WorldWideScience

Sample records for 3t3-l1 adipocyte differentiation

  1. Rubi Fructus (Rubus coreanus) Inhibits Differentiation to Adipocytes in 3T3-L1 Cells.

    Science.gov (United States)

    Jeong, Mi-Young; Kim, Hye-Lin; Park, Jinbong; An, Hyo-Jin; Kim, Sung-Hoon; Kim, Su-Jin; So, Hong-Seob; Park, Raekil; Um, Jae-Young; Hong, Seung-Heon

    2013-01-01

    Rubi Fructus (RF) is known to exert several pharmacological effects including antitumor, antioxidant, and anti-inflammatory activities. However, its antiobesity effect has not been reported yet. This study was focused on the antidifferentiation effect of RF extract on 3T3-L1 preadipocytes. When 3T3-L1 preadipocytes were differentiating into adipocytes, 10-100  μ g/mL of RF was added. Next, the lipid contents were quantified by Oil Red O staining. RF significantly reduced lipid accumulation and downregulated the expression of peroxisome proliferator-activated receptor γ (PPAR γ ), CCAAT0-enhancer-binding proteins α (C/EBP α ), adipocyte fatty acid-binding protein 2 (aP2), resistin, and adiponectin in ways that were concentration dependent. Moreover, RF markedly upregulated liver kinase B1 and AMP-activated protein kinase (AMPK). Interestingly, pretreatment with AMPK α siRNA and RF downregulated the expression of PPAR γ and C/EBP α protein as well as the adipocyte differentiation. Our study shows that RF is capable of inhibiting the differentiation of 3T3-L1 adipocytes through the modulation of PPAR γ , C/EBP α , and AMPK, suggesting that it has a potential for therapeutic application in the treatment or prevention of obesity.

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

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Jing [The First Affiliated Hospital, Jinan University, Guangzhou, Guangdong (China); Liu, Gexiu [Institute of Hematology, School of Medicine, Jinan University, Guangzhou, Guangdong (China); Yan, Guoyao [The First Affiliated Hospital, Jinan University, Guangzhou, Guangdong (China); He, Dongmei [Institute of Hematology, School of Medicine, Jinan University, Guangzhou, Guangdong (China); Zhou, Ying [The First Affiliated Hospital, Jinan University, Guangzhou, Guangdong (China); Chen, Shengting, E-mail: shengtingchen@sina.cn [The First Affiliated Hospital, Jinan University, Guangzhou, Guangdong (China)

    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.

  3. Berberine inhibits 3T3-L1 adipocyte differentiation through the PPARgamma pathway.

    Science.gov (United States)

    Huang, Cheng; Zhang, Yuebo; Gong, Zhenwei; Sheng, Xiaoyan; Li, Zongmeng; Zhang, Wei; Qin, Ying

    2006-09-22

    Berberine (BBR), a compound purified from Cortidis rhizoma, reduces serum cholesterol, triglycerides, and LDL-cholesterol of hypercholesterolemic patients and high fat diet fed animals, and increases hepatic LDLR mRNA and protein levels through a post-transcriptional mechanism. BBR also enhances the hypoglycemic action of insulin in diabetic animal models. Here, we show that BBR inhibits the differentiation of 3T3-L1 preadipocytes induced by DM and suppresses the mitotic clonal expansion of 3T3-L1 preadipocytes in a time- and dose-dependent manner. Gene expression analysis and Western blot analysis reveal that the BBR inhibits the mRNA and protein levels of adipogenesis related transcription factors PPARgamma and C/EBPalpha and their upstream regulator, C/EBPbeta. Reporter gene assays demonstrate that the full-length PPARgamma and alpha transcription activities are inhibited by BBR. Using real-time PCR, we have also found that the PPAR target genes that are involved in adipocyte differentiation, such as aP2, CD36, ACO, LPL, and other adipocyte markers, are suppressed by BBR. These studies suggest that BBR works on multiple molecular targets as an inhibitor of PPARgamma and alpha, and is a potential weight reducing, hypolipidemic, and hypoglycemic drug.

  4. Induction of adipocyte differentiation by polybrominated diphenyl ethers (PBDEs) in 3T3-L1 cells.

    Science.gov (United States)

    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.

  5. Induction of adipocyte differentiation by polybrominated diphenyl ethers (PBDEs in 3T3-L1 cells.

    Directory of Open Access Journals (Sweden)

    Emily W Y Tung

    Full Text Available 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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Boschi, Federico, E-mail: federico.boschi@univr.it [Department of Neurological and Movement Sciences, University of Verona, Strada Le Grazie 8, 37134 Verona (Italy); Department of Computer Science, University of Verona, Strada Le Grazie 15, 37134 Verona (Italy); Rizzatti, Vanni; Zamboni, Mauro [Department of Medicine, Geriatric Section, University of Verona, Piazzale Stefani 1, 37126 Verona (Italy); Sbarbati, Andrea [Department of Neurological and Movement Sciences, University of Verona, Strada Le Grazie 8, 37134 Verona (Italy)

    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

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

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, Akio, E-mail: watanabea@jfrl.or.jp [Japan Food Research Laboratories, Osaka 567-0085 (Japan); Food and Biodynamic Chemistry Laboratory, Graduate School of Agricultural Science, Tohoku University, Miyagi 981-8555 (Japan); Kato, Tsuyoshi; Ito, Yusuke; Yoshida, Izumi; Harada, Teppei; Mishima, Takashi; Fujita, Kazuhiro; Watai, Masatoshi [Japan Food Research Laboratories, Osaka 567-0085 (Japan); Nakagawa, Kiyotaka; Miyazawa, Teruo [Food and Biodynamic Chemistry Laboratory, Graduate School of Agricultural Science, Tohoku University, Miyagi 981-8555 (Japan)

    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.

  8. Mammalian ste20-like kinase and SAV1 promote 3T3-L1 adipocyte differentiation by activation of PPARγ.

    Directory of Open Access Journals (Sweden)

    Byoung Hee Park

    Full Text Available The mammalian ste20 kinase (MST signaling pathway plays an important role in the regulation of apoptosis and cell cycle control. We sought to understand the role of MST2 kinase and Salvador homolog 1 (SAV1, a scaffolding protein that functions in the MST pathway, in adipocyte differentiation. MST2 and MST1 stimulated the binding of SAV1 to peroxisome proliferator-activated receptor γ (PPARγ, a transcription factor that plays a key role in adipogenesis. The interaction of endogenous SAV1 and PPARγ was detected in differentiating 3T3-L1 adipocytes. This binding required the kinase activity of MST2 and was mediated by the WW domains of SAV1 and the PPYY motif of PPARγ. Overexpression of MST2 and SAV1 increased PPARγ levels by stabilizing the protein, and the knockdown of SAV1 resulted in a decrease of endogenous PPARγ protein in 3T3-L1 adipocytes. During the differentiation of 3T3-L1 cells into adipocytes, MST2 and SAV1 expression began to increase at 2 days when PPARγ expression also begins to increase. MST2 and SAV1 significantly increased PPARγ transactivation, and SAV1 was shown to be required for the activation of PPARγ by rosiglitazone. Finally, differentiation of 3T3-L1 cells was augmented by MST2 and SAV1 expression and inhibited by knockdown of MST1/2 or SAV1. These results suggest that PPARγ activation by the MST signaling pathway may be a novel regulatory mechanism of adipogenesis.

  9. The mixed-lineage kinase DLK is a key regulator of 3T3-L1 adipocyte differentiation.

    Directory of Open Access Journals (Sweden)

    Jean-Philippe Couture

    Full Text Available BACKGROUND: The mixed-lineage kinase (MLK family member DLK has been proposed to serve as a regulator of differentiation in various cell types; however, its role in adipogenesis has not been investigated. In this study, we used the 3T3-L1 preadipocyte cell line as a model to examine the function of DLK in adipocyte differentiation. METHODS AND FINDINGS: Immunoblot analyses and kinase assays performed on 3T3-L1 cells showed that the expression and activity of DLK substantially increase as differentiation occurs. Interestingly, DLK appears crucial for differentiation since its depletion by RNA interference impairs lipid accumulation as well as expression of the master regulators of adipogenesis C/EBPalpha and PPARgamma2 at both the mRNA and protein levels. In contrast, neither the expression nor the DNA binding activity of C/EBPbeta, an activator for C/EBPalpha and PPARgamma, is affected by DLK loss. CONCLUSIONS: Taken together, these results suggest that DLK is important for expression of mature adipocyte markers and that its action most likely takes place via regulation of C/EBPbeta transcriptional activity and/or initiation of C/EBPalpha and PPARgamma2 gene transcription.

  10. St. John's wort promotes adipocyte differentiation and modulates NF-κB activation in 3T3-L1 cells.

    Science.gov (United States)

    Hatano, Tomoko; Sameshima, Yuka; Kawabata, Mami; Yamada, Shizuo; Shinozuka, Kazumasa; Nakabayashi, Toshikatsu; Mizuno, Hideya

    2014-01-01

    St. John's wort (SJW), or Hypericum perforatum, is a perennial herb that has been used in the treatment of depression in several countries. Though its therapeutic effect on depression has been extensively studied, its influence on metabolic syndrome is yet to be fully characterized. Therefore, we investigated the effect of SJW extract on adipocyte differentiation and its anti-inflammatory effects by using 3T3-L1 preadipocytes. Oil Red O staining indicated that SJW promotes adipocyte differentiation, while immunoblots indicated that SJW increases the expression of peroxisome proliferator activated receptor γ (PPARγ), a nuclear receptor regulating adipocyte differentiation, and adiponectin, an anti-inflammatory adipokine. Furthermore, the anti-inflammatory activity of SJW was demonstrated by its inhibition of the activation of nuclear factor-κB (NF-κB), an inflammatory transcription factor. Stimulation of mature 3T3-L1 adipocytes by tumor necrosis factor-α (TNF-α) decreased the expression of the NF-κB inhibitor IκBα, and increased its phosphorylation. Treatment with SJW further decreased the TNF-α-induced perturbation in IκBα expression and phosphorylation, which indicated that SJW mediated the inhibition of NF-κB activation. In addition, SJW decreased the TNF-α-induced increase in the mRNA levels of pro-inflammatory adipokines, interleukin-6 (IL-6), and monocyte chemoattractant protein-1 (MCP-1). Collectively, our results indicate that SJW treatment could promote adipocyte differentiation probably through its anti-inflammatory activity, which in turn suggests that SJW has the potential to minimize the risk factors of metabolic syndrome.

  11. Inhibition of mitotic clonal expansion mediates fisetin-exerted prevention of adipocyte differentiation in 3T3-L1 cells.

    Science.gov (United States)

    Lee, Youngyi; Bae, Eun Ju

    2013-11-01

    Adipocytes are the key player in adipose tissue inflammation and subsequent systemic insulin resistance and its development involves complex process of proliferation and differentiation of preadipocytes. Fistein, a polyphenol flavonoid, is known to exert anti-inflammatory, anti-carcinogenic and anti-diabetic effects. In this study, we aimed to investigate the effect of fisetin on adipocyte proliferation and differentiation in 3T3-L1 preadipocyte cell line and its mechanism of action. We found that fisetin inhibits adipocyte differentiation in a concentration dependent manner, which were evidenced by Oil Red O staining and the protein expression of mature adipocyte marker genes fatty acid synthase and peroxisome proliferator-activated receptor γ. Moreover, the proliferation of preadipocytes was also markedly suppressed by treatment of fisetin for 24 and 48 h in the differentiation medium. We also found that fisetin inhibition of adipocyte differentiation was largely due to the effect on mitotic clonal expansion. Fisetin suppression of preadipocyte proliferation at early stage of differentiation was accompanied by the changes of expression of a series of cell cycle regulatory proteins. Altogether, our results suggest that the inhibition of adipocyte differentiation by fisetin may be at least in part mediated by cell cycle arrest during adipogenesis.

  12. Berberine Activates AMPK and Inhibits 3T3-L1 Adipocyte Differentiation%小檗碱激活AMPK抑制3T3-L1脂肪细胞分化

    Institute of Scientific and Technical Information of China (English)

    王宁; 张娟; 建方方; 邓儒元; 唐红菊; 刘赟; 李凤英; 王晓; 周丽斌

    2012-01-01

    目的 探讨小檗碱对3T3-L1脂肪分化的作用是否与激活腺苷酸活化蛋白激酶(AMPK)有关.方法 在3T3-L脂肪细胞分化全程加入小檗碱,以油红O染色检测3T3-L1脂肪细胞胞浆中脂肪的堆积,实时定量PCR检测过氧化物酶体增殖物激活受体γ2(PPARγ2)、CCAAT增强子结合蛋白α(CEBPα)和AMPK的mRNA表达,以Western印迹法检测AMPK和乙酰辅酶A羧化酶(ACC)的磷酸化水平.结果 小檗碱剂量依赖性地抑制3T3-L1脂肪细胞分化,10 μmol/L小檗碱几乎完全抑制胞浆中脂肪的堆积.5 μmol/L小檗碱在脂肪细胞诱导分化1、3、5、7d后均显著降低CEBPα mRNA表达(P<0.05或P<0.01),诱导分化3、5、7d时显著降低PPARγ2的mRNA表达(P<0.05或P<0.01).AMPK的mRNA水平在分化过程中未受小檗碱的明显影响,而小檗碱明显增加其蛋白磷酸化水平,其下游靶基因ACC磷酸化水平也明显增加.结论 小檗碱抑制3T3-L1脂肪细胞的分化可能与其激活AMPK有关.%Objective To investigate whether the effect of berberine ( BBR) on 3T3-L1 adipocyte differentiation is related to AMP activated protein (AMPK) activation. Methods The accumulation of lipid in the cytoplasm of differentiated 3T3-L1 adipocytes was observed by oil red 0 staining. Realtime-PCR was used to detect the mRNA ezpiesBions of PPARγ2, CEBPα, and AMPK. The phosphorylation levels of AMPK and acetyl CoA carboxylase (ACC) were detected by Western blot. Result Berberine inhibited 3T3-L1 adipocyte differentiation in a dose-dependent manner. At the concentration of 10μmol/L berberine, the accumulation of lipid in the cytoplasm of adipocytes was almost inhibited. CEBPa mRNA expression was inhibited by 5μmol/L berberine after 1,3,5, and 7day induction differentiation (P<0.05 or P<0.01) and PPARry2 mRNA expression was decreased by berberine after induction differentiation of 3,5, and 7 day (P<0.05 or P< 0.01). There were no changes of AMPK mRNA level after 3T3-IA cells were incubated with

  13. Phosphatidylcholine induces apoptosis of 3T3-L1 adipocytes

    Directory of Open Access Journals (Sweden)

    Li Hailan

    2011-12-01

    Full Text Available Abstract Background Phosphatidylcholine (PPC formulation is used for lipolytic injection, even though its mechanism of action is not well understood. Methods The viability of 3T3-L1 pre-adipocytes and differentiated 3T3-L1 cells was measured after treatment of PPC alone, its vehicle sodium deoxycholate (SD, and a PPC formulation. Western blot analysis was performed to examine PPC-induced signaling pathways. Results PPC, SD, and PPC formulation significantly decreased 3T3-L1 cell viability in a concentration-dependent manner. PPC alone was not cytotoxic to CCD-25Sk human fibroblasts at concentrations Conclusions PPC results in apoptosis of 3T3-L1 cells.

  14. Identification of suitable reference genes for quantitative RT-PCR during 3T3-L1 adipocyte differentiation.

    Science.gov (United States)

    Zhang, Juan; Tang, Hongju; Zhang, Yuqing; Deng, Ruyuan; Shao, Li; Liu, Yun; Li, Fengying; Wang, Xiao; Zhou, Libin

    2014-05-01

    Quantitative reverse transcription PCR (qRT-PCR) is becoming increasingly important in the effort to gain insight into the molecular mechanisms underlying adipogenesis. However, the expression profile of a target gene may be misinterpreted due to the unstable expression of the reference genes under different experimental conditions. Therefore, in this study, we investigated the expression stability of 10 commonly used reference genes during 3T3-L1 adipocyte differentiation. The mRNA expression levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and transferrin receptor (TFRC) significantly increased during the course of 3T3-L1 adipocyte differentiation, which was decreased by berberine, an inhibitor of adipogenesis. Three popular algorithms, GeNorm, NormFinder and BestKeeper, identified 18 ribosomal RNA and hydroxymethylbilane synthase (HMBS) as the most stable reference genes, while GAPDH and TFRC were the least stable ones. Peptidylprolyl isomerase A [PIPA (cyclophilin A)], ribosomal protein, large, P0 (36-B4), beta-2-microglobulin (B2M), α1-tubulin, hypoxanthine-guanine phosphoribosyltransferase (HPRT) and β-actin showed relatively stable expression levels. The choice of reference genes with various expression stabilities exerted a profound influence on the expression profiles of 2 target genes, peroxisome proliferator-activated receptor (PPAR)γ2 and C/EBPα. In addition, western blot analysis revealed that the increased protein expression of GAPDH was markedly inhibited by berberine during adipocyte differentiation. This study highlights the importance of selecting suitable reference genes for qRT-PCR studies of gene expression during the process of adipogenesis.

  15. Phytic acid and myo-inositol support adipocyte differentiation and improve insulin sensitivity in 3T3-L1 cells.

    Science.gov (United States)

    Kim, Jin Nam; Han, Sung Nim; Kim, Hye-Kyeong

    2014-08-01

    Phytic acid, also known as myo-inositol hexaphosphate, has been shown to lower blood glucose levels and to improve insulin sensitivity in rodents. We investigated the effects of phytic acid and myo-inositol on differentiation, insulin-stimulated glucose uptake, and lipolysis of adipocytes to test the hypothesis that the antidiabetic properties of phytic acid and myo-inositol are mediated directly through adipocytes. 3T3-L1 cells were treated with 10, 50, or 200 μmol/L of phytic acid or myo-inositol. Oil Red O staining and an intracellular triacylglycerol assay were used to determine lipid accumulation during adipocyte differentiation. Immunoblotting and real-time polymerase chain reaction (PCR) were performed to evaluate expression of transcription factors, a target protein, and insulin signaling molecules. Phytic acid and myo-inositol exposures increased lipid accumulation in a dose-dependent manner (P acid synthase increased upon treatments with phytic acid and myo-inositol (P phytic acid and myo-inositol treatments (P phytic acid and myo-inositol treatments. In fully differentiated adipocytes, phytic acid and myo-inositol reduced basal lipolysis dose dependently (P phytic acid and myo-inositol increase insulin sensitivity in adipocytes by increasing lipid storage capacity, improving glucose uptake, and inhibiting lipolysis.

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

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Seong-Il [Department of Biology, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of); Ko, Hee-Chul [Jeju Sasa Industry Development Agency, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of); Shin, Hye-Sun; Kim, Hyo-Min; Hong, Youn-Suk [Department of Biology, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of); Lee, Nam-Ho [Department of Chemistry, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of); Kim, Se-Jae, E-mail: sjkim@jejunu.ac.kr [Department of Biology, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of); Jeju Sasa Industry Development Agency, Jeju National University, Jejusi, Jeju 690-756 (Korea, Republic of)

    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.

  17. Parabens inhibit fatty acid amide hydrolase: A potential role in paraben-enhanced 3T3-L1 adipocyte differentiation.

    Science.gov (United States)

    Kodani, Sean D; Overby, Haley B; Morisseau, Christophe; Chen, Jiangang; Zhao, Ling; Hammock, Bruce D

    2016-11-16

    Parabens are a class of small molecules that are regularly used as preservatives in a variety of personal care products. Several parabens, including butylparaben and benzylparaben, have been found to interfere with endocrine signaling and to stimulate adipocyte differentiation. We hypothesized these biological effects could be due to interference with the endocannabinoid system and identified fatty acid amide hydrolase (FAAH) as the direct molecular target of parabens. FAAH inhibition by parabens yields mixed-type and time-independent kinetics. Additionally, structure activity relationships indicate FAAH inhibition is selective for the paraben class of compounds and the more hydrophobic parabens have higher potency. Parabens enhanced 3T3-L1 adipocyte differentiation in a dose dependent fashion, different from two other FAAH inhibitors URB597 and PF622. Moreover, parabens, URB597 and PF622 all failed to enhance AEA-induced differentiation. Furthermore, rimonabant, a cannabinoid receptor 1 (CB1)-selective antagonist, did not attenuate paraben-induced adipocyte differentiation. Thus, adipogenesis mediated by parabens likely occurs through modulation of endocannabinoids, but cell differentiation is independent of direct activation of CB1 by endocannabinoids.

  18. Effects of a fatty acid synthase inhibitor on adipocyte differentiation of mouse 3T3-L1 cells

    Institute of Scientific and Technical Information of China (English)

    Li-hong LIU; Xiao-kui WANG; Yuan-dong HU; Jian-lei KANG; Li-li WANG; Song LI

    2004-01-01

    AIM: To investigate the influence of C75, a fatty acid synthase inhibitor, on adipocyte differentiation. METHODS:Mouse 3T3-L1 preadipocytes were induced to differentiation by insulin, isobutylmethylxanthine, and dexamethasone.Oil red O staining was performed and activity of glycerol-3-phosphate dehydrogenase (GPDH) was measured. The level of peroxisome proliferators-activated receptor γ (PPARγ) mRNA was assayed by semi-quantitative reverse transcription PCR. RESULTS: C75 blocked the adipogenic conversion in a dose-dependent manner and the inhibitory effects occurred both in the early phases (48 h) and in the latter phases (8 d) of the process. Treatment with C75 for 8 d induced more decrease in lipid content than 48 h (P<0.01). Treatment with C75 50 mg/L for 48 h or 8 d decreased GPDH activity by 52.8 % and 31.2 % of Vehicle, respectively. Treatment with C75 10-50 mg/L for 48 h or 8 d down-regulated PPARγ mRNA expression compared with control (P<0.01). CONCLUSION: C75 blocked the adipocyte differentiation, which was related with down-regulation of PPARγ mRNA.

  19. Murine 3T3-L1 adipocyte cell differentiation model: validated reference genes for qPCR gene expression analysis.

    Directory of Open Access Journals (Sweden)

    Tatjana Arsenijevic

    Full Text Available BACKGROUND: Analysis of gene expression at the mRNA level, using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR, mandatorily requires reference genes (RGs as internal controls. However, increasing evidences have shown that RG expression may vary considerably under experimental conditions. We sought for an appropriate panel of RGs to be used in the 3T3-L1 cell line model during their terminal differentiation into adipocytes. To this end, the expression levels of a panel of seven widely used RG mRNAs were measured by qRT-PCR. The 7 RGs evaluated were ß-actin (ACTB, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, hypoxanthine phosphoribosyl-transferase I (HPRT, ATP synthase H+ transporting mitochondrial F1 complex beta subunit (ATP-5b, tyrosine 3-monooxygenase/tryptophan 5- monooxygenase activation protein, zeta polypeptide (Ywhaz, Non-POU-domain containing octamer binding protein (NoNo, and large ribosomal protein L13a (RPL. METHODOLOGY/PRINCIPAL FINDINGS: Using three Excel applications, GeNorm, NormFinder and BestKeeper, we observed that the number and the stability of potential RGs vary significantly during differentiation of 3T3-L1 cells into adipocytes. mRNA expression analyses using qRT-PCR revealed that during the entire differentiation program, only NoNo expression is relatively stable. Moreover, the RG sets that were acceptably stable were different depending on the phase of the overall differentiation process (i.e. mitotic clonal expansion versus the terminal differentiation phase. RPL, ACTB, and Ywhaz, are suitable for terminal differentiation, whereas ATP-5b and HPRT, are suitable during mitotic clonal expansion. CONCLUSION: Our results demonstrate that special attention must be given to the choice of suitable RGs during the various well defined phases of adipogenesis to ensure accurate data analysis and that the use of several RGs is absolutely required. Consequently, our data show for the first time

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

    Science.gov (United States)

    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.

  1. Insulin stimulates actin comet tails on intracellular GLUT4-containing compartments in differentiated 3T3L1 adipocytes.

    Science.gov (United States)

    Kanzaki, M; Watson, R T; Khan, A H; Pessin, J E

    2001-12-28

    Incubation of isolated GLUT4-containing vesicles with Xenopus oocyte extracts resulted in a guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) and sodium orthovanadate stimulation of actin comet tails. The in vitro actin-based GLUT4 vesicle motility was inhibited by both latrunculin B and a dominant-interfering N-WASP mutant, N-WASP/Delta VCA. Preparations of gently sheared (broken) 3T3L1 adipocytes also displayed GTP gamma S and sodium orthovanadate stimulation of actin comet tails on GLUT4 intracellular compartments. Furthermore, insulin pretreatment of intact adipocytes prior to gently shearing also resulted in a marked increase in actin polymerization and actin comet tailing on GLUT4 vesicles. In addition, the insulin stimulation of actin comet tails was completely inhibited by Clostridum difficile toxin B, demonstrating a specific role for a Rho family member small GTP-binding protein. Expression of N-WASP/Delta VCA in intact cells had little effect on adipocyte cortical actin but partially inhibited insulin-stimulated GLUT4 translocation. Taken together, these data demonstrate that insulin can induce GLUT4 vesicle actin comet tails that are necessary for the efficient translocation of GLUT4 from intracellular storage sites to the plasma membrane.

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

    Science.gov (United States)

    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.

  3. A proteomic approach for identification of secreted proteins during the differentiation of 3T3-L1 preadipocytes to adipocytes

    DEFF Research Database (Denmark)

    Kratchmarova, Irina; Kalume, Dario E; Blagoev, Blagoy;

    2002-01-01

    We have undertaken a systematic proteomic approach to purify and identify secreted factors that are differentially expressed in preadipocytes versus adipocytes. Using one-dimensional gel electrophoresis combined with nanoelectrospray tandem mass spectrometry, proteins that were specifically secre...

  4. Aspartame downregulates 3T3-L1 differentiation.

    Science.gov (United States)

    Pandurangan, Muthuraman; Park, Jeongeun; Kim, Eunjung

    2014-10-01

    Aspartame is an artificial sweetener used as an alternate for sugar in several foods and beverages. Since aspartame is 200 times sweeter than traditional sugar, it can give the same level of sweetness with less substance, which leads to lower-calorie food intake. There are reports that consumption of aspartame-containing products can help obese people lose weight. However, the potential role of aspartame in obesity is not clear. The present study investigated whether aspartame suppresses 3T3-L1 differentiation, by downregulating phosphorylated peroxisome proliferator-activated receptor γ (p-PPARγ), peroxisome proliferator-activated receptor γ (PPARγ), fatty acid-binding protein 4 (FABP4), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP1), which are critical for adipogenesis. The 3T3-L1 adipocytes were cultured and differentiated for 6 d in the absence and presence of 10 μg/ml of aspartame. Aspartame reduced lipid accumulation in differentiated adipocytes as evidenced by Oil Red O staining. qRT-PCR analysis showed that the PPARγ, FABP4, and C/EBPα mRNA expression was significantly reduced in the aspartame-treated adipocytes. Western blot analysis showed that the induction of p-PPARγ, PPARγ, SREBP1, and adipsin was markedly reduced in the aspartame-treated adipocytes. Taken together, these data suggest that aspartame may be a potent substance to alter adipocyte differentiation and control obesity.

  5. A commercial formulation of glyphosate inhibits proliferation and differentiation to adipocytes and induces apoptosis in 3T3-L1 fibroblasts.

    Science.gov (United States)

    Martini, Claudia N; Gabrielli, Matías; Vila, María del C

    2012-09-01

    Glyphosate-based herbicides are extensively used for weed control all over the world. Therefore, it is important to investigate the putative toxic effects of these formulations which include not only glyphosate itself but also surfactants that may also be toxic. 3T3-L1 fibroblasts are a useful tool to study adipocyte differentiation, this cell line can be induced to differentiate by addition of a differentiation mixture containing insulin, dexamethasone and 3-isobutyl-1-methylxanthine. We used this cell line to investigate the effect of a commercial formulation of glyphosate (GF) on proliferation, survival and differentiation. It was found that treatment of exponentially growing cells with GF for 48h inhibited proliferation in a dose-dependent manner. In addition, treatment with GF dilution 1:2000 during 24 or 48h inhibited proliferation and increased cell death, as evaluated by trypan blue-exclusion, in a time-dependent manner. We showed that treatment of 3T3-L1 fibroblasts with GF increased caspase-3 like activity and annexin-V positive cells as evaluated by flow cytometric analysis, which are both indicative of induction of apoptosis. It was also found that after the removal of GF, remaining cells were able to restore proliferation. On the other hand, GF treatment severely inhibited the differentiation of 3T3-L1 fibroblasts to adipocytes. According to our results, a glyphosate-based herbicide inhibits proliferation and differentiation in this mammalian cell line and induces apoptosis suggesting GF-mediated cellular damage. Thus, GF is a potential risk factor for human health and the environment.

  6. Cannabidiol promotes browning in 3T3-L1 adipocytes.

    Science.gov (United States)

    Parray, Hilal Ahmad; Yun, Jong Won

    2016-05-01

    Recruitment of the brown-like phenotype in white adipocytes (browning) and activation of existing brown adipocytes are currently being investigated as a means to combat obesity. Thus, a wide variety of dietary agents that contribute to browning of white adipocytes have been identified. The present study was designed to investigate the effects of cannabidiol (CBD), a major nonpsychotropic phytocannabinoid of Cannabis sativa, on induction of browning in 3T3-L1 adipocytes. CBD enhanced expression of a core set of brown fat-specific marker genes (Ucp1, Cited1, Tmem26, Prdm16, Cidea, Tbx1, Fgf21, and Pgc-1α) and proteins (UCP1, PRDM16, and PGC-1α). Increased expression of UCP1 and other brown fat-specific markers contributed to the browning of 3T3-L1 adipocytes possibly via activation of PPARγ and PI3K. In addition, CBD increased protein expression levels of CPT1, ACSL, SIRT1, and PLIN while down-regulating JNK2, SREBP1, and LPL. These data suggest possible roles for CBD in browning of white adipocytes, augmentation of lipolysis, thermogenesis, and reduction of lipogenesis. In conclusion, the current data suggest that CBD plays dual modulatory roles in the form of inducing the brown-like phenotype as well as promoting lipid metabolism. Thus, CBD may be explored as a potentially promising therapeutic agent for the prevention of obesity.

  7. TNF-alpha inhibits 3T3-L1 adipocyte differentiation without downregulating the expression of C/EBPbeta and delta.

    Science.gov (United States)

    Kurebayashi, S; Sumitani, S; Kasayama, S; Jetten, A M; Hirose, T

    2001-04-01

    Tumor necrosis factor-alpha (TNF-alpha) has been reported to inhibit adipocyte differentiation in which multiple transcription factors including CCAAT enhancer binding proteins (C/EBPs) and peroxisome proliferator-activated receptor (PPAR) gamma play an important role. Induction of C/EBPalpha and PPARgamma, which regulate the expression of many adipocyte-related genes, is dependent on the expression of C/EBPbeta and C/EBPdelta at the early phase of adipocyte differentiation. To elucidate the mechanism by which TNF-alpha inhibits adipocyte differentiation, we examined the effect of TNF-alpha on the expression of these transcription factors in mouse 3T3-L1 preadipocytes. TNF-alpha did not abrogate the induction of C/EBPbeta and C/EBPdelta in response to differentiation stimuli. In fully differentiated adipocytes, TNF-alpha rapidly induced C/EBPbeta and C/EBPdelta, whereas it downregulated the expression of C/EBPalpha and PPARgamma. Our results suggest that TNF-alpha inhibits adipocyte differentiation independently of the downregulation of C/EBPbeta and C/EBPdelta.

  8. Mango (Mangifera indica L.) peel extract fractions from different cultivars differentially affect lipid accumulation in 3T3-L1 adipocyte cells.

    Science.gov (United States)

    Taing, Meng-Wong; Pierson, Jean-Thomas; Shaw, Paul N; Dietzgen, Ralf G; Roberts-Thomson, Sarah J; Gidley, Michael J; Monteith, Gregory R

    2013-02-26

    Plant phytochemicals are increasingly recognised as sources of bioactive molecules which may have potential benefit in many health conditions. In mangoes, peel extracts from different cultivars exhibit varying effects on adipogenesis in the 3T3-L1 adipocyte cell line. In this study, the effects of preparative HPLC fractions of methanol peel extracts from Irwin, Nam Doc Mai and Kensington Pride mangoes were evaluated. Fraction 1 contained the most hydrophilic components while subsequent fractions contained increasingly more hydrophobic components. High content imaging was used to assess mango peel fraction effects on lipid accumulation, nuclei count and nuclear area in differentiating 3T3-L1 cells. For all three mango cultivars, the more hydrophilic peel fractions 1-3 inhibited lipid accumulation with greater potency than the more hydrophobic peel fractions 4. For all three cultivars, the more lipophilic fraction 4 had concentrations that enhanced lipid accumulation greater than fractions 1-3 as assessed by lipid droplet integrated intensity. The potency of this fraction 4 varied significantly between cultivars. Using mass spectrometry, five long chain free fatty acids were detected in fraction 4; these were not present in any other peel extract fractions. Total levels varied between cultivars, with Irwin fraction 4 containing the highest levels of these free fatty acids. Lipophilic components appear to be responsible for the lipid accumulation promoting effects of some mango extracts and are the likely cause of the diverse effects of peel extracts from different mango cultivars on lipid accumulation.

  9. High-dose Resveratrol Inhibits Insulin Signaling Pathway in 3T3-L1 Adipocytes

    OpenAIRE

    Lee, Haemi; Kim, Jae-Woo

    2013-01-01

    Background Insulin resistance is a major factor in the development of metabolic syndrome and is associated with central obesity and glucose intolerance. Resveratrol, a polyphenol found in fruits, has been shown to improve metabolic conditions. Although it has been widely studied how resveratrol affects metabolism, little is known about how resveratrol regulates lipogenesis with insulin signaling in 3T3-L1 adipocytes. Methods: We treated differentiated 3T3-L1 adipocytes with resveratrol to obs...

  10. 利培酮抑制3T3-L1前脂肪细胞的分化%Risperidone inhibits 3T3-L1 pre-adipocytes differentiation

    Institute of Scientific and Technical Information of China (English)

    张高丽; 张弋; 于海川; 张新雅

    2016-01-01

    Objective To investigate the influence of risperidone on differentiation of 3T3‐L1 pre‐adipocytes .Methods 3T3‐L1 pre‐adipocytes were induced to differentiate into mature adipocytes by adopting the classic hormone cocktail method and observed by the oil red O staining .Meanwhile ,the inducing medium was added with risperidone for studying its influence on 3T3‐L1 pre‐adi‐pocytes differentiation .Results 3T3‐L1 pre‐adipocytes were successfully differentiated into the mature adipocytes ,0 .1 ,1 ,10μmol/L risperidone all could inhibit the differentiation of 3T3‐L1 pre‐adipocytes .Conclusion Risperidone can inhibit the differentiation of 3T3‐L1 pre‐adipocytes .%目的:研究利培酮对3T3‐L1前脂肪细胞分化的影响。方法采用经典的激素鸡尾酒法诱导3T3‐L1前脂肪细胞分化为成熟的脂肪细胞,油红O染色观察。向诱导培养基中加入利培酮研究其对3T3‐L1前脂肪细胞分化的影响。结果用激素鸡尾酒法成功地将3T3‐L1前脂肪细胞诱导为成熟的脂肪细胞。0.1、1、10μmol/L的利培酮均能够抑制3T3‐L1前脂肪细胞的分化。结论利培酮能够抑制3T3‐L1前脂肪细胞的分化。

  11. Rosiglitazone Induces Mitochondrial Biogenesis in Differentiated Murine 3T3-L1 and C3H/10T1/2 Adipocytes.

    Science.gov (United States)

    Rong, James X; Klein, Jean-Louis D; Qiu, Yang; Xie, Mi; Johnson, Jennifer H; Waters, K Michelle; Zhang, Vivian; Kashatus, Jennifer A; Remlinger, Katja S; Bing, Nan; Crosby, Renae M; Jackson, Tymissha K; Witherspoon, Sam M; Moore, John T; Ryan, Terence E; Neill, Sue D; Strum, Jay C

    2011-01-01

    Growing evidence indicates that PPARγ agonists, including rosiglitazone (RSG), induce adipose mitochondrial biogenesis. By systematically analyzing mitochondrial gene expression in two common murine adipocyte models, the current study aimed to further establish the direct role of RSG and capture temporal changes in gene transcription. Microarray profiling revealed that in fully differentiated 3T3-L1 and C3H/10T1/2 adipocytes treated with RSG or DMSO vehicle for 1, 2, 4, 7, 24, and 48 hrs, RSG overwhelmingly increased mitochondrial gene transcripts time dependently. The timing of the increases was consistent with the cascade of organelle biogenesis, that is, initiated by induction of transcription factor(s), followed by increases in the biosynthesis machinery, and then by increases in functional components. The transcriptional increases were further validated by increased mitochondrial staining, citrate synthase activity, and O(2) consumption, and were found to be associated with increased adiponectin secretion. The work provided further insight on the mechanism of PPARγ-induced mitochondrial biogenesis in differentiated adipocytes.

  12. Borrelidin Isolated from Streptomyces sp. Inhibited Adipocyte Differentiation in 3T3-L1 Cells via Several Factors Including GATA-Binding Protein 3.

    Science.gov (United States)

    Matsuo, Hirotaka; Kondo, Yoshiyuki; Kawasaki, Takashi; Tokuyama, Shinji; Imamura, Nobutaka

    2015-01-01

    An inhibitor of 3T3-L1 adipocyte differentiation was isolated from Streptomyces sp. TK08330 and identified by spectroscopy as the 18-membered macrolide borrelidin. Treatment with 1.0 μM borrelidin suppressed intracellular lipid accumulation by 80% and inhibited the expression of adipocyte-specific genes. Borrelidin suppressed the mRNA expression of two master regulators of adipocyte differentiation, peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer binding protein (C/EBPα). Studies on well-known upstream regulators of PPARγ revealed that borrelidin down-regulated C/EBPδ mRNA expression but did not affect expression of C/EBPβ. Borrelidin increased mRNA expression of negative regulators of differentiation such as GATA-binding protein (GATA) 3, Krüppel-like factor (KLF) 3 and KLF7, as well as positive regulators, KLF4, KLF6 and KLF15, at early stages of differentiation. To elucidate a primary mediator of borrelidin differentiation inhibitory activity, small interfering RNA (siRNA) transfection experiments were performed. The mRNA expression of PPARγ, which was down-regulated by borrelidin, was not changed by KLF3 and KLF7 siRNA treatment. In contrast, expression of PPARγ in GATA-3 siRNA-treated cells was not significantly different from that of control siRNA-treated cells. Borrelidin significantly inhibited lipid accumulation in control siRNA-treated cells, and treatment with GATA-3 siRNA slightly reduced the inhibitory effect of borrelidin. These results indicate that borrelidin inhibited adipocyte differentiation partially via GATA-3.

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

    Directory of Open Access Journals (Sweden)

    Mohamad Hafizi Abu Bakar

    2014-12-01

    Full Text Available 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.

  14. Isoproterenol Increases Uncoupling, Glycolysis, and Markers of Beiging in Mature 3T3-L1 Adipocytes.

    Science.gov (United States)

    Miller, Colette N; Yang, Jeong-Yeh; England, Emily; Yin, Amelia; Baile, Clifton A; Rayalam, Srujana

    2015-01-01

    Beta-adrenergic activation stimulates uncoupling protein 1 (UCP1), enhancing metabolic rate. In vitro, most work has studied brown adipocytes, however, few have investigated more established adipocyte lines such as the murine 3T3-L1 line. To assess the effect of beta-adrenergic activation, mature 3T3-L1s were treated for 6 or 48 hours with or without isoproterenol (10 and 100 μM) following standard differentiation supplemented with thyroid hormone (T3; 1 nM). The highest dose of isoproterenol increased lipid content following 48 hours of treatment. This concentration enhanced UCP1 mRNA and protein expression. The increase in UCP1 following 48 hours of isoproterenol increased oxygen consumption rate. Further, coupling efficiency of the electron transport chain was disturbed and an enhancement of glycolytic rate was measured alongside this, indicating an attempt to meet the energy demands of the cell. Lastly, markers of beige adipocytes (protein content of CD137 and gene transcript of CITED1) were also found to be upregulated at 48 hours of isoproterenol treatment. This data indicates that mature 3T3-L1 adipocytes are responsive to isoproterenol and induce UCP1 expression and activity. Further, this finding provides a model for further pharmaceutical and nutraceutical investigation of UCP1 in 3T3-L1s.

  15. Isoproterenol Increases Uncoupling, Glycolysis, and Markers of Beiging in Mature 3T3-L1 Adipocytes.

    Directory of Open Access Journals (Sweden)

    Colette N Miller

    Full Text Available Beta-adrenergic activation stimulates uncoupling protein 1 (UCP1, enhancing metabolic rate. In vitro, most work has studied brown adipocytes, however, few have investigated more established adipocyte lines such as the murine 3T3-L1 line. To assess the effect of beta-adrenergic activation, mature 3T3-L1s were treated for 6 or 48 hours with or without isoproterenol (10 and 100 μM following standard differentiation supplemented with thyroid hormone (T3; 1 nM. The highest dose of isoproterenol increased lipid content following 48 hours of treatment. This concentration enhanced UCP1 mRNA and protein expression. The increase in UCP1 following 48 hours of isoproterenol increased oxygen consumption rate. Further, coupling efficiency of the electron transport chain was disturbed and an enhancement of glycolytic rate was measured alongside this, indicating an attempt to meet the energy demands of the cell. Lastly, markers of beige adipocytes (protein content of CD137 and gene transcript of CITED1 were also found to be upregulated at 48 hours of isoproterenol treatment. This data indicates that mature 3T3-L1 adipocytes are responsive to isoproterenol and induce UCP1 expression and activity. Further, this finding provides a model for further pharmaceutical and nutraceutical investigation of UCP1 in 3T3-L1s.

  16. Polychlorinated biphenyls (PCB 101, PCB 153 and PCB 180) alter leptin signaling and lipid metabolism in differentiated 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Ferrante, Maria C. [Department of Veterinary Medicine and Animal Productions, Federico II University of Naples, Via Delpino 1, 80137 Naples (Italy); Amero, Paola; Santoro, Anna [Department of Pharmacy, Federico II University of Naples, Via Montesano 49, 80131 Naples (Italy); Monnolo, Anna [Department of Veterinary Medicine and Animal Productions, Federico II University of Naples, Via Delpino 1, 80137 Naples (Italy); Simeoli, Raffaele; Di Guida, Francesca [Department of Pharmacy, Federico II University of Naples, Via Montesano 49, 80131 Naples (Italy); Mattace Raso, Giuseppina, E-mail: mattace@unina.it [Department of Pharmacy, Federico II University of Naples, Via Montesano 49, 80131 Naples (Italy); Meli, Rosaria, E-mail: meli@unina.it [Department of Pharmacy, Federico II University of Naples, Via Montesano 49, 80131 Naples (Italy)

    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

  17. Endoplasmic reticulum stress suppresses lipin-1 expression in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Nobuhiko, E-mail: ntkhs@hoku-iryo-u.ac.jp [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1-1-1, Midorigaoka-Higashi, Asahikawa, Hokkaido 078-8510 (Japan); Yoshizaki, Takayuki [Innovation Center, Kagoshima University, 1-21-40, Korimoto, Kagoshima 890-0065 (Japan); Hiranaka, Natsumi; Suzuki, Takeshi [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Yui, Tomoo; Akanuma, Masayoshi [Department of Fixed Prosthodontics and Oral Implantology, School of Dentistry, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Kanazawa, Kaoru [Department of Dental Anesthesiology, School of Dentistry, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Yoshida, Mika; Naito, Sumiyoshi [Department of Clinical Laboratory, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Fujiya, Mikihiro; Kohgo, Yutaka [Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1-1-1, Midorigaoka-Higashi, Asahikawa, Hokkaido 078-8510 (Japan); Ieko, Masahiro [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757, Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan)

    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.

  18. Active form Notch4 promotes the proliferation and differentiation of 3T3-L1 preadipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Peng-Yeh [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Tsai, Chong-Bin [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China); Department of Ophthalmology, Chiayi Christian Hospital, Chiayi 600, Taiwan, ROC (China); Tseng, Min-Jen, E-mail: biomjt@ccu.edu.tw [Institute of Molecular Biology and Department of Life Science, National Chung Cheng University, Chiayi 621, Taiwan, ROC (China)

    2013-01-18

    Highlights: ► Notch4IC modulates the ERK pathway and cell cycle to promote 3T3-L1 proliferation. ► Notch4IC facilitates 3T3-L1 differentiation by up-regulating proadipogenic genes. ► Notch4IC promotes proliferation during the early stage of 3T3-L1 adipogenesis. ► Notch4IC enhances differentiation during subsequent stages of 3T3-L1 adipogenesis. -- Abstract: Adipose tissue is composed of adipocytes, which differentiate from precursor cells in a process called adipogenesis. Many signal molecules are involved in the transcriptional control of adipogenesis, including the Notch pathway. Previous adipogenic studies of Notch have focused on Notch1 and HES1; however, the role of other Notch receptors in adipogenesis remains unclear. Q-RT-PCR analyses showed that the augmentation of Notch4 expression during the differentiation of 3T3-L1 preadipocytes was comparable to that of Notch1. To elucidate the role of Notch4 in adipogenesis, the human active form Notch4 (N4IC) was transiently transfected into 3T3-L1 cells. The expression of HES1, Hey1, C/EBPδ and PPARγ was up-regulated, and the expression of Pref-1, an adipogenic inhibitor, was down-regulated. To further characterize the effect of N4IC in adipogenesis, stable cells expressing human N4IC were established. The expression of N4IC promoted proliferation and enhanced differentiation of 3T3-L1 cells compared with those of control cells. These data suggest that N4IC promoted proliferation through modulating the ERK pathway and the cell cycle during the early stage of 3T3-L1 adipogenesis and facilitated differentiation through up-regulating adipogenic genes such as C/EBPα, PPARγ, aP2, LPL and HSL during the middle and late stages of 3T3-L1 adipogenesis.

  19. 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.

    Science.gov (United States)

    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.

  20. The effects of Ganoderma lucidum herba pharmacopuncture on 3T3-L1 preadipocyte differentiation

    Directory of Open Access Journals (Sweden)

    Chea-woo Lee

    2008-09-01

    Full Text Available Objective : The purpose of this study is to investigate the effects of Ganoderma lucidum herba pharmacopuncture (GHP on the adipogenesis in 3T3-L1 preadipocytes. Methods : 3T3- L1 preadipocytes were differentiated with adipogenic reagents by incubating for 2 days in the absence or presence of GHP ranging from 1 and 2%. The effect of GHP on cell proliferation of 3T3-L1 preadipocytes was investigated using MTT assay. The effect of GHP on adipogenesis was examined by Oil red O staining and measuring glycerol-3-phosphate dehydrogenase (GPDH and intracellular triglyceride (TG content. Results : Following results were obtained from the preadipocyte proliferation and adipocyte differentiation of 3T3-L1. We observed no effect of GHP on preadipocyte proliferation. GHP inhibited adipogenesis, the activity of GPDH and accumulation of intracellular TG content. Conclusions : These results suggest that GHP inhibit differentiation of preadipocyte.

  1. Inhibition of fat cell differentiation in 3T3-L1 pre-adipocytes by all-trans retinoic acid: Integrative analysis of transcriptomic and phenotypic data

    Directory of Open Access Journals (Sweden)

    Katharina Stoecker

    2017-03-01

    Full Text Available The process of adipogenesis is controlled in a highly orchestrated manner, including transcriptional and post-transcriptional events. In developing 3T3-L1 pre-adipocytes, this program can be interrupted by all-trans retinoic acid (ATRA. To examine this inhibiting impact by ATRA, we generated large-scale transcriptomic data on the microRNA and mRNA level. Non-coding RNAs such as microRNAs represent a field in RNA turnover, which is very important for understanding the regulation of mRNA gene expression. High throughput mRNA and microRNA expression profiling was performed using mRNA hybridisation microarray technology and multiplexed expression assay for microRNA quantification. After quantitative measurements we merged expression data sets, integrated the results and analysed the molecular regulation of in vitro adipogenesis. For this purpose, we applied local enrichment analysis on the integrative microRNA-mRNA network determined by a linear regression approach. This approach includes the target predictions of TargetScan Mouse 5.2 and 23 pre-selected, significantly regulated microRNAs as well as Affymetrix microarray mRNA data. We found that the cellular lipid metabolism is negatively affected by ATRA. Furthermore, we were able to show that microRNA 27a and/or microRNA 96 are important regulators of gap junction signalling, the rearrangement of the actin cytoskeleton as well as the citric acid cycle, which represent the most affected pathways with regard to inhibitory effects of ATRA in 3T3-L1 preadipocytes. In conclusion, the experimental workflow and the integrative microRNA–mRNA data analysis shown in this study represent a possibility for illustrating interactions in highly orchestrated biological processes. Further the applied global microRNA–mRNA interaction network may also be used for the pre-selection of potential new biomarkers with regard to obesity or for the identification of new pharmaceutical targets.

  2. 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.

    Science.gov (United States)

    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.

  3. 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

    Directory of Open Access Journals (Sweden)

    A. V. Ramachandran

    2011-07-01

    Full Text Available 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.

  4. Ginkgolide C Suppresses Adipogenesis in 3T3-L1 Adipocytes via the AMPK Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Chian-Jiun Liou

    2015-01-01

    Full Text Available Ginkgolide C, isolated from Ginkgo biloba leaves, is a flavone reported to have multiple biological functions, from decreased platelet aggregation to ameliorating Alzheimer disease. The study aim was to evaluate the antiadipogenic effect of ginkgolide C in 3T3-L1 adipocytes. Ginkgolide C was used to treat differentiated 3T3-L1 cells. Cell supernatant was collected to assay glycerol release, and cells were lysed to measure protein and gene expression related to adipogenesis and lipolysis by western blot and real-time PCR, respectively. Ginkgolide C significantly suppressed lipid accumulation in differentiated adipocytes. It also decreased adipogenesis-related transcription factor expression, including peroxisome proliferator-activated receptor and CCAAT/enhancer-binding protein. Furthermore, ginkgolide C enhanced adipose triglyceride lipase and hormone-sensitive lipase production for lipolysis and increased phosphorylation of AMP-activated protein kinase (AMPK, resulting in decreased activity of acetyl-CoA carboxylase for fatty acid synthesis. In coculture with an AMPK inhibitor (compound C, ginkgolide C also improved activation of sirtuin 1 and phosphorylation of AMPK in differentiated 3T3-L1 cells. The results suggest that ginkgolide C is an effective flavone for increasing lipolysis and inhibiting adipogenesis in adipocytes through the activated AMPK pathway.

  5. Traditional Herbal Formula Oyaksungi-San Inhibits Adipogenesis in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Sae-Rom Yoo

    2015-01-01

    Full Text Available Background. Oyaksungi-san (OYSGS is a herbal formula that has been used for treating cardiovascular diseases in traditional Asian medicine. Here, we investigated the antiadipogenic effect of OYSGS extract in 3T3-L1 adipose cells. Methods. 3T3-L1 preadipocytes were differentiated into adipocytes with or without OYSGS. After differentiation, we measured Oil Red O staining, glycerol-3-phosphate dehydrogenase (GPDH activity, leptin production, mRNA, and protein levels of adipogenesis-related factors. Results. OYSGS extract dramatically inhibited intracellular lipid accumulation in the differentiated adipocytes. It also significantly suppressed the (GPDH activity, triglyceride (TG content, and leptin production by reducing the expression of adipogenesis-related genes including lipoprotein lipase, fatty acid binding protein 4, CCAAT/enhancer-binding protein-alpha (C/EBP-α, and peroxisome proliferator-activated receptor gamma (PPAR-γ. Furthermore, OYSGS clearly enhanced phosphorylation of AMP-activated protein kinase (AMPK as well as its substrate acetyl CoA (ACC carboxylase. Conclusions. Our results demonstrate that OYSGS negatively controls TG accumulation in 3T3-L1 adipocytes. We suggest antiadipogenic activity of OYSGS and its potential benefit in preventing obesity.

  6. 内参基因GAPDH在3T3-L1脂肪细胞分化中的表达变化%Change of reference gene glyceraldehyde-3-phosphate dehydrogenase expression during 3T3-L1 adipocyte differentiation

    Institute of Scientific and Technical Information of China (English)

    张娟; 唐红菊; 王晓; 王宁; 邓儒元; 建方方; 刘赟; 李凤英; 周丽斌

    2012-01-01

    目的 观察甘油醛-3-磷酸脱氢酶(GAPDH)在3T3-L1脂肪细胞分化过程中表达水平是否存在变化,并与其他常用的内参基因相比较.方法 以实时定量PCR检测3T3-L1脂肪细胞分化0、1、3、5、7d几种不同常见内参基因的表达是否存在变化,并以Western印迹方法进行证实.结果 (1)内参基因GAPDH和转铁蛋白受体(TFRC)在脂肪细胞分化过程中基因表达水平逐渐明显升高,其中GAPDH mRNA 在脂肪细胞分化1、3、5、7d分别增加5.7、7.6、22.0和24.5倍(均P<0.01),β-actin、α-微管蛋白(α-tubulin)、肽酰脯氨酰异构酶(PIPA)和18S mRNA表达水平未见明显改变;采用实时定量PCR检测脂肪细胞分化的关键转录因子PPARγ2、CCAAT/增强结合蛋白(C/EBP)α和C/EBPβ的表达时,以GAPDH作内参明显低估他们的表达变化;GAPDH蛋白表达也随着脂肪细胞分化逐渐增加,β-actin、α-tubulin蛋白表达未见明显变化;(2)小檗碱明显抑制脂肪细胞分化过程中GAPDH mRNA和蛋白的表达,在脂肪细胞分化5、7d时GAPDH mRNA表达水平分别降低68.1%和66.3%(P<0.05或P<0.01),但小檗碱对其他内参基因的表达无明显改变.结论 GAPDH在3T3-L1脂肪细胞分化过程中表达增加,不适合作为内参.%Objective To observe the change of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene expression during 3T3-L1 adipocyte differentiation as well as other reference gene expressions.Methods The mRNA expressions of several common reference genes were detected by real time-PCR on day 0,1,3,5,and 7 of 3T3-L1 adipocyte differentiation.Western blot was used to confirm the protein expressions of three common reference genes.Results (1) GAPDH and transferrin receptor(TFRC) mRNA expressions were significantly increased during adipocyte differentiation.GAPDH mRNA level was increased by 5.7,7.6,22.0,and 24.5 folds on day 1,3,5,and 7 after induction of adipocyte differentiation,but no apparent changes of

  7. Fluorescence lifetime imaging of lipids during 3T3-L1 cell differentiation

    Science.gov (United States)

    Song, Young Sik; Won, Young Jae; Lee, Sang-Hak; Kim, Dug Young

    2014-03-01

    Obesity is becoming a big health problem in these days. Since increased body weight is due to increased number and size of the triglyceride-storing adipocytes, many researchers are working on differentiation conditions and processes of adipocytes. Adipocytes also work as regulators of whole-body energy homeostasis by secreting several proteins that regulate processes as diverse as haemostasis, blood pressure, immune function, angiogenesis and energy balance. 3T3-L1 cells are widely used cell line for studying adipogenesis because it can differentiate into an adipocyte-like phenotype under appropriate conditions. In this paper, we propose an effective fluorescence lifetime imaging technique which can easily distinguish lipids in membrane and those in lipid droplets. Nile red dyes are attached to lipids in 3T3-L1 cells. Fluorescence lifetime images were taken for 2 week during differentiation procedure of 3T3-L1 cells into adipocytes. We used 488 nm pulsed laser with 5MHz repetition rate and emission wavelength is 520 nm of Nile Red fluorescent dye. Results clearly show that the lifetime of Nile red in lipid droplets are smaller than those in cell membrane. Our results suggest that fluorescence lifetime imaging can be a very powerful tool to monitor lipid droplet formation in adipocytes from 3T3-L1 cells.

  8. Stevioside from Stevia rebaudiana Bertoni Increases Insulin Sensitivity in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Nabilatul Hani Mohd-Radzman

    2013-01-01

    Full Text Available Stevioside from Stevia rebaudiana has been reported to exert antihyperglycemic effects in both rat and human subjects. There have been few studies on these effects in vitro. In this paper, radioactive glucose uptake assay was implemented in order to assess improvements in insulin sensitivity in 3T3-L1 cells by elevation of glucose uptake following treatment with stevioside. Oil Red-O staining and MTT assay were utilized to confirm adipocyte differentiation and cell viability, respectively. Findings from this research showed a significant increase in absorbance values in mature adipocytes following Oil Red-O staining, confirming the differentiation process. Stevioside was noncytotoxic to 3T3-L1 cells as cell viability was reduced by a maximum of 17%, making it impossible to determine its IC50. Stevioside increased glucose uptake activities by 2.1 times (p<0.001 in normal conditions and up to 4.4 times (p<0.001 in insulin-resistant states. At times, this increase was higher than that seen in positive control group treated with rosiglitazone maleate, an antidiabetic agent. Expressions of pY20 and p-IRS1 which were measured via Western blot were improved by stevioside treatment. In conclusion, stevioside has direct effects on 3T3-L1 insulin sensitivity via increase in glucose uptake and enhanced expression of proteins involved in insulin-signalling pathway.

  9. 3,4-Oxo-isopropylidene-shikimic acid promotes adiopkine expression during murine 3T3-L1 fibroblast differentiation into adipocytes

    Directory of Open Access Journals (Sweden)

    Shifen Dong

    2014-10-01

    Conclusions: These findings demonstrated that ISA promoted adipogenesis by up-regulating expressions of C/EBP β, PPAR γ, C/EBP α, aP2 and FAS, and also stimulated adipokines during adipocyte differentiation. Further study should clarify the relationship between stimulation of adipokines and cognitive enhancing effect of ISA.

  10. Persicaria hydropiper (L.) spach and its flavonoid components, isoquercitrin and isorhamnetin, activate the Wnt/β-catenin pathway and inhibit adipocyte differentiation of 3T3-L1 cells.

    Science.gov (United States)

    Lee, Soung-Hoon; Kim, Bora; Oh, Myoung Jin; Yoon, Juyong; Kim, Hyun Yi; Lee, Kye Jong; Lee, Joo Dong; Choi, Kang-Yell

    2011-11-01

    Obesity, which is related to metabolic syndrome and is associated with liver disease, represents an epidemic problem demanding effective therapeutic strategies. Evidence shows that the Wnt/β-catenin pathway is closely associated with obesity and that small molecules regulating the Wnt/β-catenin pathway can potentially control adipogenesis related to obesity. Eleven plant extracts activating the Wnt/β-catenin pathway were screened by using HEK 293-TOP cells retaining the Wnt/β-catenin signaling reporter gene. An extract of Persicaria hydropiper (L.) Spach was found to activate Wnt/β-catenin signaling. P. hydropiper is grown worldwide in temperate climates and is found widely in Southeast Asia. The P. hydropiper extract inhibited the differentiation of adipocyte 3T3-L1 cells. Isoquercitrin and isorhamnetin, constituents of P. hydropiper, also activated Wnt/β-catenin signaling and suppressed the differentiation of 3T3-L1 cells. These results indicate that isoquercitrin in P. hydropiper suppresses the adipogenesis of 3T3-L1 cells via the inhibition of Wnt/β-catenin signaling. P. hydropiper and isoquercitrin may therefore be potential therapeutic agents for obesity and its associated disorders.

