytes, the adipocytes were isolated from the fat depots of C57BL/6J mice housed at 24°C (control group) or 10°C (cold-acclimated group) for 3 weeks. Morphological and gene expression analyses revealed that the adipocytes isolated from brown adipose tissue of both the control and cold-acclimated groups consisted mainly of brown adipocytes. These brown adipocytes contained large amounts of UCP1 and increased their oxygen consumption when stimulated with norepinephirine. Adipocytes isolated from the perigonadal white adipose tissues of both groups and the inguinal white adipose tissue of the control group were white adipocytes that showed no increase in oxygen consumption after norepinephrine stimulation. Adipocytes isolated from the inguinal white adipose tissue of the cold-acclimated group were a mixture of white and beige adipocytes, which expressed UCP1 and increased their oxygen consumption in response to norepinephrine. The UCP1 content and thermogenic ability of beige adipocytes estimated on the basis of their abundance in the cell mixture were similar to those of brown adipocytes. These results revealed that the inducible beige adipocytes have potent thermogenic ability comparable to classical brown adipocytes. PMID:24386355

  14. Enhanced accumulation of adipocytes in bone marrow stromal cells in the presence of increased extracellular and intracellular [Ca{sup 2+}

    SciTech Connect

    Hashimoto, Ryota; Katoh, Youichi; Nakamura, Kyoko; Itoh, Seigo; Iesaki, Takafumi; Daida, Hiroyuki; Nakazato, Yuji; Okada, Takao

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} enhances adipocyte accumulation in the presence of adipogenic inducers. Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} enhances both proliferation and adipocyte differentiation in BMSCs. Black-Right-Pointing-Pointer High [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub o} in BMSCs. Black-Right-Pointing-Pointer An intracellular Ca{sup 2+} chelator suppresses the enhancement in adipocyte accumulation. Black-Right-Pointing-Pointer Controlling [Ca{sup 2+}]{sub o} may govern the balance of adipocyte and osteoblast development. -- Abstract: The bone marrow stroma contains osteoblasts and adipocytes that have a common precursor: the pluripotent mesenchymal stem cell found in bone marrow stromal cells (BMSCs). Local bone marrow Ca{sup 2+} levels can reach high concentrations due to bone resorption, which is one of the notable features of the bone marrow stroma. Here, we describe the effects of high [Ca{sup 2+}]{sub o} on the accumulation of adipocytes in the bone marrow stroma. Using primary mouse BMSCs, we evaluated the level of adipocyte accumulation by measuring Oil Red O staining and glycerol-3-phosphate dehydrogenase (GPDH) activity. High [Ca{sup 2+}]{sub o} enhanced the accumulation of adipocytes following treatment with both insulin and dexamethasone together but not in the absence of this treatment. This enhanced accumulation was the result of both the accelerated proliferation of BMSCs and their differentiation into adipocytes. Using the fura-2 method, we also showed that high [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub i}. An intracellular Ca{sup 2+} chelator suppressed the enhancement in adipocyte accumulation due to increased [Ca{sup 2+}]{sub o} in BMSCs. These data suggest a new role for extracellular Ca{sup 2+} in the bone marrow stroma: increased [Ca{sup 2+}]{sub o} induces an increase in [Ca{sup 2+}]{sub i} levels, which in turn enhances the accumulation of