  11. Adipocyte differentiation of 3T3-L1 preadipocytes is dependent on lipoxygenase activity during the initial stages of the differentiation process

    DEFF Research Database (Denmark)

    Madsen, Lise; Petersen, Rasmus K; Sørensen, Morten B

    2003-01-01

    Adipocytes play a central role in whole-body energy homoeostasis. Complex regulatory transcriptional networks control adipogensis, with ligand-dependent activation of PPARgamma (peroxisome proliferator-activated receptor gamma) being a decisive factor. Yet the identity of endogenous ligands...

  12. Effects of Black Adzuki Bean (Vigna angularis Extract on Proliferation and Differentiation of 3T3-L1 Preadipocytes into Mature Adipocytes

    Directory of Open Access Journals (Sweden)

    Mina Kim

    2015-01-01

    Full Text Available 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.

  13. Effect of Tumor Necrosis Factor-α on Resistin Expression in 3T3-L1 Adipocytes and Its Mechanism

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    In order to investigate the effect of tumor necrosis factor-α (TNFα) on resistin expression in 3T3-L1 adipocytes, and further explore its mechanisms, the differentiated 3T3-L1 adipocytes were incubated with 0, 1, 10, 100 ng/mL TNFα respectively for 24 h, and then the expression of resistin was determined. The differentiated 3T3-L1 adipocytes were incubated with 100 ng/mL TNFα for 3, 6, 24 h respectively, and then the expression of resistin mRNA was analyzed.3T3-L1 adipocytes were induced to differentiate into mature adipocytes. The cells were randomly divided into 4 groups for culture. In the control group, no drugs were added. Cells of TNFα group were treated with 100 ng/mL TNFα. In Ro-31-8220 group, 5μmol/L protein kinase C inhibitor Ro-31-8220 was added. With TNFα+Ro-31-8220 group, 100 ng/mL TNFα were added 1 h after the addition of 5 μmol/L Ro-31-8220. All adipocytes were cultured for 24 h. Reverse transcriptionpolymerase chain reaction (RT-PCR) and Western blotting were employed to detect the expression of resistin gene. Our results showed that resistin protein and mRNA in 3T3-L1 adipocytes were inhibited by TNFα at different concentrations (P<0.01), and the inhibitory effect increased with the concentration (P<0.01). At the same concentrations, the inhibitory effect increased with time (P <0.01). Ro-31-8220 could inhibit its expression and the inhibitive effect remained unchanged with addition of TNFα(P>0.05). It was concluded that TNFα could inhibit the expression of resistin in 3T3-L1 adipocytes. The mechanism may be that the expression of resistin is partly controlled by protein kinase C signal conduction pathway.

  14. Effect of pycnogenol on glucose transport in mature 3T3-L1 adipocytes.

    Science.gov (United States)

    Lee, Hee-Hyun; Kim, Kui-Jin; Lee, Ok-Hwan; Lee, Boo-Yong

    2010-08-01

    Pycnogenol, a procyanidins-enriched extract of Pinus maritima bark, possesses antidiabetic properties, which improves the altered parameters of glucose metabolism that are associated with type 2 diabetes mellitus (T2DM). Since the insulin-stimulated antidiabetic activities of natural bioactive compounds are mediated by GLUT4 via the phosphatidylinositol-3-kinase (PI3K) and/or p38 mitogen activated protein kinase (p38-MAPK) pathway, the effects of pycnogenol were examined on the molecular mechanism of glucose uptake by the glucose transport system. 3T3-L1 adipocytes were treated with various concentrations of pycnogenol, and glucose uptake was examined using a non-radioisotope enzymatic assay and by molecular events associated with the glucose transport system using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The results show that pycnogenol increased glucose uptake in fully differentiated 3T3-L1 adipocytes and increased the relative abundance of both GLUT4 and Akt mRNAs through the PI3K pathway in a dose dependent manner. Furthermore, pycnogenol restored the PI3K antagonist-induced inhibition of glucose uptake in the presence of wartmannin, an inhibitor of the PI3K. Overall, these results indicate that pycnogenol may stimulate glucose uptake via the PI3K dependent tyrosine kinase pathways involving Akt. Further the results suggest that pycnogenol might be useful in maintaining blood glucose control.

  15. Octanoate and decanoate induce apoptosis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Yang, Jeong-Yeh; Rayalam, Srujana; Della-Fera, Mary Anne; Ambati, Suresh; Baile, Clifton A

    2009-10-01

    The effect of octanoate and decanoate, respectively, eight- and 10-carbon medium-chain fatty acids (MCFAs), on apoptotic signaling in 3T3-L1 adipocytes was investigated. 3T3-L1 adipocytes were treated with various concentrations of octanoate or decanoate. Cell viability, apoptosis, and expression of apoptosis-related proteins were investigated. Results indicated that both octanoate and decanoate decreased viability, increased apoptosis, and increased reactive oxygen species production. Immunoblotting analysis showed an increase in the levels of cytoplasmic cytochrome c and cleaved poly(ADP-ribose) polymerase by octanoate and decanoate. Concomitantly, we observed that pro-caspase-3 was decreased, resulting in the induced accumulation of the cleaved form of caspase-3 by both octanoate and decanoate. In addition, both octanoate and decanoate increased the expression of pro-apoptotic Bax with an accompanied decrease of anti-apoptotic Bcl-2. These results show that octanoate and decanoate mediate adipocyte apoptosis via a caspase-dependent mitochondrial pathway in 3T3-L1 adipocytes. MCFAs thus decrease adipocyte number by initiating the apoptotic process in 3T3-L1 adipocytes.

  16. 11 beta-hydroxysteroid dehydrogenase type 1 promotes differentiation of 3T3-L1 preadipocyte

    Institute of Scientific and Technical Information of China (English)

    Yun LIU; Yan SUN; Ting ZHU; Yu XIE; Jing YU; Wen-lan SUN; Guo-xian DING; Gang HU

    2007-01-01

    Aim: To investigate the relationship between 11 beta-hydroxysteroid dehydroge-nase type 1 (1 lbeta-HSD1), a potential link between obesity and type 2 diabetes,and preadipocyte differentiation. Methods: Mouse 11beta-HSD1 siRNA plasmids were transfected into 3T3-L1 preadipocytes (a cell line derived from mouse Swiss3T3 cells that were isolated from mouse embryo), for examination of the effect of targeted 11 beta-HSD1 inhibition on differentiation of 3T3-L1 cells. Dif-ferentiation was stimulated with 3-isobutyl-1-methyxanthine, insulin, and dexamethasone. The transcription level of the genes was detected by real-time PCR. Results: Lipid accumulation was significantly inhibited in cells transfected with mouse 11beta-HSD1 siRNA compared with non-transfected 3T3-L1 cells.Fewer lipid droplets were detected in the transfected cells both prior to stimulation and after stimulation with differentiation-inducing reagents. The expression of adipocyte differentiation-associated markers such as lipoprotein lipase and fatty acid synthetase were downregulated in the transfected cells. Similarly, the expres-sion of preadipocyte factor-1, an inhibitor of adipocyte differentiation, was downregulated upon stimulation of differentiation and had no changes in the transfected cells. Conclusion: 11 beta-HSD1 can promote preadipocyte differentiation. Based on this, we propose that 11 beta-HSD1 may be an important candidate mediator of obesity and obesity-induced insulin resistance.

  17. Evaluation of chylomicron effect on ASP production in 3T3-L1 adipocytes

    Institute of Scientific and Technical Information of China (English)

    Ying Gao; Danny Gauvreau; Wei Cui; Marc Lapointe; Sabina Paglialunga; Katherine Cianflone

    2011-01-01

    In the past few years,there has been increasing interest in the production and physiological role of acylation-stimu-lating protein(ASP),identical to C3adesArg,a product of the alternative complement pathway generated through C3 cleavage.Recent studies in C3(-/-)mice that are ASP deficient have demonstrated a role for ASP in postprandial triglyceride clearance and fat storage.The aim of the present study was to establish a cell model and sensitive ELISA assay for the evaluation of ASP production using 3T3-L1 adipocytes.3T3-L1 preadipocytes were differentiated into adipocytes,then cultured in different media such as serum-free(SF),Dulbecco's modified Eagle's medium(DMEM)/F12+10% fetal calf serum (FBS),and at varying concentrations of chylomicrons and insulin+chylomicrons up to 48 h.ASP production in SF and DMEM/F12+10% FBS was compared.Chylomicrons stimulated ASP production in a concen-tration- and time-dependent manner.By contrast,chylo-micron treatment had no effect on the production of C3,the precursor protein of ASP,which was constant over 48 h.Addition of insulin(100 nM)to a low-dose of chylomicrons(100 μg TG/ml)significantly increased ASP production compared with chylomicrons alone at 48 h(P<0.001).Furthermore,addition of insulin significantly increased C3 secretion at both 18 and 48 h of incubation (P<0.05,P<0.001,respectively).Overall,the proportion of ASP to C3 remained constant,indicating no change in the ratio of C3 cleaved to generate ASP.This study demonstrated that 3T3-L1 adipocyte is a useful model for the evaluation of C3 secretion and ASP production by using a sensitive mouse-specific ELISA assay.The stimulation of ASP production with chylomicrons demonstrates a physiologically relevant response,and provides a strategy for further studies on ASP production and function.

  18. Withaferin A induces apoptosis and inhibits adipogenesis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Park, Hea Jin; Rayalam, Srujana; Della-Fera, Mary Anne; Ambati, Suresh; Yang, Jeong-Yeh; Baile, Clifton A

    2008-01-01

    Withaferin A (WA), a highly oxygenated steroidal lactone that is found in the medicinal plant Withania somnifera (also called ashwagandha) has been reported to have anti-tumor, anti-angiogenesis, and pro-apoptotic activity. We investigated the effects of WA on viability, apoptosis and adipogenesis in 3T3-L1 adipocytes. Pre- and post-confluent preadipocytes and mature adipocytes were treated with WA (1-25 microM) up to 24 hrs. Viability and apoptosis were measured by CellTiter-Blue Cell Viability Assay and single strand DNA ELISA Assay, respectively. WA decreased viability and induced apoptosis in all stages of cells. Induction of apoptosis by WA in mature adipocytes was mediated by increased ERK1/2 phosphorylation and altered Bax and Bcl2 protein expression. The effect of WA on adipogenesis was examined by AdipoRed Assay after treating with WA (0.1-1 microM) during the differentiation period. WA decreased lipid accumulation in a dose-dependent manner and decreased the expression of peroxisome proliferator-activated receptor gamma, CCAAT/enhancer binding protein alpha and adipocyte fatty acid binding protein. The effects on apoptosis and lipid accumulation were also confirmed with Hoechst staining and Oil Red O staining, respectively. These results show that WA acts on adipocytes to reduce cell viability and adipogenesis and also induce apoptosis.

  19. Effects of Methylmercury exposure in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Theresa Vertigan

    2017-02-01

    Full Text Available Mercury-containing compounds are environmental pollutants that have become increasingly consequential in the Arctic regions of North America due to processes of climate change increasing their release and availability at northern latitudes. Currently, the form of mercury known to be most detrimental to human health is methylmercury, CH3Hg+, which is found in the environment and accumulates in the tissues of piscivores, including those consumed by Alaska Natives through subsistence gathering. Much is known about the neurotoxicity of methylmercury after exposure to high concentrations, but little is known about toxicity to other tissues and cell types, particularly for long-term exposure and the lower concentrations that would occur through fish consumption. Effects of methylmercury exposure on 3T3-L1 adipocytes in culture were assessed using assays for cytotoxicity and an ELISA assay for vascular endothelial growth factor (VEGF, a signaling molecule shown to be important for maintaining metabolic status in adipose tissue. Results showed that exposure to methylmercury leads to significant toxicity in adipocytes at exposures of 100 ng/mL during later stages of differentiation, but lower methylmercury concentrations produced little to no toxicity. Results also showed that VEGF secretion is elevated in adipocytes exposed to methylmercury after the process of differentiating into mature, fat-storing cells. These results provide a basis for further exploration into metabolic consequences of methylmercury exposure on specific cell types and cell models.

  20. Thyroid-stimulating hormone inhibits adipose triglyceride lipase in 3T3-L1 adipocytes through the PKA pathway.

    Directory of Open Access Journals (Sweden)

    Dongqing Jiang

    Full Text Available Thyroid-stimulating hormone (TSH has been shown to play an important role in the regulation of triglyceride (TG metabolism in adipose tissue. Adipose triglyceride lipase (ATGL is a rate-limiting enzyme controlling the hydrolysis of TG. Thus far, it is unclear whether TSH has a direct effect on the expression of ATGL. Because TSH function is mediated through the TSH receptor (TSHR, TSHR knockout mice (Tshr-/- mice (supplemented with thyroxine were used in this study to determine the effects of TSHR deletion on ATGL expression. These effects were verified in 3T3-L1 adipocytes and potential underlying mechanisms were explored. In the Tshr-/- mice, ATGL expression in epididymal adipose tissue was significantly increased compared with that in Tshr+/+ mice. ATGL expression was observed to increase with the differentiation process of 3T3-L1 preadipocytes. In mature 3T3-L1 adipocytes, TSH significantly suppressed ATGL expression at both the protein and mRNA levels in a dose-dependent manner. Forskolin, which is an activator of adenylate cyclase, suppressed the expression of ATGL in 3T3-L1 adipocytes. The inhibitory effects of TSH on ATGL expression were abolished by H89, which is a protein kinase A (PKA inhibitor. These results indicate that TSH has an inhibitory effect on ATGL expression in mature adipocytes. The associated mechanism is related to PKA activation.

  1. Berberine activates GLUT1-mediated glucose uptake in 3T3-L1 adipocytes.

    Science.gov (United States)

    Kim, So Hui; Shin, Eun-Jung; Kim, Eun-Do; Bayaraa, Tsenguun; Frost, Susan Cooke; Hyun, Chang-Kee

    2007-11-01

    It has recently been known that berberine, an alkaloid of medicinal plants, has anti-hyperglycemic effects. To explore the mechanism underlying this effect, we used 3T3-L1 adipocytes for analyzing the signaling pathways that contribute to glucose transport. Treatment of berberine to 3T3-L1 adipocytes for 6 h enhanced basal glucose uptake both in normal and in insulin-resistant state, but the insulin-stimulated glucose uptake was not augmented significantly. Inhibition of phosphatidylinositol 3-kinase (PI 3-K) by wortmannin did not affect the berberine effect on basal glucose uptake. Berberine did not augment tyrosine phosphorylation of insulin receptor (IR) and insulin receptor substrate (IRS)-1. Further, berberine had no effect on the activity of the insulin-sensitive downstream kinase, atypical protein kinase C (PKCzeta/lambda). However, interestingly, extracellular signal-regulated kinases (ERKs), which have been known to be responsible for the expression of glucose transporter (GLUT)1, were significantly activated in berberine-treated 3T3-L1 cells. As expected, the level of GLUT1 protein was increased both in normal and insulin-resistant cells in response to berberine. But berberine affected the expression of GLUT4 neither in normal nor in insulin-resistant cells. In addition, berberine treatment increased AMP-activated protein kinase (AMPK) activity in 3T3-L1 cells, which has been reported to be associated with GLUT1-mediated glucose uptake. Together, we concluded that berberine increases glucose transport activity of 3T3-L1 adipocytes by enhancing GLUT1 expression and also stimulates the GLUT1-mediated glucose uptake by activating GLUT1, a result of AMPK stimulation.

  2. Combined effects of genistein, quercetin, and resveratrol in human and 3T3-L1 adipocytes.

    Science.gov (United States)

    Park, Hea Jin; Yang, Jeong-Yeh; Ambati, Suresh; Della-Fera, Mary Anne; Hausman, Dorothy B; Rayalam, Srujana; Baile, Clifton A

    2008-12-01

    The natural compounds genistein (G), quercetin (Q), and resveratrol (R) have been reported to each exhibit anti-adipogenic activities in adipocytes and antiproliferative and pro-apoptotic activities in several cell types. We studied the combined effects of G, Q, and R on adipogenesis and apoptosis in primary human adipocytes (HAs) and 3T3-L1 murine adipocyte (MAs). Combined treatment with 6.25 microM G, 12.5 microM Q, and 12.5 microM R during the 14-day differentiation period caused an enhanced inhibition of lipid accumulation in maturing HAs that was greater than the responses to individual compounds and to the calculated additive response. Glycerol 3-phosphate dehydrogenase activity, a marker of late adipocyte differentiation, was decreased markedly in HAs treated with the combination of G+Q+R. In addition, combined treatment with 50 microM G, 100 microM Q, and 100 microM R for 3 days decreased cell viability and induced apoptosis in early- and mid- phase maturing and lipid-filled mature HAs. In contrast, no compound alone induced apoptosis. Oil Red O stain and Hoechst 33342 stain were performed to confirm the effects on lipid accumulation and apoptosis, respectively. We also determined whether MAs responded to the combination treatment similarly to HAs. As in HAs, G+Q+R treatment decreased lipid accumulation in maturing MAs and increased apoptosis in pre- and lipid-filled mature MAs more than the responses to G, Q, and R when used separately. These results show that lower concentrations of combined treatments with several natural compounds may be useful for treatments for obesity through the suppression of adipogenesis and enhanced adipocyte apoptosis.

  3. Suppressive actions of eicosapentaenoic acid on lipid droplet formation in 3T3-L1 adipocytes

    Directory of Open Access Journals (Sweden)

    Sinclair Andrew J

    2010-06-01

    Full Text Available Abstract Background Lipid droplet (LD formation and size regulation reflects both lipid influx and efflux, and is central in the regulation of adipocyte metabolism, including adipokine secretion. The length and degree of dietary fatty acid (FA unsaturation is implicated in LD formation and regulation in adipocytes. The aims of this study were to establish the impact of eicosapentaenoic acid (EPA; C20:5n-3 in comparison to SFA (STA; stearic acid, C18:0 and MUFA (OLA; oleic acid, C18:1n-9 on 3T3-L1 adipocyte LD formation, regulation of genes central to LD function and adipokine responsiveness. Cells were supplemented with 100 μM FA during 7-day differentiation. Results EPA markedly reduced LD size and total lipid accumulation, suppressing PPARγ, Cidea and D9D/SCD1 genes, distinct from other treatments. These changes were independent of alterations of lipolytic genes, as both EPA and STA similarly elevated LPL and HSL gene expressions. In response to acute lipopolysaccharide exposure, EPA-differentiated adipocytes had distinct improvement in inflammatory response shown by reduction in monocyte chemoattractant protein-1 and interleukin-6 and elevation in adiponectin and leptin gene expressions. Conclusions This study demonstrates that EPA differentially modulates adipogenesis and lipid accumulation to suppress LD formation and size. This may be due to suppressed gene expression of key proteins closely associated with LD function. Further analysis is required to determine if EPA exerts a similar influence on LD formation and regulation in-vivo.

  4. Inhibition of adipogenesis and leptin production in 3T3-L1 adipocytes by a derivative of meridianin C

    Energy Technology Data Exchange (ETDEWEB)

    Park, Yu-Kyoung [Department of Molecular Medicine, College of Medicine, Keimyung University, 1095 Dalgubeoldaero, Dalseo-gu, Daegu 704-701 (Korea, Republic of); Lee, Tae-Yoon [Department of Microbiology, College of Medicine, Yeungnam University, 170 Hyunchung-Ro, Nam-gu, Daegu 705-717 (Korea, Republic of); Choi, Jong-Soon [Division of Life Science, Korea Basic Science Institute, 169-148 Gwahakro, Yuseong-gu, Daejeon 305-333 (Korea, Republic of); Hong, Victor Sukbong [Department of Chemistry, College of Natural Sciences, Keimyung University, 1095 Dalgubeoldaero, Dalseo-gu, Daegu 704-701 (Korea, Republic of); Lee, Jinho, E-mail: jinho@gw.kmu.ac.kr [Department of Chemistry, College of Natural Sciences, Keimyung University, 1095 Dalgubeoldaero, Dalseo-gu, Daegu 704-701 (Korea, Republic of); Park, Jong-Wook, E-mail: j303nih@dsmc.or.kr [Department of Immunology, College of Medicine, Keimyung University, 1095 Dalgubeoldaero, Dalseo-gu, Daegu 704-701 (Korea, Republic of); Jang, Byeong-Churl, E-mail: jangbc123@gw.kmu.ac.kr [Department of Molecular Medicine, College of Medicine, Keimyung University, 1095 Dalgubeoldaero, Dalseo-gu, Daegu 704-701 (Korea, Republic of)

    2014-10-03

    Highlights: • Compound 7b, a meridianin C derivative, inhibits adipogenesis. • Compound 7b inhibits C/EBP-α, PPAR-γ, FAS, STAT-3, and STAT-5 in 3T3-L1 adipocytes. • Compound 7b inhibits leptin, but not adiponectin, expression in 3T3-L1 adipocytes. • Compound 7b thus may have therapeutic potential against obesity. - Abstract: Meridianin C, a marine alkaloid, is a potent protein kinase inhibitor and has anti-cancer activity. We have recently developed a series of meridianin C derivatives (compound 7a–7j) and reported their proviral integration Moloney Murine Leukemia Virus (pim) kinases’ inhibitory and anti-proliferative effects on human leukemia cells. Here we investigated the effect of these meridianin C derivatives on adipogenesis. Strikingly, among the derivatives tested, compound 7b most strongly inhibited lipid accumulation during the differentiation of 3T3-L1 preadipocytes into adipocytes. However, meridianin C treatment was largely cytotoxic to 3T3-L1 adipocytes. On mechanistic levels, compound 7b reduced not only the expressions of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), and fatty acid synthase (FAS) but also the phosphorylation levels of signal transducer and activator of transcription-3 (STAT-3) and STAT-5 during adipocyte differentiation. Moreover, compound 7b repressed leptin, but not adiponectin, expression during adipocyte differentiation. Collectively, these findings demonstrate that a meridianin C derivative inhibits adipogenesis by down-regulating expressions and/or phosphorylations of C/EBP-α, PPAR-γ, FAS, STAT-3 and STAT-5.

  5. TC10 is regulated by caveolin in 3T3-L1 adipocytes.

    Directory of Open Access Journals (Sweden)

    Dave Bridges

    Full Text Available BACKGROUND: TC10 is a small GTPase found in lipid raft microdomains of adipocytes. The protein undergoes activation in response to insulin, and plays a key role in the regulation of glucose uptake by the hormone. METHODOLOGY/PRINCIPAL FINDINGS: TC10 requires high concentrations of magnesium in order to stabilize guanine nucleotide binding. Kinetic analysis of this process revealed that magnesium acutely decreased the nucleotide release and exchange rates of TC10, suggesting that the G protein may behave as a rapidly exchanging, and therefore active protein in vivo. However, in adipocytes, the activity of TC10 is not constitutive, indicating that mechanisms must exist to maintain the G protein in a low activity state in untreated cells. Thus, we searched for proteins that might bind to and stabilize TC10 in the inactive state. We found that Caveolin interacts with TC10 only when GDP-bound and stabilizes GDP binding. Moreover, knockdown of Caveolin 1 in 3T3-L1 adipocytes increased the basal activity state of TC10. CONCLUSIONS/SIGNIFICANCE: Together these data suggest that TC10 is intrinsically active in vivo, but is maintained in the inactive state by binding to Caveolin 1 in 3T3-L1 adipocytes under basal conditions, permitting its activation by insulin.

  6. Nebivolol stimulates mitochondrial biogenesis in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Chenglin; Chen, Dongrui; Xie, Qihai [State Key Laboratory of Medical Genomics, Shanghai Key Laboratory of Vascular Biology, Department of Hypertension, Ruijin Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200025 (China); Yang, Ying, E-mail: yangying_sh@yahoo.com [Department of Endocrine and Metabolic Diseases, Shanghai Institute of Endocrine and Metabolic Diseases, Shanghai Clinical Center for Endocrine and Metabolic Diseases, Ruijin Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200025 (China); Shen, Weili, E-mail: weili_shen@hotmail.com [State Key Laboratory of Medical Genomics, Shanghai Key Laboratory of Vascular Biology, Department of Hypertension, Ruijin Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200025 (China)

    2013-08-16

    Highlights: •Nebivolol may act as a partial agonist of β3-adrenergic receptor (AR). •Nebivolol stimulates mitochondrial DNA replication and protein expression. •Nebivolol promotes mitochondrial synthesis via activation of eNOS by β3-AR. -- Abstract: Nebivolol is a third-generation β-adrenergic receptor (β-AR) blocker with additional beneficial effects, including the improvement of lipid and glucose metabolism in obese individuals. However, the underlying mechanism of nebivolol’s role in regulating the lipid profile remains largely unknown. In this study, we investigated the role of nebivolol in mitochondrial biogenesis in 3T3-L1 adipocytes. Exposure of 3T3-L1 cells to nebivolol for 24 h increased mitochondrial DNA copy number, mitochondrial protein levels and the expression of transcription factors involved in mitochondrial biogenesis, including PPAR-γ coactivator-1α (PGC-1α), Sirtuin 3 (Sirt3), mitochondrial transcription factor A (Tfam) and nuclear related factor 1 (Nrf1). These changes were accompanied by an increase in oxygen consumption and in the expression of genes involved in fatty acid oxidation and antioxidant enzymes in 3T3-L1 adipocytes, including nebivolol-induced endothelial nitric oxide synthase (eNOS), as well as an increase in the formation of cyclic guanosine monophosphate (cGMP). Pretreatment with NG-nitro-L-arginine methyl ester (l-NAME) attenuated nebivolol-induced mitochondrial biogenesis, as did the soluble guanylate cyclase inhibitor, ODQ. Treatment with nebivolol and β3-AR blocker SR59230A markedly attenuated PGC-1α, Sirt3 and manganese superoxide dismutase (MnSOD) protein levels in comparison to treatment with nebivolol alone. These data indicate that the mitochondrial synthesis and metabolism in adipocytes that is promoted by nebivolol is primarily mediated through the eNOS/cGMP-dependent pathway and is initiated by the activation of β3-AR receptors.

  7. Characterization of VAMP isoforms in 3T3-L1 adipocytes: implications for GLUT4 trafficking.

    Science.gov (United States)

    Sadler, Jessica B A; Bryant, Nia J; Gould, Gwyn W

    2015-02-01

    The fusion of GLUT4-containing vesicles with the plasma membrane of adipocytes is a key facet of insulin action. This process is mediated by the formation of functional soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes between the plasma membrane t-SNARE complex and the vesicle v-SNARE or VAMP. The t-SNARE complex consists of Syntaxin4 and SNAP23, and whereas many studies identify VAMP2 as the v-SNARE, others suggest that either VAMP3 or VAMP8 may also fulfil this role. Here we characterized the levels of expression, distribution, and association of all the VAMPs expressed in 3T3-L1 adipocytes to provide the first systematic analysis of all members of this protein family for any cell type. Despite our finding that all VAMP isoforms form SDS-resistant SNARE complexes with Syntaxin4/SNAP23 in vitro, a combination of levels of expression (which vary by >30-fold), subcellular distribution, and coimmunoprecipitation analyses lead us to propose that VAMP2 is the major v-SNARE involved in GLUT4 trafficking to the surface of 3T3-L1 adipocytes.

  8. Resveratrol metabolites modify adipokine expression and secretion in 3T3-L1 pre-adipocytes and mature adipocytes.

    Directory of Open Access Journals (Sweden)

    Itziar Eseberri

    Full Text Available OBJECTIVE: Due to the low bioavailability of resveratrol, determining whether its metabolites exert any beneficial effect is an interesting issue. METHODS: 3T3-L1 maturing pre-adipocytes were treated during differentiation with 25 µM of resveratrol or with its metabolites and 3T3-L1 mature adipocytes were treated for 24 hours with 10 µM resveratrol or its metabolites. The gene expression of adiponectin, leptin, visfatin and apelin was assessed by Real Time RT-PCR and their concentration in the incubation medium was quantified by ELISA. RESULTS: Resveratrol reduced mRNA levels of leptin and increased those of adiponectin. It induced the same changes in leptin secretion. Trans-resveratrol-3-O-glucuronide and trans-resveratrol-4'-O-glucuronide increased apelin and visfatin mRNA levels. Trans-resveratrol-3-O-sulfate reduced leptin mRNA levels and increased those of apelin and visfatin. CONCLUSIONS: The present study shows for the first time that resveratrol metabolites have a regulatory effect on adipokine expression and secretion. Since resveratrol has been reported to reduce body-fat accumulation and to improve insulin sensitivity, and considering that these effects are mediated in part by changes in the analyzed adipokines, it may be proposed that resveratrol metabolites play a part in these beneficial effects of resveratrol.

  9. Modest hypoxia significantly reduces triglyceride content and lipid droplet size in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Hashimoto, Takeshi, E-mail: thashimo@fc.ritsumei.ac.jp [Faculty of Sport and Health Science, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577 (Japan); Yokokawa, Takumi; Endo, Yuriko [Faculty of Sport and Health Science, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577 (Japan); Iwanaka, Nobumasa [Ritsumeikan Global Innovation Research Organization, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577 (Japan); Higashida, Kazuhiko [Faculty of Sport and Health Science, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577 (Japan); Faculty of Sport Science, Waseda University, 2-579-15 Mikajima, Tokorozawa, Saitama 359-1192 (Japan); Taguchi, Sadayoshi [Faculty of Sport and Health Science, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577 (Japan)

    2013-10-11

    Highlights: •Long-term hypoxia decreased the size of LDs and lipid storage in 3T3-L1 adipocytes. •Long-term hypoxia increased basal lipolysis in 3T3-L1 adipocytes. •Hypoxia decreased lipid-associated proteins in 3T3-L1 adipocytes. •Hypoxia decreased basal glucose uptake and lipogenic proteins in 3T3-L1 adipocytes. •Hypoxia-mediated lipogenesis may be an attractive therapeutic target against obesity. -- Abstract: Background: A previous study has demonstrated that endurance training under hypoxia results in a greater reduction in body fat mass compared to exercise under normoxia. However, the cellular and molecular mechanisms that underlie this hypoxia-mediated reduction in fat mass remain uncertain. Here, we examine the effects of modest hypoxia on adipocyte function. Methods: Differentiated 3T3-L1 adipocytes were incubated at 5% O{sub 2} for 1 week (long-term hypoxia, HL) or one day (short-term hypoxia, HS) and compared with a normoxia control (NC). Results: HL, but not HS, resulted in a significant reduction in lipid droplet size and triglyceride content (by 50%) compared to NC (p < 0.01). As estimated by glycerol release, isoproterenol-induced lipolysis was significantly lowered by hypoxia, whereas the release of free fatty acids under the basal condition was prominently enhanced with HL compared to NC or HS (p < 0.01). Lipolysis-associated proteins, such as perilipin 1 and hormone-sensitive lipase, were unchanged, whereas adipose triglyceride lipase and its activator protein CGI-58 were decreased with HL in comparison to NC. Interestingly, such lipogenic proteins as fatty acid synthase, lipin-1, and peroxisome proliferator-activated receptor gamma were decreased. Furthermore, the uptake of glucose, the major precursor of 3-glycerol phosphate for triglyceride synthesis, was significantly reduced in HL compared to NC or HS (p < 0.01). Conclusion: We conclude that hypoxia has a direct impact on reducing the triglyceride content and lipid droplet size via

  10. Resveratrol potentiates genistein's antiadipogenic and proapoptotic effects in 3T3-L1 adipocytes.

    Science.gov (United States)

    Rayalam, Srujana; Della-Fera, Mary Anne; Yang, Jeong-Yeh; Park, Hea Jin; Ambati, Suresh; Baile, Clifton A

    2007-12-01

    Genistein (G) and resveratrol (R) individually inhibit adipogenesis in 3T3-L1 adipocytes and induce apoptosis in cancer cells. We investigated whether the combination of G and R resulted in enhanced effects on adipogenesis, lipolysis, and apoptosis in 3T3-L1 cells. Preadipocytes and mature adipocytes were treated with G and R individually at 50 and 100 micromol/L (G100; R100) and in combination. Both in preadipocytes and mature adipocytes, G and R individually decreased cell viability dose-dependently, but G100 + R100 further decreased viability by 59 +/- 0.97% (P < 0.001) and 69.7 +/- 1.2% (P < 0.001) after 48 h compared with G100 and R100, respectively. G100 + R100 induced apoptosis 242 +/- 8.7% (P < 0.001) more than the control after 48 h, whereas G100 and R100 individually increased apoptosis only 46 +/- 9.2 and 46 +/- 7.9%, respectively. G and R did not modulate mitogen-activated protein kinase expression by themselves, but G100 + R100 increased Jun-N-terminal kinase phosphorylation by 38.8 +/- 4.4% (P < 0.001) and decreased extracellular signal-regulating kinase phosphorylation by 48 +/- 3.4% (P < 0.001). Individually, G and R at 25 micromol/L (G25; R25) decreased lipid accumulation by 30 +/- 1.7% and 20.07 +/- 4.27%, respectively (P < 0.001). However, G25 + R25 decreased lipid accumulation by 77.9 +/- 3.4% (P < 0.001). Lipolysis assay revealed that neither G25 nor R25 induced lipolysis, whereas G25 + R25 significantly increased lipolysis by 25.5 +/- 4.6%. The adipocyte-specific proteins PPARgamma and CCAAT/enhancer binding protein-alpha were downregulated after treatment with G + R, but no effect was observed with individual compounds. These results indicate that G and R in combination produce enhanced effects on inhibiting adipogenesis, inducing apoptosis, and promoting lipolysis in 3T3-L1 adipocytes. Thus, the combination of G and R is more potent in exerting antiobesity effects than the individual compounds.

  11. Suppression of lipin-1 expression increases monocyte chemoattractant protein-1 expression in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Nobuhiko, E-mail: ntkhs@hoku-iryo-u.ac.jp [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1-1-1 Midorigaoka-Higashi, Asahikawa, Hokkaido 078-8510 (Japan); Yoshizaki, Takayuki [Innovation Center, Kagoshima University, 1-21-40 Korimoto, Kagoshima 890-0065 (Japan); Hiranaka, Natsumi; Suzuki, Takeshi [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Yui, Tomoo; Akanuma, Masayasu; Oka, Kazuya [Department of Fixed Prosthodontics and Oral Implantology, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Kanazawa, Kaoru [Department of Dental Anesthesiology, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Yoshida, Mika; Naito, Sumiyoshi [Department of Clinical Laboratory, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan); Fujiya, Mikihiro; Kohgo, Yutaka [Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1-1-1 Midorigaoka-Higashi, Asahikawa, Hokkaido 078-8510 (Japan); Ieko, Masahiro [Department of Internal Medicine, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Toubetsu, Hokkaido 061-0023 (Japan)

    2011-11-11

    Highlights: Black-Right-Pointing-Pointer Lipin-1 affects lipid metabolism, adipocyte differentiation, and transcription. Black-Right-Pointing-Pointer Adipose lipin-1 expression is reduced in obesity. Black-Right-Pointing-Pointer Lipin-1 depletion using siRNA in 3T3-L1 adipocytes increased MCP-1 expression. Black-Right-Pointing-Pointer Lipin-1 is involved in adipose inflammation. -- Abstract: Lipin-1 plays a crucial role in the regulation of lipid metabolism and cell differentiation in adipocytes. Expression of adipose lipin-1 is reduced in obesity, and metabolic syndrome. However, the significance of this reduction remains unclear. This study investigated if and how reduced lipin-1 expression affected metabolism. We assessed mRNA expression levels of various genes related to adipocyte metabolism in lipin-1-depleted 3T3-L1 adipocytes by introducing its specific small interfering RNA. In lipin-1-depleted adipocytes, mRNA and protein expression levels of monocyte chemoattractant protein-1 (MCP-1) were significantly increased, although the other genes tested were not altered. The conditioned media from the cells promoted monocyte chemotaxis. The increase in MCP-1 expression was prevented by treatment with quinazoline or salicylate, inhibitors of nuclear factor-{kappa}B activation. Because MCP-1 is related to adipose inflammation and systemic insulin resistance, these results suggest that a reduction in adipose lipin-1 in obesity may exacerbate adipose inflammation and metabolism.

  12. Tea catechins modulate the glucose transport system in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ueda, Manabu; Furuyashiki, Takashi; Yamada, Kayo; Aoki, Yukiko; Sakane, Iwao; Fukuda, Itsuko; Yoshida, Ken-Ichi; Ashida, Hitoshi

    2010-11-01

    In this study, we investigated the effects of tea catechins on the translocation of glucose transporter (GLUT) 4 in 3T3-L1 adipocytes. We found that the ethyl acetate fraction of green tea extract, containing abundant catechins, most decreased insulin-induced glucose uptake activity in 3T3-L1 cells. When the cells were treated with 50 μM catechins in the absence or presence of insulin for 30 min, nongallate-type catechins increased glucose uptake activity without insulin, whereas gallate-type catechins decreased insulin-induced glucose uptake activity. (-)-Epicatechin (EC) and (-)-epigallocatechin (EGC), nongallate-type catechins, increased glucose uptake activity in the dose- and time-dependent manner, whereas (-)-catechin 3-gallate (Cg) and (-)-epigallocatechin 3-gallate (EGCg), gallate-type catechins, decreased insulin-induced glucose uptake activity in the dose- and time-dependent manner. When the cells were treated with 50 μM catechins for 30 min, EC and EGC promoted GLUT4 translocation, whereas Cg and EGCg decreased the insulin-induced translocation in the cells. EC and EGC increased phosphorylation of PKCλ/ζ without phosphorylation of insulin receptor (IR) and Akt. Wortmannin and LY294002, inhibitors for phosphatidylinositol 3'-kinase (PI3K), decreased EC- and EGC-induced glucose uptake activity in the cells. Cg and EGCg decreased phosphorylation of PKCλ/ζ in the presence of insulin without affecting insulin-induced phosphorylation of IR, and Akt. Therefore, EC and EGC promote the translocation of GLUT4 through activation of PI3K, and Cg and EGCg inhibit insulin-induced translocation of GLUT4 by the insulin signaling pathway in 3T3-L1 cells.

  13. Resveratrol induces apoptosis and inhibits adipogenesis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Rayalam, Srujana; Yang, Jeong-Yeh; Ambati, Suresh; Della-Fera, Mary Anne; Baile, Clifton A

    2008-10-01

    Resveratrol, a phytoallexin, has recently been reported to slow aging by acting as a sirtuin activator. Resveratrol also has a wide range of pharmacological effects on adipocytes. In this study, we investigated the effects of resveratrol on adipogenesis and apoptosis using 3T3-L1 cells. In mature adipocytes, 100 and 200 microM resveratrol decreased cell viability dose-dependently by 23 +/- 2.7%, and 75.3 +/- 2.8% (p < 0.0001), respectively, after 48 h treatment, and 100 microM resveratrol increased apoptosis by 76 +/- 8.7% (p < 0.0001). Resveratrol at 25 and 50 microM decreased lipid accumulation in maturing preadipocytes significantly by 43 +/- 1.27% and 94.3 +/- 0.3% (p < 0.0001) and decreased cell viability by 25 +/- 1.3% and 70.4 +/- 1.6% (p < 0.0001), respectively. In order to understand the anti-adipogenic effects of resveratrol, maturing 3T3-L1 preadipocytes were treated with 25 microM resveratrol and the change in the expression of several adipogenic transcription factors and enzymes was investigated using real-time RT-PCR. Resveratrol down-regulated the expression of PPAR gamma, C/EBP alpha, SREBP-1c, FAS, HSL, LPL and up-regulated the expression of genes regulating mitochondrial activity (SIRT3, UCP1 and Mfn2). These results indicate that resveratrol may alter fat mass by directly affecting cell viability and adipogenesis in maturing preadipocytes and inducing apoptosis in adipocytes and thus may have applications for the treatment of obesity.

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

    Science.gov (United States)

    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; Alkema, Wynand

    2013-05-01

    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.

  15. Six new chalcones from Angelica keiskei inducing adiponectin production in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ohnogi, Hiromu; Kudo, Yoko; Tahara, Kenichi; Sugiyama, Katsumi; Enoki, Tatsuji; Hayami, Shoko; Sagawa, Hiroaki; Tanimura, Yuko; Aoi, Wataru; Naito, Yuji; Kato, Ikunoshin; Yoshikawa, Toshikazu

    2012-01-01

    Angelica keiskei (Ashitaba in Japanese), a traditional herb in Japan, contains abundant prenylated chalcones. It has been reported that the chalcones from A. keiskei showed such bioactivities as anti-bacterial, anti-cancer and anti-diabetic effects. Xanthoangelol, 4-hydroxyderricin and six new chalcones were isolated in this study from an ethanol extract of A. keiskei by octadecyl silyl (ODS) and silica gel chromatography, and identified by 1D- and 2D-nuclear magnetic resonance (NMR) and high-resolution mass spectrometric analyses. The chalcones from A. keiskei markedly increased the expression of the adiponectin gene and the production of adiponectin in 3T3-L1 adipocytes. These results suggest that the chalcones from A. keiskei might be useful for preventing the metabolic syndrome.

  16. Effects of leptin on apoptosis and adipogenesis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ambati, Suresh; Kim, Hye-Kyeong; Yang, Jeong-Yeh; Lin, Ji; Della-Fera, Mary Anne; Baile, Clifton A

    2007-02-01

    Leptin has been demonstrated to induce adipose tissue apoptosis, which can contribute to the decrease of adiposity, after either central nervous system or peripheral administration. However, it is not known whether leptin acts only centrally to initiate a signal or can also act directly on adipocytes to induce apoptosis. The objective of this study was to determine the direct effect of leptin on adipocyte apoptosis and adipogenesis in vitro using 3T3-L1 cell lines. An ELISA for single stranded DNA, which is highly specific for apoptotic cells, was used to quantify apoptosis. Preconfluent preadipocytes treated with 10(-9), 10(-8), 10(-7), and 10(-6)M leptin showed inhibitory effects on cell viability, and similar observations were also found in maturing preadipocytes treated during day 0-2 and day 2-4 of maturation. After 48 h incubation with 10(-6)M leptin, LDH release was increased by 24.3% (p<0.05) in preconfluent preadipocytes and by 108.5% (p<0.01) in maturing preadipocytes. However, ssDNA analysis revealed no increased apoptosis in preconfluent or maturing preadipocytes or in mature adipocytes treated with leptin. Leptin significantly reduced lipid accumulation and GPDH activity in maturing preadipocytes, demonstrating an inhibitory effect of leptin on adipogenesis. These results indicate that leptin does not act directly to induce adipocyte apoptosis, but can act directly to inhibit maturation of preadipocytes.

  17. Anti-obesity effects of xanthohumol plus guggulsterone in 3T3-L1 adipocytes.

    Science.gov (United States)

    Rayalam, Srujana; Yang, Jeong-Yeh; Della-Fera, Mary Anne; Park, Hea Jin; Ambati, Suresh; Baile, Clifton A

    2009-08-01

    Xanthohumol (XN) and guggulsterone (GS) have each been shown to inhibit adipogenesis and induce apoptosis in adipocytes. In the present study effects of the combination of XN + GS on 3T3-L1 adipocyte apoptosis and adipogenesis were investigated. Mature adipocytes were treated with XN and GS individually and in combination. XN and GS individually decreased cell viability, but XN + GS caused an enhanced decrease in viability and potentiated induction of apoptosis. Likewise, XN + GS caused a potentiated increase in caspase-3/7 activation, whereas neither of the compounds showed any effect individually. In addition, western blot analysis revealed that XN + GS increased Bax expression and decreased Bcl-2 expression, whereas individual compounds did not show any significant effect. XN and GS both decreased lipid accumulation. Individually, XN at 1.5 microM and GS at 3.12 microM decreased lipid accumulation by 26 +/- 4.5% (P < .001) each, whereas XN1.5 + GS3.12 decreased lipid accumulation by 78.2 +/- 1.8% (P < .001). Moreover, expression of the adipocyte-specific proteins was down-regulated with XN1.5 + GS3.12, but no effect was observed with the individual compounds. Finally, XN + GS caused an enhanced stimulation of lipolysis. Thus, combination of XN and GS is more potent in exerting anti-obesity effects than additive effects of the individual compounds.

  18. Enhanced effects of guggulsterone plus 1,25(OH)2D3 on 3T3-L1 adipocytes.

    Science.gov (United States)

    Rayalam, Srujana; Della-Fera, Mary Anne; Ambati, Suresh; Boyan, Barbara; Baile, Clifton A

    2007-12-21

    Guggulsterone (GS) and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] have been shown to influence adipogenesis in 3T3-L1 cells. We investigated the ability of GS and 1,25(OH)2D3, alone and in combination to inhibit adipogenesis and induce apoptosis in 3T3-L1 adipocytes. Maturing preadipocytes were treated with 1,25(OH)2D3 in combination with GS for 6 days during differentiation. GS and 1,25(OH)2D3 each inhibited lipid accumulation, but the combination potentiated the inhibition of lipid accumulation. Apoptosis was increased by 1,25(OH)2D3 while GS had no effect, but GS + 1,25(OH)2D3 increased apoptosis more than either compound alone. Furthermore, GS + 1,25(OH)2D3 caused a potentiated decrease in the expression of aP2 and farnesoid X receptor expression more than either compound alone. In addition, 1,25(OH)2D3 increased vitamin D receptor expression after 6 days, while GS had no effect. GS + 1,25(OH)2D3, however, caused a potentiated increase in the expression of VDR. These findings show that GS potentiates 1,25(OH)2D3's anti-adipogenic and pro-apoptotic effects in maturing 3T3-L1 preadipocytes.

  19. Cell Volume Regulation and Signaling in 3T3-L1 Pre-adipocytes and Adipocytes

    DEFF Research Database (Denmark)

    Eduardsen, Kathrine; Larsen, Susanne; Novak, Ivana;

    2011-01-01

    for either RVD or RVI in pre-adipocytes. The insulin receptor (InsR) localizes to caveolae and its expression dramatically increases upon adipocyte differentiation. In pre-adipocytes, InsR and its effectors focal adhesion kinase (FAK) and extracellular signal regulated kinase (ERK1/2) localized to focal...... adhesions and were activated by a 5 min exposure to insulin (100 nM). Osmotic shrinkage transiently inhibited InsR Y(146)-phosphorylation, followed by an increase at t=15 min; a similar pattern was seen for ERK1/2 and FAK, in a manner unaffected by cholesterol depletion. In contrast, cell swelling had...... is not required for volume regulation. Given the relationship between hyperosmotic stress and insulin signaling, the finding that cell volume regulation is dramatically altered upon adipocyte differentiation may be relevant for the understanding of insulin resistance and metabolic syndrome....

  20. Ursolic acid inhibits adipogenesis in 3T3-L1 adipocytes through LKB1/AMPK pathway.

    Directory of Open Access Journals (Sweden)

    Yonghan He

    Full Text Available BACKGROUND: Ursolic acid (UA is a triterpenoid compound with multiple biological functions. This compound has recently been reported to possess an anti-obesity effect; however, the mechanisms are less understood. OBJECTIVE: As adipogenesis plays a critical role in obesity, the present study was conducted to investigate the effect of UA on adipogenesis and mechanisms of action in 3T3-L1 preadipocytes. METHODS AND RESULTS: The 3T3-L1 preadipocytes were induced to differentiate in the presence or absence of UA for 6 days. The cells were determined for proliferation, differentiation, fat accumulation as well as the protein expressions of molecular targets that regulate or are involved in fatty acid synthesis and oxidation. The results demonstrated that ursolic acid at concentrations ranging from 2.5 µM to 10 µM dose-dependently attenuated adipogenesis, accompanied by reduced protein expression of CCAAT element binding protein β (C/EBPβ, peroxisome proliferator-activated receptor γ (PPARγ, CCAAT element binding protein α (C/EBPα and sterol regulatory element binding protein 1c (SREBP-1c, respectively. Ursolic acid increased the phosphorylation of acetyl-CoA carboxylase (ACC and protein expression of carnitine palmitoyltransferase 1 (CPT1, but decreased protein expression of fatty acid synthase (FAS and fatty acid-binding protein 4 (FABP4. Ursolic acid increased the phosphorylation of AMP-activated protein kinase (AMPK and protein expression of (silent mating type information regulation 2, homolog 1 (Sirt1. Further studies demonstrated that the anti-adipogenic effect of UA was reversed by the AMPK siRNA, but not by the Sirt1 inhibitor nicotinamide. Liver kinase B1 (LKB1, the upstream kinase of AMPK, was upregulated by UA. When LKB1 was silenced with siRNA or the inhibitor radicicol, the effect of UA on AMPK activation was diminished. CONCLUSIONS: Ursolic acid inhibited 3T3-L1 preadipocyte differentiation and adipogenesis through the LKB1/AMPK

  1. mVps45 knockdown selectively modulates VAMP expression in 3T3-L1 adipocytes.

    Science.gov (United States)

    Sadler, Jessica B A; Roccisana, Jennifer; Virolainen, Minttu; Bryant, Nia J; Gould, Gwyn W

    2015-01-01

    Insulin stimulates the delivery of glucose transporter-4 (GLUT4)-containing vesicles to the surface of adipocytes. Depletion of the Sec1/Munc18 protein mVps45 significantly abrogates insulin-stimulated glucose transport and GLUT4 translocation. Here we show that depletion of mVps45 selectively reduced expression of VAMPs 2 and 4, but not other VAMP isoforms. Although we did not observe direct interaction of mVps45 with any VAMP isoform; we found that the cognate binding partner of mVps45, Syntaxin 16 associates with VAMPs 2, 4, 7 and 8 in vitro. Co-immunoprecipitation experiments in 3T3-L1 adipocytes revealed an interaction between Syntaxin 16 and only VAMP4. We suggest GLUT4 trafficking is controlled by the coordinated expression of mVps45/Syntaxin 16/VAMP4, and that depletion of mVps45 regulates VAMP2 levels indirectly, perhaps via reduced trafficking into specialized subcellular compartments.

  2. Microsomal Triglyceride Transfer Protein (MTP Associates with Cytosolic Lipid Droplets in 3T3-L1 Adipocytes.

    Directory of Open Access Journals (Sweden)

    Joseph D Love

    Full Text Available Lipid droplets are intracellular energy storage organelles composed of a hydrophobic core of neutral lipid, surrounded by a monolayer of phospholipid and a diverse array of proteins. The function of the vast majority of these proteins with regard to the formation and/or turnover of lipid droplets is unknown. Our laboratory was the first to report that microsomal triglyceride transfer protein (MTP, a lipid transfer protein essential for the assembly of triglyceride-rich lipoproteins, was expressed in adipose tissue of humans and mice. In addition, our studies suggested that MTP was associated with lipid droplets in both brown and white fat. Our observations led us to hypothesize that MTP plays a key role in lipid droplet formation and/or turnover. The objective of these studies was to gain insight into the function of MTP in adipocytes. Using molecular, biochemical, and morphologic approaches we have shown: 1 MTP protein levels increase nearly five-fold as 3T3-L1 cells differentiate into adipocytes. 2 As 3T3-L1 cells undergo differentiation, MTP moves from the juxtanuclear region of the cell to the surface of lipid droplets. MTP and perilipin 2, a major lipid droplet surface protein, are found on the same droplets; however, MTP does not co-localize with perilipin 2. 3 Inhibition of MTP activity has no effect on the movement of triglyceride out of the cell either as a lipid complex or via lipolysis. 4 MTP is found associated with lipid droplets within hepatocytes from human fatty livers, suggesting that association of MTP with lipid droplets is not restricted to adipocytes. In summary, our data demonstrate that MTP is a lipid droplet-associated protein. Its location on the surface of the droplet in adipocytes and hepatocytes, coupled with its known function as a lipid transfer protein and its increased expression during adipocyte differentiation suggest a role in lipid droplet biology.

  3. Lactobacillus plantarum LG42 Isolated from Gajami Sik-Hae Inhibits Adipogenesis in 3T3-L1 Adipocyte

    Directory of Open Access Journals (Sweden)

    Jeong-Eun Park

    2013-01-01

    Full Text Available We investigated whether lactic acid bacteria isolated from gajami sik-hae (GLAB are capable of reducing the intracellular lipid accumulation by downregulating the expression of adipogenesis-related genes in differentiated 3T3-L1 cells. The GLAB, Lactobacillus plantarum LG42, significantly decreased the intracellular triglyceride storage and the glycerol-3-phosphate dehydrogenase (GPDH activity in a dose-dependent manner. mRNA expression of transcription factors like peroxisome proliferator-activated receptor (PPAR γ and CCAAT/enhancer-binding protein (C/EBP α involved in adipogenesis was markedly decreased by the GLAB treatment. Moreover, the GLAB also decreased the expression level of adipogenic markers like adipocyte fatty acid binding protein (aP2, leptin, GPDH, and fatty acid translocase (CD36 significantly. These results suggest that the GLAB inhibits lipid accumulation in the differentiated adipocyte through downregulating the expression of adipogenic transcription factors and other specific genes involved in lipid metabolism.

  4. Berberine reduces the expression of adipogenic enzymes and inflammatory molecules of 3T3-L1 adipocyte.

    Science.gov (United States)

    Choi, Bong-Hyuk; Ahn, In-Sook; Kim, Yu-Hee; Park, Ji-Won; Lee, So-Young; Hyun, Chang-Kee; Do, Myoung-Sool

    2006-12-31

    Berberine (BBR), an isoquinoline alkaloid, has a wide range of pharmacological effects, yet its exact mechanism is unknown. In order to understand the anti-adipogenic effect of BBR, we studied the change of expression of several adipogenic enzymes of 3T3-L1 cells by BBR treatment. First, we measured the change of leptin and glycerol in the medium of 3T3-L1 cells treated with 1 micrometer, 5 micrometer and 10 micrometer concentrations of BBR. We also measured the changes of adipogenic and lipolytic factors of 3T3-L1. In 3T3-L1 cells, both leptin and adipogenic factors (SREBP-1c, C/EBP-alpha, PPAR-gamma, fatty acid synthase, acetyl-CoA carboxylase, acyl-CoA synthase and lipoprotein lipase) were reduced by BBR treatment. Glycerol secretion was increased, whereas expression of lipolytic enzymes (hormone-sensitive lipase and perilipin) mRNA was slightly decreased. Next, we measured the change of inflammation markers of 3T3-L1 cells by BBR treatment. This resulted in the down-regulation of mRNA level of inflammation markers such as TNF-alpha, IL-6, C- reactive protein and haptoglobin. Taken together, our data shows that BBR has both anti-adipogenic and anti-inflammatory effects on 3T3-L1 adipocytes, and the anti-adipogenic effect seems to be due to the down-regulation of adipogenic enzymes and transcription factors.

  5. Inhibition of adipogenesis and induction of apoptosis and lipolysis by stem bromelain in 3T3-L1 adipocytes.

    Directory of Open Access Journals (Sweden)

    Sandeep Dave

    Full Text Available The phytotherapeutic protein stem bromelain (SBM is used as an anti-obesity alternative medicine. We show at the cellular level that SBM irreversibly inhibits 3T3-L1 adipocyte differentiation by reducing adipogenic gene expression and induces apoptosis and lipolysis in mature adipocytes. At the molecular level, SBM suppressed adipogenesis by downregulating C/EBPα and PPARγ independent of C/EBPβ gene expression. Moreover, mRNA levels of adipocyte fatty acid-binding protein (ap2, fatty acid synthase (FAS, lipoprotein lipase (LPL, CD36, and acetyl-CoA carboxylase (ACC were also downregulated by SBM. Additionally, SBM reduced adiponectin expression and secretion. SBM's ability to repress PPARγ expression seems to stem from its ability to inhibit Akt and augment the TNFα pathway. The Akt-TSC2-mTORC1 pathway has recently been described for PPARγ expression in adipocytes. In our experiments, TNFα upregulation compromised cell viability of mature adipocytes (via apoptosis and induced lipolysis. Lipolytic response was evident by downregulation of anti-lipolytic genes perilipin, phosphodiestersae-3B (PDE3B, and GTP binding protein G(iα(1, as well as sustained expression of hormone sensitive lipase (HSL. These data indicate that SBM, together with all-trans retinoic-acid (atRA, may be a potent modulator of obesity by repressing the PPARγ-regulated adipogenesis pathway at all stages and by augmenting TNFα-induced lipolysis and apoptosis in mature adipocytes.

  6. Curcumin inhibits adipogenesis in 3T3-L1 adipocytes and angiogenesis and obesity in C57/BL mice.

    Science.gov (United States)

    Ejaz, Asma; Wu, Dayong; Kwan, Paul; Meydani, Mohsen

    2009-05-01

    Angiogenesis is necessary for the growth of adipose tissue. Dietary polyphenols may suppress growth of adipose tissue through their antiangiogenic activity and by modulating adipocyte metabolism. We investigated the effect of curcumin, the major polyphenol in turmeric spice, on angiogenesis, adipogenesis, differentiation, apoptosis, and gene expression involved in lipid and energy metabolism in 3T3-L1 adipocyte in cell culture systems and on body weight gain and adiposity in mice fed a high-fat diet (22%) supplemented with 500 mg curcumin/kg diet for 12 wk. Curcumin (5-20 micromol/L) suppressed 3T3-L1 differentiation, caused apoptosis, and inhibited adipokine-induced angiogenesis of human umbilical vein endothelial cells. Supplementing the high-fat diet of mice with curcumin did not affect food intake but reduced body weight gain, adiposity, and microvessel density in adipose tissue, which coincided with reduced expression of vascular endothelial growth factor (VEGF) and its receptor VEGFR-2. Curcumin increased 5'AMP-activated protein kinase phosphorylation, reduced glycerol-3-phosphate acyl transferase-1, and increased carnitine palmitoyltransferase-1 expression, which led to increased oxidation and decreased fatty acid esterification. The in vivo effect of curcumin on the expression of these enzymes was also confirmed by real-time RT-PCR in subcutaneous adipose tissue. In addition, curcumin significantly lowered serum cholesterol and expression of PPARgamma and CCAAT/enhancer binding protein alpha, 2 key transcription factors in adipogenesis and lipogenesis. The curcumin suppression of angiogenesis in adipose tissue together with its effect on lipid metabolism in adipocytes may contribute to lower body fat and body weight gain. Our findings suggest that dietary curcumin may have a potential benefit in preventing obesity.

  7. 催产素对3T3-L1脂肪细胞糖脂代谢的影响%Effect of oxytocin on glucose and lipid metabolism of 3T3-L1 adipocytes

    Institute of Scientific and Technical Information of China (English)

    朱天一; 钱唯韵; 汤冰倩; 胡浩; 俞淑琴; 孙文君; 袁国跃

    2014-01-01

    目的:观察催产素对3T3-L1脂肪细胞糖脂代谢的影响。方法3T3-L1前脂肪细胞体外培养,并诱导其分化成熟为脂肪细胞。研究催产素对脂肪细胞葡萄糖消耗量以及三酰甘油、游离脂肪酸和甘油的影响。采用实时荧光定量PCR法检测糖脂代谢相关基因GLUT-1、GLUT-4、ATGT、HSL的mRNA表达。结果与对照组比较,催产素20、50、100μg/mL组葡萄糖消耗量有所增加,且表现出剂量相关。催产素组较对照组的三酰甘油降低,而甘油和游离脂肪酸增高。催产素50μg/mL组中脂代谢相关基因HSL表达明显高于对照组,糖代谢相关基因GLUT-4 mRNA表达水平增加。结论催产素处理可减少3T3-L1细胞脂质合成、增加脂质分解作用,并可明显改善脂质积聚。%Objective To study the effect of oxytocin on glucose and lipid metabolism in 3T3-L1 adipocytes. Methods Preadipocytes from 3T3-L1 cell line were cultured in vitro and induced to differentiate to adipocytes. Mature adipocytes were treated with oxytocin. Glucose consumption, triglyceride, free fat acid, and glycerol levels were determined. The mRNA expression of differentiation marker genes such as GLUT-1, GLUT-4, ATGT, and HSL were evaluated by RT-PCR method. Results The glucose consumption in the oxytocin (20, 50, and 100μg/mL) groups were increased with dose-dependent relationship compared with control group. Triglyceride in the oxytocin group was lower than that in control group, while glycerol and free fatty acid decreased. There was significant increase of expression levels of lipid metabolism related gene HSL and sugar metabolism related gene GLUT-4 mRNA in the oxytocin (50μg/mL) group compared with control group. Conclusion Treatment of oxytocin may reduce 3T3-L1 cell lipid synthesis, increase lipid decomposition, and obviously improve lipid accumulation.

  8. Piromelatine decreases triglyceride accumulation in insulin resistant 3T3-L1 adipocytes: role of ATGL and HSL.

    Science.gov (United States)

    Wang, Ping-Ping; She, Mei-Hua; He, Ping-Ping; Chen, Wu-Jun; Laudon, Moshe; Xu, Xuan-Xuan; Yin, Wei-Dong

    2013-08-01

    Piromelatine, a novel investigational multimodal sleep medicine, is developed for the treatment of patients with primary and co-morbid insomnia. Piromelatine has been shown to inhibit weight gain and improve insulin sensitivity in high-fat/high-sucrose-fed (HFHS) rats. Considering that piromelatine has also been implicated in lowering of triglyceride levels in HFHS rats, this work elucidated whether this effect involves in the regulation of adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) in triglyceride (TG) metabolism. In this study, we investigated the effects of piromelatine and MT2 receptors inhibition on TG content, insulin-stimulated glucose uptake, and the expressions of ATGL and HSL in 3T3-L1 adipocytes preincubated in high glucose and high insulin (HGI) conditions. Our results showed that culturing 3T3-L1 adipocytes under HGI conditions increased triglyceride accumulation with concomitant decrease of ATGL and HSL expression, inducing insulin resistance in 3T3-L1 adipocytes. We also found that triglyceride accumulation was significantly inhibited and the levels of ATGL/HSL increased after melatonin or piromelatine treatment. The effects of melatonin/piromelatine (10 nM) were counteracted by pretreatment with the relatively selective MT2 receptor antagonist luzindole (100 nM). In this study, our data demonstrate that piromelatine reverses high glucose and high insulin-induced triglyceride accumulation in 3T3-L1 adipocytes, possibly through up-regulating of ATGL and HSL expression via a melatonin-dependent manner.

  9. Knockdown of LYRM1 rescues insulin resistance and mitochondrial dysfunction induced by FCCP in 3T3-L1 adipocytes.

    Science.gov (United States)

    Zhang, Min; Qin, Zhen-Ying; Dai, Yong-mei; Wang, Yu-Mei; Zhu, Guan-zhong; Zhao, Ya-Ping; Ji, Chen-Bo; Zhu, Jin-Gai; Shi, Chun-Mei; Qiu, Jie; Cao, Xin-Guo; Guo, Xi-Rong

    2014-09-01

    LYR motif-containing 1 (LYRM1) was recently discovered to be involved in adipose tissue homeostasis and obesity-associated insulin resistance. We previously demonstrated that LYRM1 overexpression might contribute to insulin resistance and mitochondrial dysfunction. Additionally, knockdown of LYRM1 enhanced insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes. We investigated whether knockdown of LYRM1 in 3T3-L1 adipocytes could rescue insulin resistance and mitochondrial dysfunction induced by the cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP), a mitochondrion uncoupler, to further ascertain the mechanism by which LYRM1 is involved in obesity-associated insulin resistance. Incubation of 3T3-L1 adipocytes with 1 µM FCCP for 12 h decreased insulin-stimulated glucose uptake, reduced intracellular ATP synthesis, increased intracellular reactive oxygen species (ROS) production, impaired insulin-stimulated Glucose transporter type 4 (GLUT4) translocation, and diminished insulin-stimulated tyrosine phosphorylation of Insulin receptor substrate-1 (IRS-1) and serine phosphorylation of Protein Kinase B (Akt). Knockdown of LYRM1 restored insulin-stimulated glucose uptake, rescued intracellular ATP synthesis, reduced intracellular ROS production, restored insulin-stimulated GLUT4 translocation, and rescued insulin-stimulated tyrosine phosphorylation of IRS-1 and serine phosphorylation of Akt in FCCP-treated 3T3-L1 adipocytes. This study indicates that FCCP-induced mitochondrial dysfunction and insulin resistance are ameliorated by knockdown of LYRM1.

  10. Radicicol, a heat shock protein 90 inhibitor, inhibits differentiation and adipogenesis in 3T3-L1 preadipocytes

    Energy Technology Data Exchange (ETDEWEB)

    He, Yonghan [Department of Nutrition and Food Hygiene, Public Health College, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Aquatic and Crop Resource Development, Life Sciences Branch, National Research Council Canada, Charlottetown, PE, Canada C1A 4P3 (Canada); State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650223 (China); Li, Ying [Department of Nutrition and Food Hygiene, Public Health College, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China); Zhang, Shuocheng [Aquatic and Crop Resource Development, Life Sciences Branch, National Research Council Canada, Charlottetown, PE, Canada C1A 4P3 (Canada); Perry, Ben [Aquatic and Crop Resource Development, Life Sciences Branch, National Research Council Canada, Charlottetown, PE, Canada C1A 4P3 (Canada); Department of Biomedical Sciences, University of Prince Edward Island, 550 University Avenue, Charlottetown, PE, Canada C1A 4P3 (Canada); Zhao, Tiantian [Aquatic and Crop Resource Development, Life Sciences Branch, National Research Council Canada, Charlottetown, PE, Canada C1A 4P3 (Canada); Department of Psychology, University of Toronto, 1265 Military Trail, Toronto, ON, Canada M1C 1A4 (Canada); Wang, Yanwen, E-mail: yanwen.wang@nrc.ca [Aquatic and Crop Resource Development, Life Sciences Branch, National Research Council Canada, Charlottetown, PE, Canada C1A 4P3 (Canada); Department of Biomedical Sciences, University of Prince Edward Island, 550 University Avenue, Charlottetown, PE, Canada C1A 4P3 (Canada); Sun, Changhao, E-mail: sun2002changhao@yahoo.com [Department of Nutrition and Food Hygiene, Public Health College, Harbin Medical University, 157 Baojian Road, Harbin 150081 (China)

    2013-06-28

    Highlights: •Radicicol suppressed intracellular fat accumulation in 3T3-L1 adipocytes. •Radicicol inhibited the expression of FAS and FABP4. •Radicicol blocked cell cycle at the G1-S phase during cell differentiation. •Radicicol inhibited the PDK1/Akt pathway in adipocyte differentiation. -- Abstract: Heat shock protein 90 (Hsp90) is involved in various cellular processes, such as cell proliferation, differentiation and apoptosis. As adipocyte differentiation plays a critical role in obesity development, the present study investigated the effect of an Hsp90 inhibitor radicicol on the differentiation of 3T3-L1 preadipocytes and potential mechanisms. The cells were treated with different concentrations of radicicol during the first 8 days of cell differentiation. Adipogenesis, the expression of adipogenic transcriptional factors, differentiation makers and cell cycle were determined. It was found that radicicol dose-dependently decreased intracellular fat accumulation through down-regulating the expression of peroxisome proliferator-activated receptor γ (PPAR{sub γ}) and CCAAT element binding protein α (C/EBP{sub α}), fatty acid synthase (FAS) and fatty acid-binding protein 4 (FABP4). Flow cytometry analysis revealed that radicicol blocked cell cycle at G1-S phase. Radicicol redcued the phosphorylation of Akt while showing no effect on β-catenin expression. Radicicol decreased the phosphorylation of phosphoinositide-dependent kinase 1 (PDK1). The results suggest that radicicol inhibited 3T3-L1 preadipocyte differentiation through affecting the PDK1/Akt pathway and subsequent inhibition of mitotic clonal expansion and the expression/activity of adipogenic transcriptional factors and their downstream adipogenic proteins.

  11. Bixin regulates mRNA expression involved in adipogenesis and enhances insulin sensitivity in 3T3-L1 adipocytes through PPAR{gamma} activation

    Energy Technology Data Exchange (ETDEWEB)

    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 [Laboratory of Molecular Function of Food, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji 611-0011 (Japan); Kawada, Teruo, E-mail: fat@kais.kyoto-u.ac.jp [Laboratory of Molecular Function of Food, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji 611-0011 (Japan)

    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.

  12. Berberine reverses free-fatty-acid-induced insulin resistance in 3T3-L1 adipocytes through targeting IKKβ

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM:To investigate the effects and molecular mechanisms of berberine on improving insulin resistance induced by free fatty acids (FFAs) in 3T3-LI adipocytes.METHODS:The model of insulin resistance in 3T3-L1 adipocytes was established by adding palmic acid (0.5 mmol/L) to the culture medium.Berberine treatment was performed at the same time.Glucose uptake rate was determined by the 2-deoxy-[3H]-Dglucose method.The levels of IkB kinase beta (IKKβ)Ser181 phosphorylation,insulin receptor substrate1(IRS-1) Ser307 phosphorylation,expression of IKKβ,IRS-1,nuclear transcription factor kappaB p65 (NF-κB p65),phosphatidylinositol-3-kinase p85(PI-3K p85) and glucose transporter 4 (GLUT4) proteins were detected by Western blotting.The distribution of NF-κB p65 proteins inside the adipocytes was observed through confocal laser scanning microscopy(CLSM).RESULTS:After the intervention of palmic acid for 24 h,the insulin-stimulated glucose transport in 3T3-L1 adipocytes was inhibited by 67%.Meanwhile,the expression of IRS-1 and PI-3K p85 protein was reduced,while the levels of IKKβ Ser181 and IRS-1 Ser307 phosphorylation,and nuclear translocation of NF-κB p65 protein were increased.However,the above indexes,which indicated the existence of insulin resistance,were reversed by berberine although the expression of GLUT4,IKKβ and total NF-κB p65 protein were not changed during this study.CONCLUSION:Insulin resistance induced by FFAs in 3T3-L1 adipocytes can be improved by berberine.Berberine reversed free-fatty-acid-induced insulin resistance in 3T3-L1 adipocytes through targeting IKKβ.

  13. Pulicaria jaubertii extract prevents triglyceride deposition in 3T3-L1 adipocytes

    Science.gov (United States)

    Currently, levels of obesity in Middle Eastern countries are increasing. Phytochemicals have anti-obesogenic properties as evidenced by prevention of adipocyte differentiation. In Yemen, Pulicaria jaubertii E.Gamal-Eldin (PJ) is a food additive and a traditional medicine. We tested the ability of ex...

  14. A resistin binding peptide selected by phage display inhibits 3T3-L1 preadipocyte differentiation

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Background Resistin, a newly discovered cysteine-rich hormone secreted mainly by adipose tissues, has been proposed to form a biochemical link between obesity and type 2 diabetes. However, the resistin receptor has not yet been identified. This study aimed to identify resistin binding proteins/receptor.Methods Three cDNA fragments with the same 11 bp 5' sequence were found by screening a cDNA phage display library of rat multiple tissues. As the reading frames of the same 11 bp 5' sequence were interrupted by a TGA stop codon, plaque lift assay was consequently used to prove the readthrough phenomenon. The stop codon in the same 11 bp 5' sequence was replaced by tryptophan, and the binding activity of the coded peptide [AWIL, which was designated as resistin binding peptide (RBP)] with resistin was identified by the confocal microscopy technique and the affinity chromatography experiment. pDual GC-resistin and pDual GC-resistin binding peptide were co-transfected into 3T3-L1 cells to confirm the function of resistin binding peptide.Results Three cDNA fragments with the same 11 bp 5' sequence were found. The TGA stop codon in reading frames of the same 11 bp 5' sequence was proved to be readthroughed. The binding activity of RBP with resistin was consequently identified. The expression of the resistin binding peptide in 3T3-L1 preadipocytes expressing pDual GC-resistin significantly inhibited the adipogenic differentiation.Conclusion RBP could effectively rescue the promoted differentiation of resistin overxepressed 3T3-L1 preadipocyte.

  15. Effects of C-reactive protein on adipokines genes expression in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Yuan, Guoyue, E-mail: yuanguoyue@hotmail.com [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Jia, Jue; Di, Liangliang [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Zhou, Libin [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China); Dong, Sijing; Ye, Jingjing; Wang, Dong; Yang, Ling; Wang, Jifang [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Li, Lianxi [Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People' s Hospital, 600, Yishan Road, Shanghai 200233 (China); Yang, Ying [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China); Mao, Chaoming [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Chen, Mingdao, E-mail: mingdaochensh@yahoo.com [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer CRP increases TNF-{alpha} and IL-6 genes expression in matured 3T3-L1 adipocytes. Black-Right-Pointing-Pointer CRP suppresses adiponectin, leptin and PPAR-{gamma} mRNA levels in matured 3T3-L1 cells. Black-Right-Pointing-Pointer Wortmannin reverses effects of CRP on adiponectin, TNF-{alpha} and leptin mRNA levels. Black-Right-Pointing-Pointer CRP may regulate IR, obesity and metabolic syndrome by this mechanism. -- Abstract: Adipose tissue is now recognized to be an important endocrine organ, secreting a variety of adipokines that are involved in the regulation of energy metabolism, insulin resistance and metabolic syndrome. C-reactive protein (CRP) is considered as one of the most sensitive markers of inflammation. A number of studies have shown that elevation of CRP concentrations is an independent predictive parameter of type 2 diabetes mellitus, which is also strongly associated with various components of the metabolic syndrome. The aim of the present study is to investigate the effects of CRP on adipokines genes expression in 3T3-L1 adipocytes. Quantitative real-time PCR analysis revealed that CRP inhibited adiponectin, leptin and peroxisome proliferator-activated receptor-gamma (PPAR-{gamma}) genes expression and raised tumor necrosis factor-{alpha} (TNF-{alpha}) and interleukin-6 (IL-6) mRNA levels in matured 3T3-L1 adipocytes in a dose and time-dependent manner. Pharmacological inhibition of phosphatidylinositol (PI)-3 kinase by wortmannin partially reversed the effects of CRP on adiponectin, TNF-{alpha} and leptin genes expression. These results collectively suggest that CRP regulates adiponectin, TNF-{alpha}, leptin, IL-6 and PPAR-{gamma} genes expression, and that might represent a mechanism by which CRP regulates insulin resistance, obesity and metabolic syndrome.

  16. Effect of Ganoderma applanatum mycelium extract on the inhibition of adipogenesis in 3T3-L1 adipocytes.

    Science.gov (United States)

    Kim, Ji-Eun; Park, Sung-Jin; Yu, Mi-Hee; Lee, Sam-Pin

    2014-10-01

    Ganoderma applanatum (GA) and related fungal species have been used for over 2000 years in China to prevent and treat various human diseases. However, there is no critical research evaluating the functionality of GA grown using submerged culture technology. This study aimed to evaluate the effects of submerged culture GA mycelium (GAM) and its active components (protocatechualdehyde [PCA]) on preadipocyte differentiation of 3T3-L1 cells. Mouse-derived preadipocyte 3T3-L1 cells were treated with differentiation inducers in the presence or absence of GAM extracts. We determined triglyceride accumulations, glycerol-3-phosphate dehydrogenase (GPDH) activities, and differentiation makers. PCA, the active component of GAM extract, was also used to treat 3T3-L1 cells. The MTT assay showed that the GAM extract (0.01-1 mg/mL) was not toxic to 3T3-L1 preadipocyte. Treatment of cells with GAM extracts and its active components significantly decreased the GPDH activity and lipid accumulation, a marker of adipogenesis, in a dose-dependent manner. Western blot analysis results showed that the protein expression levels of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP1) were inhibited by the GAM extract. In addition, adipogenic-specific genes such as perilipin, fatty acid synthase (FAS), fatty acid transport protein 1 (FATP1), and fatty acid-binding protein 4 (FABP4) decreased in a dose-dependent manner. Quantitative high-performance liquid chromatography analysis showed that the GAM extract contained 1.14 mg/g PCA. GAM extracts suppressed differentiation of 3T3-L1 preadipocytes, in part, through altered regulation of PPARγ, C/EBPα, and SREBP1. These results suggest that GAM extracts and PCA may suppress adipogenesis by inhibiting differentiation of preadipocytes.

  17. Aspirin Breaks the Crosstalk between 3T3-L1 Adipocytes and 4T1 Breast Cancer Cells by Regulating Cytokine Production.

    Directory of Open Access Journals (Sweden)

    Chia-Chien Hsieh

    Full Text Available Breast cancer is one of the most common cancers in women worldwide. The obesity process is normally accompanied by chronic, low-grade inflammation. Infiltration by inflammatory cytokines and immune cells provides a favorable microenvironment for tumor growth, migration, and metastasis. Epidemiological evidence has shown that aspirin is an effective agent against several types of cancer. The aim of this study is to investigate the anti-inflammatory and anti-cancer effects of aspirin on 3T3-L1 adipocytes, 4T1 murine breast cancer cells, and their crosstalk. The results showed that aspirin treatment inhibited differentiation and lipid accumulation by 3T3-L1 preadipocytes, and decreased the secretion of the inflammatory adipokine MCP-1 after stimulation with tumor necrosis factor (TNF-α or conditioned medium from RAW264.7 cells. In 4T1 cells, treatment with aspirin decreased cell viability and migration, possibly by suppressing MCP-1 and VEGF secretion. Subsequently, culture of 4T1 cells in 3T3-L1 adipocyte-conditioned medium (Ad-CM and co-culture of 3T3-L1 and 4T1 cells using a transwell plate were performed to clarify the relationship between these two cell lines. Aspirin exerted its inhibitory effects in the transwell co-culture system, as well as the conditioned-medium model. Aspirin treatment significantly inhibited the proliferation of 4T1 cells, and decreased the production of MCP-1 and PAI-1 in both the Ad-CM model and co-culture system. Aspirin inhibited inflammatory MCP-1 adipokine production by 3T3-L1 adipocytes and the cell growth and migration of 4T1 cells. It also broke the crosstalk between these two cell lines, possibly contributing to its chemopreventive properties in breast cancer. This is the first report that aspirin's chemopreventive activity supports the potential application in auxiliary therapy against obesity-related breast cancer development.

  18. Aspirin Breaks the Crosstalk between 3T3-L1 Adipocytes and 4T1 Breast Cancer Cells by Regulating Cytokine Production.

    Science.gov (United States)

    Hsieh, Chia-Chien; Huang, Yu-Shan

    2016-01-01

    Breast cancer is one of the most common cancers in women worldwide. The obesity process is normally accompanied by chronic, low-grade inflammation. Infiltration by inflammatory cytokines and immune cells provides a favorable microenvironment for tumor growth, migration, and metastasis. Epidemiological evidence has shown that aspirin is an effective agent against several types of cancer. The aim of this study is to investigate the anti-inflammatory and anti-cancer effects of aspirin on 3T3-L1 adipocytes, 4T1 murine breast cancer cells, and their crosstalk. The results showed that aspirin treatment inhibited differentiation and lipid accumulation by 3T3-L1 preadipocytes, and decreased the secretion of the inflammatory adipokine MCP-1 after stimulation with tumor necrosis factor (TNF)-α or conditioned medium from RAW264.7 cells. In 4T1 cells, treatment with aspirin decreased cell viability and migration, possibly by suppressing MCP-1 and VEGF secretion. Subsequently, culture of 4T1 cells in 3T3-L1 adipocyte-conditioned medium (Ad-CM) and co-culture of 3T3-L1 and 4T1 cells using a transwell plate were performed to clarify the relationship between these two cell lines. Aspirin exerted its inhibitory effects in the transwell co-culture system, as well as the conditioned-medium model. Aspirin treatment significantly inhibited the proliferation of 4T1 cells, and decreased the production of MCP-1 and PAI-1 in both the Ad-CM model and co-culture system. Aspirin inhibited inflammatory MCP-1 adipokine production by 3T3-L1 adipocytes and the cell growth and migration of 4T1 cells. It also broke the crosstalk between these two cell lines, possibly contributing to its chemopreventive properties in breast cancer. This is the first report that aspirin's chemopreventive activity supports the potential application in auxiliary therapy against obesity-related breast cancer development.

  19. Inhibition of Adipogenesis by Oligonol through Akt-mTOR Inhibition in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Jae-Yeo Park

    2014-01-01

    Full Text Available Polyphenols have recently become an important focus of study in obesity research. Oligonol is an oligomerized polyphenol, typically comprised of catechin-type polyphenols from a variety of fruits, which has been found to exhibit better bioavailability and bioreactivity than natural polyphenol compounds. Here, we demonstrated that Oligonol inhibits 3T3-L1 adipocyte differentiation by reducing adipogenic gene expression. During adipogenesis, Oligonol downregulated the mRNA levels of peroxisome proliferator-activated receptor γ (PPARγ, CCAAT/enhancer binding proteins α (C/EBPα, and δ (C/EBPδ in a dose-dependent manner and the expression of genes involved in lipid biosynthesis. The antiadipogenic effect of Oligonol appears to originate from its ability to inhibit the Akt and mammalian target of rapamycin (mTOR signaling pathway by diminishing the phosphorylation of ribosomal protein S6 kinase (p70S6K, a downstream target of mTOR and forkhead box protein O1 (Foxo1. These results suggest that Oligonol may be a potent regulator of obesity by repressing major adipogenic genes through inhibition of the Akt signaling pathway, which induces the inhibition of lipid accumulation, ultimately inhibiting adipogenesis.

  20. Alliin, a Garlic (Allium sativum Compound, Prevents LPS-Induced Inflammation in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Saray Quintero-Fabián

    2013-01-01

    Full Text Available Garlic (Allium sativum L. has been used to alleviate a variety of health problems due to its high content of organosulfur compounds and antioxidant activity. The main active component is alliin (S-allyl cysteine sulfoxide, a potent antioxidant with cardioprotective and neuroprotective actions. In addition, it helps to decrease serum levels of glucose, insulin, triglycerides, and uric acid, as well as insulin resistance, and reduces cytokine levels. However its potential anti-inflammatory effect is unknown. We examined the effects of alliin in lipopolysaccharide- (LPS- stimulated 3T3-L1 adipocytes by RT-PCR, Western blot, and microarrays analysis of 22,000 genes. Incubation of cells for 24 h with 100 μmol/L alliin prevented the increase in the expression of proinflammatory genes, IL-6, MCP-1, and Egr-1 in 3T3-L1 adipocytes exposed to 100 ng/mL LPS for 1 h. Interestingly, the phosphorylation of ERK1/2, which is involved in LPS-induced inflammation in adipocytes, was decreased following alliin treatment. Furthermore, the gene expression profile by microarrays evidentiate an upregulation of genes involved in immune response and downregulation of genes related with cancer. The present results have shown that alliin is able to suppress the LPS inflammatory signals by generating an anti-inflammatory gene expression profile and by modifying adipocyte metabolic profile.

  1. Ajoene exerts potent effects in 3T3-L1 adipocytes by inhibiting adipogenesis and inducing apoptosis.

    Science.gov (United States)

    Ambati, Suresh; Yang, Jeong-Yeh; Rayalam, Srujana; Park, Hea Jin; Della-Fera, Mary Anne; Baile, Clifton A

    2009-04-01

    This paper describes effects of several sulfur-containing compounds from garlic on the cell viability, apoptosis and adipogenesis in 3T3-L1 adipocytes. In both preadipocytes and mature adipocytes, 100 and 200 microM ajoene significantly decreased cell viability and increased apoptosis. The effect on apoptosis was further confirmed with Hoechst staining. In contrast, diallyl sulfide, diallyl disulfide, diallyl trisulfide, deoxyalliin, and allyl methyl sulfide had no significant effect on cell viability or apoptosis in either preadipocytes or mature adipocytes. In maturing preadipocytes ajoene significantly decreased lipid accumulation in a dose-dependent manner and these results were further confirmed by a decrease in lipid droplet number and lipid content through Oil Red O staining. There was no significant change in lipid accumulation in maturing preadipocytes treated with other garlic derivatives. Thus, despite the same source of origin, garlic, ajoene was the only one with potent effects on cell viability, apoptosis and adipogenesis in 3T3-L1 adipocytes.

  2. Testosterone stimulates glucose uptake and GLUT4 translocation through LKB1/AMPK signaling in 3T3-L1 adipocytes.

    Science.gov (United States)

    Mitsuhashi, Kazuteru; Senmaru, Takafumi; Fukuda, Takuya; Yamazaki, Masahiro; Shinomiya, Katsuhiko; Ueno, Morio; Kinoshita, Shigeru; Kitawaki, Jo; Katsuyama, Masato; Tsujikawa, Muneo; Obayashi, Hiroshi; Nakamura, Naoto; Fukui, Michiaki

    2016-01-01

    Decreases in serum testosterone concentrations in aging men are associated with metabolic disorders. Testosterone has been reported to increase GLUT4-dependent glucose uptake in skeletal muscle cells and cardiomyocytes. However, studies on glucose uptake occurring in response to testosterone stimulation in adipocytes are currently not available. This study was designed to determine the effects of testosterone on glucose uptake in adipocytes. Glucose uptake was assessed with 2-[(3)H] deoxyglucose in 3T3-L1 adipocytes. GLUT4 translocation was evaluated in plasma membrane (PM) sheets and PM fractions by immunofluorescence and immunoblotting, respectively. Activation of GLUT4 translocation-related protein kinases, including Akt, AMPK, LKB1, CaMKI, CaMKII, and Cbl was followed by immunoblotting. Expression levels of androgen receptor (AR) mRNA and AR translocation to the PM were assessed by real-time RT-PCR and immunoblotting, respectively. The results showed that both high-dose (100 nM) testosterone and testosterone-BSA increased glucose uptake and GLUT4 translocation to the PM, independently of the intracellular AR. Testosterone and testosterone-BSA stimulated the phosphorylation of AMPK, LKB1, and CaMKII. The knockdown of LKB1 by siRNA attenuated testosterone- and testosterone-BSA-stimulated AMPK phosphorylation and glucose uptake. These results indicate that high-dose testosterone and testosterone-BSA increase GLUT4-dependent glucose uptake in 3T3-L1 adipocytes by inducing the LKB1/AMPK signaling pathway.

  3. The role of Akt on Arsenic trioxide suppression of 3T3-L1 preadipocyte differentiation

    Institute of Scientific and Technical Information of China (English)

    Zhi Xin WANG; Chun Sun JIANG; Lei LIU; Xiao Hui WANG; Hai Jing JIN; Qiao WU; Quan CHEN

    2005-01-01

    The present study investigates the molecular details of how arsenic trioxide inhibits preadipocyte differentiation and examines the role of Akt/PKB in regulation of differentiation and apoptosis. Continual exposure of arsenic trioxide, at the clinic achievable dosage that does not induce apoptosis, suppressed 3T3-L1 cell differentiation into fat cells by inhibiting the expression of PPARγ and C/EBPα and disrupting the interaction between PPARγ and RXRα, which determines the programming of the adipogenic genes. Interestingly, if we treated the cells for 12 or 24 h and then withdrew arsenic trioxide, the cells were able to differentiate to the comparable levels of untreated cells as assayed by the activity of GAPDH, the biochemical marker of preadipocyte differentiation. Long term treatment blocked the differentiation and the activity of GAPDH could not recover to the comparable levels of untreated cells. Continual exposure of arsenic trioxide caused accumulation in G2/M phase and the accumulation of p21. We found that arsenic trioxide induced the expression and the phosphorylation of Akt/PKB and it inhibited the interaction between Akt/PKB and PPARγ. Akt/PKB inhibitor appears to block the arsenic trioxide suppression of differentiation. Our results suggested that Akt/PKB may play a role in suppression of apoptosis and negatively regulate preadipocyte differentiation.

  4. Enhancement of ajoene-induced apoptosis by conjugated linoleic acid in 3T3-L1 adipocytes.

    Science.gov (United States)

    Yang, Jeong-Yeh; Della-Fera, Mary Anne; Hausman, Dorothy B; Baile, Clifton A

    2007-06-01

    Ajoene has been shown to induce apoptosis in 3T3-L1 adipocytes. In this report the effects on apoptosis of combinations of ajoene and trans-10, cis-12 conjugated linoleic acid (t10,c12CLA) in 3T3-L1 adipocytes were investigated. Although t10,c12CLA alone had no effect, ajoene plus t10,c12CLA reduced cell viability more than ajoene alone at 24 h (59.1 vs. 85.9% of control, respectively; p<0.05). Compared to treatment with t10,c12CLA, ajoene increased apoptosis 218% after 24 h (p<0.01), whereas ajoene plus t10,c12CLA increased apoptosis 122% over that caused by ajoene alone (p<0.01). Immunoblotting analysis also indicated that ajoene plus t10,c12CLA caused a greater increase in phosphorylation of c-Jun N-terminal kinase (JNK) and Bax expression and a greater release of mitochondrial proteins (cytochrome c, AIF) than additive responses to each compound alone. Ajoene plus t10,c12CLA also increased ROS production more than that resulting from ajoene treatment alone (264 vs 204% after 40 min, respectively; p<0.01). Furthermore, the antioxidant NAC prevented ROS generation and apoptosis by ajoene plus t10,c12CLA. Interestingly, the combination of ajoene and t10,c12CLA increased NF-kappaB activation and decreased the level of phosphorylated Akt more than each compound alone. Altogether, our observations indicate that t10,c12CLA potentiates the effect of ajoene on apoptosis in 3T3-L1 adipocytes.

  5. Cultured 3T3L1 adipocytes dispose of excess medium glucose as lactate under abundant oxygen availability

    Science.gov (United States)

    Sabater, David; Arriarán, Sofía; Romero, María Del Mar; Agnelli, Silvia; Remesar, Xavier; Fernández-López, José Antonio; Alemany, Marià

    2014-01-01

    White adipose tissue (WAT) produces lactate in significant amount from circulating glucose, especially in obesity;Under normoxia, 3T3L1 cells secrete large quantities of lactate to the medium, again at the expense of glucose and proportionally to its levels. Most of the glucose was converted to lactate with only part of it being used to synthesize fat. Cultured adipocytes were largely anaerobic, but this was not a Warburg-like process. It is speculated that the massive production of lactate, is a process of defense of the adipocyte, used to dispose of excess glucose. This way, the adipocyte exports glucose carbon (and reduces the problem of excess substrate availability) to the liver, but the process may be also a mechanism of short-term control of hyperglycemia. The in vivo data obtained from adipose tissue of male rats agree with this interpretation.

  6. Berberine increases adipose triglyceride lipase in 3T3-L1 adipocytes through the AMPK pathway

    OpenAIRE

    Jiang, Dongqing; Wang, Dianhui; ZHUANG, XIANGHUA; Wang, Zhanqing; Ni, Yihong; Chen, Shihong; Sun, Fudun

    2016-01-01

    Background Obesity is closely related to the metabolism of triacylglycerol (TG) in adipocytes. Adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are rate-limiting enzymes that control the hydrolysis of TG. Effects on ATGL and HSL to increase lipolysis may counteract obesity. Berberine (BBR) is a compound derived from the Chinese medicine plant Coptis chinensis. In the present study we show the effects of BBR on ATGL and HSL and explore the potential underlying mechanisms o...

  7. Catechin and quercetin attenuate adipose inflammation in fructose-fed rats and in 3T3-L1 adipocytes

    Science.gov (United States)

    Vazquez Prieto, Marcela A.; Bettaieb, Ahmed; Rodriguez Lanzi, Cecilia; Soto, Verónica C.; Perdicaro, Diahann J.; Galmarini, Claudio R.; Haj, Fawaz G.; Miatello, Roberto M.; Oteiza, Patricia I.

    2015-01-01

    Scope This study evaluated the capacity of dietary catechin (C), quercetin (Q) and the combination of both (CQ), to attenuate adipose inflammation triggered by high fructose (HFr) consumption in rats and by tumor necrosis factor alpha (TNFα) in 3T3-L1 adipocytes. Methods and results In rats, HFr consumption for 6 wk caused dyslipidemia, insulin resistance, reduced plasma adiponectin, adiposity, and adipose tissue inflammation. Dietary supplementation with 20 mg/kg/d of C, Q and CQ improved all these parameters. In 3T3-L1 adipocytes, C and Q attenuated TNFα-induced elevated protein carbonyls, increased pro-inflammatory cytokine expression (MCP-1, resistin), and decreased adiponectin. The protective effects of C and Q on adipose inflammation are in part associated with their capacity to: i) decrease the activation of the mitogen activated kinases (MAPKs) JNK and p38; and ii) prevent the downregulation of PPARγ. In summary, C and Q, and to a larger extent the combination of both, attenuated adipose pro-inflammatory signaling cascades and regulated the balance of molecules that improve (adiponectin) or impair (TNFα, MCP-1, resistin) insulin sensitivity. Conclusion Together, these findings suggest that dietary Q and C may have potential benefits in mitigating MetS associated adipose inflammation, oxidative stress, and insulin resistance. PMID:25620282

  8. Phenyllactic Acid from Lactobacillus plantarum PromotesAdipogenic Activity in 3T3-L1 Adipocyte via Up-Regulationof PPAR-γ2

    Directory of Open Access Journals (Sweden)

    Soundharrajan Ilavenil

    2015-08-01

    Full Text Available Synthetic drugs are commonly used to cure various human ailments at present. However, the uses of synthetic drugs are strictly regulated because of their adverse effects. Thus, naturally occurring molecules may be more suitable for curing disease without unfavorable effects. Therefore, we investigated phenyllactic acid (PLA from Lactobacillus plantarum with respect to its effects on adipogenic genes and their protein expression in 3T3-L1 pre-adipocytes by qPCR and western blot techniques. PLA enhanced differentiation and lipid accumulation in 3T3-L1 cells at the concentrations of 25, 50, and 100 μM. Maximum differentiation and lipid accumulation were observed at a concentration of 100 μM of PLA, as compared with control adipocytes (p < 0.05. The mRNA and protein expression of PPAR-γ2, C/EBP‑α, adiponectin, fatty acid synthase (FAS, and SREBP-1 were increased by PLA treatment as compared with control adipocytes (p < 0.05. PLA stimulates PPAR-γ mRNA expression in a concentration dependent manner, but this expression was lesser than agonist (2.83 ± 0.014 fold of PPAR-γ2. Moreover, PLA supplementation enhances glucose uptake in 3T3-L1 pre-adipocytes (11.81 ± 0.17 mM compared to control adipocytes, but this glucose uptake was lesser than that induced by troglitazone (13.75 ± 0.95 mM and insulin treatment (15.49 ± 0.20 mM. Hence, we conclude that PLA treatment enhances adipocyte differentiation and glucose uptake via activation of PPAR-γ2, and PLA may thus be the potential candidate for preventing Type 2 Diabetes Mellitus (T2DM.

  9. Activation of AMPK by berberine promotes adiponectin multimerization in 3T3-L1 adipocytes.

    Science.gov (United States)

    Li, Yun; Wang, Pengcheng; Zhuang, Yuan; Lin, Huan; Li, Yehua; Liu, Ling; Meng, Qinghang; Cui, Ting; Liu, Jing; Li, Zhen

    2011-06-23

    Adiponectin is assembled into trimer (LMW), hexamer (MMW) and high-molecular-weight (HMW) multimer in adipocytes. The HMW adiponectin is more metabolically active and closely associated with peripheral insulin sensitivity. In this study, we reported that berberine, an isoquinoline alkaloid with insulin-sensitizing effect, inhibits the expression of adiponectin, but promotes the assembly of HMW adiponectin and increases the ratio of HMW to total adiponectin. Berberine activates AMPK. Knockdown of AMPKα1 abolishes the effect of berberine. Activation of AMPK by AICAR also increases the level of HMW adiponectin. Our study suggested that activation of AMPK by berberine promotes adiponectin multimerization.

  10. Kirenol inhibits adipogenesis through activation of the Wnt/β-catenin signaling pathway in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Mi-Bo [Department of Biomaterials Science and Engineering, Yonsei University, Seoul 120-749 (Korea, Republic of); Song, Youngwoo; Kim, Changhee [Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of); Hwang, Jae-Kwan, E-mail: jkhwang@yonsei.ac.kr [Department of Biomaterials Science and Engineering, Yonsei University, Seoul 120-749 (Korea, Republic of); Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)

    2014-03-07

    Highlights: • Kirenol inhibits the adipogenic transcription factors and lipogenic enzymes. • Kirenol stimulates the Wnt/β-catenin signaling pathway components. • Kirenol inhibits adipogenesis through activation of the Wnt/β-catenin signaling pathway. - Abstract: Kirenol, a natural diterpenoid compound, has been reported to possess anti-oxidant, anti-inflammatory, anti-allergic, and anti-arthritic activities; however, its anti-adipogenic effect remains to be studied. The present study evaluated the effect of kirenol on anti-adipogenesis through the activation of the Wnt/β-catenin signaling pathway. Kirenol prevented intracellular lipid accumulation by down-regulating key adipogenesis transcription factors [peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding proteins α (C/EBPα), and sterol regulatory element binding protein-1c (SREBP-1c)] and lipid biosynthesis-related enzymes [fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC)], as well as adipocytokines (adiponectin and leptin). Kirenol effectively activated the Wnt/β-catenin signaling pathway, in which kirenol up-regulated the expression of low density lipoprotein receptor related protein 6 (LRP6), disheveled 2 (DVL2), β-catenin, and cyclin D1 (CCND1), while it inactivated glycogen synthase kinase 3β (GSK3β) by increasing its phosphorylation. Kirenol down-regulated the expression levels of PPARγ and C/EBPα, which were up-regulated by siRNA knockdown of β-catenin. Overall, kirenol is capable of inhibiting the differentiation and lipogenesis of 3T3-L1 adipocytes through the activation of the Wnt/β-catenin signaling pathway, suggesting its potential as natural anti-obesity agent.

  11. Real-time monitoring of inflammation status in 3T3-L1 adipocytes possessing a secretory Gaussia luciferase gene under the control of nuclear factor-kappa B response element.

    Science.gov (United States)

    Nagasaki, Haruka; Yoshimura, Takeshi; Aoki, Naohito

    2012-04-13

    We have established 3T3-L1 cells possessing a secretory Gaussia luciferase (GLuc) gene under the control of nuclear factor-kappa B (NF-κB) response element. The 3T3-L1 cells named 3T3-L1-NF-κB-RE-GLuc could differentiate into adipocyte as comparably as parental 3T3-L1 cells. Inflammatory cytokines such as tumor necrosis factor (TNF)-α and interleukin (IL)-1β induced GLuc secretion of 3T3-L1-NF-κB-RE-GLuc adipocytes in a concentration- and time-dependent manner. GLuc secretion of 3T3-L1-NF-κB-RE-GLuc adipocytes was also induced when cultured with RAW264.7 macrophages and was dramatically enhanced by lipopolysaccharide (LPS)-activated macrophages. An NF-κB activation inhibitor BAY-11-7085 and an antioxidant N-acetyl cysteine significantly suppressed GLuc secretion induced by macrophages. Finally, we found that rosemary-derived carnosic acid strongly suppressed GLuc secretion induced by macrophages and on the contrary up-regulated adiponectin secretion. Collectively, by using 3T3-L1-NF-κB-RE-GLuc adipocytes, inflammation status can be monitored in real time and inflammation-attenuating compounds can be screened more conveniently.

  12. Inhibition of preadipocyte differentiation and lipid accumulation by Orengedokuto treatment of 3T3-L1 cultures.

    Science.gov (United States)

    Ikarashi, Nobutomo; Tajima, Masataka; Suzuki, Kunihiro; Toda, Takahiro; Ito, Kiyomi; Ochiai, Wataru; Sugiyama, Kiyoshi

    2012-01-01

    Obesity is a major cause of metabolic syndrome and is due to an increase in the number and hypertrophy of adipocytes. Accordingly, inhibition of the differentiation and proliferation of adipocytes may be used in the treatment and prevention of metabolic syndrome. This study investigated the effects of 50 commonly used Kampo medicines on the differentiation of 3T3-L1 preadipocytes to search for a drug with an antiobesity effect. Kampo medicines were screened, and the strongest differentiation-inhibitory effect was noted with Orengedokuto. To explore the active ingredients in Orengedokuto, the effects of four crude drug components of Orengedokuto were investigated. It was found that the differentiation-inhibitory effect of Orengedokuto was accounted for by Coptidis rhizome and Phellodendri cortex. Furthermore, berberine, a principal ingredient common to Coptidis rhizome and Phellodendri cortex, showed a differentiation-inhibitory effect. The effect of berberine involves an inhibition of the mRNA and protein expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα). Moreover, berberine inhibited lipid accumulation in adipocytes. These findings suggest that an antiobesity effect could be a new indication for Orengedokuto and that its active ingredient is berberine, with a mechanism involving the inhibition of PPARγ and C/EBPα expression.

  13. Loganin inhibits the inflammatory response in mouse 3T3L1 adipocytes and mouse model.

    Science.gov (United States)

    Li, Yang; Li, Zheng; Shi, Lei; Zhao, Chenxu; Shen, Bingyu; Tian, Ye; Feng, Haihua

    2016-07-01

    Atherosclerosis is a chronic inflammatory disease of the vascular walls. ApoCIII is an independent factor which promotes atherosclerotic processes. This study aimed to investigate whether Loganin administration inhibits the inflammatory response in vitro and in vivo. In the apoCIII-induced mouse adipocytes, the levels of cytokines, including TNF-α, MCP-1 and IL-6 were determined by enzyme-linked immunosorbent assay and their gene expressions were measured through RT-PCR. The phosphorylation of nuclear factor-κB (NF-κB) proteins was analyzed by Western blotting. Our results showed that Loganin markedly decreased TNF-α, MCP-1 and IL-6 concentrations as well as their gene expressions. Western blotting analysis indicated that Loganin suppressed the activation of NF-κB signaling. In the Tyloxapol-treated mouse model, Loganin reduced the contents of TC and TG in mouse serum. The results of Oil Red-O Staining showed that Loganin reduced the production of lipid droplets. So it is suggested that Loganin might be a potential therapeutic agent for preventing the inflammation stress in vitro and in vivo.

  14. Regulation of myosin light chain kinase during insulin-stimulated glucose uptake in 3T3-L1 adipocytes.

    Directory of Open Access Journals (Sweden)

    Shelly Woody

    Full Text Available Myosin II (MyoII is required for insulin-responsive glucose transporter 4 (GLUT4-mediated glucose uptake in 3T3-L1 adipocytes. Our previous studies have shown that insulin signaling stimulates phosphorylation of the regulatory light chain (RLC of MyoIIA via myosin light chain kinase (MLCK. The experiments described here delineate upstream regulators of MLCK during insulin-stimulated glucose uptake. Since 3T3-L1 adipocytes express two MyoII isoforms, we wanted to determine which isoform was required for insulin-stimulated glucose uptake. Using a siRNA approach, we demonstrate that a 60% decrease in MyoIIA protein expression resulted in a 40% inhibition of insulin-stimulated glucose uptake. We also show that insulin signaling stimulates the phosphorylation of MLCK. We further show that MLCK can be activated by calcium as well as signaling pathways. We demonstrate that adipocytes treated with the calcium chelating agent, 1,2-b (iso-aminophenoxy ethane-N,N,N',N'-tetra acetic acid, (BAPTA (in the presence of insulin impaired the insulin-induced phosphorylation of MLCK by 52% and the RLC of MyoIIA by 45% as well as impairing the recruitment of MyoIIA to the plasma membrane when compared to cells treated with insulin alone. We further show that the calcium ionophore, A23187 alone stimulated the phosphorylation of MLCK and the RLC associated with MyoIIA to the same extent as insulin. To identify signaling pathways that might regulate MLCK, we examined ERK and CaMKII. Inhibition of ERK2 impaired phosphorylation of MLCK and insulin-stimulated glucose uptake. In contrast, while inhibition of CaMKII did inhibit phosphorylation of the RLC associated with MyoIIA, inhibition of CAMKIIδ did not impair MLCK phosphorylation or translocation to the plasma membrane or glucose uptake. Collectively, our results are the first to delineate a role for calcium and ERK in the activation of MLCK and thus MyoIIA during insulin-stimulated glucose uptake in 3T3-L1 adipocytes.

  15. The interaction of /sup 125/I-insulin with cultured 3T3-L1 adipocytes: quantitative analysis by the hypothetical grain method

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.Y.; Carpentier, J.L.; Van Obberghen, E.; Blackett, N.M.; Grunfeld, C.; Gorden, P.; Orci, L.

    1983-07-01

    The murine 3T3-L1 fibroblast under appropriate incubation conditions differentiates into an adipocyte phenotype. This 3T3-L1 adipocyte exhibits many of the morphologic, biochemical, and insulin-responsive features of the normal rodent adipocyte. Using quantitative electron microscopic (EM) autoradiography we find that, when /sup 125/I-insulin is incubated with 3T3-L1 adipocytes, the ligand at early times of incubation localizes to the plasma membrane of the cell preferentially to microvilli and coated pits. When the incubation is continued at 37 degrees C, /sup 125/I-insulin is internalized by the cells and preferential binding to the villous surface is lost. With the internalization of the ligand, two intracellular structures become labeled, as determined by the method of hypothetical grain analysis. These include large clear, presumably endocytotic, vesicles and multivesicular bodies. Over the first hour of incubation the labeling of these structures increases in parallel, but in the second hour they diverge: the labeling of multivesicular bodies and other lysosomal forms continuing to increase and the labeling of large clear vesicles decreasing. At 3 hours limited but significant labeling occurs in small Golgi-related vesicles that have the typical distribution of GERL. The distinct morphologic features of this cell make it ideal for a quantitative morphologic analysis and allow for an unambiguous view of the sequence of events involved in receptor-mediated endocytosis of a polypeptide hormone. These events are likely to be representative of the processing of insulin by the mature rodent adipocyte.

  16. Germinated brown rice extract inhibits adipogenesis through the down-regulation of adipogenic genes in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ho, Jin-Nyoung; Son, Mi-Eun; Lim, Won-Chul; Lim, Seung-Taik; Cho, Hong-Yon

    2013-09-01

    The aim of this study was to examine the anti-adipogenic effect of germinated brown rice methanol extract (GBR) in 3T3-L1 adipocytes. The GBR inhibited adipocyte differentiation was measured by Oil Red O staining and glycerol-3-phosphate dehydrogenase (GPDH) activity in a dose-dependent manner without initiating any cytotoxicity. The mRNA levels of adipogenic transcription factors such as CCAAT/enhancer binding protein (C/EBPα), proliferator-activated receptorγ (PPARγ), and sterol regulatory element-binding protein-1c (SREBP-1c), and adipogenic genes, such as fatty acid synthase (FAS), adipocyte fatty acid-binding protein (aP2), and lipoprotein lipase (LPL), were significantly down-regulated by treatment with GBR when compared to that of untreated control cells. Moreover, tumor necrosis factor-α (TNF-α) and interlukin-6 (IL-6) mRNA expressions were attenuated by GBR in mature adipocytes. These data suggest that GBR exhibits an anti-adipogenic effect through the suppression of adipogenesis in 3T3-L1 adipocytes.

  17. Centipede grass exerts anti-adipogenic activity through inhibition of C/EBPβ, C/EBPα, and PPARγ expression and the AKT signaling pathway in 3T3-L1 adipocytes

    Directory of Open Access Journals (Sweden)

    Park Hyoung Joon

    2012-11-01

    Full Text Available Abstract Background Centipede grass (CG originates from China and South America and is reported to contain several C-glycosyl flavones and phenolic constituents, including maysin and luteolin derivatives. This study aimed to investigate, for the first time, the antiobesity activity of CG and its potential molecular mechanism in 3T3-L1 cells. Methods To study the effect of CG on adipogenesis, differentiating 3T3-L1 cells were treated every day with CG at various concentrations (0–100 μg/ml for six days. Oil-red O staining and triglyceride content assay were performed to determine the lipid accumulation in 3T3-L1 cells. The expression of mRNAs or proteins associated with adipogenesis was measured using RT-PCR and Western blotting analysis. We examined the effect of CG on level of phosphorylated Akt in 3T3-L1 cells treated with CG at various concentration s during adipocyte differentiation. Results Differentiation was investigated with an Oil-red O staining assay using CG-treated 3T3-L1 adipocytes. We found that CG suppressed lipid droplet formation and adipocyte differentiation in 3T3-L1 cells in a dose-dependent manner. Treatment of the 3T3-L1 adipocytes with CG resulted in an attenuation of the expression of adipogenesis-related factors and lipid metabolic genes. The expression of C/EBPα and PPARγ, the central transcriptional regulators of adipogenesis, was decreased by the treatment with CG. The expression of genes involved in lipid metabolism, aP2 were significantly inhibited following the CG treatment. Moreover, the CG treatment down-regulated the phosphorylation levels of Akt and GSK3β. Conclusions Taken collectively, these data indicated that CG exerts antiadipogenic activity by inhibiting the expression of C/EBPβ, C/EBPα, and PPARγ and the Akt signaling pathway in 3T3-L1 adipocytes.

  18. Over-expression of NYGGF4 inhibits glucose transport in 3T3-L1 adipocytes via attenuated phosphorylation of IRS-1 and Akt

    Institute of Scientific and Technical Information of China (English)

    Chun-mei ZHANG; Xiao-hui CHEN; Bin WANG; Feng LIU; Xia CHI; Mei-ling TONG; Yu-hui NI; Rong-hua CHEN; Xi-rong GUO

    2009-01-01

    Aim: NYGGF4 is a novel gene that is abundantly expressed in the adipose tissue of obese patients. The purpose of this study was to investigate the effects of NYGGF4 on basal and insulin-stimulated glucose uptake in mature 3T3-L1 adipocytes and to understand the underlying mechanisms. Methods: 3T3-L1 preadipocytes transfected with either an empty expression vector (pcDNA3.1Myc/His B) or an NYGGF4 expression vector were differentiated into mature adipocytes. Glucose uptake was determined by measuring 2-deoxy-D-[3H]glucose uptake into the adipocytes. Immunoblotting was performed to detect the translocation of insulin-sensitive glu-cose transporter 4 (GLUT4). Immunoblotting also was used to measure the phosphorylation and total protein contents of insulin signaling proteins such as the insulin receptor (IR), insulin receptor substrate (IRS)-I, Akt, ERK1/2, p38, and JNK. Results: NYGGF4 over-expression in 3T3-L1 adipocytes reduced insulin-stimulated glucose uptake and impaired insulin-stimulated GLUT4 translocation. It also diminished insulin-stimulated tyrosine phosphorylation of IRS-1 and serine phos-phorylation of Akt without affecting the phosphorylation of IR, ERK1/2, p38, and JNK. Conclusion: NYGGF4 regulates the functions of IRS-1 and Akt, decreases GLUT4 translocation and reduces glucose uptake in response to insulin. These observations highlight the potential role of NYGGF4 in glucose homeostasis and possibly in the pathogenesis of obesity.

  19. Pulicaria jaubertii E. Gamal-Eldin reduces triacylglyceride content and modifies cellular antioxidant pathways in 3T3-L1 adipocytes.

    Science.gov (United States)

    Al-Naqeb, Ghanya; Rousová, Jana; Kubátová, Alena; Picklo, Matthew J

    2016-06-25

    Levels of obesity in Middle Eastern countries are increasing. Phytochemicals have anti-obesogenic properties as evidenced by prevention of adipocyte differentiation and blocking triacylglyceride (TG) accumulation. In Yemen, Pulicaria jaubertii E. Gamal-Eldin (PJ) is a food additive and a traditional medicine. We tested the hypothesis that phytochemicals present in PJ inhibit adipocytic responses during differentiation of 3T3-L1 preadipocytes to adipocytes. Methanolic extracts of PJ did not block expression of fatty acid binding protein 4 (FABP4) a marker of differentiation but did inhibit TG accumulation. Treatment of 3T3-L1 preadipocytes increased NADPH:quinone oxidoreductase 1 (NQO1), a suppressor of TG accumulation. Further fractionation of the methanolic PJ extract with hexane and dichloromethane (DCM) demonstrated that bioactivity towards TG reduction and elevated expression of NQO1 and other antioxidant genes (glutamate cysteine ligase catalytic unit, glutathione disulfide reductase, glutathione peroxidase (GPx) 4 resided in the DCM fraction. Activity towards depleting GSH and elevating the expression of catalase and GPx3 were found in the DCM and hexane fractions. Analysis by gas chromatography and liquid chromatography coupled with mass spectrometry demonstrated the presence of catechin-like moieties in the DCM and methanolic fractions and suggest that these components were partially responsible for the bioactivity of these fractions. In summary, our data indicate that fractions derived PJ exhibit anti-adipogenic properties in part through the presence of catechin-like compounds.

  20. [8-hydroxy-dihydroberberine ameliorated insulin resistance induced by high FFA and high glucose in 3T3-L1 adipocytes].

    Science.gov (United States)

    Xu, Li-jun; Lu, Fu-er; Yi, Ping; Wang, Zeng-si; Wei, Shi-chao; Chen, Guang; Dong, Hui; Zou, Xin

    2009-11-01

    The purpose of the study is to investigate the effect of 8-hydroxy-dihydroberberine on insulin resistance induced by high free fatty acid (FFA) and high glucose in 3T3-L1 adipocytes and its possible molecular mechanism. Palmic acid or glucose in combination with insulin was used to induce insulin resistance in 3T3-L1 adipocytes. 8-Hydroxy-dihydroberberine and berberine were added to the cultured medium separately, which were considered as treated group and positive control group. The rate of glucose uptake was determined by 2-deoxy-[3H]-D-glucose method. The amount of glucose consumption in the medium was measured by glucose oxidase method. Cell growth and proliferation of 3T3-L1 adipocytes were detected with Cell Counting Kit-8 (CCK-8) assay. After incubated with palmic acid for 24 hours or glucose with insulin for 18 hours, the rate of glucose transport in 3T3-L1 adipocytes was inhibited by 67% and 58%, respectively. The amount of glucose consumption in 3T3-L1 adipose cells was decreased by 41% after cells were incubated with palmic acid for 24 h. However, the above changes were reversed by pretreatment with 8-hydroxy-dihydroberberine for 24 and 48 h. Significant difference existed between groups. Insulin resistance in 3T3-L1 adipocytes, which is induced by high FFA and high glucose, could be ameliorated by 8-hydroxy-dihydroberberine.

  1. Curcumin improves hypoxia induced dysfunctions in 3T3-L1 adipocytes by protecting mitochondria and down regulating inflammation.

    Science.gov (United States)

    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.

  2. Soshiho-Tang Aqueous Extract Exerts Antiobesity Effects in High Fat Diet-Fed Mice and Inhibits Adipogenesis in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Sae-Rom Yoo

    2016-01-01

    Full Text Available Soshiho-tang (SST; sho-saiko-to in Japanese; xiaochaihu-tang in Chinese has generally been used to improve liver fibrosis- and cirrhosis-related symptoms in traditional Korean medicine. Although many studies have investigated the pharmacological properties of SST, its antiobesity effect has not been elucidated. Thus, our present study was carried out to evaluate the antiobesity effect of SST using a high fat diet- (HFD induced obese mouse model and 3T3-L1 adipose cells. C57BL/6J mice were randomly divided into four groups (n=6/group, normal diet (ND, HFD-fed group, and HFD- and SST-fed groups (S200: 200 mg/kg of SST; S600: 600 mg/kg of SST and given HFD with or without SST extract for 8 weeks. 3T3-L1 preadipocytes were differentiated into adipocytes for 8 days with or without SST. In the HFD-fed obese mice, body weight and fat accumulation in adipose tissue were significantly reduced by SST administration. Compared with control-differentiated adipocytes, SST significantly inhibited lipid accumulation by decreasing the triglyceride (TG content and leptin concentration in 3T3-L1 adipocytes. SST also decreased the expression of adipogenesis-related genes including lipoprotein lipase (LPL, fatty acid binding protein 4 (FABP4, CCAAT/enhancer-binding protein-alpha (C/EBP-α, and peroxisome proliferator-activated receptor-gamma (PPAR-γ. Our findings suggest that SST has potential as a nontoxic antiobesity medication.

  3. PPARγ partial agonist GQ-16 strongly represses a subset of genes in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Milton, Flora Aparecida [Faculdade de Ciências da Saúde, Laboratório de Farmacologia Molecular, Universidade de Brasília (Brazil); Genomic Medicine, Houston Methodist Research Institute, Houston, TX (United States); Cvoro, Aleksandra [Genomic Medicine, Houston Methodist Research Institute, Houston, TX (United States); Amato, Angelica A. [Faculdade de Ciências da Saúde, Laboratório de Farmacologia Molecular, Universidade de Brasília (Brazil); Sieglaff, Douglas H.; Filgueira, Carly S.; Arumanayagam, Anithachristy Sigamani [Genomic Medicine, Houston Methodist Research Institute, Houston, TX (United States); Caro Alves de Lima, Maria do; Rocha Pitta, Ivan [Laboratório de Planejamento e Síntese de Fármacos – LPSF, Universidade Federal de Pernambuco (Brazil); Assis Rocha Neves, Francisco de [Faculdade de Ciências da Saúde, Laboratório de Farmacologia Molecular, Universidade de Brasília (Brazil); Webb, Paul, E-mail: pwebb@HoustonMethodist.org [Genomic Medicine, Houston Methodist Research Institute, Houston, TX (United States)

    2015-08-28

    Thiazolidinediones (TZDs) are peroxisome proliferator-activated receptor gamma (PPARγ) agonists that improve insulin resistance but trigger side effects such as weight gain, edema, congestive heart failure and bone loss. GQ-16 is a PPARγ partial agonist that improves glucose tolerance and insulin sensitivity in mouse models of obesity and diabetes without inducing weight gain or edema. It is not clear whether GQ-16 acts as a partial agonist at all PPARγ target genes, or whether it displays gene-selective actions. To determine how GQ-16 influences PPARγ activity on a gene by gene basis, we compared effects of rosiglitazone (Rosi) and GQ-16 in mature 3T3-L1 adipocytes using microarray and qRT-PCR. Rosi changed expression of 1156 genes in 3T3-L1, but GQ-16 only changed 89 genes. GQ-16 generally showed weak effects upon Rosi induced genes, consistent with partial agonist actions, but a subset of modestly Rosi induced and strongly repressed genes displayed disproportionately strong GQ-16 responses. PPARγ partial agonists MLR24 and SR1664 also exhibit disproportionately strong effects on transcriptional repression. We conclude that GQ-16 displays a continuum of weak partial agonist effects but efficiently represses some negatively regulated PPARγ responsive genes. Strong repressive effects could contribute to physiologic actions of GQ-16. - Highlights: • GQ-16 is an insulin sensitizing PPARγ ligand with reduced harmful side effects. • GQ-16 displays a continuum of weak partial agonist activities at PPARγ-induced genes. • GQ-16 exerts strong repressive effects at a subset of genes. • These inhibitor actions should be evaluated in models of adipose tissue inflammation.