  15. Beta-adrenergic stimulation of brown adipocyte proliferation.

    PubMed

    Géloën, A; Collet, A J; Guay, G; Bukowiecki, L J

    1988-01-01

    The mechanisms of brown adipose tissue (BAT) growth were studied by quantitative photonic radioautography using tritiated thymidine to follow mitotic activity. To identify the nature of the adrenergic pathways mediating brown adipocyte proliferation and differentiation, the effects of cold exposure (4 days at 4 degrees C) on BAT growth were compared with those induced by treating rats at 25 degrees C with norepinephrine (a mixed agonist), isoproterenol (a beta-agonist), and phenylephrine (an alpha-agonist). The drugs were continuously administrated via osmotic minipumps (0.375 mumol/h during 4 days) implanted subcutaneously. Cold exposure markedly enhanced the mitotic activity in brown adipocyte precursor cells (interstitial cells and preadipocytes) and endothelial cells forming the numerous capillaries. Norepinephrine mimicked the effects of cold exposure, not only on the mitotic activity, but also on the distribution of the labeling among the various cellular types. Isoproterenol entirely reproduced the effects of norepinephrine both on the labeling index and on the cellular type labeling frequency. In contrast, phenylephrine did not stimulate cell division. These results demonstrate that norepinephrine triggers a coordinated proliferation of brown adipocytes and endothelial cells in warm-exposed rats that is similar to that observed after cold exposure. They also suggest that cold exposure stimulates BAT growth by increasing the release of norepinephrine from sympathetic nerves and that the neurohormone activates mitoses in BAT precursor cells via beta-adrenergic pathways. PMID:2892422

  16. JAZF1 can regulate the expression of lipid metabolic genes and inhibit lipid accumulation in adipocytes

    SciTech Connect

    Ming, Guang-feng; Xiao, Di; Gong, Wei-jing; Liu, Hui-xia; Liu, Jun; Zhou, Hong-hao; Liu, Zhao-qian

    2014-03-14

    Highlights: • JAZF1 was significantly upregulated during the differentiation of 3T3-L1 preadipocytes. • JAZF1 overexpression inhibited lipid accumulation in differentiated mature 3T3-L1 adipocytes. • JAZF1 overexpression inhibited the expression of SREBP1, ACC, and FAS. • JAZF1 overexpression upregulated the expression of HSL and ATGL. • SREBP1 and JAZF1 could regulate each other in adipocytes. - Abstract: JAZF1 is a newly identified gene with unknown functions. A recent genome-wide association study showed that JAZF1 is associated with type 2 diabetes and is highly expressed in liver and adipose tissue. Studies have demonstrated that JAZF1 is the co-repressor for nuclear orphan receptor TAK1, whereas most nuclear orphan receptor family members are involved in the regulation of lipid metabolism. Therefore, JAZF1 could be closely related to glycolipid metabolism. In this study, JAZF1 was significantly upregulated during the induced differentiation process of 3T3-L1 preadipocytes. The overexpression of JAZF1 inhibited lipid accumulation in differentiated mature 3T3-L1 adipocytes and significantly inhibited the expression of SREBPl, ACC, and FAS, which were important in lipid synthesis, while upregulating the expression of key enzyme hormone-sensitive lipase in lipoclasis. Moreover, SREBPl exhibited an inhibitory function on the expression of JAZF1. SREBP1 reversed the inhibitory action on lipid accumulation of JAZF1. SREBP1 and JAZF1 were observed to regulate each other in adipocytes. Therefore, JAZF1 could regulate the expression of particular genes related to lipid metabolism and inhibit lipid accumulation in adipocytes. This result suggests that JAZF1 may be a potential target for the treatment of diseases, such as obesity and lipid metabolism disorders.

  17. Mouse white adipocytes and 3T3-L1 cells display an anomalous pattern of carnitine palmitoyltransferase (CPT) I isoform expression during differentiation. Inter-tissue and inter-species expression of CPT I and CPT II enzymes.

    PubMed Central

    Brown, N F; Hill, J K; Esser, V; Kirkland, J L; Corkey, B E; Foster, D W; McGarry, J D