  4. The effect of cultureware surfaces on functional and structural components of differentiated 3T3-L1 preadipocytes.

    Science.gov (United States)

    Pavlikova, Nela; Weiszenstein, Martin; Pala, Jan; Halada, Petr; Seda, Ondrej; Elkalaf, Moustafa; Trnka, Jan; Kovar, Jan; Polak, Jan

    2015-12-01

    Experiments using cultured primary cells or cell lines are a routine in vitro approach used across multiple biological disciplines, However, the structural and functional influences of various cultureware materials on cultured cells is not clearly understood. Surface treatments of cultureware have proven to have profound effects on cell viability and proliferation. In this study, we investigated the impact of polystyrene and fluorocarbon cultureware dishes on the proteomic profile of differentiated 3T3-L1 preadipocytes. After expansion and differentiation of cells on appropriate cultureware dishes, cell lysates were separated using two-dimensional gel electrophoresis and proteins were visualized with Coomassie blue staining. Spots with the highest differential expression between the two culture conditions were subsequently analyzed using matrix-assisted laser desorption/ionization mass spectrometry and the identified proteins were subjected to pathway analysis. We observed that 43% of all spots were differentially expressed depending on the cultureware. Pathway analysis revealed that glucose metabolism, mitochondrial structure and cell differentiation, represented by 14-3-3 protein-mediated signaling and the mitochondrial inner membrane organizing system (MINOS), were significantly affected by cultureware material. These results indicate that cultureware material can have a profound effect on key adipocyte functional pathways. These effects modifications of the cells should be reflected in the design of in vitro experiments and interpretation of their results.

  5. Coptis chinensis alkaloids exert anti-adipogenic activity on 3T3-L1 adipocytes by downregulating C/EBP-α and PPAR-γ.

    Science.gov (United States)

    Choi, Jae Sue; Kim, Ji-Hye; Ali, Md Yousof; Min, Byung-Sun; Kim, Gun-Do; Jung, Hyun Ah

    2014-10-01

    Obesity is a complex, multifactorial, and chronic disease that increases the risk for type 2 diabetes, coronary heart disease and hypertension, and has become a major worldwide health problem. Developing novel anti-obesity drugs from natural products is a promising solution to the global health problem of obesity. While screening anti-obesity potentials of natural products, the methanol extract of the rhizome of Coptis chinensis (Coptidis Rhizoma) was found to significantly inhibit adipocyte differentiation and lipid contents in 3T3-L1 cells, as assessed by Oil-Red O staining. Five known alkaloids, berberine, epiberberine, coptisine, palmatine, and magnoflorine, were isolated from the n-BuOH fraction of the methanol extract of Coptidis Rhizoma. We determined the chemical structure of these alkaloids through comparisons of published nuclear magnetic resonance (NMR) spectral data. Furthermore, we screened these alkaloids for their ability to inhibit adipogenesis over a range of concentrations (12.5-50 μM). All five Coptidis Rhizoma alkaloids significantly inhibited lipid accumulation in 3T3-L1 cells without affecting cell viability in a concentration dependent manner. In addition, the five alkaloids significantly reduced the expression levels of several adipocyte marker genes including proliferator activated receptor-γ (PPAR-γ) and CCAAT/enhancer-binding protein-α (C/EBP-α). In the present study, we found that the isolated alkaloids inhibited adipogenesis in a dose-dependent manner in 3T3-L1 cells; this inhibition was attributed to their abilities to downregulate the protein levels of the adipocyte marker proteins PPAR-γ and C/EBP-α. Thus, these results suggest that Coptidis Rhizoma extract and its isolated alkaloids may be of therapeutic interest with respect to the treatment of obesity.

  6. Theobromine inhibits differentiation of 3T3-L1 cells during the early stage of adipogenesis via AMPK and MAPK signaling pathways.

    Science.gov (United States)

    Jang, Yeon Jeong; Koo, Hyun Jung; Sohn, Eun-Hwa; Kang, Se Chan; Rhee, Dong-Kwon; Pyo, Suhkneung

    2015-07-01

    Obesity is characterized by hypertrophy and/or by the differentiation or adipogenesis of pre-existing adipocytes. In this study, we investigated the inhibitory effects of theobromine, a type of alkaloid in cocoa, on adipocyte differentiation of 3T3-L1 preadipocytes and its mechanisms of action. Theobromine inhibited the accumulation of lipid droplets, the expression of PPARγ and C/EBPα, and the mRNA expression of aP2 and leptin. The inhibition of adipogenic differentiation by theobromine occurred primarily in the early stages of differentiation. In addition, theobromine arrested the cell cycle at the G0/G1 phase and regulated the expressions of CDK2, p27, and p21. Theobromine treatment increased AMPK phosphorylation and knockdown of AMPKα1/α2 prevented the ability of theobromine to inhibit PPARγ expression in the differentiating 3T3-L1 cells. Theobromine reduced the phosphorylation of ERK and JNK. Moreover, the secretion and the mRNA level of TNF-α and IL-6 were inhibited by theobromine treatment. These data suggest that theobromine inhibits adipocyte differentiation during the early stages of adipogenesis by regulating the expression of PPARγ and C/EBPα through the AMPK and ERK/JNK signaling pathways in 3T3-L1 preadipocytes.

  7. Protein kinase A suppresses the differentiation of 3T3-L1 preadipocytes

    Institute of Scientific and Technical Information of China (English)

    Fuqiang Li; Dongmei Wang; Yiran Zhou; Bo Zhou; Yanan Yang; Hehua Chen; Jianguo Song

    2008-01-01

    cAMP and protein kinase A (PKA) are widely known as signaling molecules that are important for the induction of adipogenesis. Here we show that a strong increase in the amount of cAMP inhibits the adipogenesis of 3T3-L1 fibroblast cells. Stimulation of PKA activity suppresses adipogenesis and, in contrast, inhibition of PKA activity markedly accelerates the adipogenic process. As adipogenesis progresses, there is a significant increase in the expression level of PKA regulatory subunits and a corresponding decrease in PKA activity. Moreover, treatment of 3T3-L1 cells with epidermal growth factor (EGF) stimulates PKA activity and blocks adipogenesis. Inhibition of PKA activity abolishes this suppressive effect of EGF on adipogenesis. Moreover, activation of PKA induces serine/threonine phosphorylation, reduces tyrosine phosphorylation of insulin receptor substrate 1 (IRS-1) and the association between PKA and IRS-1. Taken together, our study demonstrates that PKA has a pivotal role in the suppression of adipogenesis. cAMP at high concentrations can suppress adipogenesis through PKA activation. These findings could be important and useful for understanding the mechanisms of adipogenesis and the relevant physiological events.

  8. Disruption of Lipid Raft Function Increases Expression and Secretion of Monocyte Chemoattractant Protein-1 in 3T3-L1 Adipocytes.

    Science.gov (United States)

    Lu, Juu-Chin; Chiang, Yu-Ting; Lin, Yu-Chun; Chang, Yu-Tzu; Lu, Chia-Yun; Chen, Tzu-Yu; Yeh, Chia-Shan

    2016-01-01

    The adipocyte is unique in its capacity to store lipids. In addition to triglycerides, the adipocyte stores a significant amount of cholesterol. Moreover, obese adipocytes are characterized by a redistribution of cholesterol with depleted cholesterol in the plasma membrane, suggesting that cholesterol perturbation may play a role in adipocyte dysfunction. We used methyl-β-cyclodextrin (MβCD), a molecule with high affinity for cholesterol, to rapidly deplete cholesterol level in differentiated 3T3-L1 adipocytes. We tested whether this perturbation altered adipocyte secretion of monocyte chemoattractant protein-1 (MCP-1), a chemokine that is elevated in obesity and is linked to obesity-associated chronic diseases. Depletion of cholesterol by MβCD increased MCP-1 secretion as well as the mRNA and protein levels, suggesting perturbation at biosynthesis and secretion. Pharmacological inhibition revealed that NF-κB, but not MEK, p38 and JNK, was involved in MβCD-stimulated MCP-1 biosynthesis and secretion in adipocytes. Finally, another cholesterol-binding drug, filipin, also induced MCP-1 secretion without altering membrane cholesterol level. Interestingly, both MβCD and filipin disturbed the integrity of lipid rafts, the membrane microdomains enriched in cholesterol. Thus, the depletion of membrane cholesterol in obese adipocytes may result in dysfunction of lipid rafts, leading to the elevation of proinflammatory signaling and MCP-1 secretion in adipocytes.

  9. Berberine attenuates cAMP-induced lipolysis via reducing the inhibition of phosphodiesterase in 3T3-L1 adipocytes.

    Science.gov (United States)

    Zhou, Libin; Wang, Xiao; Yang, Ying; Wu, Ling; Li, Fengying; Zhang, Rong; Yuan, Guoyue; Wang, Ning; Chen, Mingdao; Ning, Guang

    2011-04-01

    Berberine, a hypoglycemic agent, has been shown to decrease plasma free fatty acids (FFAs) level in insulin-resistant rats. In the present study, we explored the mechanism responsible for the antilipolytic effect of berberine in 3T3-L1 adipocytes. It was shown that berberine attenuated lipolysis induced by catecholamines, cAMP-raising agents, and a hydrolyzable cAMP analog, but not by tumor necrosis factor α and a nonhydrolyzable cAMP analog. Unlike insulin, the inhibitory effect of berberine on lipolysis in response to isoproterenol was not abrogated by wortmannin, an inhibitor of phosphatidylinositol 3-kinase, but additive to that of PD98059, an extracellular signal-regulated kinase kinase inhibitor. Prior exposure of adipocytes to berberine decreased the intracellular cAMP production induced by isoproterenol, forskolin, and 3-isobutyl-1-methylxanthine (IBMX), along with hormone-sensitive lipase (HSL) Ser-563 and Ser-660 dephosphorylation, but had no effect on perilipin phosphorylation. Berberine stimulated HSL Ser-565 as well as adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. However, compound C, an AMPK inhibitor, did not reverse the regulatory effect of berberine on HSL Ser-563, Ser-660, and Ser-565 phosphorylation, nor the antilipolytic effect of berberine. Knockdown of AMPK using RNA interference also failed to restore berberine-suppressed lipolysis. cAMP-raising agents increased AMPK activity, which was not additive to that of berberine. Stimulation of adipocytes with berberine increased phosphodiesterase (PDE) 3B and PDE4 activity measured by hydrolysis of (3)[H]cAMP. These results suggest that berberine exerts an antilipolytic effect mainly by reducing the inhibition of PDE, leading to a decrease in cAMP and HSL phosphorylation independent of AMPK pathway.

  10. A role for Rab14 in the endocytic trafficking of GLUT4 in 3T3-L1 adipocytes.

    Science.gov (United States)

    Reed, Sam E; Hodgson, Lorna R; Song, Shuang; May, Margaret T; Kelly, Eoin E; McCaffrey, Mary W; Mastick, Cynthia C; Verkade, Paul; Tavaré, Jeremy M

    2013-05-01

    Insulin enhances the uptake of glucose into adipocytes and muscle cells by promoting the redistribution of the glucose transporter isoform 4 (GLUT4) from intracellular compartments to the cell surface. Rab GTPases regulate the trafficking itinerary of GLUT4 and several have been found on immunopurified GLUT4 vesicles. Specifically, Rab14 has previously been implicated in GLUT4 trafficking in muscle although its role, if any, in adipocytes is poorly understood. Analysis of 3T3-L1 adipocytes using confocal microscopy demonstrated that endogenous GLUT4 and endogenous Rab14 exhibited a partial colocalisation. However, when wild-type Rab14 or a constitutively-active Rab14Q70L mutant were overexpressed in these cells, the colocalisation with both GLUT4 and IRAP became extensive. Interestingly, this colocalisation was restricted to enlarged 'ring-like' vesicular structures (mean diameter 1.3 µm), which were observed in the presence of overexpressed wild-type Rab14 and Rab14Q70L, but not an inactive Rab14S25N mutant. These enlarged vesicles contained markers of early endosomes and were rapidly filled by GLUT4 and transferrin undergoing endocytosis from the plasma membrane. The Rab14Q70L mutant reduced basal and insulin-stimulated cell surface GLUT4 levels, probably by retaining GLUT4 in an insulin-insensitive early endosomal compartment. Furthermore, shRNA-mediated depletion of Rab14 inhibited the transit of GLUT4 through early endosomal compartments towards vesicles and tubules in the perinuclear region. Given the previously reported role of Rab14 in trafficking between endosomes and the Golgi complex, we propose that the primary role of Rab14 in GLUT4 trafficking is to control the transit of internalised GLUT4 from early endosomes into the Golgi complex, rather than direct GLUT4 translocation to the plasma membrane.

  11. Characterization of GLUT4-containing vesicles in 3T3-L1 adipocytes by total internal reflection fluorescence microscopy

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Insulin-responsive GLUT4(glucose transporter 4) translocation plays a major role in regulating glucose uptake in adipose tissue and muscle.Whether or not there is a specialized secretory GSV(GLUT4 storage vesicle) pool,and more importantly how GSVs are translocated to the PM(plasma membrane) under insulin stimulation is still under debate.In the present study,we systematically analyzed the dynamics of a large number of single GLUT4-containing vesicles in 3T3-L1 adipocytes by TIRFM(total internal reflection fluorescence microscopy).We found that GLUT4-containing vesicles can be classified into three groups according to their mobility,namely vertical,stable,and lateral GLUT4-containing vesicles.Among these groups,vertical GLUT4-containing vesicles exclude transferrin receptors and move towards the PM specifically in response to insulin stimulation,while stable and lateral GLUT4-containing vesicles contain transferrin receptors and show no insulin responsiveness.These data demonstrate that vertical GLUT4-containing vesicles correspond to specialized secretory GSVs,which approach the PM directly and bypass the constitutive recycling pathway.

  12. Characterization of GLUT4-containing vesicles in 3T3-L1 adipocytes by total internal reflection fluorescence microscopy

    Institute of Scientific and Technical Information of China (English)

    WANG Yan; ZHANG JinZhong; CHEN Yu; JIANG Li; JI Wei; XU Tao

    2009-01-01

    Insulin-responsive GLUT4 (glucose transporter 4) translocation plays a major role in regulating glucose uptake in adipose tissue and muscle. Whether or not there is a specialized secretory GSV (GLUT4 storage vesicle) pool, and more importantly how GSVs are translocated to the PM (plasma membrane) under insulin stimulation is still under debate. In the present study, we systematically analyzed the dynamics of a large number of single GLUT4-containing vesicles in 3T3-L1 adipocytes by TIRFM (total internal reflection fluorescence microscopy). We found that GLUT4-containing vesicles can be classi fled into three groups according to their mobility, namely vertical, stable, and lateral GLUT4-containing vesicles. Among these groups, vertical GLUT4-containing vesicles exclude transferrin receptors and move towards the PM specifically in response to insulin stimulation, while stable and lateral GLUT4 containing vesicles contain transferrin receptors and show no insulin responsiveness. These data demonstrate that vertical GLUT4-containing vesicles correspond to specialized secretory GSVs, which approach the PM directly and bypass the constitutive recycling pathway.

  13. A novel IRS-1-associated protein, DGKζ regulates GLUT4 translocation in 3T3-L1 adipocytes.

    Science.gov (United States)

    Liu, TingYu; Yu, BuChin; Kakino, Mamoru; Fujimoto, Hitoshi; Ando, Yasutoshi; Hakuno, Fumihiko; Takahashi, Shin-Ichiro

    2016-10-14

    Insulin receptor substrates (IRSs) are major targets of insulin receptor tyrosine kinases. Here we identified diacylglycerol kinase zeta (DGKζ) as an IRS-1-associated protein, and examined roles of DGKζ in glucose transporter 4 (GLUT4) translocation to the plasma membrane. When DGKζ was knocked-down in 3T3-L1 adipocytes, insulin-induced GLUT4 translocation was inhibited without affecting other mediators of insulin-dependent signaling. Similarly, knockdown of phosphatidylinositol 4-phosphate 5-kinase 1α (PIP5K1α), which had been reported to interact with DGKζ, also inhibited insulin-induced GLUT4 translocation. Moreover, DGKζ interacted with IRS-1 without insulin stimulation, but insulin stimulation decreased this interaction. Over-expression of sDGKζ (short-form DGKζ), which competed out DGKζ from IRS-1, enhanced GLUT4 translocation without insulin stimulation. Taking these results together with the data showing that cellular PIP5K activity was correlated with GLUT4 translocation ability, we concluded that IRS-1-associated DGKζ prevents GLUT4 translocation in the absence of insulin and that the DGKζ dissociated from IRS-1 by insulin stimulation enhances GLUT4 translocation through PIP5K1α activity.

  14. Real-time monitoring of inflammation status in 3T3-L1 adipocytes possessing a secretory Gaussia luciferase gene under the control of nuclear factor-kappa B response element

    Energy Technology Data Exchange (ETDEWEB)

    Nagasaki, Haruka; Yoshimura, Takeshi [Department of Life Sciences, Graduate School of Bioresources, Mie University, Tsu 514-8507 (Japan); Aoki, Naohito, E-mail: n-aoki@bio.mie-u.ac.jp [Department of Life Sciences, Graduate School of Bioresources, Mie University, Tsu 514-8507 (Japan)

    2012-04-13

    Highlights: Black-Right-Pointing-Pointer Inflammation status in adipocytes can be monitored by the new assay system. Black-Right-Pointing-Pointer Only an aliquot of conditioned medium is required without cell lysis. Black-Right-Pointing-Pointer Inflammation-attenuating compounds can be screened more conveniently. -- Abstract: We have established 3T3-L1 cells possessing a secretory Gaussia luciferase (GLuc) gene under the control of nuclear factor-kappa B (NF-{kappa}B) response element. The 3T3-L1 cells named 3T3-L1-NF-{kappa}B-RE-GLuc could differentiate into adipocyte as comparably as parental 3T3-L1 cells. Inflammatory cytokines such as tumor necrosis factor (TNF)-{alpha} and interleukin (IL)-1{beta} induced GLuc secretion of 3T3-L1-NF-{kappa}B-RE-GLuc adipocytes in a concentration- and time-dependent manner. GLuc secretion of 3T3-L1-NF-{kappa}B-RE-GLuc adipocytes was also induced when cultured with RAW264.7 macrophages and was dramatically enhanced by lipopolysaccharide (LPS)-activated macrophages. An NF-{kappa}B activation inhibitor BAY-11-7085 and an antioxidant N-acetyl cysteine significantly suppressed GLuc secretion induced by macrophages. Finally, we found that rosemary-derived carnosic acid strongly suppressed GLuc secretion induced by macrophages and on the contrary up-regulated adiponectin secretion. Collectively, by using 3T3-L1-NF-{kappa}B-RE-GLuc adipocytes, inflammation status can be monitored in real time and inflammation-attenuating compounds can be screened more conveniently.

  15. Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on adipogenic differentiation and insulin-induced glucose uptake in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Hsu, Hsin-Fen [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Zhunan, Miaoli County 35053, Taiwan (China); Tsou, Tsui-Chun, E-mail: tctsou@nhri.org.tw [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Zhunan, Miaoli County 35053, Taiwan (China); Chao, How-Ran [Department of Environmental Science and Engineering, National Pingtung University of Science and Technology, Neipu 912, Pingtung, Taiwan (China); Kuo, Ya-Ting; Tsai, Feng-Yuan; Yeh, Szu-Ching [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Zhunan, Miaoli County 35053, Taiwan (China)

    2010-10-15

    Dioxin exposure has been positively associated with human type II diabetes. Because lipophilic dioxins accumulate mainly in adipose tissue, this study aimed to determine if dioxins induce metabolic dysfunction in fat cells. Using 3T3-L1 cells as an in vitro model, we analyzed the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a model dioxin, on adipogenic differentiation, glucose uptake, and lipolysis. TCDD inhibited adipogenic differentiation, as determined by using oil droplet formation and adipogenic marker gene expression, including PPAR{gamma} (peroxisome proliferator-activated receptor {gamma}), C/EBP{alpha} (CCAAT/enhancer-binding protein {alpha}), and Glut4 (glucose transporter type 4). Effects of TCDD on glucose uptake were evaluated using fully differentiated 3T3-L1 adipocytes, revealing that TCDD significantly attenuated insulin-induced glucose uptake dose dependently. Inhibition of aryl hydrocarbon receptor (AhR) by {alpha}-naphthoflavone ({alpha}-NF), an AhR inhibitor, did not prevent the inhibitory effect of TCDD on glucose uptake, suggesting that TCDD attenuates insulin-induced glucose uptake in an AhR-independent manner. Effects of TCDD on lipolysis were determined using glycerol release assay. We found that TCDD had no marked effect on isoproterenol-induced glycerol release in fully differentiated 3T3-L1 adipocytes. These results provide in vitro evidence of TCDD's effects on fat cell metabolism, suggesting dioxin exposure in development of insulin resistance and type II diabetes.

  16. Prolonged inorganic arsenite exposure suppresses insulin-stimulated AKT S473 phosphorylation and glucose uptake in 3T3-L1 adipocytes: Involvement of the adaptive antioxidant response

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Peng [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); School of Public Health, China Medical University, Shenyang 110001 (China); Hou, Yongyong; Zhang, Qiang; Woods, Courtney G.; Yarborough, Kathy; Liu, Huiyu [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Sun, Guifan [School of Public Health, China Medical University, Shenyang 110001 (China); Andersen, Melvin E. [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Pi, Jingbo, E-mail: jpi@thehamner.org [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States)

    2011-04-08

    Highlights: {yields} In 3T3-L1 adipocytes iAs{sup 3+} decreases insulin-stimulated glucose uptake. {yields} iAs{sup 3+} attenuates insulin-induced phosphorylation of AKT S473. {yields} iAs{sup 3+} activates the cellular adaptive oxidative stress response. {yields} iAs{sup 3+} impairs insulin-stimulated ROS signaling. {yields} iAs{sup 3+} decreases expression of adipogenic genes and GLUT4. -- Abstract: There is growing evidence that chronic exposure of humans to inorganic arsenic, a potent environmental oxidative stressor, is associated with the incidence of type 2 diabetes (T2D). One critical feature of T2D is insulin resistance in peripheral tissues, especially in mature adipocytes, the hallmark of which is decreased insulin-stimulated glucose uptake (ISGU). Despite the deleterious effects of reactive oxygen species (ROS), they have been recognized as a second messenger serving an intracellular signaling role for insulin action. Nuclear factor erythroid 2-related factor 2 (NRF2) is a central transcription factor regulating cellular adaptive response to oxidative stress. This study proposes that in response to arsenic exposure, the NRF2-mediated adaptive induction of endogenous antioxidant enzymes blunts insulin-stimulated ROS signaling and thus impairs ISGU. Exposure of differentiated 3T3-L1 cells to low-level (up to 2 {mu}M) inorganic arsenite (iAs{sup 3+}) led to decreased ISGU in a dose- and time-dependent manner. Concomitant to the impairment of ISGU, iAs{sup 3+} exposure significantly attenuated insulin-stimulated intracellular ROS accumulation and AKT S473 phosphorylation, which could be attributed to the activation of NRF2 and induction of a battery of endogenous antioxidant enzymes. In addition, prolonged iAs{sup 3+} exposure of 3T3-L1 adipocytes resulted in significant induction of inflammatory response genes and decreased expression of adipogenic genes and glucose transporter type 4 (GLUT4), suggesting chronic inflammation and reduction in GLUT4

  17. Butyrate and other short-chain fatty acids increase the rate of lipolysis in 3T3-L1 adipocytes

    Directory of Open Access Journals (Sweden)

    John M. Rumberger

    2014-10-01

    Full Text Available We determined the effect of butyrate and other short-chain fatty acids (SCFA on rates of lipolysis in 3T3-L1 adipocytes. Prolonged treatment with butyrate (5 mM increased the rate of lipolysis approximately 2–3-fold. Aminobutyric acid and acetate had little or no effect on lipolysis, however propionate stimulated lipolysis, suggesting that butyrate and propionate act through their shared activity as histone deacetylase (HDAC inhibitors. Consistent with this, the HDAC inhibitor trichostatin A (1 µM also stimulated lipolysis to a similar extent as did butyrate. Western blot data suggested that neither mitogen-activated protein kinase (MAPK activation nor perilipin down-regulation are necessary for SCFA-induced lipolysis. Stimulation of lipolysis with butyrate and trichostatin A was glucose-dependent. Changes in AMP-activated protein kinase (AMPK phosphorylation mediated by glucose were independent of changes in rates of lipolysis. The glycolytic inhibitor iodoacetate prevented both butyrate- and tumor necrosis factor-alpha-(TNF-α mediated increases in rates of lipolysis indicating glucose metabolism is required. However, unlike TNF-α– , butyrate-stimulated lipolysis was not associated with increased lactate release or inhibited by activation of pyruvate dehydrogenase (PDH with dichloroacetate. These data demonstrate an important relationship between lipolytic activity and reported HDAC inhibitory activity of butyrate, other short-chain fatty acids and trichostatin A. Given that HDAC inhibitors are presently being evaluated for the treatment of diabetes and other disorders, more work will be essential to determine if these effects on lipolysis are due to inhibition of HDAC.

  18. The roots of Atractylodes japonica Koidzumi promote adipogenic differentiation via activation of the insulin signaling pathway in 3T3-L1 cells

    Directory of Open Access Journals (Sweden)

    Han Yunkyung

    2012-09-01

    Full Text Available Abstract Background Type 2 diabetes (T2D is a complex metabolic disorder characterized by insulin resistance and hyperglycemia. Peroxisome proliferator-activated receptor gamma (PPARγ is a key transcription factor and plays an important role in the regulation of genes involved in adipogenic differentiation, glucose metabolism and insulin signal transduction. Methods In this study, the effects of the root extract of Atractylodes japonica Koidzumi (Atractylodis Rhizoma Alba, ARA on the differentiation of 3T3-L1 preadipocytes and the possible mechanism of glucose transport were investigated. 3T3-L1 cells were cultured with insulin and ARA extract. Results In 3T3-L1 cells, ARA extract significantly enhanced adipogenic differentiation and upregulated the expression of PPARγ genes and protein in a dose-dependent manner. ARA also promoted glucose transport by increasing the glucose transporter 4 (GLUT-4, phosphatidylinositol 3-kinase (PI3K and insulin receptor substrates-1 (IRS-1 levels. Conclusion Our results suggest that ARA extract may be an attractive therapeutic agent for managing T2D via promoting the differentiation of adipocytes with the upregulation of PPARγ levels and the activation of the insulin signaling pathway.

  19. Knockdown of NYGGF4 (PID1) rescues insulin resistance and mitochondrial dysfunction induced by FCCP in 3T3-L1 adipocytes.

    Science.gov (United States)

    Shi, Chun-Mei; Wang, Yu-Mei; Zhang, Chun-Mei; Qiu, Jie; Shen, Ya-Hui; Zhu, Jin-Gai; Chen, Lin; Xu, Guang-Feng; Zhao, Ya-Ping; Ji, Chen-Bo; Guo, Xi-Rong

    2012-11-01

    NYGGF4 is a recently identified gene that is involved in obesity-associated insulin resistance. Previous data from this laboratory have demonstrated that NYGGF4 overexpression might contribute to the development of insulin resistance (IR) and to mitochondrial dysfunction. Additionally, NYGGF4 knockdown enhanced insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes. We designed this study to determine whether silencing of NYGGF4 in 3T3-L1 adipocytes could rescue the effect of insulin sensitivity and mitochondrial function induced by the cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP), a mitochondrion uncoupler, to ascertain further the mechanism of NYGGF4 involvement in obesity-associated insulin resistance. We found that 3T3-L1 adipocytes, incubated with 5μM FCCP for 12h, had decreased levels of insulin-stimulated glucose uptake and had impaired insulin-stimulated GLUT4 translocation. Silencing also diminished insulin-stimulated tyrosinephosphorylation of IRS-1 and serine phosphorylation of Akt. This phenomenon contrasts with the effect of NYGGF4 knockdown on insulin sensitivity and describes the regulatory function of NYGGF4 in adipocytes insulin sensitivity. We next analyzed the mitochondrial function in NYGGF4-silenced adipocytes incubated with FCCP. NYGGF4 knockdown partly rescued the dissipation of mitochondrial mass, mitochondrial DNA, intracellular ATP synthesis, and intracellular reactive oxygen species (ROS) production occurred following the addition of FCCP, as well as inhibition of mitochondrial transmembrane potential (ΔΨm) in 3T3-L1 adipocytes incubated with FCCP. Collectively, our results suggested that addition of silencing NYGGF4 partly rescued the effect of insulin resistance and mitochondrial dysfunction in NYGGF4 silenced 3T3-L1 adipocytes incubated with FCCP, which might explain the involvement of NYGGF4-induced IR and the development of NYGGF4 in mitochondrial function.

  20. The Differentiation-and Proliferation-Inhibitory Effects of Sporamin from Sweet Potato in 3T3-L1 Preadipocytes

    Institute of Scientific and Technical Information of China (English)

    XIONG Zhi-dong; LI Peng-gao; MU Tai-hua

    2009-01-01

    The aim of this study was to investigate the effect of different concentrations of sporamin on the differentiation and proliferation of 3T3-L1 preadipocytes,providing the theoretical basis for the development of food to treat obesity and diabetes.The isolation and purification of sporamin from sweet potato species 55-2 were performed by ammonium sulphate precipitation in combination with ion-exchange and gel filtration chromatography.With berberine as a positive control,different concentrations of sporamin (0.000,0.125,0.025,0.250,0.500,and 1.000 mg mL-1) were used to treat 3T3-L1 preadipocytes.Intracellular fat accumulation and the degree of adipogenesis were quantified using Oil Red O staining and colorimetry.Preadipocytes differentiation was measured by 3-(4,5-dimethylthiazolyl-2-yl)-2,5-diphenyltetrazolium bromide (MTT)spectrophotometric assay.Two sporamin proteins,which were separated into sporamin A (31 kD) and sporamin B (22 kD),could be purified by ion-exchange and gel filtration chromatography.After being treated by different concentrations of sporamin,the differentiation of 3T3-L1 preadipocytes was significantly inhibited,compared with the positive control.When the sporamin solution concentration was 0.500 mg mL-1,the accumulation of lipid droplets within the cells was significantly decreased and the optical density (OD) value of the solution from destained Oil Red O reached to 0.35,which was the lowest value (P < 0.05).The proliferation of 3T3-L1 preadipocytes was significantly inhibited by treating at higher sporamin concentrations.In addition,the inhibitory effect was more obvious with the prolonged treatment time (P< 0.05).The differentiation and proliferation of 3T3-L1 preadipocytes could be inhibited significantly by the addition of higher concentration sporamin.It was,therefore,suggested that the sporamin was potentially effective for weight loss.

  1. Reduction of adipogenesis and lipid accumulation by Taraxacum officinale (Dandelion) extracts in 3T3L1 adipocytes: an in vitro study.

    Science.gov (United States)

    González-Castejón, Marta; García-Carrasco, Belén; Fernández-Dacosta, Raquel; Dávalos, Alberto; Rodriguez-Casado, Arantxa

    2014-05-01

    In this in vitro study, we have investigated the ability of Taraxacum officinale (dandelion) to inhibit adipocyte differentiation and lipogenesis in 3T3-L1 preadipocytes. HPLC analysis of the three plant extracts used in this study-leaf and root extracts and a commercial root powder-identified caffeic and chlorogenic acids as the main phenolic constituents. Oil Red O staining and triglyceride levels analysis showed decreased lipid and triglyceride accumulation, respectively. Cytotoxicity was assessed with the MTT assay showing non-toxic effect among the concentrations tested. DNA microarray analysis showed that the extracts regulated the expression of a number of genes and long non-coding RNAs that play a major role in the control of adipogenesis. Taken together, our results indicate that the dandelion extracts used in this study may play a significant role during adipogenesis and lipid metabolism, and thus, supporting their therapeutic interest as potential candidates for the treatment of obesity.

  2. Differential effects of eicosapentaenoic acid and docosahexaenoic acid in promoting the differentiation of 3T3-L1 preadipocytes.

    Science.gov (United States)

    Murali, Ganesan; Desouza, Cyrus V; Clevenger, Michelle E; Ramalingam, Ramesh; Saraswathi, Viswanathan

    2014-01-01

    The objective of this study was to determine the effects of enrichment with n-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the differentiation of 3T3-L1 preadipocytes. Enrichment with DHA but not EPA significantly increased the differentiation markers compared to control differentiated cells. DHA compared to EPA treatment led to a greater increase in adiponectin secretion and, conditioned media collected from DHA treated cells inhibited monocyte migration. Moreover, DHA treatment resulted in inhibition of pro-inflammatory signaling pathways. DHA treated cells predominantly accumulated DHA in phospholipids whereas EPA treatment led to accumulation of both EPA and its elongation product docosapentaenoic acid (DPA), an n-3 fatty acid. Of note, adding DPA to DHA inhibited DHA-induced differentiation. The differential effects of EPA and DHA on preadipocyte differentiation may be due, in part, to differences in their intracellular modification which could impact the type of n-3 fatty acids incorporated into the cells.

  3. Bisphenol-A impairs insulin action and up-regulates inflammatory pathways in human subcutaneous adipocytes and 3T3-L1 cells.

    Science.gov (United States)

    Valentino, Rossella; D'Esposito, Vittoria; Passaretti, Federica; Liotti, Antonietta; Cabaro, Serena; Longo, Michele; Perruolo, Giuseppe; Oriente, Francesco; Beguinot, Francesco; Formisano, Pietro

    2013-01-01

    Current evidence indicates that chemical pollutants may interfere with the homeostatic control of nutrient metabolism, thereby contributing to the increased prevalence of metabolic disorders. Bisphenol-A (BPA) is a lipophilic compound contained in plastic which is considered a candidate for impairing energy and glucose metabolism. We have investigated the impact of low doses of BPA on adipocyte metabolic functions. Human adipocytes derived from subcutaneous adipose tissue and differentiated 3T3-L1 cells were incubated with BPA, in order to evaluate the effect on glucose utilization, insulin sensitivity and cytokine secretion. Treatment with 1 nM BPA significantly inhibited insulin-stimulated glucose utilization, without grossly interfering with adipocyte differentiation. Accordingly, mRNA levels of the adipogenic markers PPARγ and GLUT4 were unchanged upon BPA exposure. BPA treatment also impaired insulin-activated receptor phosphorylation and signaling. Moreover, adipocyte incubation with BPA was accompanied by increased release of IL-6 and IFN-γ, as assessed by multiplex ELISA assays, and by activation of JNK, STAT3 and NFkB pathways. Treatment of the cells with the JNK inhibitor SP600125 almost fully reverted BPA effect on insulin signaling and glucose utilization. In conclusion, low doses of BPA interfere with inflammatory/insulin signaling pathways, leading to impairment of adipose cell function.

  4. Bisphenol-A impairs insulin action and up-regulates inflammatory pathways in human subcutaneous adipocytes and 3T3-L1 cells.

    Directory of Open Access Journals (Sweden)

    Rossella Valentino

    Full Text Available Current evidence indicates that chemical pollutants may interfere with the homeostatic control of nutrient metabolism, thereby contributing to the increased prevalence of metabolic disorders. Bisphenol-A (BPA is a lipophilic compound contained in plastic which is considered a candidate for impairing energy and glucose metabolism. We have investigated the impact of low doses of BPA on adipocyte metabolic functions. Human adipocytes derived from subcutaneous adipose tissue and differentiated 3T3-L1 cells were incubated with BPA, in order to evaluate the effect on glucose utilization, insulin sensitivity and cytokine secretion. Treatment with 1 nM BPA significantly inhibited insulin-stimulated glucose utilization, without grossly interfering with adipocyte differentiation. Accordingly, mRNA levels of the adipogenic markers PPARγ and GLUT4 were unchanged upon BPA exposure. BPA treatment also impaired insulin-activated receptor phosphorylation and signaling. Moreover, adipocyte incubation with BPA was accompanied by increased release of IL-6 and IFN-γ, as assessed by multiplex ELISA assays, and by activation of JNK, STAT3 and NFkB pathways. Treatment of the cells with the JNK inhibitor SP600125 almost fully reverted BPA effect on insulin signaling and glucose utilization. In conclusion, low doses of BPA interfere with inflammatory/insulin signaling pathways, leading to impairment of adipose cell function.

  5. A mutation in signal peptide of rat resistin gene inhibits differentiation of 3T3-L1 preadipocytes

    Institute of Scientific and Technical Information of China (English)

    Xi-rong GUO; Hai-xia GONG; Yan-qin GAO; Li FEI; Yu-hui NI; Rong-hua CHEN

    2004-01-01

    AIM: To detect the resistin expression of white adipose tissue in diet-induced obese (DIO) versus diet-resistant (DR) rats, and to investigate the relationship of mutated resistin and 3T3-L1 preadipocytes differentiation. METHODS:RT-PCR and Western Blot were used to detect gene/protein expression. 3T3-L1 cells were cultured, transfected,and induced to differentiation using 0.5 mmol/L 3-isobutyl-1-methyxanthine (MIX), 1 mg/L insulin, and 1μmol/Ldexamethasone. Oil red O staining was applied to detect the degree of preadipocytes differentiation. RESULTS:Expression of resistin mRNA was upregulated in DIO rats and downregulated in DR rats. However, the expression levels varied greatly within the groups. Sequencing of the resistin genes from DIO and DR rats revealed a Leu9Val (C25G) missense mutation within the signal peptide in one DR rat. The mutant resistin inhibited preadipocyte differentiation. Local experiments and Western blotting with tagged resistin fusion proteins identified both mutant and wild type proteins in the cytoplasm and secreted into the culture medium. Computer predictions using the Proscan and Subloc programs revealed four putative phosphorylation sites and a possible leucine zipper motif within the rat resistin protein. CONCLUSION: Resistin-increased differentiation may be inhibited by the mutationcontaining precursor protein, or by the mutant non-secretory resistin isoform.

  6. 小檗碱对3T3-L1脂肪细胞脂联素表达的影响%Effects of berberine on adiponectin mRNA expression in 3T3-L1 adipocyte

    Institute of Scientific and Technical Information of China (English)

    谷卫; 曾文衡; 胡海英

    2005-01-01

    目的:探讨小檗碱和胰岛素对3T3-L1脂肪细胞脂联素表达的影响.方法:检测分别经小檗碱及胰岛素处理后3T3-L1脂肪细胞脂联素表达水平改变,以β-actin为内对照,半定量法RT-PCR测定脂联素mRNA表达.结果:经小檗碱(10 μmol·L-1)干预后3T3-L1脂肪细胞脂联素表达显著增加(P<0.05),加入胰岛素后脂联素的表达受到抑制,高浓度胰岛素有显著抑制作用(P<0.05). 结论:体外实验中,小檗碱使3T3-L1脂肪细胞脂联素的表达增加,而胰岛素可抑制小檗碱增加脂联素表达的作用.

  7. A novel regulatory function of sweet taste-sensing receptor in adipogenic differentiation of 3T3-L1 cells.

    Directory of Open Access Journals (Sweden)

    Yosuke Masubuchi

    Full Text Available BACKGROUND: Sweet taste receptor is expressed not only in taste buds but also in nongustatory organs such as enteroendocrine cells and pancreatic beta-cells, and may play more extensive physiological roles in energy metabolism. Here we examined the expression and function of the sweet taste receptor in 3T3-L1 cells. METHODOLOGY/PRINCIPAL FINDINGS: In undifferentiated preadipocytes, both T1R2 and T1R3 were expressed very weakly, whereas the expression of T1R3 but not T1R2 was markedly up-regulated upon induction of differentiation (by 83.0 and 3.8-fold, respectively at Day 6. The α subunits of Gs (Gαs and G14 (Gα14 but not gustducin were expressed throughout the differentiation process. The addition of sucralose or saccharin during the first 48 hours of differentiation considerably reduced the expression of peroxisome proliferator activated receptor γ (PPARγ and CCAAT/enhancer-binding protein α (C/EBPα at Day 2, the expression of aP2 at Day 4 and triglyceride accumulation at Day 6. These anti-adipogenic effects were attenuated by short hairpin RNA-mediated gene-silencing of T1R3. In addition, overexpression of the dominant-negative mutant of Gαs but not YM-254890, an inhibitor of Gα14, impeded the effects of sweeteners, suggesting a possible coupling of Gs with the putative sweet taste-sensing receptor. In agreement, sucralose and saccharin increased the cyclic AMP concentration in differentiating 3T3-L1 cells and also in HEK293 cells heterologously expressing T1R3. Furthermore, the anti-adipogenic effects of sweeteners were mimicked by Gs activation with cholera toxin but not by adenylate cyclase activation with forskolin, whereas small interfering RNA-mediated knockdown of Gαs had the opposite effects. CONCLUSIONS: 3T3-L1 cells express a functional sweet taste-sensing receptor presumably as a T1R3 homomer, which mediates the anti-adipogenic signal by a Gs-dependent but cAMP-independent mechanism.

  8. NYGGF4基因表达沉默对3T3-L1脂肪细胞线粒体代谢的影响%Effect of NYGGF4 Gene Silencing on Mitochondrial Metabolism in 3T3 - L1 Adipocytes

    Institute of Scientific and Technical Information of China (English)

    史春梅; 季晨博; 张春梅; 朱金改; 陈玲; 郭锡熔

    2012-01-01

    目的 探讨NYGGF4(又称PID1)基因沉默对脂肪细胞线粒体代谢的影响.方法 以3T3-L1前体脂肪细胞为研究对象,体外培养稳定转染NYGGF4基因沉默载体(pGPU6/GFP/Neo-shRNA,NYGGF4-RNAi)的3T3-L1前体脂肪细胞,以转染空载体(pG-PU6/GFP/Neo-shRNA,NCshRNA)的3T3-L1前体脂肪细胞作为对照,经1-甲基-3异丁基黄嘌呤加地塞米松、胰岛素方案诱导分化为成熟脂肪细胞,采用实时定量RCR技术检测成熟脂肪细胞中线粒体代谢酶指标:己糖激酶、乙酰辅酶A羧化酶柠檬酸合成酶、肉毒碱棕榈酰转移酶1、细胞色素C基因表达水平.结果 NYGGF4表达沉默与NYGGF4基因过表达相比,可以部分逆转3T3-L1脂肪细胞中已糖激酶、乙酰辅酶A羧化酶、柠檬酸合成酶、肉毒碱棕榈酰转移酶1、细胞色素C基因表达水平,差异均有统计学意义.结论 NYGGF4基因过表达,下调3T3-L1脂肪细胞中定位于线粒体的代谢关键酶,提示NYGGF4基因的动态平衡对维持3T3-L1脂肪细胞的线粒体代谢具有重要作用.%Objective To explore the effect of knockdown NYGGF4 (PID1) gene on mitochondria] metabolism in 3T3 - LI adipocytes. Methods With 3T3 - L1 preadipocytes as a subject, which transfected with a NYGGF4 expression vector, an empty expression vector and NYGGF4 gene silencing vector and then the preadipocrtes were induced to differentiate into adipocvtes using MIX, DXM and insulin. The expression levels of metabolic enzymes in mitochondria------hexokinase (HKI) ,acetyl -CoA(ACC), citrate synthase(CS) .carnitine palmitoyl - transferasel(CPT1) ,cytochrome C(Cye — C) were detected by real -time PCR. Results Knockdown NYGGF4 could reverse the level of metabolic enzymes in mitochondria-----HKI, ACC, CS, CPT1 ,Cyc - c in NYGGF4 - overexperssion adipocytes. Conclusions In adipocytes with NYGGF4 gene overexpression, mitochondria metabolic enzymes mRNA is reversed in NYGGF4 gene silencing. It can implicate that mito

  9. Free Fatty Acids Activate Renin-Angiotensin System in 3T3-L1 Adipocytes through Nuclear Factor-kappa B Pathway

    Directory of Open Access Journals (Sweden)

    Jia Sun

    2016-01-01

    Full Text Available The activity of a local renin-angiotensin system (RAS in the adipose tissue is closely associated with obesity-related diseases. However, the mechanism of RAS activation in adipose tissue is still unknown. In the current study, we found that palmitic acid (PA, one kind of free fatty acid, induced the activity of RAS in 3T3-L1 adipocytes. In the presence of fetuin A (Fet A, PA upregulated the expression of angiotensinogen (AGT and angiotensin type 1 receptor (AT1R and stimulated the secretion of angiotensin II (ANG II in 3T3-L1 adipocytes. Moreover, the activation of RAS in 3T3-L1 adipocytes was blocked when we blocked Toll-like receptor 4 (TLR4 signaling pathway using TAK242 or NF-κB signaling pathway using BAY117082. Together, our results have identified critical molecular mechanisms linking PA/TLR4/NF-κB signaling pathway to the activity of the local renin-angiotensin system in adipose tissue.

  10. Enhanced inhibition of adipogenesis and induction of apoptosis in 3T3-L1 adipocytes with combinations of resveratrol and quercetin.

    Science.gov (United States)

    Yang, Jeong-Yeh; Della-Fera, Mary Anne; Rayalam, Srujana; Ambati, Suresh; Hartzell, Diane L; Park, Hea Jin; Baile, Clifton A

    2008-05-07

    Certain flavonoids have been shown to have specific effects on biochemical and metabolic functions of adipocytes. In this study, we investigated the effects of combinations of resveratrol and quercetin on adipogenesis and apoptosis in 3T3-L1 cells. In maturing preadipocytes resveratrol and quercetin at 25 microM individually suppressed intracellular lipid accumulation by 9.4+/-3.9% (p<0.01) and 15.9+/-2.5%, respectively, (p<0.001). The combination of resveratrol and quercetin at the same dose, however, decreased lipid accumulation by 68.6+/-0.7% (p<0.001). In addition, combinations of resveratrol and quercetin at 25 microM significantly decreased the expression of peroxisome proliferators-activated receptor gamma (PPAR gamma) and CCAAT/enhancer-binding protein (C/EBP)alpha, both of which act as key transcription factors. In mature adipocytes resveratrol and quercetin at 100 microM individually decreased viability by 18.1+/-0.6% (p<0.001) and 15.8+/-1% (p<0.001) and increased apoptosis (100 microM) by 120.5+/-8.3% (p<0.001) and 85.3+/-10% (p<0.001) at 48 h, respectively. Combinations of resveratrol and quercetin further decreased viability (73.5+/-0.9%, p<0.001) and increased apoptosis (310.3+/-9.6%, p<0.001) more than single compounds alone. The combination of resveratrol and quercetin at 100 muM increased release of cytochrome c from mitochondria to cytosol and decreased ERK 1/2 phosphorylation. Taken together, our data indicate that combinations of resveratrol and quercetin can exert potential anti-obesity effects by inhibiting differentiation of preadipocytes and inducing apoptosis of mature adipocytes.

  11. Omega-3 polyunsaturated fatty acid has an anti-oxidant effect via the Nrf-2/HO-1 pathway in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kusunoki, Chisato, E-mail: yosizaki@belle.shiga-med.ac.jp [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan); Yang, Liu; Yoshizaki, Takeshi; Nakagawa, Fumiyuki; Ishikado, Atsushi; Kondo, Motoyuki; Morino, Katsutaro; Sekine, Osamu; Ugi, Satoshi [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan); Nishio, Yoshihiko [Division of Diabetes, Metabolism and Endocrinology, Department of Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Kashiwagi, Atsunori; Maegawa, Hiroshi [Department of Medicine, Shiga University of Medical Science, Seta Tsukinowa-Cho, Otsu, Shiga 520-2192 (Japan)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Omega-3 PUFA has a direct anti-oxidant effect in adipocytes. Black-Right-Pointing-Pointer EPA and DHA induce HO-1 expression in 3T3-L1 adipocytes. Black-Right-Pointing-Pointer Omega-3 PUFA and its end-product, 4-HHE, activates the Nrf-2/HO-1 pathway. Black-Right-Pointing-Pointer Omega-3 PUFA protects against oxidative stress-induced cytotoxicity. -- Abstract: Oxidative stress is produced in adipose tissue of obese subjects and has been associated with obesity-related disorders. Recent studies have shown that omega-3 polyunsaturated fatty acid ({omega}3-PUFA) has beneficial effects in preventing atherosclerotic diseases and insulin resistance in adipose tissue. However, the role of {omega}3-PUFA on adipocytes has not been elucidated. In this study, 3T3-L1 adipocytes were treated with {omega}3-PUFA and its metabolites, eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), or 4-hydroxy hexenal (4-HHE). {omega}3-PUFA and its metabolites dose-dependently increased mRNA and protein levels of the anti-oxidative enzyme, heme oxygenase-1 (HO-1); whereas no changes in the well-known anti-oxidant molecules, superoxide dismutase, catalase, and glutathione peroxidase, were observed. Knockdown of nuclear factor erythroid 2-related factor 2 (Nrf-2) significantly reduced EPA, DHA or 4-HHE-induced HO-1 mRNA and protein expression. Also, pretreatment with {omega}3-PUFA prevented H{sub 2}O{sub 2}-induced cytotoxicity in a HO-1 dependent manner. In conclusion, treatment with EPA and DHA induced HO-1 through the activation of Nrf-2 and prevented oxidative stress in 3T3-L1 adipocytes. This anti-oxidant defense may be of high therapeutic value for clinical conditions associated with systemic oxidative stress.

  12. Inhibitory effect of cajanonic acid A on lipogenesis and lipolysis in 3 T3-L1 adipocytes%树豆酮酸A抑制3 T3-L1细胞脂肪合成与分解的作用研究

    Institute of Scientific and Technical Information of China (English)

    秦佑; 杨瑞仪; 陈梅果; 沈小玲; 胡英杰

    2016-01-01

    Aim To investigate the effects of cajanonic acid A (CAA) on lipid metabolism in murine 3T3-L1 adipocytes. Methods 3T3-L1 cells induced to differ-entiated into mature adipocytes were treated with CAA in different dosages for 48 h, then total lipids as well as triglyceride, free fatty acid and glycerol were meas-ured. The expression levels of genes related to lipid metabolism were quantitatively analyzed by real-time fluorescent quantitative polymearase chain reaction ( RTFQ-PCR) . Results Total lipids and triglyceride in 3T3-L1 adipocytes were markedly reduced by CAA. The release of free fatty acid and glycerol was lower than that of control. This coincided with decreased mRNA levels of the key enzymes involved in de novo lipogenesis ( acetyl CoA carboxylase and fatty acid syn-thase) , fatty acid uptake ( lipoprotein lipase) , and li-polysis ( hormone sensitive lipase and adipose triglycer-ide lipase ) . While the expression of fatty acid oxida-tive genes including acyl CoA oxidase and carnitine palmitoyl transferase1 was increased after CAA treat-ment. Conclusion CAA may inhibit lipogenesis and lipolysis,reduce circulating free fatty acid and improve the lipid metabolism in adipocytes by regulating gene expressions.%目的:探讨树豆酮酸A( cajanonic acid A,CAA)对小鼠3 T3-L1脂肪细胞脂质代谢的影响及其机制。方法诱导分化成熟的3T3-L1脂肪细胞加入不同浓度的 CAA作用48 h后,进行总脂肪、甘油三酯、游离脂肪酸和甘油含量测定。实时荧光定量PCR法检测脂质代谢相关基因的表达。结果CAA明显降低3T3-L1脂肪细胞总脂肪和甘油三酯含量,抑制游离脂肪酸和甘油的释放,下调乙酰辅酶A羧化酶( acetyl CoA carboxylase,ACC)、脂肪酸合成酶( fatty acid synthase, FAS)、脂蛋白脂酶( lipoprotein lipase,LPL)、激素敏感性脂肪酶( hormone sensitive lipase,HSL)和脂肪甘油三酯脂酶( adi-pose triglyceride lipase,ATGL)基因的表达,

  13. OM2, a Novel Oligomannuronate-Chromium(III) Complex, Promotes Mitochondrial Biogenesis and Lipid Metabolism in 3T3-L1 Adipocytes via the AMPK-PGC1α Pathway

    OpenAIRE

    Jiejie Hao; Cui Hao; Lijuan Zhang; Xin Liu; Xiaolin Zhou; Yunlou Dun; Haihua Li; Guangsheng Li; Xiaoliang Zhao; Yuanyuan An; Jiankang Liu; Guangli Yu

    2015-01-01

    Background In our previous studies, we prepared novel oligomannuronate-chromium(III) complexes (OM2, OM4) from marine alginate, and found that these compounds sensitize insulin action better than oligomannuronate(OM), chromium, and metformin in C2C12 skeletal muscle cells. In the present study, we studied their effects on mitochondrial biogenesis, lipid metabolism, and the underlying molecular mechanisms in differentiated 3T3-L1 adipocytes. Methodology/Principal Findings We firstly used the p...

  14. Ethanol extracts of chickpeas alter the total lipid content and expression levels of genes related to fatty acid metabolism in mouse 3T3-L1 adipocytes

    Science.gov (United States)

    Shinohara, Shigeo; Gu, Yuanjun; Yang, Ying; Furuta, Yasuo; Tanaka, Masahiko; Yue, Xiaohua; Wang, Weiqing; Kitano, Masaru; Kimura, Hiroshi

    2016-01-01

    Desi-type chickpeas, which have long been used as a natural treatment for diabetes, have been reported to lower visceral adiposity, dyslipidemia and insulin resistance induced by a chronic high-fat diet in rats. In this study, in order to examine the effects of chickpeas of this type in an in vitro system, we used the 3T3-L1 mouse cell line, a subclone of Swiss 3T3 cells, which can differentiate into cells with an adipocyte-like phenotype, and we used ethanol extracts of chickpeas (ECP) instead of chickpeas. Treatment of the 3T3-L1 cells with ECP led to a decrease in the lipid content in the cells. The desaturation index, defined as monounsaturated fatty acids (MUFAs)/saturated fatty acids (SFAs), was also decreased by ECP due to an increase in the cellular content of SFAs and a decrease in the content of MUFAs. The decrease in this index may reflect a decreased reaction from SFA to MUFA, which is essential for fat storage. To confirm this hypothesis, we conducted a western blot analysis, which revealed a reduction in the amount of stearoyl-CoA desaturase 1 (SCD1), a key enzyme catalyzing the reaction from SFA to MUFA. We observed simultaneous inactivations of enzymes participating in lipogenesis, i.e., liver kinase B1 (LKB1), acetyl-CoA carboxylase (ACC) and AMPK, by phosphorylation, which may lead to the suppression of reactions from acetyl-CoA to SFA via malonyl-CoA in lipogenesis. We also investigated whether lipolysis is affected by ECP. The amount of carnitine palmitoyltransferase 1 (CPT1), an enzyme important for the oxidation of fatty acids, was increased by ECP treatment. ECP also led to an increase in uncoupling protein 2 (UCP2), reported as a key protein for the oxidation of fatty acids. All of these results obtained regarding lipogenesis and fatty acid metabolism in our in vitro system are consistent with the results previously shown in rats. We also examined the effects on SCD1 and lipid contents of ethanol extracts of Kabuli-type chickpeas, which are

  15. Trans10,cis15 18:2 Isolated from Beef Fat Does Not Have the Same Anti-Adipogenic Properties as Trans10,cis12-18:2 in 3T3-L1 Adipocytes.

    Science.gov (United States)

    Vahmani, Payam; Meadus, William J; Rolland, David C; Duff, Pascale; Dugan, Michael E R

    2016-11-01

    During ruminal biohydrogenation of α-linolenic acid, a non-conjugated non-methylene interrupted dienoic acid is formed containing a t10 double bond, namely t10,c15-18:2. The present study was designed to examine whether t10,c15-18:2 would exert similar anti-adipogenic effects compared to t10,c12-18:2 in 3T3-L1 adipocytes. Differentiated 3T3-L1 adipocytes were treated with 35 or 70 µM of LNA, t10,c12-18:2, t10,c15-18:2, or bovine serum albumin (BSA) vehicle control for 120 h. Cellular triacylglycerol and protein were quantified using commercial colorimetric kits. Cells were analyzed for fatty acid composition and gene expression using gas chromatography and quantitative PCR, respectively. Trans10,cis12-18:2 decreased (P 18:1). Trans10,cis12 also decreased (P 18:2 did not affect the gene expression and cellular content of the TAG, SFA, c-MUFA, or SCD1 indices in adipocytes. Our findings suggest that t10,c15-18:2, despite having structural similarity to t10,c12-18:2 (presence of a trans-10 double bond), does not exert anti-adipogenic effects in 3T3-L1 adipocytes.

  16. The inhibition of inflammatory molecule expression on 3T3-L1 adipocytes by berberine is not mediated by leptin signaling

    OpenAIRE

    Choi, Bong-Hyuk; Kim, Yu-Hee; Ahn, In-Sook; Ha, Jung-Heun; Byun, Jae-Min; Do, Myoung-Sool

    2009-01-01

    In our previous study, we have shown that berberine has both anti-adipogenic and anti-inflammatory effects on 3T3-L1 adipocytes, and the anti-adipogenic effect is due to the down-regulation of adipogenic enzymes and transcription factors. Here we focused more on anti-inflammatory effect of berberine using real time RT-PCR and found it changes expressions of adipokines. We hypothesized that anti-adipogenicity of berberine mediates anti-inflammtory effect and explored leptin as a candidate medi...

  17. Trans-10,cis-12 conjugated linoleic acid (CLA) interferes with lipid droplet accumulation during 3T3-L1 preadipocyte differentiation.

    Science.gov (United States)

    Yeganeh, Azadeh; Taylor, Carla G; Tworek, Leslee; Poole, Jenna; Zahradka, Peter

    2016-07-01

    In this study, we hypothesize that the biologically active isomers of conjugated linoleic acid (CLA), cis-9,trans-11 (c9,t11) and trans-10,cis-12 (t10,c12) CLA, have different effects on early and late stages 3T3-L1 preadipocyte differentiation. Both c9-t11 and t10-c12CLA stimulated early stage pre-adipocyte differentiation (day 2), while t10-c12CLA inhibited late differentiation (day 8) as determined by lipid droplet numbers and both perilipin-1 levels and phosphorylation state. At day 8, the adipokines adiponectin, chemerin and adipsin were all reduced in t10-c12CLA treated cells versus control cells. Immunofluorescence microscopy showed perilipin-1 was present solely on lipid droplets on day 8 in t10-c12 treated 3T3-L1 cells, whereas preilipin-1 was also located in the perinuclear region in control and c9-t11 treated cells. The t10-c12CLA isomer also decreased levels of hormone-sensitive lipase and inhibited lipolysis. These findings indicate that the decrease in lipid droplets caused by t10-c12CLA is the result of an inhibition of lipid droplet production during adipogenesis rather than a stimulation of lipolysis. Additionally, treatment with Gö6976 blocked the effect of t10-c12CLA on perilipin-1 phosphorylation, implicating PKCα in perilipin-1 phosphorylation, and thus a regulator of triglyceride catabolism. These data are supported by evidence that t10-c12CLA activated PKCα. These are the first data to show that CLA isomers can affect lipid droplet dynamics in adipocytes through PKCα.

  18. Data from proteomic characterization of the role of Snail1 in murine mesenchymal stem cells and 3T3-L1 fibroblasts differentiation

    Directory of Open Access Journals (Sweden)

    A. Peláez-García

    2015-09-01

    Full Text Available The transcription factor (TF Snail1 is a major inducer of the epithelial–mesenchymal transition (EMT during embryonic development and cancer progression. Ectopic expression of Snail in murine mesenchymal stem cells (mMSC abrogated their differentiation to osteoblasts or adipocytes. We used either stable isotopic metabolic labeling (SILAC for 3T3-L1 cells or isobaric labeling with tandem mass tags (TMT for mMSC stably transfected cells with Snail1 or control. We carried out a proteomic analysis on the nuclear fraction since Snail is a nuclear TF that mediates its effects mainly through the regulation of other TFs. Proteomics data have been deposited in ProteomeXchange via the PRIDE partner repository with the dataset identifiers PXD001529 and PXD002157 (Vizcaino et al., 2014 [1]. Data are associated with a research article published in Molecular and Cellular Proteomics (Pelaez-Garcia et al., 2015 [2].

  19. Melatonin rescues 3T3-L1 adipocytes from FFA-induced insulin resistance by inhibiting phosphorylation of IRS-1 on Ser307.

    Science.gov (United States)

    She, Meihua; Hou, Hongjie; Wang, Zongbao; Zhang, Chi; Laudon, Moshe; Yin, Weidong

    2014-08-01

    Melatonin is biosynthesized in the pineal gland and secreted into the bloodstream. Evidences indicate a role of melatonin in the regulation of glucose metabolism. The objective of this study was to investigate the effect of melatonin on insulin sensitivity in insulin resistant adipocytes. Following a preincubation with melatonin or vehicle for 30 min, insulin resistant cells of 3T3-L1 adipocytes were induced by palmitic acids (300 μM, 6 h). Our results showed that palmitic acids inhibited both the basal and insulin-stimulated uptake of [(3)H]-2-Deoxyglucose, down-regulated the levels of IRS-1 and GLUT-4. However, compared to the vehicle group, melatonin pre-treatment increased significantly the uptake of [(3)H]-2-Deoxyglucose as well as the level of GLUT-4, and decreased phosphorylated IRS-1 (Ser307) although total IRS-1 did not change significantly. These data suggest that palmitic acids impair insulin signal via down-regulating the expressions of IRS-1 and GLUT-4; whereas melatonin can ameliorate insulin sensitivity by inhibiting Ser307 phosphorylation in IRS-1 and increasing GLUT-4 expressions in insulin resistant 3T3-L1 adipocytes. We conclude that melatonin regulates the insulin sensitivity and glucose homeostasis via inhibiting Ser-phosphorylation and improving function of IRS-1.

  20. Arachidonic acid has a dominant effect to regulate lipogenic genes in 3T3-L1 adipocytes compared to omega-3 fatty acids

    Directory of Open Access Journals (Sweden)

    Hitesh Vaidya

    2015-03-01

    Full Text Available Background: The effects of long-chain n-3 and n-6 polyunsaturated fatty acids (PUFA on the regulation of adipocytes metabolism are well known. These fatty acids are generally consumed together in our diets; however, the metabolic regulation of adipocytes in the presence of these fatty acids when given together is not known. Objective: To investigate the effects of n-3 PUFA and arachidonic acid (AA, an n-6 PUFA, on the regulation of adipogenic and lipogenic genes in mature 3T3-L1 adipocytes. Methods: 3T3-L1 adipocytes were incubated in the presence or absence of 100 µM of eicosapentaenoic acid, EPA; docosahexaenoic acid, DHA; docosapentaenoic acid, DPA and AA, either alone or AA+n-3 PUFA; control cells received bovine serum albumin alone. The mRNA expression of adipogenic and lipogenic genes was measured. The fatty acid composition of adipocytes was analyzed using gas chromatography. Results: Individual n-3 PUFA or AA had no effect on the mRNA expression of peroxisome-proliferator-activated receptor-γ; however, AA+EPA and AA+DPA significantly increased (P<0.05 the expression compared to control cells (38 and 42%, respectively. AA and AA+EPA increased the mRNA expression of acetyl-CoA carboxylase 1 (P<0.05. AA treatment decreased the mRNA expression of stearoyl-CoA desaturase (SCD1 (P<0.01, while n-3 PUFA, except EPA, had no effect compared to control cells. AA+DHA and AA+DPA inhibited SCD1 gene expression (P<0.05 suggesting a dominant effect of AA. Fatty acids analysis of adipocytes revealed a higher accretion of AA compared to n-3 PUFA. Conclusions: Our findings reveal that AA has a dominant effect on the regulation of lipogenic genes in adipocytes.

  1. Buckwheat (Fagopyrum esculentum M. Sprout Treated with Methyl Jasmonate (MeJA Improved Anti-Adipogenic Activity Associated with the Oxidative Stress System in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Jeong-Ho Lim

    2013-01-01

    Full Text Available Buckwheat sprouts contain various bioactive compounds including rutin which have a number of biological activities. We have previously shown that buckwheat sprouts (TBWE treated with methyl jasmonate (MeJA significantly increased the amount of phenolics and the antioxidant activity. The aim of this study was to demonstrate the effect of TBWE on anti-adipogenesis and pro-oxidant enzyme in 3T3-L1 adipocytes. We also evaluated the anti-oxidative activity of TBWE in adipocytes by using the nitroblue tetrazolium assay. Our data showed that TBWE markedly inhibited adipocyte differentiation and ROS production in 3T3-L1 cells compared with control groups. Moreover, TBWE has strongly shown the inhibition of adipogenic transcription factor as well as pro-oxidant enzymes. Together, we demonstrate that the MeJA treatment significantly increased the amount of phenolic compound, resulting in the suppression of adipogenesis and ROS production in the 3T3-L1 cells. These findings indicate that TBWE has the potential for anti-adipogenesis activity with anti-oxidative properties.

  2. Buckwheat (Fagopyrum esculentum M.) sprout treated with methyl jasmonate (MeJA) improved anti-adipogenic activity associated with the oxidative stress system in 3T3-L1 adipocytes.

    Science.gov (United States)

    Lee, Young-Jun; Kim, Kui-Jin; Park, Kee-Jai; Yoon, Bo-Ra; Lim, Jeong-Ho; Lee, Ok-Hwan

    2013-01-11

    Buckwheat sprouts contain various bioactive compounds including rutin which have a number of biological activities. We have previously shown that buckwheat sprouts (TBWE) treated with methyl jasmonate (MeJA) significantly increased the amount of phenolics and the antioxidant activity. The aim of this study was to demonstrate the effect of TBWE on anti-adipogenesis and pro-oxidant enzyme in 3T3-L1 adipocytes. We also evaluated the anti-oxidative activity of TBWE in adipocytes by using the nitroblue tetrazolium assay. Our data showed that TBWE markedly inhibited adipocyte differentiation and ROS production in 3T3-L1 cells compared with control groups. Moreover, TBWE has strongly shown the inhibition of adipogenic transcription factor as well as pro-oxidant enzymes. Together, we demonstrate that the MeJA treatment significantly increased the amount of phenolic compound, resulting in the suppression of adipogenesis and ROS production in the 3T3-L1 cells. These findings indicate that TBWE has the potential for anti-adipogenesis activity with anti-oxidative properties.

  3. 小檗碱对3T3-L1脂肪细胞内脂素表达的影响%Effect of berberine on visfatin expression in 3T3-L1 adipocytes

    Institute of Scientific and Technical Information of China (English)

    刘毅; 王文健; 李佑生; 马宇滢; 何燕铭; 何春燕; 陈伟华; 应健

    2007-01-01

    目的 观察小檗碱对3T3-L1脂肪细胞内脂素(visfatin)表达的影响和探讨小檗碱改善胰岛素抵抗的机制.方法 采用RT-PCR法检测不同浓度小檗碱和不同作用时间后,3T3-L1脂肪细胞内脂素mRNA的表达,并以Western印迹方法检测其蛋白水平的表达.结果 0~10 μmol/L小檗碱剂量依赖性地增加内脂素mRNA的表达,其中10 μmol/L时最明显,是空白对照组的4.96倍,其后随着小檗碱浓度的进一步增加,内脂素mRNA的表达反而降低;10 μmol/L的小檗碱作用3 h后内脂素mRNA的表达开始明显增强,至作用12 h时表达增强最明显,是基础组的4.57倍,随着作用时间的延长内脂素mRNA的表达虽然有所下降,但到48 h时内脂素mRNA的表达仍比空白对照组明显增高;5、10、20 μmol/L的小檗碱均可使内脂素蛋白的表达增加1.13、2.46、2.34倍,与空白对照组相比差异有统计学意义(P<0.05或P<0.01).结论 在一定浓度范围内小檗碱可剂量和时间依赖性地促进离体脂肪细胞内脂素mRNA及蛋白表达.小檗碱对内脂素表达的调节作用可能是其改善机体胰岛素抵抗、降低血糖的机制之一.

  4. 4,4'-Dichlorodiphenyltrichloroethane (DDT) and 4,4'-dichlorodiphenyldichloroethylene (DDE) promote adipogenesis in 3T3-L1 adipocyte cell culture.

    Science.gov (United States)

    Kim, Jonggun; Sun, Quancai; Yue, Yiren; Yoon, Kyong Sup; Whang, Kwang-Youn; Marshall Clark, J; Park, Yeonhwa

    2016-07-01

    4,4'-Dichlorodiphenyltrichloroethane (DDT), a chlorinated hydrocarbon insecticide, was extensively used in the 1940s and 1950s. DDT is mainly metabolically converted into 4,4'-dichlorodiphenyldichloroethylene (DDE). Even though most countries banned DDT in the 1970s, due to the highly lipophilic nature and very stable characteristics, DDT and its metabolites are present ubiquitously in the environment, including food. Recently, there are publications on relationships between exposure to insecticides, including DDT and DDE, and weight gain and altered glucose homeostasis. However, there are limited reports regarding DDT or DDE and adipogenesis, thus we investigated effects of DDT and DDE on adipogenesis using 3T3-L1 adipocytes. Treatment of DDT or DDE resulted in increased lipid accumulation accompanied by increased expression of CCAAT/enhancer-binding protein α (C/EBPα), peroxisome-proliferator activated receptor-γ (PPARγ), fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), adipose triglyceride lipase, and leptin. Moreover, treatment of DDT or DDE increased protein levels of C/EBPα, PPARγ, AMP-activated protein kinase-α (AMPKα), and ACC, while significant decrease of phosphorylated forms of AMPKα and ACC were observed. These finding suggest that increased lipid accumulation caused by DDT and DDE may mediate AMPKα pathway in 3T3-L1 adipocytes.

  5. Hop and Acacia Phytochemicals Decreased Lipotoxicity in 3T3-L1 Adipocytes, db/db Mice, and Individuals with Metabolic Syndrome

    Directory of Open Access Journals (Sweden)

    Deanna M. Minich

    2010-01-01

    Full Text Available The plant-based compounds rho-iso-alpha acids (RIAA from Humulus lupulus (hops and proanthocyanidins (PAC from Acacia nilotica have been shown to modulate insulin signaling in vitro. We investigated their effects on triglyceride (TG deposition in 3T3-L1 adipocytes, glucose and insulin in obese mouse models, and metabolic syndrome markers in adults with metabolic syndrome. The combination of RIAA and PAC synergistically increased TG content and adiponectin secretion in 3T3-L1 adipocytes under hyperinsulinemic conditions and reduced glucose or insulin in obese mice. In a clinical trial, tablets containing 100 mg RIAA and 500 mg PAC or placebo were administered to metabolic syndrome subjects (3 tablets/day, n=35; 6 tablets/day, n=34; or placebo, n=35 for 12 weeks. Compared to placebo, subjects taking 3 tablets daily showed greater reductions in TG, TG  :  HDL, fasting insulin, and HOMA scores. The combination of RIAA  :  PAC at 1  :  5 (wt  :  wt favorably modulates dysregulated lipids in insulin resistance and metabolic syndrome.

  6. PPARα agonist fenofibrate attenuates TNF-α-induced CD40 expression in 3T3-L1 adipocytes via the SIRT1-dependent signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Weirong [Department of Pharmacology, Cardiovascular Research Center, School of Medicine, Xi' an Jiaotong University, Xi' an, Shaanxi 710061 (China); Lin, Qinqin [Physical Education College, Yanshan University, Qinhuangdao, Hebei 066004 (China); Lin, Rong, E-mail: linrong63@yahoo.com.cn [Department of Pharmacology, Cardiovascular Research Center, School of Medicine, Xi' an Jiaotong University, Xi' an, Shaanxi 710061 (China); Zhang, Jiye [Faculty of Pharmacy, School of Medicine, Xi' an Jiaotong University, Xi' an, Shaanxi 710061 (China); Ren, Feng; Zhang, Jianfeng; Ji, Meixi; Li, Yanxiang [Department of Pharmacology, Cardiovascular Research Center, School of Medicine, Xi' an Jiaotong University, Xi' an, Shaanxi 710061 (China)

    2013-06-10

    The ligand-activated transcription factor peroxisome proliferator-activated receptor-α (PPARα) participates in the regulation of cellular inflammation. More recent studies indicated that sirtuin1 (SIRT1), a NAD{sup +}-dependent deacetylase, regulates the inflammatory response in adipocytes. However, whether the role of PPARα in inflammation is mediated by SIRT1 remains unclear. In this study, we aimed to determine the effect of PPARα agonist fenofibrate on the expressions of SIRT1 and pro-inflammatory cytokine CD40 and underlying mechanisms in 3T3-L1 adipocytes. We found that fenofibrate inhibited CD40 expression and up-regulated SIRT1 expression in tumor necrosis factor-α (TNF-α)-stimulated adipocytes, and these effects of fenofibrate were reversed by PPARα antagonist GW6471. Moreover, SIRT1 inhibitors sirtinol/nicotinamide (NAM) or knockdown of SIRT1 could attenuate the effect of fenofibrate on TNF-α-induced CD40 expression in adipocytes. Importantly, NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) augmented the effect of fenofibrate on CD40 expression in adipocytes. Further study found that fenofibrate decreased the expression of acetylated-NF-κB p65 (Ac-NF-κB p65) in TNF-α-stimulated adipocytes, and the effect of fenofibrate was abolished by SIRT1 inhibition. In addition, fenofibrate up-regulated SIRT1 expression through AMPK in TNF-α-stimulated adipocytes. Taken together, these findings indicate that PPARα agonist fenofibrate inhibits TNF-α-induced CD40 expression in 3T3-L1 adipocytes via the SIRT1-dependent signaling pathway. -- Highlights: • Fenofibrate up-regulates SIRT1 expression in TNF-α-stimulated adipocytes. • Fenofibrate inhibits CD40 expression through SIRT1 in adipocytes. • The effects of fenofibrate on CD40 and SIRT1 expressions are dependent on PPARα. • Fenofibrate inhibits CD40 expression via SIRT1-dependent deacetylation of NF-κB. • Fenofibrate increases SIRT1 expression through PPARα and AMPK in adipocytes.

  7. 8-Hydroxy-dihydroberberine ameliorated insulin resistance induced by high FFA and high glucose in 3T3-L1 adipocytes%8-羟基二氢小檗碱改善高FFA和高糖诱导的3T3-L1脂肪细胞胰岛素抵抗的作用

    Institute of Scientific and Technical Information of China (English)

    徐丽君; 陆付耳; 易屏; 王增四; 魏世超; 陈广; 董慧; 邹欣

    2009-01-01

    @@ 我国应用小檗碱临床治疗2型糖尿病已有30多年的经验,虽确有疗效,但其药效强度不够,始终不能成为治疗糖尿病的主导药物.小檗碱的口服吸收率太低是其降糖效果不佳的主要原因.%The purpose of the study is to investigate the effect of 8-hydroxy-dihydroberberine on insulin resistance induced by high free fatty acid (FFA) and high glucose in 3T3-L1 adipoeytes and its possible molecular mechanism. Palmic acid or glucose in combination with insulin was used to induce insulin resistance in 3T3-L1 adipocytes. 8-Hydroxy-dihydroberberine and berberine were added to the cultured medium separately, which were considered as treated group and positive control group. The rate of glucose uptake was determined by 2-deoxy-[~3H]-D-glucose method. The amount of glucose consumption in the medium was measured by glucose oxidase method. Cell growth and proliferation of 3T3-L1 adipocytes were detected with Cell Counting Kit-8 (CCK-8) assay. After incubated with palmic acid for 24 hours or glucose with insulin for 18 hours, the rate of glucose transport in 3T3-L1 adipocytes was inhibited by 67% and 58%, respectively. The amount of glucose consumption in 3T3-L1 adipose cells was decreased by 41% after cells were incubated with palmic acid for 24 h. However, the above changes were reversed by pretreatment with 8-hydroxy-dihydroberberine for 24 and 48 h. Significant difference existed between groups. Insulin resistance in 3T3-L1 adipocytes, which is induced by high FFA and high glucose, could be ameliorated by 8-hydroxy-dihydroberberine.

  8. Regulation of myosin IIA and filamentous actin during insulin-stimulated glucose uptake in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Stall, Richard; Ramos, Joseph; Kent Fulcher, F.; Patel, Yashomati M., E-mail: ympatel@uncg.edu

    2014-03-10

    Insulin stimulated glucose uptake requires the colocalization of myosin IIA (MyoIIA) and the insulin-responsive glucose transporter 4 (GLUT4) at the plasma membrane for proper GLUT4 fusion. MyoIIA facilitates filamentous actin (F-actin) reorganization in various cell types. In adipocytes F-actin reorganization is required for insulin-stimulated glucose uptake. What is not known is whether MyoIIA interacts with F-actin to regulate insulin-induced GLUT4 fusion at the plasma membrane. To elucidate the relationship between MyoIIA and F-actin, we examined the colocalization of MyoIIA and F-actin at the plasma membrane upon insulin stimulation as well as the regulation of this interaction. Our findings demonstrated that MyoIIA and F-actin colocalized at the site of GLUT4 fusion with the plasma membrane upon insulin stimulation. Furthermore, inhibition of MyoII with blebbistatin impaired F-actin localization at the plasma membrane. Next we examined the regulatory role of calcium in MyoIIA-F-actin colocalization. Reduced calcium or calmodulin levels decreased colocalization of MyoIIA and F-actin at the plasma membrane. While calcium alone can translocate MyoIIA it did not stimulate F-actin accumulation at the plasma membrane. Taken together, we established that while MyoIIA activity is required for F-actin localization at the plasma membrane, it alone is insufficient to localize F-actin to the plasma membrane. - Highlights: • Insulin induces colocalization of MyoIIA and F-actin at the cortex in adipocytes. • MyoIIA is necessary but not sufficient to localize F-actin at the cell cortex. • MyoIIA-F-actin colocalization is regulated by calcium and calmodulin.

  9. Catabolism of Branched Chain Amino Acids Contributes Significantly to Synthesis of Odd-Chain and Even-Chain Fatty Acids in 3T3-L1 Adipocytes.

    Science.gov (United States)

    Crown, Scott B; Marze, Nicholas; Antoniewicz, Maciek R

    2015-01-01

    The branched chain amino acids (BCAA) valine, leucine and isoleucine have been implicated in a number of diseases including obesity, insulin resistance, and type 2 diabetes mellitus, although the mechanisms are still poorly understood. Adipose tissue plays an important role in BCAA homeostasis by actively metabolizing circulating BCAA. In this work, we have investigated the link between BCAA catabolism and fatty acid synthesis in 3T3-L1 adipocytes using parallel 13C-labeling experiments, mass spectrometry and model-based isotopomer data analysis. Specifically, we performed parallel labeling experiments with four fully 13C-labeled tracers, [U-13C]valine, [U-13C]leucine, [U-13C]isoleucine and [U-13C]glutamine. We measured mass isotopomer distributions of fatty acids and intracellular metabolites by GC-MS and analyzed the data using the isotopomer spectral analysis (ISA) framework. We demonstrate that 3T3-L1 adipocytes accumulate significant amounts of even chain length (C14:0, C16:0 and C18:0) and odd chain length (C15:0 and C17:0) fatty acids under standard cell culture conditions. Using a novel GC-MS method, we demonstrate that propionyl-CoA acts as the primer on fatty acid synthase for the production of odd chain fatty acids. BCAA contributed significantly to the production of all fatty acids. Leucine and isoleucine contributed at least 25% to lipogenic acetyl-CoA pool, and valine and isoleucine contributed 100% to lipogenic propionyl-CoA pool. Our results further suggest that low activity of methylmalonyl-CoA mutase and mass action kinetics of propionyl-CoA on fatty acid synthase result in high rates of odd chain fatty acid synthesis in 3T3-L1 cells. Overall, this work provides important new insights into the connection between BCAA catabolism and fatty acid synthesis in adipocytes and underscores the high capacity of adipocytes for metabolizing BCAA.

  10. Catabolism of Branched Chain Amino Acids Contributes Significantly to Synthesis of Odd-Chain and Even-Chain Fatty Acids in 3T3-L1 Adipocytes.

    Directory of Open Access Journals (Sweden)

    Scott B Crown

    Full Text Available The branched chain amino acids (BCAA valine, leucine and isoleucine have been implicated in a number of diseases including obesity, insulin resistance, and type 2 diabetes mellitus, although the mechanisms are still poorly understood. Adipose tissue plays an important role in BCAA homeostasis by actively metabolizing circulating BCAA. In this work, we have investigated the link between BCAA catabolism and fatty acid synthesis in 3T3-L1 adipocytes using parallel 13C-labeling experiments, mass spectrometry and model-based isotopomer data analysis. Specifically, we performed parallel labeling experiments with four fully 13C-labeled tracers, [U-13C]valine, [U-13C]leucine, [U-13C]isoleucine and [U-13C]glutamine. We measured mass isotopomer distributions of fatty acids and intracellular metabolites by GC-MS and analyzed the data using the isotopomer spectral analysis (ISA framework. We demonstrate that 3T3-L1 adipocytes accumulate significant amounts of even chain length (C14:0, C16:0 and C18:0 and odd chain length (C15:0 and C17:0 fatty acids under standard cell culture conditions. Using a novel GC-MS method, we demonstrate that propionyl-CoA acts as the primer on fatty acid synthase for the production of odd chain fatty acids. BCAA contributed significantly to the production of all fatty acids. Leucine and isoleucine contributed at least 25% to lipogenic acetyl-CoA pool, and valine and isoleucine contributed 100% to lipogenic propionyl-CoA pool. Our results further suggest that low activity of methylmalonyl-CoA mutase and mass action kinetics of propionyl-CoA on fatty acid synthase result in high rates of odd chain fatty acid synthesis in 3T3-L1 cells. Overall, this work provides important new insights into the connection between BCAA catabolism and fatty acid synthesis in adipocytes and underscores the high capacity of adipocytes for metabolizing BCAA.

  11. Mechanisms of TNF-α-induced insulin resistance in 3T3-L1 adipocytes%肿瘤坏死因子α诱导3T3-L1脂肪细胞胰岛素抵抗的机制

    Institute of Scientific and Technical Information of China (English)

    黄雌友; 吴卫国; 王卓平; 姚伟峰; 王雯

    2015-01-01

    Objective To demonstrate the roles of hydrogen sulfide (H2S) on TNF-α-induced insulin re-sistance in 3T3-L1 adipocytes. Methods The differentiated 3T3-L1 adipocytes were incubated without or with 100 nmol/L insulin for 30 min periods. For experiments with TNF-αtreatment, different concentrations of TNF-α(0, 10 ng/ml, 50 ng/ml, 100 ng/ml) were added for 24 h and then treated with 100 nmol/L insulin for 30 min. For the inter-vention group, 3T3-L1 adipocytes were pretreated with the cystathionineγ-lyase (CSE) inhibitor DL-propargylgly-cine (PPG, 10 mmol/L) or beta-cyano-L-alanine (BCA, 2 mmol/L) for 30 min prior to treatment with 50 ng/ml of TNF-α for 24 h and insulin stimulation (100 nmol/L, 30 min). The glucose consumption in the medium was mea-sured by glucose oxidase method, and the glucose uptake were detected by 2-[3H]-DG method. The content of H2S in culture supernatant was measured by the methylene blue method. Results Compared with the control group, in-sulin-induced glucose consumption [(32.43 ± 2.60)% vs (5.31 ± 1.87)%, P<0.01)] and uptake [(7.91 ± 0.31) cpm vs (1.38±0.13) cpm, P<0.01)] in 3T3-L1 adipocytes were significantly higher. After treatment with TNF-α(10 ng/ml, 50 ng/ml, 100 ng/ml), the insulin-induced glucose consumption [(23.27 ± 1.41)%, (14.73 ± 3.76)%, (7.81 ± 1.56)% vs (32.43±2.60)%, P<0.01] and uptake [(5.86±0.95) cpm, (2.60±0.35) cpm, (1.87±0.17) cpm vs (7.91±0.31) cpm, P<0.01)] were significantly decreased in a dose-dependent manner, while endogenous H2S generation increased in a dose-de-pendent manner [(0.06 ± 0.002) mmol/(g·protein), (0.11 ± 0.003) mmol/(g·protein), (0.14 ± 0.005) mmol/(g·protein) vs (0.17±0.001) mmol/(g·protein), P<0.01)]. Pretreatment with the CSE inhibitor PPG (10 mmol/L) or BCA (2 mmol/L) for 30 min respectively resulted in significant increase in insulin-induced glucose consumption [(24.52 ± 2.63)%, (27.84±1.82)%vs (14.73±3.76)%, P<0.01] and uptake [(7.22±0.27) cpm, (7.96±0.19) cpm vs

  12. 4-Hydroxyisoleucine ameliorates an insulin resistant-like state in 3T3-L1 adipocytes by regulating TACE/TIMP3 expression

    Directory of Open Access Journals (Sweden)

    Gao F

    2015-10-01

    Full Text Available Feng Gao,1 Wen Du,1,4 Mohammad Ishraq Zafar,1 Raja Adeel Shafqat,2 Liumeng Jian,1 Qin Cai,1 Furong Lu3 1Department of Endocrinology, Union Hospital, 2Department of Medicine, Tongji Hospital, 3Department of Integrated Traditional Chinese and Western Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, 4Chengdu First People’s Hospital, Sichuan, People’s Republic of China Background: Obesity-associated insulin resistance (IR is highly correlated with soluble tumor necrosis factor-α (sTNF-α, which is released from transmembranous TNF-α by TNF-α converting enzyme (TACE. In vivo, TACE activity is suppressed by tissue inhibitor of metalloproteinase 3 (TIMP3. Agents that can interact with TACE/TIMP3 to improve obesity-related IR would be highly valuable. In the current study, we assessed whether (2S,3R,4S-4-hydroxyisoleucine (4-HIL could modulate TACE/TIMP3 and ameliorate an obesity-induced IR-like state in 3T3-L1 adipocytes. Materials and methods: 3T3-L1 adipocytes were incubated in the presence of 25 mM glucose and 0.6 nM insulin to induce an IR-like state, and were then treated with different concentrations of 4-HIL or 10 µM pioglitazone (positive control. The glucose uptake rate was determined using the 2-deoxy-[3H]-d-glucose method, and the levels of sTNF-α in the cell supernatant were determined using ELISA. The protein expression of TACE, TIMP3, and insulin signaling-related molecules was measured using western blotting. Results: Exposure to high glucose and insulin for 18 hours increased the levels of sTNF-α in the cell supernatant. The phosphorylation of insulin receptor substrate-1 (IRS-1 Ser307 and Akt Ser473 was increased, whereas the protein expression of IRS-1, Akt, and glucose transporter-4 was decreased. The insulin-induced glucose uptake was reduced by 67% in 3T3-L1 adipocytes, which indicated the presence of an IR-like state. The above indexes, which demonstrated the

  13. The Effect of Pericellular Oxygen Levels on Proteomic Profile and Lipogenesis in 3T3-L1 Differentiated Preadipocytes Cultured on Gas-Permeable Cultureware.

    Directory of Open Access Journals (Sweden)

    Martin Weiszenstein

    Full Text Available Pericellular oxygen concentration represents an important factor in the regulation of cell functions, including cell differentiation, growth and mitochondrial energy metabolism. Hypoxia in adipose tissue has been associated with altered adipokine secretion profile and suggested as a possible factor in the development of type 2 diabetes. In vitro experiments provide an indispensable tool in metabolic research, however, physical laws of gas diffusion make prolonged exposure of adherent cells to desired pericellular O2 concentrations questionable. The aim of this study was to investigate the direct effect of various O2 levels (1%, 4% and 20% O2 on the proteomic profile and triglyceride accumulation in 3T3-L1 differentiated preadipocytes using gas-permeable cultureware. Following differentiation of cells under desired pericellular O2 concentrations, cell lysates were subjected to two-dimensional gel electrophoresis and protein visualization using Coomassie blue staining. Spots showing differential expression under hypoxia were analyzed using matrix-assisted laser desorption/ionization mass spectrometry. All identified proteins were subjected to pathway analysis. We observed that protein expression of 26 spots was reproducibly affected by 4% and 1% O2 (17 upregulated and 9 downregulated. Pathway analysis showed that mitochondrial energy metabolism and triglyceride synthesis were significantly upregulated by hypoxia. In conclusion, this study demonstrated the direct effects of pericellular O2 levels on adipocyte energy metabolism and triglyceride synthesis, probably mediated through the reversed tricarboxylic acid cycle flux.

  14. The Effect of Pericellular Oxygen Levels on Proteomic Profile and Lipogenesis in 3T3-L1 Differentiated Preadipocytes Cultured on Gas-Permeable Cultureware

    Science.gov (United States)

    Weiszenstein, Martin; Pavlikova, Nela; Elkalaf, Moustafa; Halada, Petr; Seda, Ondrej; Trnka, Jan; Kovar, Jan; Polak, Jan

    2016-01-01

    Pericellular oxygen concentration represents an important factor in the regulation of cell functions, including cell differentiation, growth and mitochondrial energy metabolism. Hypoxia in adipose tissue has been associated with altered adipokine secretion profile and suggested as a possible factor in the development of type 2 diabetes. In vitro experiments provide an indispensable tool in metabolic research, however, physical laws of gas diffusion make prolonged exposure of adherent cells to desired pericellular O2 concentrations questionable. The aim of this study was to investigate the direct effect of various O2 levels (1%, 4% and 20% O2) on the proteomic profile and triglyceride accumulation in 3T3-L1 differentiated preadipocytes using gas-permeable cultureware. Following differentiation of cells under desired pericellular O2 concentrations, cell lysates were subjected to two-dimensional gel electrophoresis and protein visualization using Coomassie blue staining. Spots showing differential expression under hypoxia were analyzed using matrix-assisted laser desorption/ionization mass spectrometry. All identified proteins were subjected to pathway analysis. We observed that protein expression of 26 spots was reproducibly affected by 4% and 1% O2 (17 upregulated and 9 downregulated). Pathway analysis showed that mitochondrial energy metabolism and triglyceride synthesis were significantly upregulated by hypoxia. In conclusion, this study demonstrated the direct effects of pericellular O2 levels on adipocyte energy metabolism and triglyceride synthesis, probably mediated through the reversed tricarboxylic acid cycle flux. PMID:27023342

  15. The Effect of Pericellular Oxygen Levels on Proteomic Profile and Lipogenesis in 3T3-L1 Differentiated Preadipocytes Cultured on Gas-Permeable Cultureware.

    Science.gov (United States)

    Weiszenstein, Martin; Pavlikova, Nela; Elkalaf, Moustafa; Halada, Petr; Seda, Ondrej; Trnka, Jan; Kovar, Jan; Polak, Jan

    2016-01-01

    Pericellular oxygen concentration represents an important factor in the regulation of cell functions, including cell differentiation, growth and mitochondrial energy metabolism. Hypoxia in adipose tissue has been associated with altered adipokine secretion profile and suggested as a possible factor in the development of type 2 diabetes. In vitro experiments provide an indispensable tool in metabolic research, however, physical laws of gas diffusion make prolonged exposure of adherent cells to desired pericellular O2 concentrations questionable. The aim of this study was to investigate the direct effect of various O2 levels (1%, 4% and 20% O2) on the proteomic profile and triglyceride accumulation in 3T3-L1 differentiated preadipocytes using gas-permeable cultureware. Following differentiation of cells under desired pericellular O2 concentrations, cell lysates were subjected to two-dimensional gel electrophoresis and protein visualization using Coomassie blue staining. Spots showing differential expression under hypoxia were analyzed using matrix-assisted laser desorption/ionization mass spectrometry. All identified proteins were subjected to pathway analysis. We observed that protein expression of 26 spots was reproducibly affected by 4% and 1% O2 (17 upregulated and 9 downregulated). Pathway analysis showed that mitochondrial energy metabolism and triglyceride synthesis were significantly upregulated by hypoxia. In conclusion, this study demonstrated the direct effects of pericellular O2 levels on adipocyte energy metabolism and triglyceride synthesis, probably mediated through the reversed tricarboxylic acid cycle flux.

  16. The impact of cholesteryl ester transfer protein on glucose metabolism in 3T3-L1 adipocytes%胆固醇酯转运蛋白对3T3-L1脂肪细胞糖代谢的影响

    Institute of Scientific and Technical Information of China (English)

    朱晓慧; 常毅娜; 付真真; 郭雯; 高贝贝; 符金香; 陈晓丽; 周红文

    2014-01-01

    3T3-L1 adipocytes stably expressing different levels of human cholesterol ester transfer protein (CETP) were constructed and identified.Glucose uptake and glucose transporter 4 (GLUT4) protein levels of these cells were also measured.Insulin-stimulated 2-deoxyglucose uptake was significantly higher in 3T3-L1 adipocytes which expressed high,medium,and low levels of CETP than that in control ceils,and the elevated levels of glucose uptake were positively related with CETP expression in a dose-dependent manner.After insulin stimulation,there was no difference in GLUT4 protein expression among control cell and those expressing CETP.CETP plays a role in the regulation of glucose metabolism in adipocytes.%建立人胆固醇酯转运蛋白(CETP)不同表达水平的3T3-L1脂肪前体细胞株,并进行鉴定,测定葡萄糖摄取率以及葡萄糖转运体4(GLUT4)蛋白表达水平.与对照组相比,表达CETP的3T3-L1脂肪细胞葡萄糖摄取显著增高,且此作用与CETP表达量呈正相关.胰岛素刺激后,稳定表达CETP的3T3-L1脂肪细胞GLUT4蛋白表达水平与对照组相比无显著差异.推测CETP可能通过调节脂肪细胞内胆固醇含量的变化而促进脂肪细胞的糖代谢.

  17. Nuclear phosphoproteome analysis of 3T3-L1 preadipocyte differentiation reveals system-wide phosphorylation of transcriptional regulators

    DEFF Research Database (Denmark)

    Rabiee, Atefeh; Schwämmle, Veit; Sidoli, Simone;

    2017-01-01

    HIGHLIGHTS: Mass spectrometry (MS) based quantitative proteomics and phosphoproteomics applied to monitor the alteration of nuclear proteins during the early stages (4 hours) of preadipocyte differentiation. A total of 4072 proteins including 2434 phosphorylated proteins identified, a majority......), in particular phosphorylation, play a major role in activating and propagating signals within TR networks upon induction of adipogenesis by extracellular stimulus. We applied mass spectrometry (MS) based quantitative proteomics and phosphoproteomics to monitor the alteration of nuclear proteins during the early....... New insights into phosphorylation-dependent signaling networks that impact on nuclear proteins and controls adipocyte differentiation and cell fate. Adipocytes (fat cells) are important endocrine and metabolic cells critical for systemic insulin sensitivity. Both adipose excess and insufficiency...

  18. Uncoupling of 3T3-L1 gene expression from lipid accumulation during adipogenesis

    OpenAIRE

    Temple, Karla A.; Basko, Xheni; Allison, Margaret B.; Brady, Matthew J.

    2007-01-01

    Adipocyte differentiation comprises altered gene expression and increased triglyceride storage. To investigate the interdependency of these two events, 3T3-L1 cells were differentiated in the presence of glucose or pyruvate. All adipocytic proteins examined were similarly increased between the two conditions. In contrast, 3T3-L1 adipocytes differentiated with glucose exhibited significant lipid accumulation, which was largely suppressed in the presence of pyruvate. Subsequent addition of gluc...

  19. 小檗碱对白介素-6诱导胰岛素抵抗3T3-L1脂肪细胞脂联素表达的影响%Effect of Berberine on Gene Expression of Adiponectin by IL-6 Induced Insulin Resistance in 3T3-L1 Adipocyte

    Institute of Scientific and Technical Information of China (English)

    章常华; 李冰涛; 屈飞; 魏学鑫; 余日跃; 徐国良

    2012-01-01

    Objective; To explore the effect of berberine on adiponectin gene mRNA expression in 3T3-Ll adipocyte of insulin resistant induced by interleukin-6 (IL-6). Method; To build 3T3-L1 adipocyte model of insulin resistant induced by interleukin-6. 3T3-L1 adipocytes were exposed to 20 μg -L-1 IL-6 for 48 h, the 3T3-L1 adipocytes with insulin resistant were divided into six groups randomly; normal control groups, model groups, pioglitazone (50 (μmol -L-1 ) , and low, middle, hight doses of berberine groups (10, 20, 50 (μmol -L-1) , the amount of glucose consumption was measured by detecting the glucose content in cell culture supernatants with glucose oxidase assay, the effect of berberine on glucose uptake of 3T3-L1 adipocytes was observed, and cell model of insulin resistant was identified. mRNA expression of adiponection gene was determined by real-time quantitative reverse-transcription polymerase chain reaction. Result; There was statistical difference between model group and normal control group in their amount of glucose consumption and gene expression of adiponectin, which was decreased markedly in model group (P < 0. 05 ). Pioglitazone (50 (μmol -L-1 ) , and low, middle, high doses of berberine groups (10, 20, 50 (μmol -L-1 ) markedly increased both amount of glucose consumption and gene expression of adiponectin. Conclusion; Berberine can increase mRNA expression of adiponection gene and improve insulin resistant in 3T3-L1 adipocytes.%目的:观察小檗碱对白介素-6(IL-6)诱导胰岛素抵抗的3T3-L1脂肪细胞脂联素表达的影响.方法:选用IL-6诱导3T3-L1脂肪细胞胰岛素抵抗(IR)模型.以20 μg·L-1IL-6培养48 h,将3T3-L1脂肪细胞随机分为正常对照组、模型组、吡格列酮组(50μmol·L-1)和小檗碱高、中、低剂量组(10,20,50 μmol·L-1),以葡萄糖氧化酶法测定葡萄糖消耗量,观察小檗碱对脂肪细胞葡萄糖摄取的影响,鉴定IR模型;采用实时荧光定量PCR技术测定脂肪细胞脂联

  20. Glutamine, insulin and glucocorticoids regulate glutamine synthetase expression in C2C12 myotubes, Hep G2 hepatoma cells and 3T3 L1 adipocytes.

    Science.gov (United States)

    Wang, Yanxin; Watford, Malcolm

    2007-04-01

    The cell-specific regulation of glutamine synthetase expression was studied in three cell lines. In C2C12 myotubes, glucocorticoids increased the abundance of both glutamine synthetase protein and mRNA. Culture in the absence of glutamine also resulted in very high glutamine synthetase protein abundance but mRNA levels were unchanged. Glucocorticoids also increased the abundance of glutamine synthetase mRNA in Hep G2 hepatoma cells but this was not reflected in changes in protein abundance. Culture of Hep G2 cells without glutamine resulted in very high levels of protein, again with no change in mRNA abundance. Insulin was without effect in both C2C12 and Hep G2 cells. In 3T3 L1 adipocytes glucocorticoids increased the abundance of both glutamine synthetase mRNA and protein, insulin added alone had no effect but in the presence of glucocorticoids resulted in lower mRNA levels than seen with glucocorticoids alone, although protein levels remained high under such conditions. In contrast to the other cell lines glutamine synthetase protein levels were relatively unchanged by culture in the absence of glutamine. The results support the hypothesis that in myocytes, and hepatomas, but not in adipocytes, glutamine acts to moderate glutamine synthetase induction by glucocorticoids.

  1. Myosin IIA participates in docking of Glut4 storage vesicles with the plasma membrane in 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Le Thi Kim, E-mail: ngocanh@nutr.med.tokushima-u.ac.jp [Department of Nutrition and Metabolism, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8503 (Japan); Hosaka, Toshio [Department of Public Health and Applied Nutrition, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Harada, Nagakatsu; Jambaldorj, Bayasgalan; Fukunaga, Keiko; Nishiwaki, Yuka [Department of Nutrition and Metabolism, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8503 (Japan); Teshigawara, Kiyoshi [Clinical Research Center for Diabetes, Tokushima University Hospital, 2-50-1 Kuramoto-cho, Tokushima 770-8503 (Japan); Sakai, Tohru [Department of Public Health and Applied Nutrition, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima (Japan); Nakaya, Yutaka [Department of Nutrition and Metabolism, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8503 (Japan); Funaki, Makoto, E-mail: m-funaki@clin.med.tokushima-u.ac.jp [Clinical Research Center for Diabetes, Tokushima University Hospital, 2-50-1 Kuramoto-cho, Tokushima 770-8503 (Japan)

    2010-01-01

    In adipocytes and myocytes, insulin stimulation translocates glucose transporter 4 (Glut4) storage vesicles (GSVs) from their intracellular storage sites to the plasma membrane (PM) where they dock with the PM. Then, Glut4 is inserted into the PM and initiates glucose uptake into these cells. Previous studies using chemical inhibitors demonstrated that myosin II participates in fusion of GSVs and the PM and increase in the intrinsic activity of Glut4. In this study, the effect of myosin IIA on GSV trafficking was examined by knocking down myosin IIA expression. Myosin IIA knockdown decreased both glucose uptake and exposures of myc-tagged Glut4 to the cell surface in insulin-stimulated cells, but did not affect insulin signal transduction. Interestingly, myosin IIA knockdown failed to decrease insulin-dependent trafficking of Glut4 to the PM. Moreover, in myosin IIA knockdown cells, insulin-stimulated binding of GSV SNARE protein, vesicle-associated membrane protein 2 (VAMP2) to PM SNARE protein, syntaxin 4 was inhibited. These data suggest that myosin IIA plays a role in insulin-stimulated docking of GSVs to the PM in 3T3-L1 adipocytes through SNARE complex formation.

  2. Cinnamon extract enhances glucose uptake in 3T3-L1 adipocytes and C2C12 myocytes by inducing LKB1-AMP-activated protein kinase signaling.

    Directory of Open Access Journals (Sweden)

    Yan Shen

    Full Text Available We previously demonstrated that cinnamon extract (CE ameliorates type 1 diabetes induced by streptozotocin in rats through the up-regulation of glucose transporter 4 (GLUT4 translocation in both muscle and adipose tissues. This present study was aimed at clarifying the detailed mechanism(s with which CE increases the glucose uptake in vivo and in cell culture systems using 3T3-L1 adipocytes and C2C12 myotubes in vitro. Specific inhibitors of key enzymes in insulin signaling and AMP-activated protein kinase (AMPK signaling pathways, as well as small interference RNA, were used to examine the role of these kinases in the CE-induced glucose uptake. The results showed that CE stimulated the phosphorylation of AMPK and acetyl-CoA carboxylase. An AMPK inhibitor and LKB1 siRNA blocked the CE-induced glucose uptake. We also found for the first time that insulin suppressed AMPK activation in the adipocyte. To investigate the effect of CE on type 2 diabetes in vivo, we further performed oral glucose tolerance tests and insulin tolerance tests in type 2 diabetes model rats administered with CE. The CE improved glucose tolerance in oral glucose tolerance tests, but not insulin sensitivity in insulin tolerance test. In summary, these results indicate that CE ameliorates type 2 diabetes by inducing GLUT4 translocation via the AMPK signaling pathway. We also found insulin antagonistically regulates the activation of AMPK.

  3. Fisetin Suppresses Lipid Accumulation in Mouse Adipocytic 3T3-L1 Cells by Repressing GLUT4-Mediated Glucose Uptake through Inhibition of mTOR-C/EBPα Signaling.

    Science.gov (United States)

    Watanabe, Marina; Hisatake, Mitsuhiro; Fujimori, Ko

    2015-05-27

    3,7,3',4'-Tetrahydroxyflavone (fisetin) is a flavonoid found in vegetables and fruits having broad biological activities. Here the effects of fisetin on adipogenesis and its regulatory mechanism in mouse adipocytic 3T3-L1 cells are studied. Fisetin inhibited the accumulation of intracellular lipids and lowered the expression of adipogenic genes such as peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein (C/EBP) α and fatty acid-binding protein 4 (aP2) during adipogenesis. Moreover, the mRNA levels of genes such as acetyl-CoA carboxylase, fatty acid synthase, and stearoyl-CoA desaturase involved in the fatty acid biosynthesis (lipogenesis) were reduced by the treatment with fisetin. The expression level of the glucose transporter 4 (GLUT4) gene was also decreased by fisetin, resulting in down-regulation of glucose uptake. Furthermore, fisetin inhibited the phosphorylation of the mammalian target of rapamycin (mTOR) and that of p70 ribosomal S6 kinase, a target of the mTOR complex, the inhibition of which was followed by a decreased mRNA level of the C/EBPα gene. The results obtained from a chromatin immunoprecipitation assay demonstrated that the ability of C/EBPα to bind to the GLUT4 gene promoter was reduced by the treatment with fisetin, which agreed well with those obtained when 3T3-L1 cells were allowed to differentiate into adipocytes in medium in the presence of rapamycin, an inhibitor for mTOR. These results indicate that fisetin suppressed the accumulation of intracellular lipids by inhibiting GLUT4-mediated glucose uptake through inhibition of the mTOR-C/EBPα signaling in 3T3-L1 cells.

  4. Glycerol Production from Glucose and Fructose by 3T3-L1 Cells: A Mechanism of Adipocyte Defense from Excess Substrate.

    Directory of Open Access Journals (Sweden)

    María del Mar Romero

    Full Text Available Cultured adipocytes (3T3-L1 produce large amounts of 3C fragments; largely lactate, depending on medium glucose levels. Increased glycolysis has been observed also in vivo in different sites of rat white adipose tissue. We investigated whether fructose can substitute glucose as source of lactate, and, especially whether the glycerol released to the medium was of lipolytic or glycolytic origin. Fructose conversion to lactate and glycerol was lower than that of glucose. The fast exhaustion of medium glucose was unrelated to significant changes in lipid storage. Fructose inhibited to a higher degree than glucose the expression of lipogenic enzymes. When both hexoses were present, the effects of fructose on gene expression prevailed over those of glucose. Adipocytes expressed fructokinase, but not aldolase b. Substantive release of glycerol accompanied lactate when fructose was the substrate. The mass of cell triacylglycerol (and its lack of change could not justify the comparatively higher amount of glycerol released. Consequently, most of this glycerol should be derived from the glycolytic pathway, since its lipolytic origin could not be (quantitatively sustained. Proportionally (with respect to lactate plus glycerol, more glycerol was produced from fructose than from glucose, which suggests that part of fructose was catabolized by the alternate (hepatic fructose pathway. Earlier described adipose glycerophophatase activity may help explain the glycolytic origin of most of the glycerol. However, no gene is known for this enzyme in mammals, which suggests that this function may be carried out by one of the known phosphatases in the tissue. Break up of glycerol-3P to yield glycerol, may be a limiting factor for the synthesis of triacylglycerols through control of glycerol-3P availability. A phosphatase pathway such as that described may have a potential regulatory function, and explain the production of glycerol by adipocytes in the absence of

  5. Nymphaea nouchali Burm. f. hydroalcoholic seed extract increases glucose consumption in 3T3-L1 adipocytes through activation of peroxisome proliferator-activated receptor gamma and insulin sensitization

    Directory of Open Access Journals (Sweden)

    Mabel Parimala

    2015-01-01

    Full Text Available Nymphaea nouchali Burm. f. (Family - Nymphaeaceae is a well-known medicinal plant used in the Indian ayurvedic system of medicine for treating diabetes. The seeds especially have been prescribed for diabetes. The hydroalcoholic extract of N. nouchali seeds has been demonstrated to possess anti-hyperglycemic effects in diabetic rats, but the functional mechanism remains unknown. The nuclear receptor, peroxisome proliferator-activated receptor gamma (PPARγ is noted to play an important role in glucose and lipid homeostasis. This study was hence focused in evaluating the effect of the extract on PPARγ activation, adipocyte differentiation, and glucose consumption in 3T3-L1 cells. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide (MTT, followed by adipogenesis assay using Oil Red O technique. Glucose consumption of preadipocytes and adipocytes in the presence of the extract was also determined. Real-time polymerase chain reaction was performed to identify the expression of genes involved in glucose consumption in the adipocytes. MTT assay confirmed the extract to be nontoxic, and Oil Red O staining confirmed enhanced adipocyte differentiation of 3T3-L1 cells in a dose-dependent manner. The extract also increased the expression of PPARγ target gene, which in turn enhanced the expression of GLUT-4. The data, therefore, suggests that N. nouchali seed extract promotes adipocyte differentiation and glucose consumption by inducing PPARγ activation, which in turn increases mRNA GLUT-4 expression and subsequently enhances insulin-responsiveness in insulin target tissues.

  6. Nymphaea nouchali Burm. f. hydroalcoholic seed extract increases glucose consumption in 3T3-L1 adipocytes through activation of peroxisome proliferator-activated receptor gamma and insulin sensitization.

    Science.gov (United States)

    Parimala, Mabel; Debjani, M; Vasanthi, Hannah Rachel; Shoba, Francis Gricilda

    2015-01-01

    Nymphaea nouchali Burm. f. (Family - Nymphaeaceae) is a well-known medicinal plant used in the Indian ayurvedic system of medicine for treating diabetes. The seeds especially have been prescribed for diabetes. The hydroalcoholic extract of N. nouchali seeds has been demonstrated to possess anti-hyperglycemic effects in diabetic rats, but the functional mechanism remains unknown. The nuclear receptor, peroxisome proliferator-activated receptor gamma (PPARγ) is noted to play an important role in glucose and lipid homeostasis. This study was hence focused in evaluating the effect of the extract on PPARγ activation, adipocyte differentiation, and glucose consumption in 3T3-L1 cells. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), followed by adipogenesis assay using Oil Red O technique. Glucose consumption of preadipocytes and adipocytes in the presence of the extract was also determined. Real-time polymerase chain reaction was performed to identify the expression of genes involved in glucose consumption in the adipocytes. MTT assay confirmed the extract to be nontoxic, and Oil Red O staining confirmed enhanced adipocyte differentiation of 3T3-L1 cells in a dose-dependent manner. The extract also increased the expression of PPARγ target gene, which in turn enhanced the expression of GLUT-4. The data, therefore, suggests that N. nouchali seed extract promotes adipocyte differentiation and glucose consumption by inducing PPARγ activation, which in turn increases mRNA GLUT-4 expression and subsequently enhances insulin-responsiveness in insulin target tissues.

  7. Role of 11-beta-hydroxysteroid dehydrogenase type 1 in differentiation of 3T3-L1 cells and in rats with diet-induced obesity

    Institute of Scientific and Technical Information of China (English)

    Yun LIU; Wen-lan SUN; Yan SUN; Gang HU; Guo-xian DING

    2006-01-01

    Aim: To observe the roles of 11-beta-hydroxysteroid dehydrogenase type 1 (11β-HSD1) in in vitro preadipocyte differentiation and in rats with diet-induced obesity (DIO). Methods: Protein expression of 11β-HSD1 in the process of 3T3-L1 cell differentiation and in various tissues of the rats were detected by Western blot analysis; expression of 11β-HSD1 mRNA and glucocorticoid receptor (GR) and other marker genes of preadipocyte differentiation were detected by using real-time PCR. Results: Lipid droplets in 3T3-L1 cells accumulated and increased after stimulation. A dramatically elevated protein level of 11β-HSD1, especially in the late stages of 3T3-L1 cell differentiation, was detected. The relative mRNA levels of 11β-HSD1, GR and cell differentiation markers LPL, aP2, and FAS were upregulated, and Pref-1 was downregulated during the differentiation. In DIO rats, bodyweight, visceral adipose mass index and the protein expression of 11β-HSD1 increased, especially in adipose tissue, brain and muscles. Serum insulin, triglyceride, total cholesterol and 1oW-density lipoprotein cholesterol were found to be increased in DIO rats, but without any obvious changes in blood glucose or tumor necrosis factor-αlevels. Conclusion: 11β-HSD1 may promote preadipocyte differentiation, and may be involved in the development of obesity.

  8. Purification and Characterization of Aporphine Alkaloids from Leaves of Nelumbo nucifera Gaertn and Their Effects on Glucose Consumption in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Chengjun Ma

    2014-02-01

    Full Text Available Aporphine alkaloids from the leaves of Nelumbo nucifera Gaertn are substances of great interest because of their important pharmacological activities, particularly anti-diabetic, anti-obesity, anti-hyperlipidemic, anti-oxidant, and anti-HIV’s activities. In order to produce large amounts of pure alkaloid for research purposes, a novel method using high-speed counter-current chromatography (HSCCC was developed. Without any initial cleanup steps, four main aporphine alkaloids, including 2-hydroxy-1-methoxyaporphine, pronuciferine, nuciferine and roemerine were successfully purified from the crude extract by HSCCC in one step. The separation was performed with a simple two-phase solvent system composed of n-hexane-ethyl acetate-methanol-acetonitrile-water (5:3:3:2.5:5, v/v/v/v/v. In each operation, 100 mg crude extracts was separated and yielded 6.3 mg of 2-hydroxy-1-methoxyaporphine (95.1% purity, 1.1 mg of pronuciferine (96.8% purity, 8.5 mg of nuciferine (98.9% purity, and 2.7 mg of roemerine (97.4% respectively. The chemical structure of four aporphine alkaloids are identified by means of electrospray ionization MS (ESI-MS and nuclear magnetic resonance (NMR analysis. Moreover, the effects of four separated aporphine alkaloids on insulin-stimulated glucose consumption were examined in 3T3-L1 adipocytes. The results showed that 2-hydroxy-1-methoxyaporphine and pronuciferine increased the glucose consumption significantly as rosiglitazone did.

  9. The inhibition of inflammatory molecule expression on 3T3-L1 adipocytes by berberine is not mediated by leptin signaling.

    Science.gov (United States)

    Choi, Bong-Hyuk; Kim, Yu-Hee; Ahn, In-Sook; Ha, Jung-Heun; Byun, Jae-Min; Do, Myoung-Sool

    2009-01-01

    In our previous study, we have shown that berberine has both anti-adipogenic and anti-inflammatory effects on 3T3-L1 adipocytes, and the anti-adipogenic effect is due to the down-regulation of adipogenic enzymes and transcription factors. Here we focused more on anti-inflammatory effect of berberine using real time RT-PCR and found it changes expressions of adipokines. We hypothesized that anti-adipogenicity of berberine mediates anti-inflammtory effect and explored leptin as a candidate mediator of this signaling. We studied this hypothesis by western blot analysis, but our results showed that berberine has no effect on the phosphorylations of STAT-3 and ERK which have important roles on leptin signaling. These results led us to conclude that the anti-inflammatory effect of berberine is not mediated by the inhibition of leptin signal transduction. Moreover, we have found that berberine down-regulates NF-kappaB signaling, one of the inflammation-related signaling pathway, through western blot analysis. Taken together, the anti-inflammatory effect of berberine is not mediated by leptin, and berberine induces anti-inflammatory effect independent of leptin signaling.

  10. Cucumis melo ssp. Agrestis var. Agrestis Ameliorates High Fat Diet Induced Dyslipidemia in Syrian Golden Hamsters and Inhibits Adipogenesis in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Kripa Shankar

    2015-01-01

    Full Text Available Background: Cucumis melo ssp. agrestis var. agrestis (CMA is a wild variety of C. melo. This study aimed to explore anti-dyslipidemic and anti-adipogenic potential of CMA. Materials and Methods: For initial anti-dyslipidemic and antihyperglycemic potential of CMA fruit extract (CMFE, male Syrian golden hamsters were fed a chow or high-fat diet with or without CMFE (100 mg/kg. Further, we did fractionation of this CMFE into two fractions namely; CMA water fraction (CMWF and CMA hexane fraction (CMHF. Phytochemical screening was done with liquid chromatography-mass spectrometry LC- (MS/MS and direct analysis in real time-MS to detect active compounds in the fractions. Further, high-fat diet fed dyslipidemic hamsters were treated with CMWF and CMHF at 50 mg/kg for 7 days. Results: Oral administration of CMFE and both fractions (CMWF and CMHF reduced the total cholesterol, triglycerides, low‐density lipoprotein cholesterol, and very low‐density lipoprotein-cholesterol levels in high fat diet-fed dyslipidemic hamsters. CMHF also modulated expression of genes involved in lipogenesis, lipid metabolism, and reverse cholesterol transport. Standard biochemical diagnostic tests suggested that neither of fractions causes any toxicity to hamster liver or kidneys. CMFE and CMHF also decreased oil-red-O accumulation in 3T3-L1 adipocytes. Conclusion: Based on these results, it is concluded that CMA possesses anti-dyslipidemic and anti-hyperglycemic activity along with the anti-adipogenic activity.

  11. Effects of aqueous extracts of raw pu-erh tea and ripened pu-erh tea on proliferation and differentiation of 3T3-L1 preadipocytes.