    1997-01-01

    The outer mitochondrial membrane enzyme carnitine palmitoyltransferase I (CPT I) represents the initial and regulated step in the beta-oxidation of fatty acids. It exists in at least two isoforms, denoted L (liver) and M (muscle) types, with very different kinetic properties and sensitivities to malonyl-CoA. Here we have examined the relative expression of the CPT I isoforms in two different models of adipocyte differentiation and in a number of rat tissues. Adipocytes from mice, hamsters and humans were also evaluated. Primary monolayer cultures of undifferentiated rat preadipocytes expressed solely L-CPT I, but significant levels of M-CPT I emerged after only 3 days of differentiation in vitro; in the mature cell M-CPT I predominated. In sharp contrast, the murine 3T3-L1 preadipocyte expressed essentially exclusively L-CPT I, both in the undifferentiated state and throughout the differentiation process in vitro. This was also true of the mature mouse white fat cell. Fully developed adipocytes from the hamster and human behaved similarly to those of the rat. Thus the mouse white fat cell differs fundamentally from those of the other species examined in terms of tis choice of a key regulatory enzyme in fatty acid metabolism. In contrast, brown adipose tissue from all three rodents displayed the same isoform profiles, each expressing overwhelmingly M-CPT I. Northern blot analysis of other rat tissues established L-CPT I as the dominant isoform not only in liver but also in kidney, lung, ovary, spleen, brain, intestine and pancreatic islets. In addition to its primacy in skeletal muscle, heart and fat, M-CPT I was also found to dominate the testis. The same inter-tissue isoform pattern (with the exception of white fat) was found in the mouse. Taken together, the data bring to light an intriguing divergence between white adipocytes of the mouse and other mammalian species. They also raise a cautionary note that should be considered in the choice of animal model used

  18. Inhibition of inflammatory signaling pathways in 3T3-L1 adipocytes by apolipoprotein A-I.

    PubMed

    Sultana, Afroza; Cochran, Blake J; Tabet, Fatiha; Patel, Mili; Torres, Luisa Cuesta; Barter, Philip J; Rye, Kerry-Anne

    2016-06-01

    Activation of inflammatory signaling pathways links obesity with metabolic disorders. TLR4-mediated activation of MAPKs and NF-κB are 2 such pathways implicated in obesity-induced inflammation. Apolipoprotein A-I (apoA-I) exerts anti-inflammatory effects on adipocytes by effluxing cholesterol from the cells via the ATP binding cassette transporter A1 (ABCA1). It is not known if these effects involve inhibition of inflammatory signaling pathways by apoA-I. This study asks if apoA-I inhibits activation of MAPKs and NF-κB in mouse 3T3-L1 adipocytes and whether this inhibition is ABCA1 dependent. Incubation of differentiated 3T3-L1 adipocytes with apoA-I decreased cell surface expression of TLR4 by 16 ± 2% and synthesis of the TLR4 adaptor protein, myeloid differentiation primary response 88, by 24 ± 4% in an ABCA1-dependent manner. ApoA-I also inhibited downstream activation of MAPKs, such as ERK, p38MAPK, and JNK, as well as expression of proinflammatory adipokines in bacterial LPS-stimulated 3T3-L1 adipocytes in an ABCA1-dependent manner. ApoA-I, by contrast, suppressed nuclear localization of the p65 subunit of NF-κB by 30 ± 3% in LPS-stimulated 3T3-L1 adipocytes in an ABCA1-independent manner. In conclusion, apoA-I inhibits TLR4-mediated inflammatory signaling pathways in adipocytes by preventing MAPK and NF-κB activation.-Sultana, A., Cochran, B. J., Tabet, F., Patel, M., Cuesta Torres, L., Barter, P. J., Rye, K.-A. Inhibition of inflammatory signaling pathways in 3T3-L1 adipocytes by apolipoprotein A-I. PMID:26965683

  19. Metabolic interplay between white, beige, brown adipocytes and the liver.

    PubMed

    Scheja, Ludger; Heeren, Joerg

    2016-05-01

    In mammalian evolution, three types of adipocytes have developed, white, brown and beige adipocytes. White adipocytes are the major constituents of white adipose tissue (WAT), the predominant store for energy-dense triglycerides in the body that are released as fatty acids during catabolic conditions. The less abundant brown adipocytes, the defining parenchymal cells of brown adipose tissue (BAT), internalize triglycerides that are stored intracellularly in multilocular lipid droplets. Beige adipocytes (also known as brite or inducible brown adipocytes) are functionally very similar to brown adipocytes and emerge in specific WAT depots in response to various stimuli including sustained cold exposure. The activation of brown and beige adipocytes (together referred to as thermogenic adipocytes) causes both the hydrolysis of stored triglycerides as well as the uptake of lipids and glucose from the circulation. Together, these fuels are combusted for heat production to maintain body temperature in mammals including adult humans. Given that heating by brown and beige adipocytes is a very-well controlled and energy-demanding process which entails pronounced shifts in energy fluxes, it is not surprising that an intensive interplay exists between the various adipocyte types and parenchymal liver cells, and that this influences systemic metabolic fluxes and endocrine networks. In this review we will emphasize the role of hepatic factors that regulate the metabolic activity of white and thermogenic adipocytes. In addition, we will discuss the relevance of lipids and hormones that are secreted by white, brown and beige adipocytes regulating liver metabolism in order to maintain systemic energy metabolism in health and disease. PMID:26829204