    Science.gov (United States)

    Cao, Zhen-Hui; Yang, Hui; He, Zhan-Long; Luo, Cheng; Xu, Zhi-Qiang; Gu, Da-Hai; Jia, Jun-Jing; Ge, Chang-Rong; Lin, Qiu-Ye

    2013-08-01

    Pu-erh tea has shown anti-obesity effects but little is known about its effect on proliferation and differentiation of preadipocytes. This study investigated the effects of the aqueous extracts of raw pu-erh tea and ripened pu-erh tea on proliferation and differentiation of murine 3T3-L1 preadiopocytes. We examined dose and time effects of both aqueous extracts on proliferation of 3T3-L1 preadipocytes. The contents of triglycerides in cytoplasm and the mRNA expression of critical transcriptional factors involved in differentiation were determined. Cytotoxicity and apoptosis rate of preadipocytes by pu-erh tea extracts treatment were test for toxic and pro-apoptotic effects. Both aqueous extracts of pu-erh tea inhibited the proliferation of 3T3-L1 preadipocytes at the selected time points. At lower concentration of raw pu-erh tea extracts (less than 300 µg/ml) and ripened pu-erh tea extracts (less than 350 µg/ml), no significant cytotoxic and pro-apoptotic were observed. Ripened pu-erh tea was more effective with lower IC50 than raw pu-erh tea. Both extracts suppressed the differentiation and down-regulated the gene expression of peroxisome proliferator-activated receptor-γ and CCAAT/enhancer binding proteins-α. Therefore, these results indicate that both aqueous extracts of pu-erh tea can inhibit proliferation and differentiation with ripened pu-erh tea more potent. Polyphenol rich in both extracts may play a role in the inhibition of proliferation and differentiation of 3T3-L1 preadipocytes.

  12. Effect of Liupao Tea on Glucose and Lipid Metabolism in Palmitate-induced Insulin Resistance 3T3-L1 Adipocytes%六堡茶对胰岛素抵抗3T3-L1脂肪细胞糖脂代谢的影响

    Institute of Scientific and Technical Information of China (English)

    滕翠琴; 刘仲华; 龚受基; 彭雨轩; 马蕊

    2014-01-01

    To investigate the effect of Liupao tea on glucose and lipid metabolism in palmitate-induced insulin resistance 3T3-L1 adipocytes. Glucose intake decreased and insulin resistance was induced in 3T3-L1 adipocytes after incubation in 1mmol/L palmitate for 24 hours. Compared with the model group, the concentrations of glucose and NEFA decreased along with the increase of Liupao tea concentrations in medium. The mRNA expression of key enzyme for glycolysis of glucose (Hexokinase, 6-phosphofructokinase-1, and pyruvatekinase) and the Lipid-Metabolism-Related Enzymes (Sn-glycerol-3-phosphate acyltransferase) significantly increased in the Liupao tea groups comparing with model sample. Meanwhile the glucose transporter 4 mRNA expression was also increased and the acetyl-CoA carboxylase, protein tyrosine phosphatase 1B mRNA expression was decreased. Low doses of Liupao tea reduced carnitine palmitoyl transferase I mRNA expression while high doses significantly raised it. Research proved that Liupao tea were able to increase glucose uptake and activate the glucolipid metabolic pathways.Liupao tea showed the effect of improving insulin resistance induced by palmitate in 3T3 -L1 adipocytes.%采用软脂酸诱导3T3-L1脂肪细胞胰岛素抵抗后,给予不同剂量的六堡茶水提物治疗,探讨六堡茶对胰岛素抵抗3T3-L1脂肪细胞的糖脂代谢的影响。结果表明,1 mmol·L-1的软脂酸对3T3-L1脂肪细胞作用24 h后,葡萄糖摄取量减少,建立胰岛素抵抗模型。六堡茶对模型细胞干预24 h 后,与模型组比较,细胞培养基的残留液中葡萄糖和游离脂肪酸的含量降低;对糖脂代谢相关关键酶(己糖激酶、6-磷酸果糖激酶-1、丙酮酸激酶、甘油-3-磷酸酰基转移酶)和葡萄糖转运子基因表达都有上调作用,并下调乙酰辅酶 A 羧化酶、蛋白酪氨酸磷酸酯酶1B基因表达。低剂量的六堡茶下调肉碱脂酰转移酶-I基因的表达,但是高剂量能

  13. Regulation of apelin and its receptor expression in adipose tissues of obesity rats with hypertension and cultured 3T3-L1 adipocytes.

    Science.gov (United States)

    Wu, Hongxian; Cheng, Xian Wu; Hao, Changning; Zhang, Zhi; Yao, Huali; Murohara, Toyoaki; Dai, Qiuyan

    2014-01-01

    The apelin/APJ system has been implicated in obesity-related hypertension. We investigated the mechanism responsible for the pathogenesis of obesity-related hypertension with a special focus on the crosstalk between AngII/its type 1 receptor (AT1R) signaling and apelin/APJ expression. Sprague-Dawley rats fed a high-fat (obesity-related hypertension, OH) or normal-fat diet (NF) for 15 weeks were randomly assigned to one of two groups and administered vehicle or perindopril for 4 weeks. Compared to the NF rats, the OH rats showed lower levels of plasma apelin and apelin/APJ mRNAs of perirenal adipose tissues, and these changes were restored by perindopril. Administration of the AT1R antagonist olmesartan resulted in the restoration of the reduction of apelin and APJ expressions induced by AngII for 48 h in 3T3-L1 adipocytes. Among several inhibitors for extracellular signal-regulated kinases 1/2 (ERK1/2) PD98059, p38 mitogen-activated protein kinase (p38MAPK) SB203580 and phosphatidylinositol 3-kinase (PI3K) LY294002, the latter showed an additive effect on AngII-mediated inhibitory effects. In addition, the levels of p-Akt, p-ERK and p38MAPK proteins were decreased by long-term treatment with AngII (120 min), and these changes were restored by Olmesartan. Apelin/APJ appears to be impaired in obesity-related hypertension. The AngII inhibition-mediated beneficial effects are likely attributable, at least in part, to restoration of p38/ERK-dependent apelin/APJ expression in diet-induced obesity-related hypertension.

  14. Inhibitory effects of harpagoside on TNF-α-induced pro-inflammatory adipokine expression through PPAR-γ activation in 3T3-L1 adipocytes.

    Science.gov (United States)

    Kim, Tae Kon; Park, Kyoung Sik

    2015-12-01

    Obesity is closely associated with increased production of pro-inflammatory adipokines, including interleukin (IL)-6, plasminogen activator inhibitor (PAI)-1, and adipose-tissue-derived monocyte chemoattractant protein (MCP)-1, which contribute to chronic and low-grade inflammation in adipose tissue. Harpagoside, a major iridoid glycoside present in devil's claw, has been reported to show anti-inflammatory activities by suppression of lipopolysaccharide (LPS)-induced production of inflammatory cytokines in murine macrophages. The present study is aimed to investigate the effects of harpagoside on both tumor necrosis factor (TNF)-α-induced inflammatory adipokine expression and its underlying signaling pathways in differentiated 3T3-L1 cells. Harpagoside significantly inhibited TNF-α-induced mRNA synthesis and protein production of the atherogenic adipokines including IL-6, PAI-1, and MCP-1. Further investigation of the molecular mechanism revealed that pretreatment with harpagoside activated peroxisome proliferator-activated receptor (PPAR)-γ. These findings suggest that the clinical application of medicinal plants which contain harpagoside may lead to a partial prevention of obesity-induced atherosclerosis by attenuating inflammatory responses.

  15. Aucubin, a naturally occurring iridoid glycoside inhibits TNF-α-induced inflammatory responses through suppression of NF-κB activation in 3T3-L1 adipocytes.

    Science.gov (United States)

    Park, Kyoung Sik

    2013-06-01

    Obesity is closely associated with a state of chronic, low-grade inflammation characterized by abnormal cytokine production and activation of inflammatory signaling pathways in adipose tissue. Tumor necrosis factor (TNF)-α is chronically elevated in adipose tissues of obese rodents and humans. Increased levels of TNF-α are implicated in the induction of atherogenic adipokines, such as plasminogen activator inhibitor (PAI)-1, adipose-tissue-derived monocyte chemoattractant protein (MCP)-1, and interleukin (IL)-6. Aucubin, an iridoid glycoside existing in medicinal plants, has been reported to show an anti-inflammatory activity by suppression of TNF-α production in murine macrophages. The present study is aimed to investigate the effects of aucubin on TNF-α-induced atherogenic changes of the adipokines in differentiated 3T3-L1 cells. Aucubin significantly inhibited TNF-α-induced secretion and mRNA synthesis of the atherogenic adipokines including PAI-1, MCP-1, and IL-6. Further investigation of the molecular mechanism revealed that pretreatment with aucubin suppressed extracellular signal-regulated kinase (ERK) activation, inhibitory kappa Bα (IκBα) degradation, and subsequent nuclear factor kappa B (NF-κB) activation. These findings suggest that aucubin may improve obesity-induced atherosclerosis by attenuating TNF-α-induced inflammatory responses.

  16. Doses of Quercetin in the Range of Serum Concentrations Exert Delipidating Effects in 3T3-L1 Preadipocytes by Acting on Different Stages of Adipogenesis, but Not in Mature Adipocytes

    Directory of Open Access Journals (Sweden)

    Itziar Eseberri

    2015-01-01

    Full Text Available Scope. To determine whether doses of quercetin in the range of serum concentrations exert any effect on triacylglycerol accumulation in maturing preadipocytes and mature adipocytes. The influence on the expression of adipogenic markers as well as on gene expression and activity of enzymes involved in triacylglycerol metabolism were assessed. Methods and Results. 3T3-L1 preadipocytes were treated during differentiation and mature adipocytes for 24 hours with low doses (0.1–10 µM of quercetin. Triacylglycerol content in both cell types and free fatty acid and glycerol in the incubation medium of mature adipocytes were measured spectrophotometrically. Gene and protein expression were assessed by RT-PCR and Western blot. LPL and FAS activities were quantified. During differentiation quercetin reduced triacylglycerol content at doses from 0.5 to 10 µM. 1 µM of quercetin reduced C/EBPβ gene expression, SREBP1 mature protein levels, and PPARγ gene expression. 10 µM of quercetin reduced LPL gene expression and PPARγ and SREBP1c expression. In mature adipocytes, only 10 µM of quercetin reduced triacylglycerol content. Lipogenic FAS expression and activity were reduced at this dose. Conclusion. Quercetin, in the range of serum concentrations, is able to inhibit adipogenesis, but higher doses, at least 10 µM, are needed to reduce fat accumulation in mature adipocytes.

  17. Effect of Catalpol 、Berberine and Their Combination on The Glycometabolism and Differentiation of 3T3-L1 Adipocytes%梓醇和小檗碱及其配伍对脂肪细胞糖代谢和分化的影响

    Institute of Scientific and Technical Information of China (English)

    刘芳芳; 杨明玮

    2014-01-01

    目的:观察中药提取物梓醇和小檗碱及其配伍对3T3-L1脂肪细胞增殖,葡萄糖代谢及细胞分化的影响,初步探讨药物改善胰岛素抵抗(IR)的可能机制.方法:采用四甲基偶氮唑盐(MTT)方法检测3T3-L1前脂肪细胞的增殖;以葡萄糖氧化酶法检测药物干预后培养液中葡萄糖消耗量,并以油红O染色检测细胞分化过程中胞浆脂质的堆积.结果:与对照组比较,小檗碱组使前脂肪细胞MTT值增加29%,而小檗碱加梓醇组、梓醇组均无差异;梓醇、小檗碱及其配伍组分别使成熟脂肪细胞的葡萄糖消耗增加64%、76.5%和98%;小檗碱处理脂肪细胞胞浆脂质堆积明显低于对照组,梓醇组与对照组无差异.结论:小檗碱能促进前脂肪细胞增殖,增加葡萄糖消耗,抑制脂肪细胞分化;梓醇能促进葡萄糖吸收;其作用均不依赖胰岛素的存在.直接增加葡萄糖的吸收可能是这两种药物降低血糖,改善胰岛素抵抗的机制之一.

  18. 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

    Directory of Open Access Journals (Sweden)

    Xiang-Zhu Xie

    2016-01-01

    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/PKA-C/EBPβ signaling pathway may participate in it.

  19. Heat Shock Protein Augmentation of Angelica gigas Nakai Root Hot Water Extract on Adipogenic Differentiation in Murine 3T3-L1 Preadipocytes.

    Science.gov (United States)

    Lumbera, Wenchie Marie L; Dela Cruz, Joseph; Yang, Seung-Hak; Hwang, Seong Gu

    2016-03-01

    There is a high association of heat shock on the alteration of energy and lipid metabolism. The alterations associated with thermal stress are composed of gene expression changes and adaptation through biochemical responses. Previous study showed that Angelica gigas Nakai (AGN) root extract promoted adipogenic differentiation in murine 3T3-L1 preadipocytes under the normal temperature condition. However, its effect in heat shocked 3T3-L1 cells has not been established. In this study, we investigated the effect of AGN root hot water extract in the adipogenic differentiation of murine 3T3-L1 preadipocytes following heat shock and its possible mechanism of action. Thermal stress procedure was executed within the same stage of preadipocyte confluence (G0) through incubation at 42°C for one hour and then allowed to recover at normal incubation temperature of 37°C for another hour before AGN treatment for both cell viability assay and Oil Red O. Cell viability assay showed that AGN was able to dose dependently (0 to 400 μg/mL) increase cell proliferation under normal incubation temperature and also was able to prevent cytotoxicity due to heat shock accompanied by cell proliferation. Confluent preadipocytes were subjected into heat shock procedure, recovery and then AGN treatment prior to stimulation with the differentiation solution. Heat shocked preadipocytes exhibited reduced differentiation as supported by decreased amount of lipid accumulation in Oil Red O staining and triglyceride measurement. However, those heat shocked preadipocytes that then were given AGN extract showed a dose dependent increase in lipid accumulation as shown by both evaluation procedures. In line with these results, real-time polymerase chain reaction (RT-PCR) and Western blot analysis showed that AGN increased adipogenic differentiation by upregulating heat shock protection related genes and proteins together with the adipogenic markers. These findings imply the potential of AGN in heat

  20. Effects of varying degrees of intermittent hypoxia on proinflammatory cytokines and adipokines in rats and 3T3-L1 adipocytes.

    Directory of Open Access Journals (Sweden)

    Qing He

    Full Text Available OBJECTIVES: Intermittent hypoxia (IH, resulted from recurring episodes of upper airway obstruction, is the hallmark feature and the most important pathophysiologic pathway of obstructive sleep apnea (OSA. IH is believed to be the most important factor causing systemic inflammation. Studies suggest that insulin resistance (IR is positively associated with OSA. In this study, we hypothesized that the recurrence of IH might result in cellular and systemic inflammation, which was manifested through the levels of proinflammatory cytokines and adipokines after IH exposure, and because IR is linked with inflammation tightly, this inflammatory situation may implicate an IR status. METHODS: We developed an IH 3T3-L1 adipocyte and rat model respectively, recapitulating the nocturnal oxygen profile in OSA. In IH cells, nuclear factor kappa B (NF-κB DNA binding reactions, hypoxia-inducible factor-1α (HIF-1α, glucose transporter-1 (Glut-1, necrosis factor alpha (TNF-α, interleukin (IL -6, leptin, adiponectin mRNA transcriptional activities and protein expressions were measured. In IH rats, blood glucose, insulin, TNF-α, IL-6, leptin and adiponectin levels were analyzed. RESULTS: The insulin and blood glucose levels in rats and NF-κB DNA binding activities in cells had significantly statistical results described as severe IH>moderate IH>mild IH>sustained hypoxia>control. The mRNA and protein levels of HIF-1α and Glut-1 in severe IH group were the highest. In cellular and animal models, both the mRNA and protein levels of TNF-α, IL-6 and leptin were the highest in severe IH group, when the lowest in severe IH group for adiponectin. CONCLUSIONS: Oxidative stress and the release of pro-inflammatory cytokines/adipokines, which are the systemic inflammatory markers, are associated with IH closely and are proportional to the severity of IH. Because IR and glucose intolerance are linked with inflammation tightly, our results may implicate the clinical

  1. OM2, a Novel Oligomannuronate-Chromium(III Complex, Promotes Mitochondrial Biogenesis and Lipid Metabolism in 3T3-L1 Adipocytes via the AMPK-PGC1α Pathway.

    Directory of Open Access Journals (Sweden)

    Jiejie Hao

    Full Text Available In our previous studies, we prepared novel oligomannuronate-chromium(III complexes (OM2, OM4 from marine alginate, and found that these compounds sensitize insulin action better than oligomannuronate(OM, chromium, and metformin in C2C12 skeletal muscle cells. In the present study, we studied their effects on mitochondrial biogenesis, lipid metabolism, and the underlying molecular mechanisms in differentiated 3T3-L1 adipocytes.We firstly used the pGL3-PGC1α and pGL3-ATGL promoter plasmids to compare their effects on PGC1α and ATGL transcription activities. Then mitochondrial biogenesis was quantified by transmission electron microscopy and MitoTracker staining. Mitochondrial oxygen consumption and fatty acid oxidation were measured by an oxygen biosensor system and ³H-labelled water scintillation. The mitochondrial DNA and mRNA involved in mitochondrial biogenesis and lipid oxidation were evaluated by real-time PCR. AMPK together with other protein expression levels were measured by western blotting. The inhibitor compound C and siRNA of PGC1α were used to inhibit the OM2-induced AMPK-PGC1α signaling pathway. And we found that OM2 stimulated AMPK-PGC1α pathway in the 3T3-L1 adipocytes, which were correlated with induced mitochondrial biogenesis, improved mitochondrial function, and reduced lipid accumulation by enhanced fatty acid β-oxidation and augmented ATGL protein expression.Our data indicated that the marine oligosaccharide-derived OM2 might represent a novel class of molecules that could be useful for type 2 diabetes prevention and treatment by up-regulating AMPK-PGC1α signaling pathway.

  2. Effect of conjugated linoleic acid supplementation on lipoprotein lipase activity in 3T3-L1 adipocyte culture Efeito da suplementação com ácido linoléico conjugado sobre a atividade da lípase lipoprotéica em cultura de adipócitos 3T3-L1

    Directory of Open Access Journals (Sweden)

    Adriana Prais Botelho

    2009-10-01

    Full Text Available Supplementation with conjugated linoleic acid may reduce fat body mass and increase lean body mass in various species. Some studies have demonstrated that conjugated linoleic acid reduces body fat, in part, by inhibiting the activity of lipoprotein lipase in adipocytes. The objective of this work was to study the effect of conjugated linoleic acid supplementation on lipoprotein lipase activity in 3T3-L1 adipocyte culture. 3T3-L1 adipocytes received linoleic acid (group C or conjugated linoleic acid (group AE, supplemented with AdvantEdge® CLA, and group CO, supplemented with CLA One® in concentrations of 1 mmol/L. Heparin-releasable lipoprotein lipase activity was analyzed by means of a 3T3-L1 adipocyte culture. After 7 days, heparin-releasable lipoprotein lipase activity was lower in the groups AE and CO supplemented with conjugated linoleic acid. These results suggest that one of the mechanisms by which CLA is capable of reducing body fat is by reducing lipoprotein lipase activity.A suplementação com ácido linoléico conjugado pode reduzir a gordura corporal e aumentar a massa magra em diferentes espécies. Alguns estudos têm demonstrado que o ácido linoléico conjugado reduz a gordura corporal, por meio da inibição da atividade de lípase lipoprotéica em adipócitos. O objetivo deste estudo foi avaliar o efeito da suplementação com uma mistura de isômeros do ácido linoléico conjugado sobre a atividade da lípase lipoprotéica em cultura de adipócitos 3T3-L1. Os adipócitos 3T3-L1 receberam ácido linoléico (grupo controle ou ácido linoléico conjugado (grupo AE, suplementado com AdvantEdge® CLA, e grupo CO, suplementado com CLA One® na concentração de 1 mmol/L. A atividade de lípase lipoprotéica livre de heparina foi analisada pela média da cultura de adipócitos. Após 7 dias, a atividade da lípase lipoprotéica livre de heparina mostrou menores valores nos grupos AE e CO, suplementados com ácido linol

  3. G9a is transactivated by C/EBPβ to facilitate mitotic clonal expansion during 3T3-L1 preadipocyte differentiation.

    Science.gov (United States)

    Li, Shu-Fen; Guo, Liang; Qian, Shu-Wen; Liu, Yuan; Zhang, You-You; Zhang, Zhi-Chun; Zhao, Yue; Shou, Jian-Yong; Tang, Qi-Qun; Li, Xi

    2013-05-01

    In 3T3-L1 preadipocyte differentiation, the CCAAT/enhancer-binding protein-β (C/EBPβ) is an important early transcription factor that activates cell cycle genes during mitotic clonal expansion (MCE), sequentially activating peroxisome proliferator-activated receptor-γ (PPARγ) and C/EBPα during terminal differentiation. Although C/EBPβ acquires its DNA binding activity via dual phosphorylation at about 12-16 h postinduction, the expression of PPARγ and C/EBPα is not induced until 36-72 h. The delayed expression of PPARγ and C/EBPα ensures the progression of MCE, but the mechanism responsible for the delay remains elusive. We provide evidence that G9a, a major euchromatic methyltransferase, is transactivated by C/EBPβ and represses PPARγ and C/EBPα through H3K9 dimethylation of their promoters during MCE. Inhibitor- or siRNA-mediated G9a downregulation modestly enhances PPARγ and C/EBPα expression and adipogenesis in 3T3-L1 preadipocytes. Conversely, forced expression of G9a impairs the accumulation of triglycerides. Thus, this study elucidates an epigenetic mechanism for the delayed expression of PPARγ and C/EBPα.

  4. Uncoupling of 3T3-L1 gene expression from lipid accumulation during adipogenesis.

    Science.gov (United States)

    Temple, Karla A; Basko, Xheni; Allison, Margaret B; Brady, Matthew J

    2007-02-06

    Adipocyte differentiation comprises altered gene expression and increased triglyceride storage. To investigate the interdependency of these two events, 3T3-L1 cells were differentiated in the presence of glucose or pyruvate. All adipocytic proteins examined were similarly increased between the two conditions. In contrast, 3T3-L1 adipocytes differentiated with glucose exhibited significant lipid accumulation, which was largely suppressed in the presence of pyruvate. Subsequent addition of glucose to the latter cells restored lipid accumulation and acute rates of insulin-stimulated lipogenesis. These data indicate that extracellular energy is required for induction of adipocytic proteins, while only glucose sustained the parallel increase in triglyceride storage.

  5. Effect of globular domain of adiponectin on glycolysis in 3T3-L1 adipocytes%gAd对3T3-L1脂肪细胞葡萄糖分解代谢关键酶表达的影响

    Institute of Scientific and Technical Information of China (English)

    王彦; 廉如; 陈思思; 高婷婷; 吴彬; 陈显久; 杨静

    2013-01-01

    目的 通过检测3T3-L1脂肪细胞内糖酵解关键酶的表达情况,探讨课题组前期制备的脂联素球状结构域(gAd)对3T3-L1脂肪细胞葡萄糖分解代谢的影响.方法 用课题组前期制备的gAd(质量浓度依次为10、50、100、300、1 000 ng/mL)干预分化成熟的3T3-L1脂肪细胞,干预结束后以实时荧光定量聚合酶链反应(RT-PCR)法检测各组细胞糖酵解关键酶包括己糖激酶、6-磷酸果糖激酶-1及丙酮酸激酶转录表达情况,以Western blot法检测各组细胞丙酮酸激酶翻译表达情况.结果 gAd干预组(5个浓度)己糖激酶的转录表达[(2.02±0.16)、(3.47±0.29)、(4.22±0.33)、(5.83±0.45)、(6.65±0.51)倍]显著高于对照组(1.00±0.00)(F=125.789,P<0.001),6-磷酸果糖激酶-1和丙酮酸激酶的转录表达也显著高于对照组(F分别为85.399和113.661,P均<0.001),同时丙酮酸激酶的翻译表达(12.430%±0.800%、1.700%±3.010%、26.570%±1.114%、52.600%±2.910%、71.130%±10.600%)也显著高于对照组(8.000%±1.610%)(F=161.007,P<0.01).结论 gAd促进3T3-L1脂肪细胞摄取胞外葡萄糖的同时,激活了细胞内葡萄糖的分解供能代谢途径.%Objective To investigate the effect of globular domain of adiponectin(gAd) on glycolysis in 3T3-L1 adipocytes by detecting expression of the key enzymes in the process of glycolysis. Methods Matured 3T3-L1 adipocytes were intervened with pre-prepared gAd(mass concentrations of 10, 50, 100, 300, and 1 000 ng/mL, respectively) and the expression of several key enzymes for glycolysis such as hexokinase, 6-phosphofructokinase 1 and pyruvate kinase were determined by real-time polymerase chain reaction(RT-PCR). Pyruvate kinase protein level was also determined by Western blot. Results The level of hexokinase mRNA in the gAd intervention group was significantly higher than those of the control group[(2.02 ± 0.16), (3.47 ± 0.29), (4.22 ± 0.33), (5.83 ± 0.45) and (6.65 ± 0

  6. Ascofuranone stimulates expression of adiponectin and peroxisome proliferator activated receptor through the modulation of mitogen activated protein kinase family members in 3T3-L1, murine pre-adipocyte cell line

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Young-Chae, E-mail: ycchang@cu.ac.kr [Research Institute of Biomedical Engineering and Department of Medicine, Catholic University of Daegu School of Medicine, Daegu 705-718 (Korea, Republic of); Cho, Hyun-Ji, E-mail: hjcho.dr@gmail.com [Research Institute of Biomedical Engineering and Department of Medicine, Catholic University of Daegu School of Medicine, Daegu 705-718 (Korea, Republic of)

    2012-06-08

    Highlights: Black-Right-Pointing-Pointer Ascofuranone increases expression of adiponectin and PPAR{gamma}. Black-Right-Pointing-Pointer Inhibitors for MEK and JNK increased the expression of adiponectin and PPAR{gamma}. Black-Right-Pointing-Pointer Ascofuranone significantly suppressed phosho-ERK, while increasing phospho-p38. -- Abstract: Ascofuranone, an isoprenoid antibiotic, was originally isolated as a hypolipidemic substance from a culture broth of the phytopathogenic fungus, Ascochyta visiae. Adiponectin is mainly synthesized by adipocytes. It relieves insulin resistance by decreasing the plasma triglycerides and improving glucose uptake, and has anti-atherogenic properties. Here, we found that ascofuranone increases expression of adiponectin and PPAR{gamma}, a major transcription factor for adiponectin, in 3T3-L1, murine pre-adipocytes cell line, without promoting accumulation of lipid droplets. Ascofuranone induced expression of adiponectin, and increases the promoter activity of adiponectin and PPRE, PPAR response element, as comparably as a PPAR{gamma} agonist, rosiglitazone, that stimulates lipid accumulation in the preadipocyte cell line. Moreover, inhibitors for MEK and JNK, like ascofuranone, considerably increased the expression of adiponectin and PPAR{gamma}, while a p38 inhibitor significantly suppressed. Ascofuranone significantly suppressed ERK phosphorylation, while increasing p38 phosphorylation, during adipocyte differentiation program. These results suggest that ascofuranone regulates the expression of adiponectin and PPAR{gamma} through the modulation of MAP kinase family members.

  7. Effects of Berberine on Inflammatory Factors, Adipokines and Fatty Acid Metabolism in 3T3-L1 Adipocytes%小檗碱对3T3-L1脂肪细胞炎症因子、脂肪因子及脂肪酸代谢的影响

    Institute of Scientific and Technical Information of China (English)

    李萍; 岳晶晶; 张达; 牛文彦; 何庆

    2014-01-01

    Objective To observe the effects of berberine on inflammatory factors, adipokines and fatty acid metabo-lism in 3T3-L1 adipocytes, and to investigate the molecular mechanism underlying berberine’s role of improving insulin re-sistance. Methods mRNA level of inflammatory molecules, adipokines, key enzymes and protein in fatty acid metabolism in 3T3-L1 cells were determined by quantitative real time polymerase chain reaction (qRT-PCR) after cells were treated with different concentrations of berberine (0, 5, 10, 20, 40μmol/L) for 24 hours and with 10μmol/L berberine at different du-rations (0,4,8,24,48 h). These factors mainly included interleukin-6 (IL-6), tumor necrosis factor-α(TNF-α), leptin, adipo-nectin, visfatin, fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), adipose triglyceride lipase (ATGL) and adipocyte fatty acid binding protein (AFABP). Results In 3T3-L1 adipocytes, transcription level of IL-6, TNF-α, leptin, FAS, AT-GL, AFABP reduced with addition of berberine dosage at 10~40μmol/L(P0.05). Tran-scription level of IL-6, TNF-α, leptin, AFABP, ATGL, FAS decreased with time after 10μmol/L berberine intervention (8-48 h) compared with the control group(P 0.05). Conclusion mRNA level of inflammatory factors, adipokines, key enzymes and protein in fatty acid metabolism in 3T3-L1 adipocytes can be affected by berberine and this effect depend on its dose and time . This might be the mechanisms underlying berber- ine to improve insulin resistance.%目的:观察小檗碱对3T3-L1脂肪细胞炎症因子、脂肪因子及脂肪酸代谢的影响,探讨小檗碱改善胰岛素抵抗的分子机制。方法3T3-L1脂肪细胞经不同浓度小檗碱(0、5、10、20和40μmol/L)处理24 h后,采用qRT-PCR技术检测相关因子mRNA表达水平,包括白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、瘦素、脂联素和内脂素以及脂肪酸合酶(FAS)、乙酰辅酶A羧化酶(ACC)、脂肪组织甘油三酯

  8. The anti-obesity effects of a tuna peptide on 3T3-L1 adipocytes are mediated by the inhibition of the expression of lipogenic and adipogenic genes and by the activation of the Wnt/β-catenin signaling pathway.

    Science.gov (United States)

    Kim, Young-Min; Kim, In-Hye; Choi, Jeong-Wook; Lee, Min-Kyeong; Nam, Taek-Jeong

    2015-08-01

    The differentiation of 3T3-L1 cells into adipocytes involves the activation of an organized system of obesity-related genes, of which those encoding CCAAT/enhancer-binding proteins (C/EBPs) and the Wnt-10b protein may play integral roles. In a previous study of ours, we found that a specific peptide found in tuna (sequence D-I-V-D-K-I-E-I; termed TP-D) inhibited 3T3-L1 cell differentiation. In the present study, we observed that the expression of expression of C/EBPs and Wnt-10b was associated with obesity. The initial step of 3T3-L1 cell differentiation involved the upregulation of C/EBP-α expression, which in turn activated various subfactors. An upstream effector of glycogen synthase kinase-3β (GSK-3β) inhibited Wnt-10b expression in 3T3-L1 adipocytes. In a previous study of ours, we sequenced the tuna peptide via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and quadrupole time-of-flight mass spectrometry (Q-TOF MS/MS) and confirmed the anti-obesity effects thereof in 3T3-L1 adipocytes. In the present study, we demonstrate that TP-D inhibits C/EBP and promotes Wnt-10b mRNA expression, thus activating the Wnt pathway. The inhibition of lipid accumulation was measured using a glucose and triglyceride (TG) assay. Our results confirmed that TP-D altered the expression levels of C/EBP-related genes in a dose-dependent manner and activated the Wnt signaling pathway. In addition, we confirmed that total adiponectin and high-molecular weight (HMW) adiponectin levels were reduced by treatment with TP-D. These data indicate that TP-D inhibits adipocyte differentiation through the inhibition of C/EBP genes and the subsequent activation of the Wnt/β-catenin signaling pathway.

  9. The anti-obesity effects of a tuna peptide on 3T3-L1 adipocytes are mediated by the inhibition of the expression of lipogenic and adipogenic genes and by the activation of the Wnt/β-catenin signaling pathway

    Science.gov (United States)

    KIM, YOUNG-MIN; KIM, IN-HYE; CHOI, JEONG-WOOK; LEE, MIN-KYEONG; NAM, TAEK-JEONG

    2015-01-01

    The differentiation of 3T3-L1 cells into adipocytes involves the activation of an organized system of obesity-related genes, of which those encoding CCAAT/enhancer-binding proteins (C/EBPs) and the Wnt-10b protein may play integral roles. In a previous study of ours, we found that a specific peptide found in tuna (sequence D-I-V-D-K-I-E-I; termed TP-D) inhibited 3T3-L1 cell differentiation. In the present study, we observed that the expression of expression of C/EBPs and Wnt-10b was associated with obesity. The initial step of 3T3-L1 cell differentiation involved the upregulation of C/EBP-α expression, which in turn activated various subfactors. An upstream effector of glycogen synthase kinase-3β (GSK-3β) inhibited Wnt-10b expression in 3T3-L1 adipocytes. In a previous study of ours, we sequenced the tuna peptide via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and quadrupole time-of-flight mass spectrometry (Q-TOF MS/MS) and confirmed the anti-obesity effects thereof in 3T3-L1 adipocytes. In the present study, we demonstrate that TP-D inhibits C/EBP and promotes Wnt-10b mRNA expression, thus activating the Wnt pathway. The inhibition of lipid accumulation was measured using a glucose and triglyceride (TG) assay. Our results confirmed that TP-D altered the expression levels of C/EBP-related genes in a dose-dependent manner and activated the Wnt signaling pathway. In addition, we confirmed that total adiponectin and high-molecular weight (HMW) adiponectin levels were reduced by treatment with TP-D. These data indicate that TP-D inhibits adipocyte differentiation through the inhibition of C/EBP genes and the subsequent activation of the Wnt/β-catenin signaling pathway. PMID:26046125

  10. Bavachin from Psoralea corylifolia Improves Insulin-Dependent Glucose Uptake through Insulin Signaling and AMPK Activation in 3T3-L1 Adipocytes

    Directory of Open Access Journals (Sweden)

    Hyejin Lee

    2016-04-01

    Full Text Available The fruit of Psoralea corylifolia L. (Fabaceae (PC, known as “Bo-Gol-Zhee” in Korea has been used as traditional medicine. Ethanol and aqueous extracts of PC have an anti-hyperglycemic effect by increasing plasma insulin levels and decreasing blood glucose and total plasma cholesterol levels in type 2 diabetic rats. In this study, we purified six compounds from PC and investigated their anti-diabetic effect. Among the purified compounds, bavachin most potently accumulated lipids during adipocyte differentiation. Intracellular lipid accumulation was measured by Oil Red-O (ORO cell staining to investigate the effect of compounds on adipogenesis. Consistently, bavachin activated gene expression of adipogenic transcriptional factors, proliferator-activated receptorγ (PPARγ and CCAAT/enhancer binding protein-α (C/EBPα. Bavachin also increased adiponectin expression and secretion in adipocytes. Moreover, bavachin increased insulin-induced glucose uptake by differentiated adipocytes and myoblasts. In differentiated adipocytes, we found that bavachin enhanced glucose uptake via glucose transporter 4 (GLUT4 translocation by activating the Akt and 5′AMP-activated protein kinase (AMPK pathway in the presence or absence of insulin. These results suggest that bavachin from Psoralea corylifolia might have therapeutic potential for type 2 diabetes by activating insulin signaling pathways.

  11. The combination of resveratrol and CLA does not increase the delipidating effect of each molecule in 3T3-L1 adipocytes La combinación de resveratrol y CLA no incrementa el efecto hipolipemiante de cada molécula en adipocitos 3T3-L1

    Directory of Open Access Journals (Sweden)

    A. Lasa

    2011-10-01

    Full Text Available Introduction: Trans-10, cis-12 conjugated linoleic acid (CLA and resveratrol have been shown to reduce TG content in cultured 3T3-L1 adipocyte acting on different pathways. In recent years, the method of simultaneously targeting several signal transduction pathways with multiple natural products in order to achieve additive or synergistic effects has been tested. However, the combined effect of both molecules on lipid metabolism has not been described before. Objective: The aim of the present work was to analyze the effect of the combination of trans-10, cis-12 CLA and resveratrol on TG accumulation as well as on FAS, HSL and ATGL expression in 3T3-L1 mature adipocytes, in order to assess a potential interaction between both molecules. Methods: For this purpose, 3T3-L1 mature adipocytes were treated with the two molecules, both separately and combined, in 10 and 100 μM for 20 hours. TG content and FAS, ATGL and HSL expression were measured by spectrophotometry and Real Time RT-PCR respectively. Results: Both doses of CLA and 100 M resveratrol decreased TG content in mature adipocytes. The combination of both molecules reduced TG accumulation to the same extent as each one separately. No change in FAS and HSL mRNA levels after CLA and resveratrol treatment was observed. ATGL was not modified by CLA but it was increased by resveratrol and by the combination. This combination did not increase the effect caused by resveratrol on its own. Conclusion: Lipolysis increase via ATGL is involved in the TG reduction induced by resveratrol and the combination of both molecules. The combination of these two molecules does not increase the efficacy of each molecule separately in mature adipocytes and thus it does not represent an advantage for obesity treatment or prevention.Introducción: Se ha demostrado que el ácido linoleico trans-10, cis-12 conjugado (ALC y el resveratrol reducen el contenido de TG en el adipocito 3T3-L1 cultivado actuando sobre

  12. Induction of mitochondrial uncoupling enhances VEGF₁₂₀ but reduces MCP-1 release in mature 3T3-L1 adipocytes: possible regulatory mechanism through endogenous ER stress and AMPK-related pathways.

    Science.gov (United States)

    Miyokawa-Gorin, Kaoru; Takahashi, Kazuto; Handa, Keiko; Kitahara, Atsuko; Sumitani, Yoshikazu; Katsuta, Hidenori; Tanaka, Toshiaki; Nishida, Susumu; Yoshimoto, Katsuhiko; Ohno, Hideki; Ishida, Hitoshi

    2012-03-01

    Although white adipocytes contain a larger number of mitochondria per cytoplasmic volume, adipocyte mitochondrial uncoupling to reduce the efficiency of ATP production on cellular function including secretory regulation of bioactive molecules such as VEGF and MCP-1 remains to be elucidated. Here we induce mitochondrial uncoupling under hypoxia-independent conditions in mature 3T3-L1 adipocytes using a metabolic uncoupler, dinitrophenol (DNP). MCP-1 release was significantly decreased by 26% (poxidative stress was observed. Treatment with thapsigargin, which can induce exogenous endoplasmic reticulum (ER) stress, clearly attenuated MCP-1 release (pmetabolic syndrome and type 2 diabetes.

  13. Pycnogenol® inhibits lipid accumulation in 3T3-L1 adipocytes with the modulation of reactive oxygen species (ROS) production associated with antioxidant enzyme responses.

    Science.gov (United States)

    Lee, Ok-Hwan; Seo, Min-Jung; Choi, Hyeon-Son; Lee, Boo-Yong

    2012-03-01

    Pycnogenol® is a group of flavonoids with antioxidant effects. Adipogenesis is the process of adipocyte differentiation. It causes the increase of lipids as well as ROS (reactive oxygen species). Lipid accumulation and ROS production were determined in 3 T3-L1 adipocyte, and the effect of Pycnogenol® was evaluated. Lipid accumulation was elevated in adipocyte treated with hydrogen peroxide, one of the ROS. Pycnogenol® showed an inhibitory effect on the lipid accumulation and ROS production during the adipogenesis. We also investigated the molecular events associated with ROS production and lipid accumulation. Our results showed that Pycnogenol® inhibited the mRNA expression of pro-oxidant enzymes, such as NOX4 (NADPH (nicotinamide adenine dinucleotide phosphate hydrogen) oxidase 4), and the NADPH-producing G6PDH (glucose-6-phosphate dehydrogenase) enzyme. In addition, Pycnogenol® suppressed the mRNA abundance of adipogenic transcription factors, PPAR-γ (peroxisome proliferator-activated receptor γ) and C/EBP-α (CCAAT/enhancer binding protein α), and their target gene, aP2 (adipocyte protein 2) responsible for fatty acid transportation. On the other hand, Pycnogenol® increased the abundance of antioxidant proteins such as Cu/Zn-SOD (copper-zinc superoxide dismutase), Mn-SOD (manganese superoxide dismutase), GPx (glutathione peroxidase) and GR (glutathione reductase). Our results suggest that Pycnogenol® inhibits lipid accumulation and ROS production by regulating adipogenic gene expression and pro-/antioxidant enzyme responses in adipocytes.

  14. Two chalcones, 4-hydroxyderricin and xanthoangelol, stimulate GLUT4-dependent glucose uptake through the LKB1/AMP-activated protein kinase signaling pathway in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ohta, Mitsuhiro; Fujinami, Aya; Kobayashi, Norihiro; Amano, Akiko; Ishigami, Akihito; Tokuda, Harukuni; Suzuki, Nobutaka; Ito, Fumitake; Mori, Taisuke; Sawada, Morio; Iwasa, Koichi; Kitawaki, Jo; Ohnishi, Katsunori; Tsujikawa, Muneo; Obayashi, Hiroshi

    2015-07-01

    4-Hydroxyderricin (4HD) and xanthoangelol (XAG) are major components of n-hexane/ethyl acetate (5:1) extract of the yellow-colored stem juice of Angelica keiskei. 4-Hydroxyderricin and XAG have been reported to increase glucose transporter 4 (GLUT4)-dependent glucose uptake in 3T3-L1 adipocytes, but the detailed mechanism of this phenomenon remains unknown. This present study was aimed at clarifying the detailed mechanism by which 4HD and XAG increase GLUT4-dependent glucose uptake in 3T3-L1 adipocytes. Both 4HD and XAG increased glucose uptake and GLUT4 translocation to the plasma membrane. 4-Hydroxyderricin and XAG also stimulated the phosphorylation of 5' adenosine monophosphate-activated protein kinase (AMPK) and its downstream target acetyl-CoA carboxylase. In addition, phosphorylation of liver kinase B1 (LKB1), which acts upstream of AMPK, was also increased by 4HD and XAG treatment. Small interfering RNA knockdown of LKB1 attenuated 4HD- and XAG-stimulated AMPK phosphorylation and suppressed glucose uptake. These findings demonstrate that 4HD and XAG can increase GLUT4-dependent glucose uptake through the LKB1/AMPK signaling pathway in 3T3-L1 adipocytes.

  15. Comparative Study of Inducing Insulin Resistance in 3T3-L1 Adipocytes with Three Different Methods%三种不同方法诱导3T3-L1脂肪细胞胰岛素抵抗的对比研究

    Institute of Scientific and Technical Information of China (English)

    陈立; 杨明炜; 库宝庆; 莫阔; 汪继敏; 王晓翠; 马聪

    2012-01-01

    Objective To observe the glucose consumption and Glut4 expression in inducing insulin resistance in 3T3 - L1 adipocytes with three methods ( dexamethasone, free fatty acids, high glucose and hyperinsulinism ) . Methods Three methods were used to induce insulin resistance in 3T3 - L1 adipocytes, and the glucose consumption and Glut4 expression were detected in each group in 12 h, 24 h, 36 h, 48 h and 60 h. Results Within 24 hours, compared with the normal control group, the glucose consumption in dexamethasone group, free fatty acids group and high glucose and hyperinsulinism group decreased significantly ( P 0. 05 ) . The glucose consumption in dexamethasone group decreased significantly compared with normal control group ( P < 0. 05 );compared with the dexamethasone group, the glucose consumption in high glucose and hyperinsulinism group decreased significantly ( P < 0. 05 ) . Compared with the normal control group, the Glut4 expression in dexamethasone group and free fatty acids group both decreased significantly ( P < 0. 01 ), and compared with dexamethasone group and free fatty acids group, the Glut4 expression in high glucose and hyperinsulinism group decreased significantly ( P < 0. 01 ) . Conclusion High glucose and hyperinsulinism can induce stronger insulin resistance in 3T3 -L1 adipocytes, and this may result from the inhibition of Glut4 expression.%目的 对比地塞米松、游离脂肪酸、高糖高胰岛素3种不同方法诱导的胰岛素抵抗3T3-L1脂肪细胞的葡萄糖消耗量及葡萄糖转运子4(Glut4)的表达.方法 采用3种不同方法诱导3T3-L1脂肪细胞产生胰岛素抵抗,分别检测各组细胞12、24、36、48、60 h时的葡萄糖消耗量及Glut4蛋白的表达.结果 在24 h内,地塞米松组、游离脂肪酸组、高糖高胰岛素组细胞的葡萄糖消耗量与正常对照组比较均显著降低,差异有统计学意义(P<0.05);在24 h以后,游离脂肪酸组葡萄糖消耗量与正常对照组比

  16. Curcumin, a Potential Inhibitor of Up-regulation of TNF-alpha and IL-6 Induced by Palmitate in 3T3-L1 Adipocytes through NF-kappaB and JNK Pathway

    Institute of Scientific and Technical Information of China (English)

    SHAO-LING WANG; YING EI; YING WEN; YAN-FENG CHEN; LI-XIN NA; SONG-TAO LI; CHANG-HAO SUN

    2009-01-01

    Objective To investigate the attenuating effect of curcumin, an anti-inflammatory compound derived from dietary spice turmeric (Curcuma longa) on the pro-inflammatory insulin-resistant state in 3T3-L1 adipocytes. Methods Glucose uptake rate was determined with the [3H] 2-deoxyglucose uptake method. Expressions of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were measured by quantitative RT-PCR analysis and ELISA. Nuclear transcription factor kappaB p65 (NF-κB p65) and mitogen-activated protein kinase (MAPKs) were detected by Western blot assay. Results The basal glucose uptake was not altered, and curcumin increased the insulin-stimulated glucose uptake in 3T3-L1 cells. Curcumin suppressed the transcription and secretion of TNF-α and IL-6 induced by palmitate in a concentration-dependent manner. Palmitate induced nuclear translocation of NF-kB. The activities of Jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase1/2 (ERK1/2) and p38MAPK decreased in the presence of curcumin. Moreover, pretreatment with SP600125 (inhibitor of JNK) instead of PD98059 or SB203580 (inhibitor of ERK 1/2 or p38MAPK, respectively) decreased the up-regulation of TNF-α induced by palmitate. Conclusion Curcumin reverses palmitate-induced insulin resistance state in 3T3-L1 adipocytes through the NF-kB and JNK pathway.

  17. Biogenesis, characterization, and the effect of vicenin-gold nanoparticles on glucose utilization in 3T3-L1 adipocytes: a bioinformatic approach to illuminate its interaction with PTP 1B and AMPK.

    Science.gov (United States)

    Chockalingam, Shivashri; Thada, Rajarajeshwari; Dhandapani, Ramesh Kumar; Panchamoorthy, Rajasekar

    2015-01-01

    This study reported the synthesis of Vicenin-2 gold nanoparticles (VN-AuNPs) and evaluated their effect on the glucose utilization efficiency of 3T3-L1 adipocytes. The VN-AuNPs were characterized by microscopic, DLS and spectral analysis. The bio-reducing efficiency of Vicenin-2 (VN) was computed and confirmed by HPLC analysis. The stability of VN-AuNPs in various physiological media was explored. The cytotoxicity and glucose uptake assays were performed in 3T3-L1 adipocytes. The docking of VN with PTP1B and AMPK was also performed. The color change and UV absorption at 537 nm preliminarily confirmed the VN reduced gold nanoparticles. The VN-AuNPs appeared as spherical particles (57 nm) and face centered cubic crystals under TEM and XRD analysis, respectively. Its zeta potential was found to be -6.53 mV. The FT-IR spectra of VN and its AuNPs confirmed its stability. The computed reducing potential of VN was similar to the extent of VN utilized during the synthesis of VN-AuNPs. The VN-AuNPs showed a remarkable stability in different physiological media. At 100 µM concentration, VN-AuNPs displayed 78.21% cell viability. A concentration dependent increase in glucose uptake was noted in 3T3-L1 adipocytes when incubated with VN-AuNPs. The docking data revealed a strong interaction of VN with the binding pockets of PTP1B and AMPK. This demonstrates that the fabricated VN-AuNPs might enhance the intracellular VN availability mediated cellular glucose utilization and this would serve as a novel nanodrug for the management of diabetes.

  18. Fisetin up-regulates the expression of adiponectin in 3T3-L1 adipocytes via the activation of silent mating type information regulation 2 homologue 1 (SIRT1)-deacetylase and peroxisome proliferator-activated receptors (PPARs).

    Science.gov (United States)

    Jin, Taewon; Kim, Oh Yoen; Shin, Min-Jeong; Choi, Eun Young; Lee, Sung Sook; Han, Ye Sun; Chung, Ji Hyung

    2014-10-29

    Adiponectin, an adipokine, has been described as showing physiological benefits against obesity-related malfunctions and vascular dysfunction. Several natural compounds that promote the expression and secretion of adipokines in adipocytes could be useful for treating metabolic disorders. This study investigated the effect of fisetin, a dietary flavonoid, on the regulation of adiponectin in adipocytes using 3T3-L1 preadipocytes. The expression and secretion of adiponectin increased in 3T3-L1 cells upon treatment with fisetin in a dose-dependent manner. Fisetin-induced adiponectin secretion was inhibited by peroxisome proliferator-activated receptor (PPAR) antagonists. It was also revealed that fisetin increased the activities of PPARs and silent mating type information regulation 2 homologue 1 (SIRT1) in a dose-dependent manner. Furthermore, the up-regulation of adiponectin and the activation of PPARs induced by fisetin were prevented by a SIRT1 inhibitor. Fisetin also promoted deacetylation of PPAR γ coactivator 1 (PGC-1) and its interaction with PPARs. SIRT knockdown by siRNA significantly decreased both adiponectin production and PPARs-PGC-1 interaction. These results provide evidence that fisetin promotes the gene expression of adiponectin through the activation of SIRT1 and PPARs in adipocytes.

  19. The Rab4 effector Rabip4 plays a role in the endocytotic trafficking of Glut 4 in 3T3-L1 adipocytes

    NARCIS (Netherlands)

    Mari, Muriel; Monzo, Pascale; Kaddai, Vincent; Keslair, Frédérique; Gonzalez, Teresa; Le Marchand-Brustel, Yannick; Cormont, Mireille

    2006-01-01

    Insulin regulates glucose uptake in the adipocytes by modulating Glut 4 localization, a traffic pathway involving the endocytic small GTPases Rab4, Rab5, and RabThe expression of the Rab4 effector Rabip4 leads to a 30% increase in glucose uptake and Glut 4 translocation in the presence of insulin, w

  20. Effects of Daizongfang Formula on the Glucose-Lipid Metabolism in 3 T3-L1 Adipocytes%代综方对3T3-L1脂肪细胞糖脂代谢的作用研究

    Institute of Scientific and Technical Information of China (English)

    朱晓云; 王朋倩; 刘喜明

    2015-01-01

    Objective To study the effects of Daizongfang formula( DZF)on the glucose-lipid me-tabolism and its possible regulatory mechanism in 3T3 -L1 adipocyte. Methods Mature adipocytes were treated with different concentration of DZF and Rosiglitazone( Ros)for 48 hours,then the concentration of glu-cose,glycerol and free fatty acid in the culture supernatant and the content of triglyceride( TG)in the cell were detected. RT-PCR was used to detect the mRNA expression of glucose transporter 4(GLUT-4),peril-ipin and hormone sensitive lipase( HSL). Results All the DZF of different concentration significantly in-creased the consumption of glucose(P﹤0. 01),increased the concentration of the glycerol in culture super-natant and reduced the accumulation of intracellular TG(P﹤0. 05 or P﹤0. 05),while it had no effect on free fatty acids(P﹥0. 05). Ros increased the consumption of glucose(P﹤0. 01),increased the accumulation of intracellular TG(P﹤0. 05),while it had no effect on glycerol and free fatty acids in the culture superna-tant(P﹥0. 05). All the DZF groups raised the mRNA expression of GLUT-4 and HSL differently,lowered the mRNA expression of Perilipin significantly(P﹤0. 01). Ros raised mRNA expression of GLUT-4,HSL, Perilipin. Conclusion DZF can change the dynamic balance of lipid synthesis and katabolism,and it can promote glucose consumption of adipocytes and increase katabolism of intracellular TG. The mechanism of DZF is different from Ros in increasing the accumulation of intracellular TG.%目的:研究代综方( DZF)对3T3-L1脂肪细胞糖脂代谢的作用及可能的调控机制。方法分化成熟的脂肪细胞给予不同浓度DZF或对照药物罗格列酮( Ros)干预48 h,检测培养上清中葡萄糖消耗量、甘油、游离脂肪酸( NEFA)浓度及胞内甘油三酯( TG)含量;应用反转录-聚合酶链反应(RT-PCR)检测葡萄糖转运体4(GLUT -4)、脂滴包被蛋白(perilipin)、激素敏感性脂肪

  1. Potassium channel tetramerisation domain containing 15 regulates preadipocyte differentiation%KCTD15基因调控3T3-L1脂肪前体细胞分化的研究

    Institute of Scientific and Technical Information of China (English)

    徐景; 赵旭; 杨莹; 徐梓辉

    2013-01-01

    Objective To study the effect of potassium channel tetramerisation domain containing 15 (KCTD15) gene on preadipocyte differentiation.Methods The expression of KCTD15 gene during 3T3-L1 preadipocyte differentiation was detected by semi-quantitative reverse transcriptase PCR.After transferring KCTD15 siRNA into the preadipocytes,the cell morphology was observed during preadipocyte differentiation by oil red O staining,and the level of triglyceride was examined by assay kit.The expression of adipogenesis genes,peroxisome proliferator-activated receptor (PPAR) γ,CCAAT/enhancer-binding protein (C/EBP) α,C/EBPβ and C/EBPδ was detected by semi-quantitative reverse transcriptase PCR.Results The expression of KCTD15 gene was decreased during 3T3-L1 cell differentiation.KCTD15 gene knockdown inhibited the differentiation and lipid accumulation of 3T3-L1 cells,and there was no significant change in the expression of PPARγ,C/EBPα,C/EBPβ and C/EBPδ.Conclusion KCTD15 gene deficiency leads to the inhibition of 3T3-L1 preadipocyte differentiation at early stage.%目的 探讨含钾通道四聚化结构域15(KCTD15)基因在3T3-L1脂肪前体细胞分化过程中的作用.方法 ①采用半定量逆转录PCR检测在3T3-L1脂肪前体细胞分化过程中KCTD15 mRNA表达变化.②在3T3-L1脂肪前体细胞增殖早期通过RNA干扰技术靶向敲低KCTD15基因的表达,在靶向敲低KCTD15基因后的转染KCTD15 siRNA 48 h后通过半定量逆转录PCR验证KCTD15基因的敲低效果.用油红O染色法观察KCTD15敲低后3T3-L1细胞第0天和第10天的细胞形态学改变.③采用半定量逆转录PCR检测KCTD15基因敲低后PPARγ、C/EBPα、C/EBPβ、C/EBPδ成脂基因的变化.结果 在3T3-L1脂肪前体细胞分化过程中,KCTD15 mRNA表达水平逐渐降低(P<0.05);KCTD15敲低能显著抑制3T3-L1脂肪前体细胞分化;KCTD15敲低后PPARγ、C/EBPα、C/EBPβ、C/EBPδ成脂基因无明显变化.结论 在分化早期阶段敲低KCTD15

  2. 小檗碱对3T3-L1脂肪细胞炎症因子分泌和炎症信号通路的影响%Effects of Berberine on the Secretion of Inflammatory Factors of 3T3-L1 Adipocytes and Inflammation Signal Path

    Institute of Scientific and Technical Information of China (English)

    于希忠; 刘佳; 程罗根; 尚文斌

    2010-01-01

    目的 观察小檗碱对3T3-L1脂肪细胞炎症因子分泌和表达的作用及其分子机制.方法 将3T3-L1脂肪细胞用10μmol/L的小檗碱干预后取上清检测肿瘤坏死因子(TNF-α)、白介素6(IL-6)、脂联素的含量.另外,以10 ng/mLTNF-α诱导3T3-L1脂肪细胞产生胰岛素抵抗模型,予以10μmol/L小檗碱进行干预,western blotting法检测脂肪细胞IKK-β的表达及IKK-β(ser181)的磷酸化水平;实时定量PCR(QRTPCR)法检测TNF-α、IL-6、CRP、MCP-1及脂联素mRNA的表达.结果 10μmol/L的小檗碱能够降低基础状态下3T3-L1脂肪细胞TNF-α和IL-6的分泌,但是没有增加脂联素分泌的作用.10 ng/mL TNF-α作用24 h使3T3-L1脂肪细胞IKK-β的表达没有明显改变,但是IKK-β(ser181)的磷酸化水平明显增加,经小檗碱干预后显著下降.QRTPCR检测结果显示10 ng/mL TNF-α作用24 h使3T3-L1脂肪细胞TNF-α、IL-6、CRP、MCP-1的mRNA表达增加,经小檗碱干预后这些炎症因子的表达显著下降.结论 小檗碱可能通过作用于IKK-β降低脂肪细胞炎症因子的分泌,改善胰岛素抵抗.

  3. Quantification of hormone sensitive lipase phosphorylation and colocalization with lipid droplets in murine 3T3L1 and human subcutaneous adipocytes via automated digital microscopy and high-content analysis.

    Science.gov (United States)

    McDonough, Patrick M; Ingermanson, Randall S; Loy, Patricia A; Koon, Erick D; Whittaker, Ross; Laris, Casey A; Hilton, Jeffrey M; Nicoll, James B; Buehrer, Benjamin M; Price, Jeffrey H

    2011-06-01

    Lipolysis in adipocytes is associated with phosphorylation of hormone sensitive lipase (HSL) and translocation of HSL to lipid droplets. In this study, adipocytes were cultured in a high-throughput format (96-well dishes), exposed to lipolytic agents, and then fixed and labeled for nuclei, lipid droplets, and HSL (or HSL phosphorylated on serine 660 [pHSLser660]). The cells were imaged via automated digital fluorescence microscopy, and high-content analysis (HCA) methods were used to quantify HSL phosphorylation and the degree to which HSL (or pHSLser660) colocalizes with the lipid droplets. HSL:lipid droplet colocalization was quantified through use of Pearson's correlation, Mander's M1 Colocalization, and the Tanimoto coefficient. For murine 3T3L1 adipocytes, isoproterenol, Lys-γ3-melanocyte stimulating hormone, and forskolin elicited the appearance and colocalization of pHSLser660, whereas atrial natriuretic peptide (ANP) did not. For human subcutaneous adipocytes, isoproterenol, forskolin, and ANP activated HSL phosphorylation/colocalization, but Lys-γ3-melanocyte stimulating hormone had little or no effect. Since ANP activates guanosine 3',5'-cyclic monophosphate (cGMP)-dependent protein kinase, HSL serine 660 is likely a substrate for cGMP-dependent protein kinase in human adipocytes. For both adipocyte model systems, adipocytes with the greatest lipid content displayed the greatest lipolytic responses. The results for pHSLser660 were consistent with release of glycerol by the cells, a well-established assay of lipolysis, and the HCA methods yielded Z' values >0.50. The results illustrate several key differences between human and murine adipocytes and demonstrate advantages of utilizing HCA techniques to study lipolysis in cultured adipocytes.

  4. 小鼠前脂肪细胞3T3-L1培养与四联诱导分化方法的探讨%Discussion on cultivation and methodology of four-drug combination-induced differentiation in mouse preadipocytes 3T3-L1 cells

    Institute of Scientific and Technical Information of China (English)

    孙慧誌; 田德润; 孟洁; 赵楠; 韩洁; 甘椿椿; 王勇

    2016-01-01

    Objective To optimize and establish the methodology for culturing and inducing differentiation of mouse preadipocytes 3T3-L1. Methods The mouse cells 3T3-L1 were incubated in DMEM medium contained with 10%FBS, during which the incubation medium was refreshed every 2 to 3 days. Two methods were used to introduce differentiation, including three-drug combination group and four-drug combination group. The protocol of mediumⅠin three-drug combination group including insulin 10 mg/L, IBMX 0.5 mmol/L and DEX 1.0μmol/L. The protocol of mediumⅠin four-drug combination group including indometacin 0.1 mmol/L based on those of three-drug combination group. Both of them were incubated for 2 days and continuous for 2 times. And medium Ⅱ included insulin 10 mg/L for 2-day culturing and continuous for 2 times. Oil red O staining was used to observe the morphological changes of two groups of cells before and after treatment under inverted microscope. Results Mouse preadipocytes 3T3-L1 appeared in good conditions and grew in a paving stone fashion. These cells covered homogeneously the bottom of incubators, the culture medium refreshed every 2 days. The results of four-drug combination group were better than those of three-drug combination group. After three-drug combination induced differentiation, there was no significant change in cell morphology. Comparing with three-drug combination induced differentiation, four-drug combination was successfully achieved in over 90% of the cell inducing, which were round-shaped, with jacinth ester droplets by oil-red O staining. Conclusion We have optimized the method for culturing and inducing differentiation of mouse preadipocytes 3T3-L1 by adding indometacin on the basis of the three-drug combination induced differentiation.%目的:改进小鼠前脂肪细胞3T3-L1的培养并诱导分化为成熟脂肪细胞的方法。方法使用含有10%胎牛血清(FBS)的高糖型DMEM液体培养基常规培养小鼠前脂肪细胞,2

  5. The Herbal Medicine KBH-1 Inhibits Fat Accumulation in 3T3-L1 Adipocytes and Reduces High Fat Diet-Induced Obesity through Regulation of the AMPK Pathway.

    Directory of Open Access Journals (Sweden)

    Ji-Hye Lee

    Full Text Available The aim of this study was to investigate whether a novel formulation of an herbal extract, KBH-1, has an inhibitory effect on obesity. To determine its anti-obesity effects and its underlying mechanism, we performed anti-obesity-related experiments in vitro and in vivo. 3T3-L1 preadipocytes were analyzed for lipid accumulation as well as the protein and gene expression of molecular targets involved in fatty acid synthesis. To determine whether KBH-1 oral administration results in a reduction in high-fat diet (HFD-induced obesity, we examined five groups (n = 9 of C57BL/6 mice as follows: 10% kcal fat diet-fed mice (ND, 60% kcal fat diet-fed mice (HFD, HFD-fed mice treated with orlistat (tetrahydrolipstatin, marketed under the trade name Xenical, HFD-fed mice treated with 150 mg/kg KBH-1 (KBH-1 150 and HFD-fed mice treated with 300 mg/kg KBH-1 (KBH-1 300. During adipogenesis of 3T3-L1 cells in vitro, KBH-1 significantly reduced lipid accumulation and down-regulated the expression of master adipogenic transcription factors, including CCAAT/enhancer binding protein (C/EBP β, C/EBP α and peroxisome proliferation-activity receptor (PPAR γ, which led to the suppression of the expression of several adipocyte-specific genes and proteins. KBH-1 also markedly phosphorylated the adenosine monophosphate-activated protein kinase (AMPK and acetyl-CoA carboxylase (ACC. In addition, KBH-1-induced the inhibition effect on lipid accumulation and AMPK-mediated signal activation were decreased by blocking AMPK phosphorylation using AMPK siRNA. Furthermore, daily oral administration of KBH-1 resulted in dose-dependent decreases in body weight, fat pad mass and fat tissue size without systemic toxicity. These results suggest that KBH-1 inhibits lipid accumulation by down-regulating the major transcription factors of the adipogenesis pathway by regulating the AMPK pathway in 3T3-L1 adipocytes and in mice with HFD-induced obesity. These results implicate KBH-1, a

  6. Effects of Different Species Bovine Sera on Proliferation and Differentiation in 3T3L1 Preadipocyte%不同品种牛血清对3T3L1前脂肪细胞增殖与分化的影响

    Institute of Scientific and Technical Information of China (English)

    冯丽萍; 宋忠峰; 施琼; 李文波; 罗晓瑜; 曹兵海

    2011-01-01

    研究不同品种牛血清对3T3-L1细胞增殖与分化的影响,采用模型细胞体外培养法模拟牛脂肪组织生长环境,为牛大理石纹肉早期选择提供一种可能性的方法.抽取17头秦川牛和28头秦杂牛血清,先制备没灭活和灭活血清组培养细胞,MTT法检测细胞增殖相对数,油红O检测脂肪含量,再用不同品种牛血清培养细胞,检测细胞增殖相对数和脂肪含量,以及用比色法测定三磷酸甘油脱氢酶(GPDH)和脂肪酸合成酶(FAS)活性.结果表明,灭活血清培养细胞的增殖相对数量在分化培养第2和4天与分化脂肪含量在第2、4、6和8天都极显著高于没灭活血清组(P<0.01);秦杂牛血清培养细胞的数量在第2和4天显著高于秦川牛(P<0.05),秦川牛血清培养细胞分化的脂肪含量在第8天显著高于秦杂牛(P<0.05),其他天数没有显著性差异(P>0.05);分化第8天的细胞内GPDH和FAS酶活两牛种间没有显著性差异(P>0.05).结果显示自制血清灭活比没灭活的更利于细胞的培养;牛血清品种是影响前脂肪细胞增殖与分化的因素,秦杂牛血清可能更有利于前脂肪细胞的增殖,而秦川牛血清可能更有利于前脂肪细胞的分化,但仍需进一步研究.%The purpose of this study was to know the effects of different species bovine sera on the proliferation and differentiation of 3T3-L1 preadipocytes. In order to provide the possible method of early selection for marbling, we use cell model in vitro culture to simulate the environment of bovine adipose tissue. Collected the bovine sera of different species from seventeen Qinchuan cattle and twenty-eight Qinchuan- Angus hybrid cattle. One part of sera were heat inactivated. The other were activated. The relatively cell number, which stands for proliferation of 3T3-L1 cells, was assayed by MTT colorimetry. Fat content and enzyme activities,which contain glycerol phosphate dehydrogenase (GPDH) and fatty acid

  7. Coculture with BJ fibroblast cells inhibits the adipogenesis and lipogenesis in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Hyun Jeong [Department of Biochemistry, Kwandong University College of Medicine, Gangneung, Gangwondo 210-701 (Korea, Republic of); Park, Sahng Wook [Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Kim, Hojeong [Department of Anatomy, Kwandong University College of Medicine, Gangneung, Gangwondo 210-701 (Korea, Republic of); Park, Sang-Kyu, E-mail: 49park@kd.ac.kr [Department of Biochemistry, Kwandong University College of Medicine, Gangneung, Gangwondo 210-701 (Korea, Republic of); Yoon, Dojun, E-mail: mozart@kd.ac.kr [Department of Biochemistry, Kwandong University College of Medicine, Gangneung, Gangwondo 210-701 (Korea, Republic of)

    2010-02-19

    Mouse or human fibroblasts are commonly used as feeder cells to prevent differentiation in stem or primary cell culture. In the present study, we addressed whether fibroblasts can affect the differentiation of adipocytes. We found that the differentiation of 3T3-L1 preadipocytes was strongly suppressed when the cells were cocultured with human fibroblast (BJ) cells. BrdU incorporation analysis indicated that mitotic clonal expansion, an early event required for 3T3-L1 cell adipogenesis, was not affected by BJ cells. The 3T3-L1 cell expression levels of peroxisome proliferator-activated receptor {gamma}2, CCAAT/enhancer-binding protein alpha (C/EBP{alpha}), sterol regulatory element binding protein-1c, and Krueppel-like factor 15, but not those of C/EBP{beta} or C/EBP{delta}, were decreased by coculture with BJ cells. When mature 3T3-L1 adipocytes were cocultured with BJ cells, their lipid contents were significantly reduced, with decreased fatty acid synthase expression and increased phosphorylated form of acetyl-CoA carboxylase 1. Our data indicate that coculture with BJ fibroblast cells inhibits the adipogenesis of 3T3-L1 preadipocytes and decreases the lipogenesis of mature 3T3-L1 adipocytes.

  8. High-density lipoprotein and apolipoprotein A-I inhibit palmitate-induced translocation of toll-like receptor 4 into lipid rafts and inflammatory cytokines in 3T3-L1 adipocytes.

    Science.gov (United States)

    Yamada, Hodaka; Umemoto, Tomio; Kawano, Mikihiko; Kawakami, Masanobu; Kakei, Masafumi; Momomura, Shin-Ichi; Ishikawa, San-E; Hara, Kazuo

    2017-03-04

    Saturated fatty acids (SFAs) activate toll-like receptor 4 (TLR4) signal transduction in macrophages and are involved in the chronic inflammation accompanying obesity. High-density lipoprotein (HDL) and apolipoprotein A-I (apoA-I) produce anti-inflammatory effects via reverse cholesterol transport. However, the underlying mechanisms by which HDL and apoA-I inhibit inflammatory responses in adipocytes remain to be determined. Here we examined whether palmitate increases the translocation of TLR4 into lipid rafts and whether HDL and apoA-I inhibit inflammation in adipocytes. Palmitate exposure (250 μM, 24 h) increased interleukin-6 and tumor necrosis factor-α gene expressions and translocation of TLR4 into lipid rafts in 3T3-L1 adipocytes. Pretreatment with HDL and apoA-I (50 μg/mL, 6 h) suppressed palmitate-induced inflammatory cytokine expression and TLR4 translocation into lipid rafts. Moreover, HDL and apoA-I inhibited palmitate-induced phosphorylation of nuclear factor-kappa B. HDL showed an anti-inflammatory effect via ATP-binding cassette transporter G1 and scavenger receptor class B, member 1, whereas apoA-I showed an effect via ATP-binding cassette transporter A1. These results demonstrated that HDL and apoA-I reduced palmitate-potentiated TLR4 trafficking into lipid rafts and its related inflammation in adipocytes via these specific transporters.

  9. 小檗碱改善3T3-L1脂肪细胞胰岛素抵抗的分子机制%Molecular mechanism for berberine to improve insulin resistance in 3T3-L1 adipocytes

    Institute of Scientific and Technical Information of China (English)

    易屏; 陆付耳; 陈广; 徐丽君; 王开富

    2007-01-01

    目的 研究小檗碱对游离脂肪酸诱导的3T3-L1脂肪细胞胰岛素抵抗的作用,探讨小檗碱改善胰岛素抵抗的分子机制.方法 以0.5 mmol/L软脂酸诱导3T3-L1脂肪细胞产生胰岛素抵抗,予以小檗碱进行干预,同时以阿司匹林作为阳性对照,用葡萄糖氧化酶法检测培液中的葡萄糖消耗量,以2-脱氧-[3H]-D-葡萄糖摄入法观察葡萄糖的转运率,用Western印迹检测IκB激酶β(IKKβ)、胰岛素受体底物1(IRS-1)、磷酸肌醇3激酶p85(PI-3K p85),葡萄糖转运子4(Glut4)的蛋白表达和IKKβ 181位丝氨酸(IKKβ Ser181)、IRS-1 307位丝氨酸(IRS-1 Ser307)的磷酸化.结果 0.5 mmol/L软脂酸作用24 h使3T3-L1脂肪细胞葡萄糖消耗降低41%,胰岛素刺激的葡萄糖转运抑制67%,IKKβ Ser181和IRS-1 Ser307的磷酸化增加,IRS-1和PI-3K p85蛋白的表达减少;同时加入小檗碱或阿司匹林则可逆转上述效应.但软脂酸、小檗碱、阿司匹林对3T3-L1脂肪细胞IKKβ蛋白、Glut4蛋白的表达无明显影响.结论 小檗碱可以明显改善游离脂肪酸诱导的胰岛素抵抗,其分子机制可能是通过抑制IKKβ Ser181磷酸化实现的.

  10. Molecular mechanism of berberine on improving insulin resistance of 3T3-L1 adipocytes induced by high glucose through IKKβ Ser 181 phosphorylation%小檗碱抑制IKKβ Ser 181磷酸化改善高糖诱导的3T3-L1脂肪细胞胰岛素抵抗的分子机制

    Institute of Scientific and Technical Information of China (English)

    易屏; 陆付耳; 陈广; 徐丽君; 王开富

    2008-01-01

    目的 研究小檗碱对高糖诱导的3T3-L1脂肪细胞胰岛素抵抗的作用,探讨小檗碱改善胰岛素抵抗的分子机制.方法 以25 mmol/L葡萄糖加0.6 nmol/L胰岛素诱导3T3-L1脂肪细胞产生胰岛素抵抗,予以小檗碱进行干预,同时以阿司匹林作为阳性对照,以2-脱氧-[3H]-D-葡萄糖摄人法观察葡萄糖的转运率,用Western blotting 检测IKKβ蛋白,IKKβSer 181磷酸化,IRS-1蛋白,IRS-1 Ser 307磷酸化,PI-3K p85蛋白,GLUT4蛋白的表达.结果 25 mmol/L葡萄糖加0.6 nmol/L胰岛素作用18 h使3T3-L1脂肪细胞胰岛素刺激的葡萄糖转运抑制60%,IKKβ Ser 181磷酸化、IRS-1 Ser 307磷酸化的表达增加,IRS-1和PI-3K p85蛋白的表达减少;同时加入小檗碱或阿司匹林则可逆转上述效应.但高糖、小檗碱、阿司匹林对3T3-L1脂肪细胞IKKl3蛋白、GLUT4蛋白的表达无明显影响.结论 小檗碱可以明显改善高糖诱导的胰岛素抵抗,其分子机制可能是小檗碱通过抑制IKKβ Ser 181磷酸化,使IRS-1丝氨酸残基磷酸化减少而酪氨酸残基磷酸化增加,调节胰岛素信号蛋白的表达来实现的.

  11. mdm-2 gene amplification in 3T3-L1 preadipocytes.

    Science.gov (United States)

    Berberich, S J; Litteral, V; Mayo, L D; Tabesh, D; Morris, D

    1999-05-01

    In this study the regulation of the murine double minute-2 (mdm-2) gene was examined in NIH 3T3-L1 preadipocytes. The 3T3-L1 cell line, under proper conditions, has the capacity to differentiate from fibroblasts into adipocytes [15]. A recent report demonstrated that mdm-2 overexpression could block myogenesis [12]. While examining the regulation of the mdm-2 gene during adipogenesis, it was discovered that 3T3-L1 cells possess a 36-fold elevation of mdm-2 mRNA relative to A31 cells, another immortalized Balb/c 3T3 fibroblast cell line that lacks the capacity to differentiate. Based on Southern blot analysis, the increase in mdm-2 mRNA was the result of a mdm-2 gene amplification. The level of Mdm-2 protein in undifferentiated 3T3-L1 cells was elevated relative to A31 fibroblasts and resulted from translation of mRNA transcripts initiating from the p53-independent P1 promoter. We also examined how mdm-2 and p53 levels changed as undifferentiated fibroblasts converted to adipocytes. While mdm-2 mRNA levels remained elevated, p53 mRNA, protein, and DNA-binding activity decreased. These results suggest that adipogenesis is unaffected by elevated Mdm-2 levels and that the overexpression of mdm-2 mRNA is predominantly p53 independent.

  12. Prolonged insulin stimulation down-regulates GLUT4 through oxidative stress-mediated retromer inhibition by a protein kinase CK2-dependent mechanism in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ma, Jinhui; Nakagawa, Yuko; Kojima, Itaru; Shibata, Hiroshi

    2014-01-03

    Although insulin acutely stimulates glucose uptake by promotion of GLUT4 translocation from intracellular compartments to the plasma membrane in adipocytes and muscles, long term insulin stimulation causes GLUT4 depletion that is particularly prominent in the insulin-responsive GLUT4 storage compartment. This effect is caused mainly by accelerated lysosomal degradation of GLUT4, although the mechanism is not fully defined. Here we show that insulin acutely induced dissociation of retromer components from the low density microsomal membranes of 3T3-L1 adipocytes that was accompanied by disruption of the interaction of Vps35 with sortilin. This insulin effect was dependent on the activity of protein kinase CK2 but not phosphatidylinositol 3-kinase or extracellular signal-regulated kinase 1/2. Knockdown of Vps26 decreased GLUT4 to a level comparable with that with insulin stimulation for 4 h. Vps35 with a mutation in the CK2 phosphorylation motif (Vps35-S7A) was resistant to insulin-induced dissociation from the low density microsomal membrane, and its overexpression attenuated GLUT4 down-regulation with insulin. Furthermore, insulin-generated hydrogen peroxide was an upstream mediator of the insulin action on retromer and GLUT4. These results suggested that insulin-generated oxidative stress switches the GLUT4 sorting direction to lysosomes through inhibition of the retromer function in a CK2-dependent manner.

  13. Effects of berberine on PI-3K p85 protein expression in insulin-resistant cell model in 3T3-L1 adipocytes%小檗碱对3T3-L1胰岛素抵抗细胞模型PI-3K p85蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    易屏; 陆付耳; 陈广; 徐丽君; 董慧; 王开富

    2008-01-01

    目的:研究小檗碱对3T3-L1胰岛素抵抗细胞模型PI-3K p85蛋白表达的影响,探讨小檗碱改善胰岛素抵抗的分子机制.方法:分别以0.5 mmol/L软脂酸与25 mmol/L葡萄糖加0.6 nmmol/L胰岛素诱导3T3-L1脂肪细胞产生胰岛素抵抗,予以小檗碱进行干预,同时以阿司匹林作为阳性对照,以2-脱氧-[3H]-D-葡萄糖摄入法观察葡萄糖的转运率,用Western blot检测PI-3K p85蛋白的表达.结果:0.5 mmol/L软脂酸作用24 h或25 mmol/L葡萄糖加0.6 nmmol/L胰岛素作用18 h分别使3T3-L1脂肪细胞胰岛素刺激的葡萄糖转运抑制67%和60%,Westem blot显示PI-3K p85蛋白表达减少,与正常对照组比较有统计学意义(P<0.01);同时加入小檗碱则可逆转上述效应使P1-3K p85蛋白表达增加,与模型组比较有明显差异(P<0.01),并且PI-3K p85蛋白的表达与小檗碱的剂量和作用时间呈依赖关系.结论:小檗碱可以明显改善游离脂肪酸和高糖诱导的胰岛素抵抗,其分子机制可能与小檗碱提高PI-3K p85蛋白的表达有关.

  14. Effects of retinoic acid and hydrogen peroxide on sterol regulatory element-binding protein-1a activation during adipogenic differentiation of 3T3-L1 cells

    OpenAIRE

    Eldaim, Mabrouk A. Abd; Okamatsu-Ogura, Yuko; Terao, Akira; Kimura, Kazuhiro

    2010-01-01

    Both retinoic acid (RA) and oxidative stress (H2O2) increased transcription and cleavage of membrane-bound sterol regulatory element-binding protein (SREBP)-1, leading to enhanced transcription of fatty acid synthase (FAS) in hepatoma cells. On the other hand, RA and H2O2 decreased and increased lipogenesis in adipocytes, respectively, although roles of SREBP-1 activation in these effects remain to be elucidated. To elucidate its involvement, we examined the activation of SREBP...

  15. Effect of catalpol, berberine, and their combination on insulin resistant 3T3-L1 adipocytes%梓醇与小檗碱及其配伍对胰岛素抵抗3T3-L1脂肪细胞的影响

    Institute of Scientific and Technical Information of China (English)

    刘芳芳; 杨明炜; 王晓强; 王开富; 陆付耳

    2007-01-01

    目的 观察梓醇与小檗碱及其配伍对地塞米松诱导的胰岛素抵抗3T3-L1脂肪细胞葡萄糖消耗、转运及这一过程中过氧化物体增殖物激活受体(PPAR-γ)mRNA表达的影响.方法 采用地塞米松诱导胰岛素抵抗细胞模型,分别给予罗格列酮、小檗碱、梓醇、梓醇+小檗碱进行干预,以葡萄糖氧化酶法检测培养液中葡萄糖消耗量,以2-脱氧-[3H]-D-葡萄糖摄入法观察葡萄糖的转运率,以RT-PCR检测PPAR-γmRNA的表达.结果 含或不含10 nmol/L胰岛素的条件下,梓醇、小檗碱及其配伍组胰岛素抵抗脂肪细胞的葡萄糖消耗量和转运率较模型组明显改善(P<0.05、0.01),配伍组效应优于梓醇、小檗碱单药组(P<0.05、0.01);且小檗碱组及配伍组PPAR-γmRNA的表达降低(P<0.05、0.01).结论 梓醇、小檗碱均能增加葡萄糖消耗和转运,改善胰岛素抵抗,其作用不依赖胰岛素的存在,且小檗碱及两药配伍还能下调脂肪细胞PPAR-γ mRNA的表达水平,提示梓醇、小檗碱改善胰岛素抵抗的作用机制可能与罗格列酮不同.

  16. Citrus aurantium flavonoids inhibit adipogenesis through the Akt signaling pathway in 3T3-L1 cells

    Directory of Open Access Journals (Sweden)

    Kim Gon-Sup

    2012-04-01

    Full Text Available Abstract Background Obesity is a health hazard that is associated with a number of diseases and metabolic abnormalities, such as type-2 diabetes, hypertension, dyslipidemia, and coronary heart disease. In the current study, we investigated the effects of Citrus aurantium flavonoids (CAF on the inhibition of adipogenesis and adipocyte differentiation in 3T3-L1 cells. Methods During adipocyte differentiation, 3T3-L1 cells were treated with 0, 10, and 50 μg/ml CAF, and then the mRNA and protein expression of adipogenesis-related genes was assayed. We examined the effect of CAF on level of phosphorylated Akt in 3T3-L1 cells treated with CAF at various concentrations during adipocyte differentiation. Results The insulin-induced expression of C/EBPβ and PPARγ mRNA and protein were significantly down-regulated in a dose-dependent manner following CAF treatment. CAF also dramatically decreased the expression of C/EBPα, which is essential for the acquisition of insulin sensitivity by adipocytes. Moreover, the expression of the aP2 and FAS genes, which are involved in lipid metabolism, decreased dramatically upon treatment with CAF. Interestingly, CAF diminished the insulin-stimulated serine phosphorylation of Akt (Ser473 and GSK3β (Ser9, which may reduce glucose uptake in response to insulin and lipid accumulation. Furthermore, CAF not only inhibited triglyceride accumulation during adipogenesis but also contributed to the lipolysis of adipocytes. Conclusions In the present study, we demonstrate that CAF suppressed adipogenesis in 3T3-L1 adipocytes. Our results indicated that CAF down-regulates the expression of C/EBPβ and subsequently inhibits the activation of PPARγ and C/EBPα. The anti-adipogenic activity of CAF was mediated by the inhibition of Akt activation and GSK3β phosphorylation, which induced the down-regulation of lipid accumulation and lipid metabolizing genes, ultimately inhibiting adipocyte differentiation.

  17. Increased Oxidative Stress in Cultured 3T3-L1 Cells was Attenuated by Berberine Treatment.

    Science.gov (United States)

    Dong, Shi-Fen; Yasui, Naomi; Negishb, Hiroko; Kishimoto, Aya; Sun, Jian-Ning; Ikeda, Katsumi

    2015-06-01

    The 3T3-L1 cell line is one of the most well-characterized and reliable models for studying adipocytes. Increased oxidative stress in accumulated fat was found in 3T3-L1 cells. Berberine, an isoquinoline alkaloid, could suppress fat deposition in 3T3-L1 cells; however, whether berberine suppresses increased oxidative stress is not well known. In this study, we observed the effect of berberine on increased oxidative stress in 3T3-L1 cells. 3T3-L1 cells were cultured and treated with berberine (5-20 μM) from day 3 to day 8. We confirmed that berberine markedly inhibited fat accumulation and lipid droplets in 3T3-L1 adipocytes and decreased triglyceride content. Berberine inhibited increased oxidative stress in 3T3-L1 cells by suppressing reactive oxygen species (ROS) production, and increased glutathione peroxidase (GPx) gene expression and GPx activity. Berberine also markedly reduced adipokines secreted by adipocytes, including leptin and resistin.

  18. Hydrogen sulfide promotes adipogenesis in 3T3L1 cells.

    Directory of Open Access Journals (Sweden)

    Chin-Yi Tsai

    Full Text Available The effect of hydrogen sulfide (H2S on differentiation of 3T3L1-derived adipocytes was examined. Endogenous H2S was increased after 3T3L1 differentiation. The expression of the H2S-synthesising enzymes, cystathionine γ-lyase (CSE, cystathionine β-synthase (CBS and 3-mercaptopyruvate sulfurtransferase (3-MST, was increased in a time-dependent manner during 3T3L1 differentiation. Expression of genes associated with adipogenesis related genes including fatty acid binding protein 4 (FABP4/aP2, a key regulator of this process, was increased by GYY4137 (a slow-releasing H2S donor compound and sodium hydrosulfide (NaHS, a classical H2S donor but not by ZYJ1122 or time-expired NaHS. Furthermore expression of these genes were reduced by aminooxyacetic acid (AOAA, CBS inhibitor, DL-propargylglycine (PAG, CSE inhibitor as well as by CSE small interference RNA (siCSE and siCBS. The size and number of lipid droplets in mature adipocytes was significantly increased by both GYY4137 and NaHS, which also impaired the ability of CL316,243 (β3-agonist to promote lipolysis in these cells. In contrast, AOAA and PAG had the opposite effect. Taken together, we show that the H2S-synthesising enzymes CBS, CSE and 3-MST are endogenously expressed during adipogenesis and that both endogenous and exogenous H2S modulate adipogenesis and adipocyte maturation.

  19. Effect of globular domain of adiponectin on pentose phosphate pathway key enzyme of 3T3 adipocyte%脂联素球状结构域对3T3-L1脂肪细胞磷酸戊糖途径关键酶表达的影响

    Institute of Scientific and Technical Information of China (English)

    王彦; 廉如; 陈思思; 高婷婷; 吴彬; 陈显久

    2012-01-01

    Objective To investigate the effect of globular domain of adiponectin (gAd) on pentose phosphate pathway key enzyme of 3T3-L1 adipocytes. Methods Matured 3T3-L1 adipocytes were intervened with gAd, and realtime PCR (RT-PCR) was used to determine the transcription level of glucose-6-phosphatase (G6PD), the key enzyme in pentose phosphate pathway; the changes of transcription level of G6PD were expressed in changes of mRNA levels. All the data were analyzed statistically. Results G6PD transcription level was not different between gAd group and control group. Conclusions gAd promotes the intake of glucose by 3T3-L1 cells, and the catabolism of glucose taken probably was not via the pentose phosphate pathway.Objective To investigate the effect of globular domain of adiponectin (gAd) on pentose phosphate pathway key enzyme of 3T3-L1 adipocytes. Methods Matured 3T3-L1 adipocytes were intervened with gAd, and realtime PCR (RT-PCR) was used to determine the transcription level of glucose-6-phosphatase (G6PD), the key enzyme in pentose phosphate pathway; the changes of transcription level of G6PD were expressed in changes of mRNA levels. All the data were analyzed statistically. Results G6PD transcription level was not different between gAd group and control group. Conclusions gAd promotes the intake of glucose by 3T3-L1 cells, and the catabolism of glucose taken probably was not via the pentose phosphate pathway.%目的:通过探讨脂联素球状结构域(gAd)对3T3-L1脂肪细胞磷酸戊糖途径关键酶表达的影响,进而探讨gAd促进脂肪细胞摄取的葡萄糖是否经磷酸戊糖途径代谢.方法:用gAd干预分化成熟的3T3-L1脂肪细胞,干预结束后测定细胞残液的葡萄糖浓度,并以实时荧光定量PCR(RT-PCR)法检测各组细胞磷酸戊糖途径关键酶葡萄糖-6-磷酸酶(G6PD)转录水平的表达情况,进行统计学分析.结果:各实验组细胞残液中葡萄糖浓度均显著低于对照组(均P<O.01);各实验组G6PD

  20. The aporphine alkaloid boldine induces adiponectin expression and regulation in 3T3-L1 cells.

    Science.gov (United States)

    Yu, Bangning; Cook, Carla; Santanam, Nalini

    2009-10-01

    Adiponectin is an adipokine secreted by differentiated adipocytes. Clinical studies suggest a negative correlation between oxidative stress and adiponectin levels in patients with metabolic syndrome or cardiovascular disease. Natural compounds that can prevent oxidative stress mediated inhibition of adiponectin may be potentially therapeutic. Boldine, an aporphine alkaloid abundant in the medicinal plant Peumus boldus, is a powerful antioxidant. The current study demonstrates the effects of boldine on the expression of adiponectin and its regulators, CCAAT/enhancer binding protein-alpha (C/EBPalpha) and peroxisome proliferator-activated receptor (PPAR)-gamma, in 3T3-L1 cells. Differentiated 3T3-L1 adipocytes were exposed to either hydrogen peroxide (H(2)O(2)) (100 microM) or tumor necrosis factor-alpha (TNFalpha) (1 ng/mL) for 24 hours in the presence or absence of increasing concentrations of boldine (5-100 microM). Quantitative polymerase chain reaction showed that both the oxidants decreased the mRNA levels of adiponectin, PPARgamma, and C/EBPalpha to half of the control levels. Boldine, at all concentrations, counteracted the inhibitory effect of H(2)O(2) or TNFalpha and increased the expression of adiponectin and its regulators. The effect of boldine on adiponectin expression was biphasic, with the lower concentrations (5-25 microM) having a larger inductive effect compared to higher concentrations (50-100 microM). Boldine treatment alone in the absence of H(2)O(2) or TNFalpha was also able to induce adiponectin at the inductive phase of adipogenesis. Peroxisome proliferator response element-luciferase promoter transactivity analysis showed that boldine interacts with the PPAR response element and could potentially modulate PPAR responsive genes. Our results indicate that boldine is able to modulate the expression of adiponectin and its regulators in 3T3-L1 cells and has the potential to be beneficial in obesity-related cardiovascular disease.

  1. Effect of cortisol on calpains in the C2C12 and 3T3-L1 cells.

    Science.gov (United States)

    Muthuraman, Pandurangan; Ravikumar, Sambandam; Muthuviveganandavel, Veerappan; Kim, Jongpil

    2014-03-01

    The present study was carried out to understand the effect of cortisol on calpain system in the C2C12 and 3T3-L1 adipocyte cells under co-culture system. Cells were co-cultured by using transwell inserts with a 0.4 μm porous membrane to separate C2C12 and 3T3-L1 preadipocyte cells. Each cell type was grown independently on the transwell plates. Following cell differentiation, inserts containing 3T3-L1 cells were transferred to C2C12 plates. Ten microgram per milliliter of cortisol was added to the medium. Following treatment for 3 days, the cells in the lower well were harvested for analysis. Calpains such as μ-calpain, m-calpain, and calpastatin were selected for the analysis. RT-PCR results indicated the significant increase in the mRNA expression of μ-calpain, m-calpain, and calpastatin. In addition, the confocal microscopical investigation indicated the cortisol treatment increases calpain expression in the C2C12 and 3T3-L1 cells. Taking all these together, cortisol treatment with co-culture system shows most reliable status of calpains expression in the cells, which is quite distinct from one-dimensional monocultured cells.

  2. Molecular mechanism of berberine improving insulin resistance induced by high glucose in 3T3-L1 adipocytes through targeting NF-кB p65%小檗碱抑制核因子NF-кB p65的核转位改善高糖诱导的3T3-L1细胞胰岛素抵抗的分子机制

    Institute of Scientific and Technical Information of China (English)

    易屏; 陆付耳; 陈广; 徐丽君; 王开富

    2008-01-01

    目的:观察小檗碱对高糖诱导的3T3-L1脂肪细胞胰岛素抵抗模型核因子NF-кB p65表达及转位的影响,探讨小檗碱改善胰岛素抵抗的分子生物学机制.方法:以25mmol·L-1葡萄糖加0.6nml·L-1胰岛素诱导3T3-L1脂肪细胞产生胰岛素抵抗,以小檗碱进行干预,同时阿司匹林作为阳性对照,以2-脱氧-[3H]-D-葡萄糖摄入法推算葡萄糖的转运率,用Western blot检测3T3-L1脂肪细胞总NF-кB p65蛋白及核NF-кB p65蛋白的表达,激光扫描共聚焦(CLSM)对NF-кB p65蛋白进行定位显示.结果:25mmol·L-1葡萄糖加0.6nmol·L-1胰岛素作用18h后使3T3-L1脂肪细胞胰岛素刺激的葡萄糖转运率抑制60%, Western blot显示核NF-кB p65蛋白表达明显增加, CLSM显示NF-кB p65核转位增加;同时加入小檗碱或阿司匹林则可逆转上述效应.但高糖、小檗碱、阿司匹林对3T3-L1脂肪细胞总NF-кB p65蛋白的表达无明显影响.结论:小檗碱可以改善高糖诱导的胰岛素抵抗,其分子机制可能与小檗碱抑制NF-кB p65的核转位有关.

  3. 小檗碱抑制核因子NF-κB p65表达及转位改善游离脂肪酸诱导的3T3-L1细胞胰岛素抵抗的分子机制%Molecular Mechanism of Berberine in Improving Insulin Resistance Induced by Free Fatty Acid through Inhibiting Nuclear Trascription Factor-κB p65 in 3T3-L1 Adipocytes

    Institute of Scientific and Technical Information of China (English)

    易屏; 陆付耳; 陈广; 徐丽君; 王开富

    2007-01-01

    目的 观察小檗碱对游离脂肪酸诱导的3T3-L1脂肪细胞胰岛素抵抗模型核因子(nclcar transcription factor kappaB,NF-κB p65)表达及转位的影响,探讨小檗碱改善胰岛素抵抗的分子机制.方法 以0.5 mmol/L软脂酸诱导3T3-L1脂肪细胞产生胰岛素抵抗,予以小檗碱进行干预,同时以阿司匹林作为阳性对照,用葡萄糖氧化酶法检测培液中的葡萄糖消耗量,以2-脱氧-[3H]-D-葡萄糖摄入法观察葡萄糖的转运率,用Western blot检测3T3-L1脂肪细胞总NF-κB p65蛋白及核NF-κB p65蛋白的表达,用激光扫描共聚焦显微镜(CLSM)对NF-κB p65进行定位显示.结果 0.5 mmol/L软脂酸作用24 h使3T3-L1脂肪细胞葡萄糖消耗降低41%,胰岛素刺激的葡萄糖转运抑制67%,NF-κB p65蛋白表达明显增加,CLSM显示NF-κBp65核转位增加;同时加入小檗碱或阿司匹林则可逆转上述效应.但软脂酸、小檗碱、阿司匹林对3T3-L1脂肪细胞总NF-κB p65蛋白的表达无明显影响.结论 小檗碱可以改善游离脂肪酸诱导的胰岛素抵抗,其分子机制可能是小檗碱通过抑制NF-κB的活化转位调节相关基因的表达而起作用.

  4. Neu-p11 ameliorates insulin resisitance in 3T3-L1 adipocytes based on ATGL/HSL and its underlying mechanism%ATGL/HSL角度下解析Neu-p11改善胰岛素抵抗作用机制

    Institute of Scientific and Technical Information of China (English)

    王平平; 佘美华; Laudon Moshe; 尹卫东

    2013-01-01

    Aim To explore the possible role of adipose tissue triglyceride enzyme ( ATGL ) and hormonesensitive lipase ( HSL ) in high glucose and insulin ( HGI ) - induced insulin resistance in 3T3-L1 adipocytes and the underlying mechanisms. Methods 3T3-L1 adipocytes were administered with HGI for 24 h to induce insulin resistance. Glucose uptake and the quantitative determination of triglycerides were designed for detection indicators. Protein expressions were detected by Western blot. Results HGI incubating resulted in decreased insulin-stimulated glucose uptake and a significant increase in TG content in fat cells, with a concomitant decrease in ATGL and HSL protein expression. The Neu-p11 intervention reversed the effects of HGI on fat cells, while luzindole counteracted the effect of Neu-pll. Conclusions Neu-p11 might inhibit TG deposition in insulin-resistant 3T3-L1 adipocytes via MT2 receptor -dependent manner, at least in part by increasing triglyceride hydrolysis, resulting from enhancing ATGL and HSL levels.%目的 探讨脂肪组织甘油三酯酶(adipose triglyceride lipase,ATGL)及激素敏感性脂肪酶(hormone-sensitive lipase,HSL)在褪黑素非选择性受体激动剂Neu-p11改善高糖高胰岛素(high glucose and insulin,HGI)诱导的3T3-L1脂肪细胞胰岛素抵抗(insulin resistance,IR)中的作用及机制.方法 培养3T3-L1脂肪细胞,HGI诱导IR模型.以葡萄糖消耗量及细胞内甘油三酯(triglyceride,TG)定量测定作为检测指标,Western blot检测蛋白水平的表达情况.结果 HGI孵育减少脂肪细胞葡萄糖摄取,促进细胞内TG积聚,同时伴有ATGL及HSL的蛋白表达下调.Neu-p11干预逆转了HGI对脂肪细胞的作用效应,而MT2竞争性拮抗剂luzindole却拮抗了Neu-p11的上述效应.结论 Neu-p11以MT2受体依赖性方式抑制IR脂肪细胞TG沉积,可能与其上调ATGL、HSL蛋白的表达,促进TG水解相关.

  5. Hydroxytyrosol Inhibits Cannabinoid CB1 Receptor Gene Expression in 3T3-L1 Preadipocyte Cell Line.

    Science.gov (United States)

    Tutino, Valeria; Orlando, Antonella; Russo, Francesco; Notarnicola, Maria

    2016-02-01

    The 3T3-L1 preadipocyte cell line is a well characterized cell model for studying the adipocyte status and the molecular mechanisms involved in differentiation of these cells. 3T3-L1 preadipocytes have the ability to synthesize and degrade endocannabinoid anandamide (AEA) and their differentiation into adipocytes increases the expression of cannabinoid (CB1) and PPAR-γ receptors. Clinically, the blocking stimulation of the endocannabinoid pathway has been one of the first approaches proposed to counteract the obesity and obesity-associated diseases (such as diabetes, metabolic syndrome and cancer). In this connection, here we studied in cultured 3T3-L1 pre-adipocytes the effects of n-3-PUFA, α-Linolenic acid (OM-3), n-6-PUFA, Linoleic acid (OM-6), and hydroxytyrosol (HT) on the expression of CB1 receptor gene and the adipogenesis-related genes PPAR-γ, Fatty Acid Synthase (FAS) and Lipoprotein Lipase (LPL). HT was able to inhibit 3T3-L1 cell differentiation by down-regulating cell proliferation and CB1 receptor gene expression. HT exhibited anti-adipogenic effects, whereas OM-3 and OM-6 exerted an inhibitory action on cell proliferation associated with an induction of the preadipocytes differentiation and CB1 receptor gene expression. Moreover, the expression of FAS and LPL genes resulted increased after treatment with both HT and OM-3 and OM-6. The present study points out that the intake of molecules such as HT, contained in extra virgin olive oil, may be considered also in view of antiobesity and antineoplastic properties by acting directly on the adipose tissue and modulating CB1 receptor gene transcription.

  6. Specific Labeling of Mouse 3T3-L1 Preadipocyte Cell Line with Green Fluorescent Protein%小鼠3T3-L1前脂肪细胞系的特异性标记

    Institute of Scientific and Technical Information of China (English)

    张崇本; 张晓兰; 李成健; 成俊英; 吴显荣

    2004-01-01

    A vector of paP2-promoter-EGFP was constructed and introduced into mouse 3T3-L1 preadipocyte cells, a cell line derived from mouse Swiss3T3 cells that were isolated from mouse embryo, to make the cells labelled with enhanced green fluorescent protein (EGFP) whose expression was controlled by the promoter of adipose-specific gene aP2. The cells were then induced to differentiate and the expression of aP2 was detected by EGFP-microscopy and RT-PCR assays. The EGFP gene was transferred into the mouse 3T3-L1 preadipocyte cells, and EGFP expression and lipid accumulation were observed during differentiation. The expression of aP2 was stable and similar to the expression of EGFP. A preadipocyte cell line expressing EGFP was obtained under the control of the promoter of adipocyte-specific expression gene aP2, and the preadipocyte cell line was specifically labelled. The cell line provides a powerful approach for the research of adipocyte differentiation and for the screening of anti-obesity and anti-diabetes drugs.%用增强绿色荧光蛋白特异性标记小鼠3T3-L1前脂肪细胞系.构建pap2-promoter-EGFP载体,电穿孔转染小鼠3T3-L1前脂肪细胞,显微荧光观察和RT-PCR确认aP2基因的内源表达.EGFP基因转入3T3-L1前脂肪细胞,观察到细胞分化过程中EGFP表达和脂肪积累.RT-PCR分析表明,EGFP代表了稳定而真实的aP2基因的内源性表达.建立了由脂肪组织特异表达基因aP2的表达控制的EGFP标记的小鼠3T3-L1前脂肪细胞系,目前尚未见用同样方法对前脂肪细胞进行特异性标记.该细胞系将为脂肪细胞分化机理研究以及为抗肥胖症和抗糖尿病药物筛选提供有力工具.

  7. Effect of catalpol,berberine,and their combination on expression of Glut4 protein and C-Cbl associated protein in insulin resistant 3T3-L1 adipocytes%小檗碱与梓醇及其配伍对胰岛素抵抗3T3-L1脂肪细胞葡萄糖转运子4蛋白及C-Cb1相关蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    陈立; 杨明炜; 汪忠煜; 刘艳娟; 陆付耳; 黄光英

    2008-01-01

    目的 观察梓醇与小檗碱及其配伍对胰岛素抵抗3T3-L1脂肪细胞葡萄糖消耗及这一过程中葡萄糖转运子4(Glut4)和C-Cb1相关蛋白(CAP)表达的影响.方法 采用高糖联合高胰岛素诱导3T3-L1脂肪细胞产生胰岛素抵抗(IR),分别给予小檗碱、梓醇、小檗碱+梓醇、盐酸罗格列酮进行干预,以葡萄糖氧化酶法检测培养液中葡萄糖消耗量.以Western Blotting法检测Glut4和CAP蛋白的表达.结果 与模型组相比,小檗碱能增加培养液中葡萄糖的消耗.但对Glut4蛋白的表达无影响;梓醇、小檗碱+梓醇均能显著增加培养液中葡萄糖的消耗,并使细胞中Glut4蛋白的表达增强.且小檗碱+梓醇组的效应优于梓醇组及小檗碱组;与模型组相比,小檗碱与梓醇及其配伍对CAP的表达没有显著性影响.结论 小檗碱、梓醇及其配伍能改善IR 3T3-L1脂肪细胞的胰岛素活性,其作用机制与罗格列酮不同.

  8. Heterologous expression of C. elegans fat-1 decreases the n-6/n-3 fatty acid ratio and inhibits adipogenesis in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    An, Lei, E-mail: anleim@yahoo.com.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Pang, Yun-Wei, E-mail: yunweipang@126.com [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Gao, Hong-Mei, E-mail: Gaohongmei_123@yahoo.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Research Unit for Animal Life Sciences, Animal Resource Science Center, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Ibaraki-Iwama 319-0206 (Japan); Tao, Li, E-mail: Eunice8023@yahoo.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin 130118 (China); Miao, Kai, E-mail: miaokai7@163.com [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); Wu, Zhong-Hong, E-mail: wuzhh@cau.edu.cn [Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Science and Technology, China Agricultural University, Beijing 100193 (China); and others

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer Expression of C. elegans fat-1 reduces the n-6/n-3 PUFA ratio in 3T3-L1 cells. Black-Right-Pointing-Pointer fat-1 inhibits the proliferation and differentiation of 3T3-L1 preadipocytes. Black-Right-Pointing-Pointer fat-1 reduces lipid deposition in 3T3-L1 adipocytes. Black-Right-Pointing-Pointer The lower n-6/n-3 ratio induces apoptosis in 3T3-L1 adipocytes. -- Abstract: In general, a diet enriched in polyunsaturated fatty acids (PUFAs) inhibits the development of obesity and decreases adipose tissue. The specific impacts of n-3 and n-6 PUFAs on adipogenesis, however, have not been definitively determined. Traditional in vivo and in vitro supplementation studies have yielded inconsistent or even contradictory results, which likely reflect insufficiently controlled experimental systems. Caenorhabditiselegans fat-1 gene encodes an n-3 fatty acid desaturase, and its heterologous expression represents an effective method both for altering the n-6/n-3 PUFA ratio and for evaluating the biological effects of n-3 and n-6 PUFAs. We sought to determine whether a reduced n-6/n-3 ratio could influence adipogenesis in 3T3-L1 cells. Lentivirus-mediated introduction of the fat-1 gene into 3T3-L1 preadipocytes significantly reduced the n-6/n-3 ratio and inhibited preadipocyte proliferation and differentiation. In mature adipocytes, fat-1 expression reduced lipid deposition, as measured by Oil Red O staining, and induced apoptosis. Our results indicate that a reduced n-6/n-3 ratio inhibits adipogenesis through several mechanisms and that n-3 PUFAs more effectively inhibit adipogenesis (but not lipogenesis) than do n-6 PUFAs.

  9. 3T3-L1 preadipocytes exhibit heightened monocyte-chemoattractant protein-1 response to acute fatty acid exposure.

    Science.gov (United States)

    Dordevic, Aimee L; Konstantopoulos, Nicky; Cameron-Smith, David

    2014-01-01

    Preadipocytes contribute to the inflammatory responses within adipose tissue. Whilst fatty acids are known to elicit an inflammatory response within adipose tissue, the relative contribution of preadipocytes and mature adipocytes to this is yet to be determined. We aimed to examine the actions of common dietary fatty acids on the acute inflammatory and adipokine response in 3T3-L1 preadipocytes and differentiated mature adipocytes. Gene expression levels of key adipokines in 3T3-L1 preadipocytes and adipocytes were determined following incubation with palmitic acid, myristic acid or oleic acid and positive inflammatory control, lipopolysaccharide for 2 and 4 h. Inflammatory kinase signalling was assessed by analysis of nuclear factor-κB, p38-mitogen-activated protein kinase and c-jun amino-terminal kinase phosphorylation. Under basal conditions, intracellular monocyte chemoattractant protein-1 and interleukin-6 gene expression levels were increased in preadipocytes, whereas mature adipocytes expressed increased gene expression levels of leptin and adiponectin. Fatty acid exposure at 2 and 4 h increased both monocyte chemoattractant protein-1 and interleukin-6 gene expression levels in preadipocytes to greater levels than in mature adipocytes. There was an accompanying increase of inhibitor of κB-α degradation and nuclear factor-κB (p65) (Ser536) phosphorylation with fatty acid exposure in the preadipocytes only. The current study points to preadipocytes rather than the adipocytes as the contributors to both immune cell recruitment and inflammatory adipokine secretion with acute increases in fatty acids.

  10. Effect of Berberine,Catalpol and Their Combination on the Expression of Glut-4,IRS-1,IRS-1 Ser307 Phosphorylation in Insulin Resistant 3T3-L1 Adipocytes%小檗碱与梓醇及其配伍对胰岛素抵抗3T3-L1脂肪细胞Glut-4、IRS-1、IRS-1 Ser307磷酸化蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    汪忠煜; 杨明炜; 陈立; 刘艳娟; 陆付耳; 黄光英

    2008-01-01

    目的: 观察梓醇与小檗碱及其配伍对胰岛素抵抗3T3-L1脂肪细胞葡萄糖消耗及这一过程中葡萄糖转运子-4(Glut-4)、胰岛素受体底物-1(IRS-1)和胰岛素受体底物-1丝氨酸307(IRS-1 Ser307)磷酸化蛋白表达的影响.方法: 采用高糖联合高胰岛素诱导3T3-L1脂肪细胞产生胰岛素抵抗,分别给予小檗碱、梓醇、小檗碱+梓醇、盐酸罗格列酮进行干预,以葡萄糖氧化酶法检测培养液中葡萄糖消耗量,以Western Blot法检测蛋白的表达.结果: 与模型组相比,小檗碱能增加培养液中葡萄糖的消耗(P

  11. The 3T3-L1 adipocyte glycogen proteome

    OpenAIRE

    Stapleton, David; Nelson, Chad; Parsawar, Krishna; Flores-Opazo, Marcelo; McClain, Donald; Parker, Glendon

    2013-01-01

    Background Glycogen is a branched polysaccharide of glucose residues, consisting of α-1-4 glycosidic linkages with α-1-6 branches that together form multi-layered particles ranging in size from 30 nm to 300 nm. Glycogen spatial conformation and intracellular organization are highly regulated processes. Glycogen particles interact with their metabolizing enzymes and are associated with a variety of proteins that intervene in its biology, controlling its structure, particle size and sub-cellula...

  12. Effects of Berberine on Cell Proliferation, Peroxisome Proliferation Activated Receptorγ, CAAT/Enhan-cer Binding Protein mRNA and Protein Expression in 3T3-L1 Pre-adipocytes%小檗碱对3T3-L1前脂肪细胞增殖及分化相关基因PPARγ、C/EBPαmRNA和蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    刘毅; 娄少颖; 何燕铭; 陈伟华; 应健; 王文健

    2008-01-01

    目的 探讨小檗碱对脂肪细胞增殖、分化的影响及其机制.方法 以XTT法检测3T3-L1前脂肪细胞的增殖;油红O染色并通过比色定量分析检测313-L1前脂肪细胞分化过程中胞浆脂质的堆积;采用Real-time PCR和蛋白质免疫印迹(Western blotting)技术检测脂肪细胞分化相关基因过氧化物体增殖剂活化受体γ(peroxisome proliferator activated receptor,PPARγ)、CAAT/增强子结合蛋白α(CAAT/enhancer bindingprotein,C/EBPα)mRNA以及蛋白的表达.结果 浓度低于10 μmol/L小檗碱干预24 h,对脂肪细胞增殖的影响不明显(与空白对照组比较,P>0.05),20、40、80 μmol/L小檗碱在作用24 h后即表现出明显的抑制效应;不同浓度的小檗碱作用48、72 h后对脂肪细胞的增殖亦表现出抑制效应,且有一定的量效关系,即小檗碱浓度越高抑制作用越明显,与空白对照组比较差异有统计学意义(P<0.05,P<0.01);10 μmol/L小檗碱处理的前脂肪细胞,分化后胞浆中脂滴明显减少,分化相关基因PPARγmRNA、C/EBPα mRNA和蛋白的表达亦减少,与空白对照组和罗格列酮干预组比较差异有统计学意义(P<0.05,P<0.01).结论 小檗碱能够抑制前脂肪细胞的增殖和分化,减少脂肪细胞分化过程中脂质的堆积,机制可能与其抑制脂肪细胞分化相关基因PPAHγ/、C/EBPα mRNA和蛋白表达有关,实验为小檗碱防治肥胖等代谢相关性疾病提供了依据.

  13. Overexpression of Runx2 and MKP-1 stimulates transdifferentiation of 3T3-L1 preadipocytes into bone-forming osteoblasts in vitro.

    Science.gov (United States)

    Takahashi, Tomihisa

    2011-04-01

    Runx2, a transcription factor, is essential for osteoblastic differentiation, bone formation, and maintenance. We examined the effect of Runx2 on transdifferentiation of 3T3-L1 preadipocytes into functional, mature osteoblasts. Forced expression of exogenous Runx2 using a retroviral gene-delivery system showed increases of alkaline phosphatase (ALP) activity and expression of the osteoblastic marker genes osteocalcin (OC), bone sialoprotein (BSP), and osterix (Osx), accompanied by low-level matrix mineralization. In contrast, adipocytic differentiation was completely blocked with downregulation of adipogenic transcription factors PPARγ2, C/EBPα, and C/EBPδ. Treatment of dexamethasone (Dex), a synthetic glucocorticoid, stimulated the formation of mineralized nodules in Runx2-overexpressing 3T3-L1 cells with increases of ALP, OC, BSP, and Osx expression. Here, we focused on a dual specific phosphatase, mitogen-activated protein kinase (MKP-1), since Dex significantly increased MKP-1 expression in Runx2-overexpressing 3T3-L1 cells. Forced expression of exogenous MKP-1 resulted in accumulation of robust matrix mineralization in parallel with induction of ALP activity and expression of OC, BSP, and Osx in Runx2-overexpressing 3T3-L1 cells. These results suggest that simultaneous overexpression of Runx2 and MKP-1 is effective for transdifferentiation of preadipocytes into fully differentiated bone-forming osteoblasts and provide a novel strategy for cell-based therapeutic applications requiring significant numbers of osteogenic cells to synthesize mineralized constructs for the treatment of large bone defects.

  14. THE COMBINED EFFECTS OF CATECHINS AND CAFFEINE ON CELLULAR PROLIFERATION AND LIPID METABOLISM IN 3T3-L1 CELLS%儿茶素和咖啡碱组合对3T3-L1细胞增殖及脂肪代谢的影响

    Institute of Scientific and Technical Information of China (English)

    郑国栋; 邱阳阳; 张清峰; 徐峰

    2013-01-01

    目的 研究对儿茶素和咖啡碱对3T3-L1细胞的增殖及脂肪代谢的影响.方法 采用四甲基偶氮唑盐比色法(MTT)检测对3T3-L1细胞增殖的影响;3T3-L1细胞诱导分化8d后,对各组细胞进行油红O染色并测定细胞内甘油三酯(TG)含量;细胞分化12d后,添加儿茶素和咖啡碱组合或同时添加去甲肾上腺素(NA)作用24h,分析各组细胞内脂肪分解.结果 儿茶素能明显抑制3T3-L1细胞的增殖;儿茶素和咖啡碱组合能明显抑制3T3-L1细胞分化后,细胞内TG的沉积,且在相同儿茶素浓度下,咖啡碱浓度越高抑制效果越明显.咖啡碱明显提高NA诱导成熟脂肪细胞脂解的能力,且呈剂量效应关系.结论 儿茶素和咖啡碱组合能够抑制脂肪细胞增殖和甘油三酯积聚,咖啡碱促进激素诱导脂肪细胞中脂肪分解.%Objective To investigate the combined effects of catechins and caffeine on cells proliferation and lipid metabolism in 3T3-L1 cells. Method MTT colorimetry was used to detect the effects of catechins and caffeine combination on the proliferation of 3T3-L1 cells. The differentiation of 3T3-L1 cells was induced for 8 d, then the adipocytes were stained by oil Red O, and the level of triglyceride (TG) was measured. The lipolytic effect of catechins and caffeine combination in presence or absence of noradrenaline (NA) for 24 h on 3T3-L1 cells was analyzed on the 12 th day after differentiation. Results Catechins significantly inhibited 3T3-L1 cells proliferation. Catechins and caffeine combination remarkably decreased TG accumulation after differentiation of 3T3-L1 cells, and the higher caffeine concentration was better when combined with the same catechins dose. Caffeine significantly improved NA-induced lipolysis in mature adipocytes. Conclusion Catechins and caffeine combination might inhibit cells proliferation and TG accumulation in 3T3-L1 cells. Caffeine promotes hormone-induced lipolysis in adipocytes.

  15. Effects of Berberine on Adipose Tissues and Kidney Function in 3T3-L1 Cells and Spontaneously Hypertensive Rats.

    Science.gov (United States)

    Kishimoto, Aya; Dong, Shi-Fen; Negishi, Hiroko; Yasui, Naomi; Sun, Jian-Ning; Ikeda, Katsumi

    2015-09-01

    We aimed to investigate the effect of berberine on adipose tissues, as well as its effect on renal injury in 3T3-L1 cells and spontaneously hypertensive rats. 3T3-L1 cells were cultured and treated with berberine (5-20 pM) from days 3 to 8. Berberine added to the cultured medium could significantly down-regulate transcription factors, including CCAAT/enhancer binding protein β, CCAAT/enhancer binding protein a, and peroxisome pro liferator-activated receptor y, and suppress peroxisome proliferator-activated receptor target genes, such as adipocyte fatty acid binding protein and fatty acid synthase, and inhibit 3T3-Ll fibroblast differentiation to adipocytes. Male spontaneously hypertensive rats received either 150 mg/day of berberine or saline orally for 8 weeks. Compared with the control, berberine-treated rats exhibited significant reductions in body weight gain (p Berberine-treated rats significantly decreased urinary albumin excretion, a marker of renal injury (p berberine decreased the adipose tissues weight and attenuated renal injury in spontaneously hypertensive rats. Based on these results, berberine has an important role in regulating adipose tissues. These results suggest the protective effect of berberine on metabolic syndrome related diseases, such as renal injury.

  16. Flavonol acylglycosides from flower of Albizia julibrissin and their inhibitory effects on lipid accumulation in 3T3-L1 cells.

    Science.gov (United States)

    Yahagi, Tadahiro; Daikonya, Akihiro; Kitanaka, Susumu

    2012-01-01

    Obesity is a serious health problem worldwide. We investigated the anti-obesity effect of the flower of Albizia julibrissin DURAZZ. (Leguminosae). A 90% EtOH extract of the flower inhibited adipogenesis in 3T3-L1 preadipocytes, as well as the activity of glycerol-3-phosphate dehydrogenase (GPDH) activity. New flavonol acylglycosides (1-4) and eighteen known compounds (5-22) were isolated by bioassay-directed fractionation. These new glycosides were elucidated to be 3″-(E)-p-coumaroylquercitrin (1), 3″-(E)-feruloylquercitrin (2), 3″-(E)-cinnamoylquercitrin (3), and 2″-(E)-cinnamoylquercitrin (4) on the basis of spectroscopic and chemical analysis. These compounds inhibited adipogenesis in 3T3-L1 preadipocytes. In particular, 2 exhibited potent inhibitory effects on triglyceride accumulation. Furthermore, GPDH activity was inhibited by 2. Additionally, 2 inhibited glucose uptake in 3T3-L1 adipocytes. These results indicate that the 90% EtOH extract and compounds isolated from the flower of A. julibrissin inhibit adipogenesis in 3T3-L1 preadipocytes and may have anti-obesity effect through the inhibition of preadipocyte differentiation.

  17. Fisetin induces Sirt1 expression while inhibiting early adipogenesis in 3T3-L1 cells.

    Science.gov (United States)

    Kim, Sang Chon; Kim, Yoo Hoon; Son, Sung Wook; Moon, Eun-Yi; Pyo, Suhkneung; Um, Sung Hee

    2015-11-27

    Fisetin (3,7,3',4'-tetrahydroxyflavone) is a naturally found flavonol in many fruits and vegetables and is known to have anti-aging, anti-cancer and anti-viral effects. However, the effects of fisetin on early adipocyte differentiation and the epigenetic regulator controlling adipogenic transcription factors remain unclear. Here, we show that fisetin inhibits lipid accumulation and suppresses the expression of PPARγ in 3T3-L1 cells. Fisetin suppressed early stages of preadipocyte differentiation, and induced expression of Sirt1. Depletion of Sirt1 abolished the inhibitory effects of fisetin on intracellular lipid accumulation and on PPARγ expression. Mechanistically, fisetin facilitated Sirt1-mediated deacetylation of PPARγ and FoxO1, and enhanced the association of Sirt1 with the PPARγ promoter, leading to suppression of PPARγ transcriptional activity, thereby repressing adipogenesis. Lowering Sirt1 levels reversed the effects of fisetin on deacetylation of PPARγ and increased PPARγ transactivation. Collectively, our results suggest the effects of fisetin in increasing Sirt1 expression and in epigenetic control of early adipogenesis.