  20. White-to-brite conversion in human adipocytes promotes metabolic reprogramming towards fatty acid anabolic and catabolic pathways

    PubMed Central

    Barquissau, V.; Beuzelin, D.; Pisani, D.F.; Beranger, G.E.; Mairal, A.; Montagner, A.; Roussel, B.; Tavernier, G.; Marques, M.-A.; Moro, C.; Guillou, H.; Amri, E.-Z.; Langin, D.

    2016-01-01

    Objective Fat depots with thermogenic activity have been identified in humans. In mice, the appearance of thermogenic adipocytes within white adipose depots (so-called brown-in-white i.e., brite or beige adipocytes) protects from obesity and insulin resistance. Brite adipocytes may originate from direct conversion of white adipocytes. The purpose of this work was to characterize the metabolism of human brite adipocytes. Methods Human multipotent adipose-derived stem cells were differentiated into white adipocytes and then treated with peroxisome proliferator-activated receptor (PPAR)γ or PPARα agonists between day 14 and day 18. Gene expression profiling was determined using DNA microarrays and RT-qPCR. Variations of mRNA levels were confirmed in differentiated human preadipocytes from primary cultures. Fatty acid and glucose metabolism was investigated using radiolabelled tracers, Western blot analyses and assessment of oxygen consumption. Pyruvate dehydrogenase kinase 4 (PDK4) knockdown was achieved using siRNA. In vivo, wild type and PPARα-null mice were treated with a β3-adrenergic receptor agonist (CL316,243) to induce appearance of brite adipocytes in white fat depot. Determination of mRNA and protein levels was performed on inguinal white adipose tissue. Results PPAR agonists promote a conversion of white adipocytes into cells displaying a brite molecular pattern. This conversion is associated with transcriptional changes leading to major metabolic adaptations. Fatty acid anabolism i.e., fatty acid esterification into triglycerides, and catabolism i.e., lipolysis and fatty acid oxidation, are increased. Glucose utilization is redirected from oxidation towards glycerol-3-phophate production for triglyceride synthesis. This metabolic shift is dependent on the activation of PDK4 through inactivation of the pyruvate dehydrogenase complex. In vivo, PDK4 expression is markedly induced in wild-type mice in response to CL316,243, while this increase is blunted

  1. Glyphosate Inhibits PPAR Gamma Induction and Differentiation of Preadipocytes and is able to Induce Oxidative Stress.

    PubMed

    Martini, Claudia N; Gabrielli, Matías; Brandani, Javier N; Vila, María Del C

    2016-08-01

    Glyphosate-based herbicides (GF) are extensively used for weed control. Thus, it is important to investigate their putative toxic effects. We have reported that GF at subagriculture concentrations inhibits proliferation and differentiation to adipocytes of 3T3-L1 fibroblasts. In this investigation, we evaluated the effect of GF on genes upregulated during adipogenesis. GF was able to inhibit the induction of PPAR gamma, the master gene in adipogenesis but not C/EBP beta, which precedes PPAR gamma activation. GF also inhibited differentiation and proliferation of another model of preadipocyte: mouse embryonic fibroblasts. In exponentially growing 3T3-L1 cells, GF increased lipid peroxidation and the activity of the antioxidant enzyme, superoxide dismutase. We also found that proliferation was inhibited with lower concentrations of GF when time of exposure was extended. Thus, GF was able to inhibit proliferation and differentiation of preadipocytes and to induce oxidative stress, which is indicative of its ability to alter cellular physiology. PMID:27044015

  2. Olanzapine promotes the accumulation of lipid droplets and the expression of multiple perilipins in human adipocytes.

    PubMed

    Nimura, Satomi; Yamaguchi, Tomohiro; Ueda, Koki; Kadokura, Karin; Aiuchi, Toshihiro; Kato, Rina; Obama, Takashi; Itabe, Hiroyuki

    2015-11-27

    Second generation antipsychotics are useful for the treatment of schizophrenia, but concerns have been raised about the side effects of diabetes mellitus and obesity. Olanzapine, especially, is associated with more weight gain than the others. It has been reported that olanzapine promotes adipocyte-differentiation in rodents both in vivo and in vitro. In this study the effects of antipsychotics on human adipocytes were investigated by using human mesenchymal stem cells (hMSCs). When hMSCs were differentiated and treated with various antipsychotics, olanzapine and clozapine increased intracellular lipids. Olanzapine induced lipid accumulation in a dose-dependent manner. Proteomic analysis revealed that PLIN4 and several enzymes for lipid metabolism were increased in the hMSCs after olanzapine treatment. During adipocyte differentiation, olanzapine increased the protein expression of PLIN1, PLIN2 and PLIN4. These proteins are known to be associated with the initial stage of lipid droplet formation. Immunocytochemistry showed that olanzapine increased and enlarged the lipid droplets coated with PLIN1 and PLIN2 while PLIN4 was largely distributed in the cytosol. mRNA expression of PLIN2, but not PLIN1 or PLIN4, was increased by olanzapine. On the other hand, olanzapine did not alter the mRNA level of transcription regulators involved in adipocyte-differentiation or adipokines. The present study shows that olanzapine induced transient PLIN2 expression in hMSCs that could result in an accumulation of lipid droplets and overexpression of PLIN1 and PLIN4, providing information of possible interest for olanzapine-induced weight gain. PMID:26471304

  3. 3D Tissue Formation of Unilocular Adipocytes in Hydrogel Microfibers.

    PubMed

    Hsiao, Amy Y; Okitsu, Teru; Teramae, Hiroki; Takeuchi, Shoji

    2016-03-01

    Adipose tissue, an active metabolic and endocrine organ mainly composed of unilocular adipocytes, is implicated in various obesity related diseases. Developing morphologically and functionally accurate in vitro models of the adipose tissue is therefore critically important for basic biological studies, drug screening/testing, and clinical implants to advance the understanding and treatment of these diseases. However, current adipose tissue engineering technologies either cannot replicate the unilocular morphologies of mature adipocytes, or lack the ease of monitoring, handling, and scaling up required in the above mentioned applications. This paper presents the differentiation of adipose derived stem cells (ADSCs) to mature adipocytes in highly observable and highly handleable 3D fiber shaped constructs exhibiting morphologies and functions of native adipose tissues. Using the cell fiber technology, ADSCs were encapsulated in hydrogel microfibers, allowed to form into fiber shaped constructs, and differentiated to mature unilocular adipocytes. These adipocyte fibers are observed and maintained for up to 91 d, and secretion of adipose tissue-specific factor, adiponectin, is further confirmed. The handleability of the adipocyte fibers is demonstrated by assembling the adipocyte fibers into doll shaped constructs. Such highly observable, highly handleable, and scalable characteristics of the adipocyte fibers make them suitable for biological studies, high-throughput drug screening/testing, and clinical applications. PMID:26680212

  4. Id4, a new candidate gene for senile osteoporosis, acts as a molecular switch promoting osteoblast differentiation.

    PubMed

    Tokuzawa, Yoshimi; Yagi, Ken; Yamashita, Yzumi; Nakachi, Yutaka; Nikaido, Itoshi; Bono, Hidemasa; Ninomiya, Yuichi; Kanesaki-Yatsuka, Yukiko; Akita, Masumi; Motegi, Hiromi; Wakana, Shigeharu; Noda, Tetsuo; Sablitzky, Fred; Arai, Shig