  18. Isorhamnetin represses adipogenesis in 3T3-L1 cells.

    Science.gov (United States)

    Lee, Jongsung; Jung, Eunsun; Lee, Jienny; Kim, Saebom; Huh, Sungran; Kim, Youngsoo; Kim, Yongwoo; Byun, Sang Yo; Kim, Yeong-Shik; Park, Deokhoon

    2009-02-01

    Adipocyte dysfunction is strongly associated with the development of obesity, which is a major risk factor for many disorders including diabetes, hypertension, and heart disease. It is generally accepted that the regulation of adipogenesis or adipokines expression prevents obesity. In this study, we show that isorhamnetin inhibits adipocyte differentiation, as evidenced by reduced triglyceride (TG) accumulation and glycerol-3-phosphate dehydrogenase (GPDH) activity. At the molecular level, the mRNA expression levels of peroxidase proliferator-activated receptor-gamma (PPAR-gamma) and CCAAT/enhancer-binding protein-alpha (C/EBP-alpha), which are the major adipogenic transcription factors, were markedly reduced by isorhamnetin. However, the mRNA levels of C/EBP-beta and -delta, the upstream regulators of PPAR-gamma and C/EBP-alpha, were not reduced by isorhamnetin. Moreover, the mRNA levels of PPAR-gamma target genes such as lipoprotein lipase (LPL), CD36, aP2, and liver X receptor-alpha (LXR-alpha) were downregulated by isorhamnetin. We also showed that isorhamnetin inhibits the expression and secretion of adiponectin, and the results of adiponectin promoter assays suggest the inhibition of PPAR-gamma expression as a possible mechanism underlying the isorhamnetin-mediated effects. Taken together, these results indicate that isorhamnetin inhibits adipogenesis through downregulation of PPAR-gamma and C/EBP-alpha.

  19. Berberine Alleviates Olanzapine-Induced Adipogenesis via the AMPKα–SREBP Pathway in 3T3-L1 Cells

    OpenAIRE

    Yanjie Li; Xiaomin Zhao; Xiyu Feng; Xuemei Liu; Chao Deng; Chang-Hua Hu

    2016-01-01

    The aim of this study was to investigate the mechanisms underlying the inhibitory effects of berberine (BBR) on olanzapine (OLZ)-induced adipogenesis in a well-replicated 3T3-L1 cell model. Oil-Red-O (ORO) staining showed that BBR significantly decreased OLZ-induced adipogenesis. Co-treatment with OLZ and BBR decreased the accumulation of triglyceride (TG) and total cholesterol (TC) by 55.58% ± 3.65% and 49.84% ± 8.31%, respectively, in 3T3-L1 adipocytes accompanied by reduced expression of S...

  20. Traditional Korean Herbal Formula Samsoeum Attenuates Adipogenesis by Regulating the Phosphorylation of ERK1/2 in 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Soo-Jin Jeong

    2015-01-01

    Full Text Available Adipogenesis is the cell differentiation process from preadipocytes into adipocytes and the critical action in the development of obesity. In the present study, we conducted in vitro analyses to investigate the inhibitory effects of Samsoeum (SSE, a traditional herbal decoction. SSE had no significant cytotoxic effect against either the undifferentiated or differentiated 3T3-L1 cells. Oil Red O staining results showed that SSE significantly inhibited fat accumulation in adipocytes. SSE treatment consistently reduced the intracellular triglyceride content in the cells. SSE significantly inactivated glycerol-3-phosphate dehydrogenase (GPDH, a major link between carbohydrate and lipid metabolisms in 3T3-L1 adipocytes, and markedly inhibited the production of leptin, an important adipokine, in differentiated cells. SSE markedly suppressed the mRNA expression of the adipogenesis-related genes peroxisome proliferator-activated receptor-gamma (PPAR-γ, CCAAT/enhancer binding protein-alpha (C/EBP-α, fatty acid synthase (FAS, lipoprotein lipase (LPL, and fatty acid binding protein 4 (FABP4. Importantly, SSE increased the phosphorylation of ERK1/2, but not p38 MAPK and JNK, in adipose cells. Overall, our results indicate that SSE exerts antiadipogenic activity and modulates expressions of adipogenesis-related genes and ERK1/2 activation in adipocytes.

  1. RKIP phosphorylation-dependent ERK1 activation stimulates adipogenic lipid accumulation in 3T3-L1 preadipocytes overexpressing LC3.

    Science.gov (United States)

    Hahm, Jong Ryeal; Ahmed, Mahmoud; Kim, Deok Ryong

    2016-09-01

    3T3-L1 preadipocytes undergo adipogenesis in response to treatment with dexamethaxone, 1-methyl-3-isobutylxanthine, and insulin (DMI) through activation of several adipogenic transcription factors. Many autophagy-related proteins are also highly activated in the earlier stages of adipogenesis, and the LC3 conjugation system is required for formation of lipid droplets. Here, we investigated the effect of overexpression of green fluorescent protein (GFP)-LC3 fusion protein on adipogenesis. Overexpression of GFP-LC3 in 3T3-L1 preadipocytes using poly-l-lysine-assisted adenoviral GFP-LC3 transduction was sufficient to produce intracellular lipid droplets. Indeed, GFP-LC3 overexpression stimulated expression of some adipogenic transcription factors (e.g., C/EBPα or β, PPARγ, SREBP2). In particular, SREBP2 was highly activated in preadipocytes transfected with adenoviral GFP-LC3. Also, phosphorylation of Raf kinase inhibitory protein (RKIP) at serine 153, consequently stimulating extracellular-signal regulated kinase (ERK)1 activity, was significantly increased during adipogenesis induced by either poly-l-lysine-assisted adenoviral GFP-LC3 transduction or culture in the presence of dexamethasone, 1-methyl-3-isobutylxanthine, and insulin. Furthermore, RKIP knockdown promoted ERK1 and PPARγ activation, and significantly increased the intracellular accumulation of triacylglycerides in DMI-induced adipogenesis. In conclusion, GFP-LC3 overexpression in 3T3-L1 preadipocytes stimulates adipocyte differentiation via direct modulation of RKIP-dependent ERK1 activity.

  2. 13-Methylberberine, a berberine analogue with stronger anti-adipogenic effects on mouse 3T3-L1 cells

    OpenAIRE

    Chow, Yit-Lai; Sogame, Mami; Sato, Fumihiko

    2016-01-01

    Lipid metabolism modulation is a main focus of metabolic syndrome research, an area in which many natural and synthetic chemicals are constantly being screened for in vitro and in vivo activity. Berberine, a benzylisoquinoline plant alkaloid, has been extensively investigated for its anti-obesity effects and as a potential cholesterol and triglyceride-lowering drug. We screened 11 protoberberine and 2 benzophenanthridine alkaloids for their anti-adipogenic effects on 3T3-L1 adipocytes and fou...

  3. Regulation of glucose transport in the NIH 3T3 L1 preadipocyte cell line by TCDD.

    OpenAIRE

    1994-01-01

    This study examined the changes in cellular glucose uptake induced by 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) as measured by quantification of intracellular radioactivity in the NIH 3T3 L1 preadipocyte cell line after a 30-minute incubation with the non-metabolizable radioactive analogue of glucose, 3-O-methyl-D-[1-3H] glucose. Treatment of differentiated NIH 3T3 L1 cells with TCDD produced a time- and dose-dependent decrease in the cellular uptake of glucose. Treatment of cells for 3 hr w...

  4. 13-Methylberberine, a berberine analogue with stronger anti-adipogenic effects on mouse 3T3-L1 cells.

    Science.gov (United States)

    Chow, Yit-Lai; Sogame, Mami; Sato, Fumihiko

    2016-12-05

    Lipid metabolism modulation is a main focus of metabolic syndrome research, an area in which many natural and synthetic chemicals are constantly being screened for in vitro and in vivo activity. Berberine, a benzylisoquinoline plant alkaloid, has been extensively investigated for its anti-obesity effects and as a potential cholesterol and triglyceride-lowering drug. We screened 11 protoberberine and 2 benzophenanthridine alkaloids for their anti-adipogenic effects on 3T3-L1 adipocytes and found that 13-methylberberine exhibited the most potent activity. 13-Methylberberine down-regulated the expression of the main adipocyte differentiation transcription factors, peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT enhancer binding protein alpha (C/EBPα), as well as their target genes. PPARγ, C/EBPα, and sterol regulatory element binding protein 1 (SREBP-1) protein levels were reduced, and this lipid-reducing effect was attenuated by an AMP-activated protein kinase (AMPK) inhibitor, indicating that the effect of this compound requires the AMPK signaling pathway. Decreased Akt phosphorylation suggested reduced de novo lipid synthesis. C-13 methyl substitution of berberine increased its accumulation in treated cells, suggesting that 13-methylberberine has improved absorption and higher accumulation compared to berberine. Our findings suggest that 13-methylberberine has potential as an anti-obesity drug.

  5. A Quantified Ginseng (Panax ginseng C.A. Meyer Extract Influences Lipid Acquisition and Increases Adiponectin Expression in 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Chia-Rou Yeo

    2011-01-01

    Full Text Available A Panax ginseng extract (PGE with a quantified amount of ginsenosides was utilized to investigate its potential to inhibit proliferation, influence lipid acquisition and adiponectin expression in 3T3-L1 cells. Seven fingerprint ginsenosides were quantified using high performance liquid chromatography and their respective molecular weights were further confirmed via LC-ESI-MS analysis from four different extraction methods. Extraction using methanol under reflux produced significantly higher amounts of ginsenosides. The methanol extract consisted of Rg1 (47.40 ± 4.28 mg/g, dry weight of extract, Re (61.62 ± 5.10 mg/g, Rf (6.14 ± 0.28 mg/g, Rb1 (21.73 ± 1.29 mg/g, Rc (78.79 ± 4.15 mg/g, Rb2 (56.80 ± 3.79 mg/g, Rd (5.90 ± 0.41 mg/g. MTT analysis showed that PGE had a concentrationdependent cytotoxic effect on 3T3-L1 preadipocyte and the LC50 value was calculated to be 18.2 ± 5 μg/mL. Cell cycle analysis showed minimal changes in all four phases. Differentiating adipocytes treated with ginseng extract had a visible decrease in lipid droplets formation measured by Oil red O staining. Consequently, triglycerides levels in media significantly (P < 0.05 decreased by 39.5% and 46.1% when treated at concentrations of 1 μg/mL and 10 μg/mL compared to untreated control cells. Western blot analysis showed that the adiponectin protein expression was significantly (P < 0.05 increased at 10 μg/mL, but not at 1 μg/mL. A quantified PGE reduced the growth of 3T3-L1 cells, down-regulated lipid accumulation and up-regulated adiponectin expression in the 3T3-L1 adipocyte cell model.

  6. Effects of parabens on adipocyte differentiation.

    Science.gov (United States)

    Hu, Pan; Chen, Xin; Whitener, Rick J; Boder, Eric T; Jones, Jeremy O; Porollo, Aleksey; Chen, Jiangang; Zhao, Ling

    2013-01-01

    Parabens are a group of alkyl esters of p-hydroxybenzoic acid that include methylparaben, ethylparaben, propylparaben, butylparaben, and benzylparaben. Paraben esters and their salts are widely used as preservatives in cosmetics, toiletries, food, and pharmaceuticals. Humans are exposed to parabens through the use of such products from dermal contact, ingestion, and inhalation. However, research on the effects of parabens on health is limited, and the effects of parabens on adipogenesis have not been systematically studied. Here, we report that (1) parabens promote adipogenesis (or adipocyte differentiation) in murine 3T3-L1 cells, as revealed by adipocyte morphology, lipid accumulation, and mRNA expression of adipocyte-specific markers; (2) the adipogenic potency of parabens is increased with increasing length of the linear alkyl chain in the following potency ranking order: methyl- parabens, and the structurally related benzoic acid (without the OH group) are inactive in promoting 3T3-L1 adipocyte differentiation; (3) parabens activate glucocorticoid receptor and/or peroxisome proliferator-activated receptor γ in 3T3-L1 preadipocytes; however, no direct binding to, or modulation of, the ligand binding domain of the glucocorticoid receptor by parabens was detected by glucocorticoid receptor competitor assays; and lastly, (4) parabens, butyl- and benzylparaben in particular, also promote adipose conversion of human adipose-derived multipotent stromal cells. Our results suggest that parabens may contribute to obesity epidemic, and the role of parabens in adipogenesis in vivo needs to be examined further.

  7. Co-culture of C2C12 and 3T3-L1 preadipocyte cells alters the gene expression of calpains, caspases and heat shock proteins.

    Science.gov (United States)

    Pandurangan, Muthuraman; Jeong, Dawoon; Amna, Touseef; Van Ba, Hoa; Hwang, Inho

    2012-10-01

    The present study was carried out to understand the co-culture effect of C2C12 and 3T3-L1 preadipocyte cells on calpain, caspase, and heat shock protein (Hsp) systems. Calpains, caspases, and heat shock proteins play critical roles in the growth and development of mammalian cells. Cells were co-cultured using transwell inserts with a 0.4-μm porous membrane to separate C2C12 and 3T3-L1 preadipocyte cells. Each cell type was grown independently on the transwell plates. Following cell differentiation, inserts containing 3T3-L1 cells were transferred to C2C12 plates and inserts containing C2C12 transferred to 3T3-L1 plates. Following co-culture for 24 and 48 h, the cells in the lower well were harvested for analysis. Calpains include μ-calpain, m-calpain, and their specific inhibitor calpastatin. The expression pattern of μ-calpain did not change in the co-cultured C2C12 and 3T3-L1 cells, whereas m-capain mRNA expression significantly reduced in the 48-h co-cultured 3T3-L1 cells. Calpastatin mRNA expression significantly increased in the 48-h co-cultured C2C12 cells. Caspase-7 mRNA expression did not change in the 24- and 48-h co-cultured C2C12 and 3T3-L1 cells. Caspase-3 mRNA expression significantly reduced in the 24- and 48-h co-cultured 3T3-L1 cells; caspase-9 mRNA had a significant reduction only at 48 h, whereas caspase-9 mRNA expression significantly increased in the 48-h co-cultured C2C12 cells. Hsp27 and Hsp90 mRNA expressions are significantly reduced in the 24- and 48-h co-cultured C2C12 and 3T3-L1 cells, whereas Hsp70 mRNA expression significantly increased in the 48-h co-cultured 3T3-L1 cells. The co-culture reflects three-dimensional views of C2C12 and 3T3-L1 cell types as in vivo, which is quite distinct from the one-dimensional monocultured C2C12 and 3T3-L1 cells.

  8. 黄芪多糖和小檗碱对3T3-L1脂肪细胞糖代谢及细胞分化的影响%Effect of Astragalus Polysaccharides and Berberine on Carbohydrate Metabolism and Cell Differentiation in 3T3-L1 Adipocytes

    Institute of Scientific and Technical Information of China (English)

    王树海; 王文健; 汪雪峰; 陈伟华

    2004-01-01

    目的比较黄芪多糖和小檗碱对脂肪细胞糖代谢和细胞分化的影响,并分析其改善糖代谢的可能机制.方法检测黄芪多糖和小檗碱干预的脂肪细胞对3H-葡萄糖的摄取,对黄芪多糖和小檗碱干预分化的细胞进行油红O染色并通过比色定量分析脂肪细胞分化程度.逆转录多聚酶联反应(RT-PCR)检测脂肪细胞分化相关基因过氧化物体增殖剂活化受体γ(PPARγ)、CAAT/增强子结合蛋白α(C/EBPα)mRNA的表达.结果黄芪多糖和小檗碱组葡萄糖摄取率分别为正常组的109.3%和182.7%;黄芪多糖明显促进分化及其PPARγmRNA表达,与对照组比较,差异有显著性(P<0.01);小檗碱则明显抑制细胞分化及PPARγ、C/EBPαmRNA表达,与对照组比较,差异有显著性(P<0.01).结论黄芪多糖促进脂肪细胞葡萄糖摄取及细胞分化和PPARγmRNA表达,而小檗碱促进葡萄糖摄取但抑制脂肪细胞分化和PPARγ及C/EBPαmRNA的表达.

  9. Traditional medicine yanggyuksanhwa-tang inhibits adipogenesis and suppresses proliferator-activated receptor gamma expression in 3T3-L1 cells

    Directory of Open Access Journals (Sweden)

    Soo-Jin Jeong

    2015-01-01

    Full Text Available Background: Yanggyuksanhwa-tang (YGSHT is a specific traditional Korean herbal formula for Soyangin according to Sasang constitutional philosophy. Although its biological activities against inflammation and cerebral infarction have been reporting, there is no information about the adipogenic activity of YGSHT. In the present study, we investigated the anti adipogenic activity of YGSHT to evaluate effects of YGSHT on adipogenesis in vitro. Materials and Methods: Using 3T3 L1 preadipocytes, we induced the cellular differentiation into adipocytes by adding insulin. Anti adipogenic activity of YGSHT was measured by oil red O staining, triglyceride assay, glycerol 3 phosphate dehydrogenase (GPDH activity test, and leptin assay. Results: YGSHT extract had no significant cytotoxicity in preadipocytes or differentiated adipocytes. YGSHT reduced the number of lipid droplets and content of triglyceride in adipose cells. YGSHT also significantly inhibited GPDH activity and decreased leptin production compared with control adipocytes. Down regulation of peroxisome proliferator activated receptor gamma (PPAR g expression at the messenger RNA level was observed in YGSHT treated adipocytes. Conclusion: Taken together, our data suggest that YGSHT has potential as an anti-obesity drug candidate.

  10. Mango fruit peel and flesh extracts affect adipogenesis in 3T3-L1 cells.

    Science.gov (United States)

    Taing, Meng-Wong; Pierson, Jean-Thomas; Hoang, Van L T; Shaw, Paul N; Dietzgen, Ralf G; Gidley, Michael J; Roberts-Thomson, Sarah J; Monteith, Gregory R

    2012-08-01

    Obesity is associated with many chronic disease states, such as diabetes mellitus, coronary disease and certain cancers, including those of the breast and colon. There is a growing body of evidence that links phytochemicals with the inhibition of adipogenesis and protection against obesity. Mangoes (Mangifera indica L.) are tropical fruits that are rich in a diverse array of bioactive phytochemicals. In this study, methanol extracts of peel and flesh from three archetypal mango cultivars; Irwin, Nam Doc Mai and Kensington Pride, were assessed for their effects on a 3T3-L1 pre-adipocyte cell line model of adipogenesis. High content imaging was used to assess: lipid droplets per cell, lipid droplet area per cell, lipid droplet integrated intensity, nuclei count and nuclear area per cell. Mango flesh extracts from the three cultivars did not inhibit adipogenesis; peel extracts from both Irwin and Nam Doc Mai, however, did so with the Nam Doc Mai extract most potent at inhibiting adipogenesis. Peel extract from Kensington Pride promoted adipogenesis. The inhibition of adipogenesis by Irwin (100 μg mL(-1)) and Nam Doc Mai peel extracts (50 and 100 μg mL(-1)) was associated with an increase in the average nuclear area per cell; similar effects were seen with resveratrol, suggesting that these extracts may act through pathways similar to resveratrol. These results suggest that differences in the phytochemical composition between mango cultivars may influence their effectiveness in inhibiting adipogenesis, and points to mango fruit peel as a potential source of nutraceuticals.

  11. Dehydrodiconiferyl alcohol isolated from Cucurbita moschata shows anti-adipogenic and anti-lipogenic effects in 3T3-L1 cells and primary mouse embryonic fibroblasts.

    Science.gov (United States)

    Lee, Junghun; Kim, Donghyun; Choi, Jonghyun; Choi, Hyounjeong; Ryu, Jae-Ha; Jeong, Jinhyun; Park, Eun-Jin; Kim, Seon-Hee; Kim, Sunyoung

    2012-03-16

    A water-soluble extract from the stems of Cucurbita moschata, code named PG105, was previously found to contain strong anti-obesity activities in a high fat diet-induced obesity mouse model. One of its biological characteristics is that it inhibits 3T3-L1 adipocyte differentiation. To isolate the biologically active compound(s), conventional solvent fractionation was performed, and the various fractions were tested for anti-adipogenic activity using Oil Red O staining method. A single spot on thin layer chromatography of the chloroform fraction showed a potent anti-adipogenic activity. When purified, the structure of its major component was resolved as dehydrodiconiferyl alcohol (DHCA), a lignan, by NMR and mass spectrometry analysis. In 3T3-L1 cells, synthesized DHCA significantly reduced the expression of several adipocyte marker genes, including peroxisome proliferator-activated receptor γ (Pparg), CCAAT/enhancer-binding protein α (Cebpa), fatty acid-binding protein 4 (Fabp4), sterol response element-binding protein-1c (Srebp1c), and stearoyl-coenzyme A desaturase-1 (Scd), and decreased lipid accumulation without affecting cell viability. DHCA also suppressed the mitotic clonal expansion of preadipocytes (an early event of adipogenesis), probably by suppressing the DNA binding activity of C/EBPβ, and lowered the production level of cyclinA and cyclin-dependent kinase 2 (Cdk2), coinciding with the decrease in DNA synthesis and cell division. In addition, DHCA directly inhibited the expression of SREBP-1c and SCD-1. Similar observations were made, using primary mouse embryonic fibroblasts. Taken together, our data indicate that DHCA may contain dual activities, affecting both adipogenesis and lipogenesis.

  12. In vitro and in vivo enhancement of adipogenesis by Italian ryegrass (Lolium multiflorum in 3T3-L1 cells and mice.

    Directory of Open Access Journals (Sweden)

    Mariadhas Valan Arasu

    Full Text Available Adipogenesis is very much important in improving the quality of meat in animals. The aim of the present study was to investigate the in vitro and in vivo adipogenesis regulation properties of Lolium multiflorum on 3T3-L1 pre-adipocytes and mice. Chemical composition of petroleum ether extract of L. multiflorum (PET-LM confirmed the presence of fatty acids, such as α-linolenic acid, docosahexaenoic acid, oleic acid, docosatetraenoic acid, and caprylic acid, as the major compounds. PET-LM treatment increased viability, lipid accumulation, lipolysis, cell cycle progression, and DNA synthesis in the cells. PET-LM treatment also augmented peroxysome proliferator activated receptor (PPAR-γ2, CCAAT/enhancer binding protein-α, adiponectin, adipocyte binding protein, glucose transporter-4, fatty acid synthase, and sterol regulatory element binding protein-1 expression at mRNA and protein levels in differentiated adipocytes. In addition, mice administered with 200 mg/kg body weight PET-LM for 8 weeks showed greater body weight than control mice. These findings suggest that PET-LM facilitates adipogenesis by stimulating PPARγ-mediated signaling cascades in adipocytes which could be useful for quality meat development in animals.

  13. 小鼠3T3-L1前脂肪细胞系的增强绿色荧光蛋白标记%The Labeling of 3T3-L1 Preadipocyte Cells with Enhanced Green Fluorescent Protein

    Institute of Scientific and Technical Information of China (English)

    李成建; 成俊英; 张晓岚; 张崇本

    2004-01-01

    A cell model is desired for adipocyte differentiation investigation and for high-throughput screening of anti-obesity and anti-diabetes molecules from chemical resources due to the world wide epidemic of obesity and diabetes. In order to establish such a cell model, a plasmid of pPPARγ2-promoter-EGFP was constructed by inserting a 660bp sequence of mouse PPARγ2 promoter into the AseⅠ and KpnⅠ sites of pEGFP-N3 and transferred into 3T3-L1 preadipocyte cells. The cells were induced to differentiate and the expression of PPARγ2 was detected by the microscopic observation of EGFP and by RT-PCR assays. The results showed that the EGFP gene expression patterns were similar to that of pPPARγ2's, which indicated that the EGFP gene was transferred into the mouse 3T3-L1 preadipocyte cells, and its expression was under the control of pPPARγ2 promoter. RT-PCR assays showed that the EGFP expression authentically represented the stable expression of PPARγ2. In conclusion, a preadipocyte cell line expressing EGFP under the control of the promoter of adipocyte-specific expression gene PPARγ2 was generated. The cell line provides a powerful approach for the research of adipocyte differentiation and for the high-throughput screening of anti-obesity and anti-diabetes chemicals.%细胞模型是研究细胞分化原理以及进行高通量筛选的有效工具.为了建立特异性标记的脂肪细胞分化模型,构建了包括脂肪细胞分化特异性表达基因PPARγ2的启动子在内的载体(pPPARγ2-promoter-EGFP),用电穿孔方法转染小鼠3T3-L1 前脂肪细胞,用显微荧光观察和RT-PCR确认PPARγ2基因的内源表达.结果显示,EGFP基因成功转入3T3-L1前脂肪细胞,观察到细胞分化过程中EGFP表达和脂肪积累,RT-PCR分析表明EGFP代表了稳定而真实的PPARγ2基因的内源性表达.建立了由脂肪组织特异表达基因PPARγ2的表达控制的EGFP标记的小鼠3T3-L1前脂肪细胞系,目前国内外尚未见用同样

  14. 绿茶成分对3T3-L1细胞的细胞增殖及脂肪代谢的影响%Effects of Green Tea Components on Cell Proliferation and Lipid Metabolism in 3T3-L1 Cells

    Institute of Scientific and Technical Information of China (English)

    郑国栋; 徐峰; 吴少福; 张清峰; 邱阳阳

    2012-01-01

    目的:研究绿茶成分——儿茶素、咖啡碱和茶氨酸对3T3-L1前脂肪细胞的细胞增殖及脂肪代谢的影响.方法:测定不同浓度儿茶素、咖啡碱和荼氨酸对3T3-L1细胞增殖的影响,确定无毒性浓度.在分化诱导液中添加各绿茶成分后对3T3-L1细胞进行96h诱导分化,分化后第6天测定脂肪细胞中甘油三酯(TG)含量.细胞分化后第9天,单独添加绿茶成分或同时添加去甲肾上腺素(NA)24 h,分析对细胞中脂肪分解的影响.结果:添加20 μg/mL以上质量浓度的儿茶素能显著抑制3T3-L1细胞增殖,但质量浓度40,80 μg/mL的儿茶素对3T3-L1细胞有毒性作用,而160 μg/mL咖啡碱和茶氨酸对细胞增殖无明显影响.20 μg/mL儿茶素能显著抑制3T3-L1细胞中TG的合成,而咖啡碱和茶氨酸对细胞脂肪沉积无明显影响.与单独添加NA相比,同时添加咖啡碱能显著促进NA诱导细胞中脂肪分解的能力.结论:儿荼素抑制脂肪细胞的增殖和脂肪沉积,咖啡碱促进激素诱导脂肪分解.绿茶成分中儿茶素和咖啡碱对脂肪细胞内的脂肪代谢有调节作用.%Objective: To investigate the effects of green tea components, catechins, caffeine and theanine on 3T3-L1 cells proliferation and fat metabolism. Methods: 3T3—L1 cells were cultured in DMEM contained different concentrations of catechins, caffeine or theanine, and analyzed on cells proliferation and cytotoxicity. Then the differentiation of 3-T3—L1 cells were induced with these green tea components at noncytotoxic concentration for 96 hours. 6th day after differentiation, triglycerides (TG) in 3T3-L1 cells was measured. Lipolytic effect of green tea components in the present or absent of noradrenaline (NA) for 24 hours in 3T3-L1 cells was analyzed on 9th day after differentiation. Results: Above 20 μg/mL catechins significantly inhibited the proliferation of 3T3—L1 cells, but there was cytotoxic effect 40 and 80 μg/ mL. However, caffeine and

  15. Ginseng (Panax quinquefolius Reduces Cell Growth, Lipid Acquisition and Increases Adiponectin Expression in 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Chia-Rou Yeo

    2011-01-01

    Full Text Available An American ginseng (Panax quinquefolius extract (GE that contained a quantifiable amount of ginsenosides was investigated for the potential to inhibit proliferation, affect the cell cycle, influence lipid acquisition and adiponectin expression in 3T3-L1 cells. Six fingerprint ginsenosides were quantified by high performance liquid chromatography and the respective molecular weights were confirmed by LC-ESI-MS analysis. The extract contained Rg1 (347.3 ± 99.7 μg g−1, dry weight, Re (8280.4 ± 792.3 μg g−1, Rb1 (1585.8 ± 86.8 μg g−1, Rc (32.9 ± 8 μg g−1, Rb2 (62.6 ± 10.6 μg g−1 and Rd (90.4 ± 3.2 μg g−1. The GE had a dose-dependent effect on 3T3-L1 cell growth, the LC50 value was determined to be 40.3 ± 5 μg ml−1. Cell cycle analysis showed modest changes in the cell cycle. No significant changes observed in both G1 and G2/M phases, however there was a significant decrease (P<.05 in the S phase after 24 and 48 h treatment. Apoptotic cells were modest but significantly (P<.05 increased after 48 h (3.2 ± 1.0% compared to untreated control cells (1.5 ± 0.1%. Lipid acquisition was significantly reduced (P<.05 by 13 and 22% when treated at concentrations of 20.2 and 40.3 μg ml−1 compared to untreated control cells. In relation to adiponectin activation, western blot analysis showed that the protein expression was significantly (P<.05 increased at concentrations tested. A quantified GE reduced the growth of 3T3-L1 cells, down-regulated the accumulation of lipid and up-regulated the expression of adiponectin in the 3T3-L1 adipocyte cell model.

  16. Bisdemethoxycurcumin Inhibits Adipogenesis in 3T3-L1 Preadipocytes and Suppresses Obesity in High-Fat Diet-Fed C57BL/6 Mice.

    Science.gov (United States)

    Lai, Ching-Shu; Chen, Ying-Yi; Lee, Pei-Sheng; Kalyanam, Nagabhushanam; Ho, Chi-Tang; Liou, Wen-Shiung; Yu, Roch-Chui; Pan, Min-Hsiung

    2016-02-01

    Obesity is caused by excessive accumulation of body fat and is closely related to complex metabolic diseases. Adipogenesis is a key process that is required in adipocyte hypertrophy in the development of obesity. Curcumin (Cur) has been reported to inhibit adipocyte differentiation, but the inhibitory effects of other curcuminoids present in turmeric, such as demethoxycurcumin (DMC) and bisdemethoxycurcumin (BDMC), on adipogenesis have not been investigated. Here, we investigated the effects of curcuminoids on adipogenesis and the molecular mechanisms of adipocyte differentiation. Among three curcuminoids, BDMC was the most effective suppressor of lipid accumulation in adipocytes. BDMC suppressed adipogenesis in the early stage primarily through attenuation of mitotic clonal expansion (MCE). In BDMC-treated preadipocytes, cell cycle arrest at the G0/G1 phase was found after initiation of adipogenesis and was accompanied by downregulation of cyclin A, cyclin B, p21, and mitogen-activated protein kinase (MAPK) signaling. The protein levels of the adipogenic transcription factors peroxisome proliferator-activated receptor (PPAR)γ and CCAAT/enhancer-binding proteins (C/EBP)α were also reduced by BDMC treatment. Furthermore, 0.5% dietary BDMC (w/w) significantly lowered body weight gain and adipose tissue mass in high-fat diet (HFD)-fed mice. The results of H&E staining showed that dietary BDMC reduced hypertrophy in adipocytes. These results demonstrate for the first time that BDMC suppressed adipogenesis in 3T3-L1 adipocytes and prevented HFD-induced obesity. Our results suggest that BDMC has the potential to prevent obesity.

  17. Salicortin-Derivatives from Salix pseudo-lasiogyne Twigs Inhibit Adipogenesis in 3T3-L1 Cells via Modulation of C/EBPα and SREBP1c Dependent Pathway

    Directory of Open Access Journals (Sweden)

    Hong Pyo Kim

    2013-08-01

    Full Text Available Obesity is reported to be associated with excessive growth of adipocyte mass tissue as a result of increases in the number and size of adipocytes differentiated from preadipocytes. To search for anti-adipogenic phytochemicals, we screened for inhibitory activities of various plant sources on adipocyte differentiation in 3T3-L1 preadipocytes. Among the sources, a methanolic extract of Salix pseudo-lasiogyne twigs (Salicaceae reduced lipid accumulation in a concentration-dependent manner. During our search for anti-adipogenic constituents from S. pseudo-lasiogyne, five salicortin derivatives isolated from an EtOAc fraction of this plant and bearing 1-hydroxy-6-oxo-2-cyclohexene-carboxylate moieties, namely 2′,6′-O-acetylsalicortin (1, 2′-O-acetylsalicortin (2, 3′-O-acetylsalicortin (3, 6′-O-acetylsalicortin (4, and salicortin (5, were found to significantly inhibit adipocyte differentiation in 3T3-L1 cells. In particular, 2′,6′-O-acetylsalicortin (1 had the most potent inhibitory activity on adipocyte differentiation, with an IC50 value of 11.6 μM, and it significantly down-regulated the expressions of CCAAT/enhancer binding protein α (C/EBPα and sterol regulatory element binding protein 1 (SREBP1c. Furthermore, 2′,6′-O-acetylsalicortin (1 suppressed mRNA expression levels of C/EBPβ during the early stage of adipocyte differentiation and stearoyl coenzyme A desaturase 1 (SCD-1, acetyl-CoA carboxylase (ACC, and fatty acid synthase (FAS expression, target genes of SREBP1c. In the present study, we demonstrate that the anti-adipogenesis mechanism of 2′,6′-O-acetylsalicortin (1 may be mediated via down-regulation of C/EBPα and SREBP1c dependent pathways. Through their anti-adipogenic activity, salicortin derivatives may be potential novel therapeutic agents against obesity.

  18. Berberine increases expression of GATA-2 and GATA-3 during inhibition of adipocyte differentiation.

    Science.gov (United States)

    Hu, Y; Davies, G E

    2009-09-01

    It is known that a number of transcription factors are key regulators in the complex process of adipocyte differentiation including peroxisome proliferator activated receptor gamma (PPARgamma) and the CCAAT enhancer binding protein alpha (C/EBPalpha). Studies have demonstrated that in pre-adipocyte 3T3-L1 cells constitutive expression of the DNA binding proteins GATA-2 and GATA-3 results in protein/protein interactions with C/EBPalpha resulting in down regulation of PPARgamma and subsequent suppressed adipocyte differentiation with cells trapped at the pre-adipocyte stage. Thus it appears that GATA-2 and GATA-3 are of critical importance in regulating adipocyte differentiation through molecular interactions with PPARgamma and C/EBPalpha. Recent reports suggest that berberine, an isoquinoline derivative alkaloid isolated from many medicinal herbs prevents differentiation of 3T3-L1 cells via a down regulation of PPARgamma and C/EBPalpha expression. The aim of this study was to determine the effect of berberine on GATA-2 and 3 gene and protein expression levels during differentiation of 3T3-L1 cells. MTT (Methylthiazolyldiphenyl-tetrazolium bromide) was used to detect the cytotoxic effects of berberine on the viability of 3T3-L1 cells during proliferation and differentiation. Differentiation of 3T3-L1 cells was monitored by Oil Red O staining and RT-PCR of PPARgamma and C/EBPalpha and the expression of GATA-2 and 3 was determined by RT-PCR and Western Blot. Results show that following treatment with 8microM berberine the mRNA and protein expression levels of GATA-2 and 3 were elevated and accompanied by inhibited adipocyte differentiation. These results may lead to the use of berberine to target the induction of specific genes such as GATA-2 and GATA-3 which affect adipocyte differentiation.

  19. Effect of Mangiferin and Mahanimbine on Glucose Utilization in 3T3-L1 cells

    Science.gov (United States)

    Kumar, B Dinesh; Krishnakumar, K; Jaganathan, Saravana Kumar; Mandal, Mahitosh

    2013-01-01

    Background: Stem barks of Mangifera indica contain a rich content of mangiferin (xanthone glucoside), whereas Murraya koenigii leaves contain rich sources of mahanimbine (carbazole alkaloid) and used traditionally for the treatment of diabetes. Objective: To investigate the effects of mangiferin (xanthone glucoside) and mahanimbine (carbazole alkaloid) on glucose utilization in 3T3-L1 cells. Materials and Methods: Mangiferin was isolated from stem barks of Mangifera indica and mahanimbine was isolated from Murraya koenigii leaves. These isolated compounds were subjected to MTT assay and glucose utilization test with 3T3-L1 cells. Results: Treatment of the 3T3-L1 cells with mangiferin and mahanimbine increased the glucose utilization in a dose-dependent manner. At a concentration of 1 mM, mangniferin showed 2-fold increase in glucose utilization compared with untreated control. In case of mahanimbine, the observed effect at 1 mM was almost equivalent to positive control (insulin at 1 μM). Moreover, MTT assay showed that both of these compounds were less toxic at a concentration of 1 mM (nearly 75% cells are viable). Conclusion: The present results indicated that these natural products (mangiferin and mahanimbine) exhibited potential ethnomedical uses in management of diabetes. PMID:23661997

  20. Benzyl butyl phthalate promotes adipogenesis in 3T3-L1 preadipocytes: A High Content Cellomics and metabolomic analysis.

    Science.gov (United States)

    Yin, Lei; Yu, Kevin Shengyang; Lu, Kun; Yu, Xiaozhong

    2016-04-01

    Benzyl butyl phthalate (BBP) has been known to induce developmental and reproductive toxicity. However, its association with dysregulation of adipogenesis has been poorly investigated. The present study aimed to examine the effect of BBP on the adipogenesis, and to elucidate the underlying mechanisms using the 3T3-L1 cells. The capacity of BBP to promote adipogenesis was evaluated by multiple staining approaches combined with a High Content Cellomics analysis. The dynamic changes of adipogenic regulatory genes and proteins were examined, and the metabolite profile was identified using GC/MC based metabolomic analysis. The High Content analysis showed BBP in contrast with Bisphenol A (BPA), a known environmental obesogen, increased lipid droplet accumulation in a similar dose-dependent manner. However, the size of the lipid droplets in BBP-treated cells was significantly larger than those in cells treated with BPA. BBP significantly induced mRNA expression of transcriptional factors C/EBPα and PPARγ, their downstream genes, and numerous adipogenic proteins in a dose and time-dependent manner. Furthermore, GC/MC metabolomic analysis revealed that BBP exposure perturbed the metabolic profiles that are associated with glyceroneogenesis and fatty acid synthesis. Altogether, our current study clearly demonstrates that BBP promoted the differentiation of 3T3-L1 through the activation of the adipogenic pathway and metabolic disturbance.

  1. Berberine Alleviates Olanzapine-Induced Adipogenesis via the AMPKα-SREBP Pathway in 3T3-L1 Cells.

    Science.gov (United States)

    Li, Yanjie; Zhao, Xiaomin; Feng, Xiyu; Liu, Xuemei; Deng, Chao; Hu, Chang-Hua

    2016-11-09

    The aim of this study was to investigate the mechanisms underlying the inhibitory effects of berberine (BBR) on olanzapine (OLZ)-induced adipogenesis in a well-replicated 3T3-L1 cell model. Oil-Red-O (ORO) staining showed that BBR significantly decreased OLZ-induced adipogenesis. Co-treatment with OLZ and BBR decreased the accumulation of triglyceride (TG) and total cholesterol (TC) by 55.58% ± 3.65% and 49.84% ± 8.31%, respectively, in 3T3-L1 adipocytes accompanied by reduced expression of Sterol regulatory element binding proteins 1 (SREBP1), fatty acid synthase (FAS), peroxisome proliferator activated receptor-γ (PPARγ), SREBP2, low-density lipoprotein receptor (LDLR), and hydroxymethylglutaryl-coenzyme A reductase (HMGR) genes compared with OLZ alone. Consistently, the co-treatment downregulated protein levels of SREBP1, SREBP2, and LDLR by 57.71% ± 9.42%, 73.05% ± 11.82%, and 59.46% ± 9.91%, respectively. In addition, co-treatment reversed the phosphorylation level of AMP-activated protein kinase-α (AMPKα), which was reduced by OLZ, determined via the ratio of pAMPKα:AMPKα (94.1%) compared with OLZ alone. The results showed that BBR may prevent lipid metabolism disorders caused by OLZ by reversing the degree of SREBP pathway upregulated and the phosphorylation of AMPKα downregulated. Collectively, these results indicated that BBR could be used as a potential adjuvant to prevent dyslipidemia and obesity caused by the use of second-generation antipsychotic medication.

  2. Berberine Alleviates Olanzapine-Induced Adipogenesis via the AMPKα–SREBP Pathway in 3T3-L1 Cells

    Science.gov (United States)

    Li, Yanjie; Zhao, Xiaomin; Feng, Xiyu; Liu, Xuemei; Deng, Chao; Hu, Chang-Hua

    2016-01-01

    The aim of this study was to investigate the mechanisms underlying the inhibitory effects of berberine (BBR) on olanzapine (OLZ)-induced adipogenesis in a well-replicated 3T3-L1 cell model. Oil-Red-O (ORO) staining showed that BBR significantly decreased OLZ-induced adipogenesis. Co-treatment with OLZ and BBR decreased the accumulation of triglyceride (TG) and total cholesterol (TC) by 55.58% ± 3.65% and 49.84% ± 8.31%, respectively, in 3T3-L1 adipocytes accompanied by reduced expression of Sterol regulatory element binding proteins 1 (SREBP1), fatty acid synthase (FAS), peroxisome proliferator activated receptor-γ (PPARγ), SREBP2, low-density lipoprotein receptor (LDLR), and hydroxymethylglutaryl-coenzyme A reductase (HMGR) genes compared with OLZ alone. Consistently, the co-treatment downregulated protein levels of SREBP1, SREBP2, and LDLR by 57.71% ± 9.42%, 73.05% ± 11.82%, and 59.46% ± 9.91%, respectively. In addition, co-treatment reversed the phosphorylation level of AMP-activated protein kinase-α (AMPKα), which was reduced by OLZ, determined via the ratio of pAMPKα:AMPKα (94.1%) compared with OLZ alone. The results showed that BBR may prevent lipid metabolism disorders caused by OLZ by reversing the degree of SREBP pathway upregulated and the phosphorylation of AMPKα downregulated. Collectively, these results indicated that BBR could be used as a potential adjuvant to prevent dyslipidemia and obesity caused by the use of second-generation antipsychotic medication. PMID:27834848

  3. Berberine Alleviates Olanzapine-Induced Adipogenesis via the AMPKα–SREBP Pathway in 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Yanjie Li

    2016-11-01

    Full Text Available The aim of this study was to investigate the mechanisms underlying the inhibitory effects of berberine (BBR on olanzapine (OLZ-induced adipogenesis in a well-replicated 3T3-L1 cell model. Oil-Red-O (ORO staining showed that BBR significantly decreased OLZ-induced adipogenesis. Co-treatment with OLZ and BBR decreased the accumulation of triglyceride (TG and total cholesterol (TC by 55.58% ± 3.65% and 49.84% ± 8.31%, respectively, in 3T3-L1 adipocytes accompanied by reduced expression of Sterol regulatory element binding proteins 1 (SREBP1, fatty acid synthase (FAS, peroxisome proliferator activated receptor-γ (PPARγ, SREBP2, low-density lipoprotein receptor (LDLR, and hydroxymethylglutaryl-coenzyme A reductase (HMGR genes compared with OLZ alone. Consistently, the co-treatment downregulated protein levels of SREBP1, SREBP2, and LDLR by 57.71% ± 9.42%, 73.05% ± 11.82%, and 59.46% ± 9.91%, respectively. In addition, co-treatment reversed the phosphorylation level of AMP-activated protein kinase-α (AMPKα, which was reduced by OLZ, determined via the ratio of pAMPKα:AMPKα (94.1% compared with OLZ alone. The results showed that BBR may prevent lipid metabolism disorders caused by OLZ by reversing the degree of SREBP pathway upregulated and the phosphorylation of AMPKα downregulated. Collectively, these results indicated that BBR could be used as a potential adjuvant to prevent dyslipidemia and obesity caused by the use of second-generation antipsychotic medication.

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

    NARCIS (Netherlands)

    Fleuren, W.W.M.; Linssen, M.M.; Toonen, E.J.M.; Zon, G.C. van der; Guigas, B.; Vlieg, J. de; Dokter, W.H.; Ouwens, D.M.; Alkema, W.

    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 t

  5. Resveratrol inhibits lipogenesis of 3T3-L1 and SGBS cells by inhibition of insulin signaling and mitochondrial mass increase.

    Science.gov (United States)

    Li, Shuijie; Bouzar, Célia; Cottet-Rousselle, Cécile; Zagotta, Ivana; Lamarche, Frédéric; Wabitsch, Martin; Tokarska-Schlattner, Malgorzata; Fischer-Posovszky, Pamela; Schlattner, Uwe; Rousseau, Denis

    2016-06-01

    Resveratrol is attracting much interest because of its potential to decrease body weight and increase life span, influencing liver and muscle function by increasing mitochondrial mass and energy expenditure. Even though resveratrol was already shown to reduce the adipose tissue mass in animal models, its effects on mitochondrial mass and network structure in adipocytes have not yet been studied. For this purpose, we investigated the effect of resveratrol on mitochondrial mass increase and remodeling during adipogenic differentiation of two in vitro models of adipocyte biology, the murine 3T3-L1 cell line and the human SGBS cell strain. We confirm that resveratrol inhibits lipogenesis in differentiating adipocytes, both mouse and human. We further show that this is linked to inhibition of the normally observed mitochondrial mass increase and mitochondrial remodeling. At the molecular level, the anti-lipogenic effect of resveratrol seems to be mediated by a blunted expression increase and an inhibition of acetyl-CoA carboxylase (ACC). This is one of the consequences of an inhibited insulin-induced signaling via Akt, and maintained signaling via AMP-activated protein kinase. The anti-lipogenic effect of resveratrol is further modulated by expression levels of mitochondrial ATAD3, consistent with the emerging role of this protein as an important regulator of mitochondrial biogenesis and lipogenesis. Our data suggest that resveratrol acts on differentiating preadipocytes by inhibiting insulin signaling, mitochondrial biogenesis, and lipogenesis, and that resveratrol-induced reduction of mitochondrial biogenesis and lipid storage contribute to adipose tissue weight loss in animals and humans.

  6. Effect of xanthohumol and isoxanthohumol on 3T3-L1 cell apoptosis and adipogenesis.

    Science.gov (United States)

    Yang, Jeong-Yeh; Della-Fera, Mary Anne; Rayalam, Srujana; Baile, Clifton A

    2007-11-01

    Xanthohumol (XN), the chalcone from beer hops has several biological activities. XN has been shown to induce apoptosis in cancer cells and also has been reported to be involved in lipid metabolism. Based on these studies and our previous work with natural compounds, we hypothesized that XN and its isomeric flavanone, isoxanthohumol (IXN), would induce apoptosis in adipocytes through the mitochondrial pathway and would inhibit maturation of preadipocytes. Adipocytes were treated with various concentrations of XN or IXN. In mature adipocytes both XN and IXN decreased viability, increased apoptosis and increased ROS production, XN being more effective. Furthermore, the antioxidants ascorbic acid and 2-mercaptoethanol prevented XN and IXN-induced ROS generation and apoptosis. Immunoblotting analysis showed an increase in the levels of cytoplasmic cytochrome c and cleaved poly (ADP-ribose) polymerase (PARP) by XN and IXN. Concomitantly, we observed activation of the effectors caspase-3/7. In maturing preadipocytes both XN and IXN were effective in reducing lipid content, XN being more potent. Moreover, the major adipocyte marker proteins such as PPARgamma, C/EBPalpha, and aP2 decreased after treatment with XN during the maturation period and that of DGAT1 decreased after treatment with XN and IXN. Taken together, our data indicate that both XN and IXN inhibit differentiation of preadipocytes, and induce apoptosis in mature adipocytes, but XN is more potent.

  7. Transcriptional and epigenetic mechanisms underlying enhanced in vitro adipocyte differentiation by the brominated flame retardant BDE-47

    DEFF Research Database (Denmark)

    Kamstra, Jorke H; Hruba, Eva; Blumberg, Bruce;

    2014-01-01

    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 mech......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...

  8. Effect of Ichnocarpus frutescens (L.) R.Br. hexane extract on preadipocytes viability and lipid accumulation in 3T3-L1 cells

    Institute of Scientific and Technical Information of China (English)

    M Saravanan; S Ignacimuthu

    2013-01-01

    Objective: To investigate the crude extracts of Ichnocarpus frutescens (I. frutescens) for antiobesity effect. Methods: Leaves of I. frutescens were sequentially extracted with hexane, ethyl acetate, and methanol and their effect on viability of 3T3-L1 preadipocytes were evaluated. Based on this the apoptosis on preadipocytes was confirmed by DNA fragmentation and LDH (Lactate dehydrogenase) leakage assays. Anti-adipogenesis was performed by oil red O (ORO) staining and free glycerol release in the medium of differentiated adipocytes. Results: The hexane extract of I. frutescens (IFHE) inhibited cell viability in a time- and dose-related manner. An increased release of LDH, as a marker of membrane integrity, was observed at a dose of 200 μg/mL. The discontinuous DNA fragments on agarose gel electrophoresis showed the apoptotic effect of the IFHE. Morphological observations of cells stained with ORO showed a decrease in cellular lipid content at the concentrations tested compared to the induced control cells. In the experiment of lipolytic activity, treatment with IFHE enhanced glycerol secretion with the rates of approximately 28%, 55%, and 46% at the concentrations of 100, 200 and 300 μg/mL, respectively. Conclusions:The observed properties clearly revealed the medicinal property of I. frutescens in the treatment of obesity.

  9. ATF3 represses PPARγ expression and inhibits adipocyte differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Min-Kyung; Jung, Myeong Ho, E-mail: jung0603@pusan.ac.kr

    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

  10. Insulin like growth factor-1/insulin bypasses Pref-1/FA1-mediated inhibition of adipocyte differentiation

    DEFF Research Database (Denmark)

    Zhang, Hongbin; Nøhr, Jane; Jensen, Charlotte Harken;

    2003-01-01

    of Pref-1/FA1 in 3T3-L1 or 3T3-F442A cells inhibited adipocyte differentiation when insulin or insulin-like growth factor-1 (IGF-1) was omitted from the differentiation mixture. We demonstrate that the level of the mature form of the IGF-1 receptor is reduced and that IGF-1-dependent activation of p42/p44......, and adipocyte differentiation in a dose-dependent manner. Udgivelsesdato: 2003-Jun-6......Pref-1 is a highly glycosylated Delta-like transmembrane protein containing six epidermal growth factor-like repeats in the extracellular domain. Pref-1 is abundantly expressed in preadipocytes, but expression is down-regulated during adipocyte differentiation. Forced expression of Pref-1 in 3T3-L1...

  11. Effects of different fatty acids and dietary lipids on adiponectin gene expression in 3T3-L1 cells and C57BL/6J mice adipose tissue.

    Science.gov (United States)

    Bueno, Allain Amador; Oyama, Lila Missae; de Oliveira, Cristiane; Pisani, Luciana Pelegrini; Ribeiro, Eliane Beraldi; Silveira, Vera Lucia Flor; Oller do Nascimento, Cláudia Maria

    2008-01-01

    Obesity is positively correlated to dietary lipid intake, and the type of lipid may play a causal role in the development of obesity-related pathologies. A major protein secreted by adipose tissue is adiponectin, which has antiatherogenic and antidiabetic properties. The aim of this study was to evaluate the effects of four different high-fat diets (enriched with soybean oil, fish oil, coconut oil, or lard) on adiponectin gene expression and secretion by the white adipose tissue (WAT) of mice fed on a selected diet for either 2 (acute treatment) or 60 days (chronic treatment). Additionally, 3T3-L1 adipocytes were treated for 48 h with six different fatty acids: palmitic, linoleic, eicosapentaenoic (EPA), docosahexaenoic (DHA), lauric, or oleic acid. Serum adiponectin concentration was reduced in the soybean-, coconut-, and lard-enriched diets in both groups. Adiponectin gene expression was lower in retroperitoneal WAT after acute treatment with all diets. The same reduction in levels of adiponectin gene expression was observed in epididymal adipose tissue of animals chronically fed soybean and coconut diets and in 3T3-L1 cells treated with palmitic, linoleic, EPA, and DHA acids. These results indicate that the intake of certain fatty acids may affect serum adiponectin levels in mice and adiponectin gene expression in mouse WAT and 3T3-L1 adipocytes. The effects appear to be time dependent and depot specific. It is postulated that the downregulation of adiponectin expression by dietary enrichment with soybean oil or coconut oil may contribute to the development of insulin resistance and atherosclerosis.

  12. Flavonoids from Triticum aestivum inhibit adipogenesis in 3T3-L1 cells by upregulating the insig pathway.

    Science.gov (United States)

    Poudel, Barun; Nepali, Sarmila; Xin, Mingjie; Ki, Hyeon-Hui; Kim, Young-Ho; Kim, Dae-Ki; Lee, Young-Mi

    2015-08-01

    The present study aimed to compare the potential anti-adipogenic effects and underlying mechanisms of the luteolin, isoscoparin and isoorientin flavonoids, purified from Triticum aestivum sprout (TA) in 3T3-L1 cells. The cells were treated with different concentrations of flavonoids for 8 days and the lipid accumulation was assessed using Oil-Red-O staining. The expression levels of the transcription factors and the genes involved in adipogenesis in the cells were assessed by reverse transcription-quantitative polymerase chain reaction and western blotting. The results demonstrated that 10 μM luteolin, isoscoparin or isoorientin inhibited lipid deposition in the cells by 74, 63 and 65%, respectively. The flavonoids also significantly inhibited the transcriptional regulators of adipogenesis, including peroxisome proliferator-activated receptor-γ, CAAT/enhancer binding protein-α and sterol regulatory element binding protein (SREBP)-1c, compared with the control cells. Similarly, there was a significant downregulation of the adipocyte specific markers associated with lipid metabolism, including activating protein-2, fatty acid synthase, hormone-sensitive lipase and lipoprotein lipase, in the flavonoid treated cells. Notably, the cells treated with the flavonoids demonstrated increased expression levels of the insulin-induced genes, insig-1 and insig-2, which may have inhibited the activation of the adipogenic transcription factor, SREBP, eventually leading to the inhibition of adipogenesis. Taken together, these results revealed that the flavonoids from TA possessed an inhibitory effect on adipogenesis through downregulation of adipogenic transcription factors and genes associated with lipid metabolism, and the upregulation of insig 1 and 2, suggesting that the flavonoids from TA may be potential therapeutic agents for the prevention and treatment of obesity.

  13. N(ϵ) -Carboxymethyllysine Increases the Expression of miR-103/143 and Enhances Lipid Accumulation in 3T3-L1 Cells.

    Science.gov (United States)

    Holik, Ann-Katrin; Lieder, Barbara; Kretschy, Nicole; Somoza, Mark M; Held, Sandra; Somoza, Veronika

    2016-10-01

    Advanced glycation endproducts, formed in vivo, but also by the Maillard reaction upon thermal treatment of foods, have been associated with the progression of pathological conditions such as diabetes mellitus. In addition to the accumulation with age, exogenous AGEs are introduced into the circulation from dietary sources. In this study, we investigated the effects of addition of free N(ϵ) -carboxymethyllysine (CML), a well-characterized product of the Maillard reaction, on adipogenesis in 3T3-L1 preadipocytes. Treatment with 5, 50, or 500 μM CML resulted in increased lipid accumulation to similar extents, by 11.5 ± 12.6%, 12.9 ± 8.6%, and 12.8 ± 8.5%, respectively. Long-term treatment with 500 μM CML during adipogenesis resulted in increases in miR-103 and miR-143 levels, two miRNAs described to be involved in impaired glucose homeostasis and increased lipid accumulation. Furthermore, the expression of genes associated with these miRNAs, consisting of Akt1, PI3k, and Cav1 was regulated by CML. Short-term treatment of mature 3T3-L1 adipocytes with CML resulted in decreased basal glucose uptake. These results, indicate that the addition of protein-free CML to 3T3-L1 cells influence parameters associated with adipogenesis and glucose homeostasis at transcriptional, and functional level; this indicates that free CML derived from exogenous sources, in addition to protein-bound CML may be relevant in this context. J. Cell. Biochem. 117: 2413-2422, 2016. © 2016 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.

  14. Soy pinitol acts partly as an insulin sensitizer or insulin mediator in 3T3-L1 preadipocytes

    OpenAIRE

    Do, Gyeong-Min; Choi, Myung-Sook; Kim, Hye-Jin; Woo, Myung-Nam; Lee, Mi-Kyung; Jeon, Seon-Min

    2007-01-01

    The blood glucose-lowering property of pinitol is mediated via the insulin signaling pathway. This study was carried out to evaluate the effects of soy pinitol on adipogenesis in a 3T3-L1 cell line; 3T3-L1 preadipocytes were treated with pinitol (0–1 mM) together with insulin for 9 days. The regulation of lipid metabolism was assessed by oil-red-O staining of intracellular lipids and real-time PCR of adipogenesis-related factors. The inhibition of cell proliferation was estimated by MTT assay...

  15. 10e12z CLA alters adipocyte differentiation and adipocyte cytokine expression and induces macrophage proliferation.

    Science.gov (United States)

    Belda, Benjamin J; Thompson, Jerry T; Eser, Pinar O; Vanden Heuvel, John P

    2012-05-01

    The trans-10, cis-12 (10e12z) conjugated linoleic acid (CLA) isomer of CLA is responsible for loss of lipid storage or adipose tissue in vitro or in vivo. This isomer also induces inflammatory signaling in both mouse and human adipocytes in vitro. However, when these events occur and whether they are significant enough to affect other cell types are unclear. In these experiments, the 3T3-L1 cell line has been used to examine the interaction between inflammatory signaling and decreased differentiation or lipid storage induced by 10e12z CLA. In assays measuring both lipid accumulation and gene expression, differentiating 3T3-L1 cells exhibit concurrent induction of inflammatory signaling, as measured by cyclooxygenase-2 expression, and a decrease in adipocyte marker gene expression. Furthermore, in fully differentiated adipocytes, as identified in microarray assays and confirmed with real-time polymerase chain reaction, 10e12z CLA also significantly affected expression of both matrix metalloprotein-3 (MMP-3), collagen VI α 3 ColVI alpha 3 (VIα3) and the cytokine epiregulin, demonstrating that the effects of 10e12z broadly impact adipocyte function. In agreement with other experimental systems, 10e12z CLA inhibited RAW 264.7 cell proliferation; however, in response to adipocyte-conditioned media, 10e12z-CLA-treated adipocytes induced proliferation of this cell line, suggesting that the effect of 10e12z CLA is context dependent. These results are largely consistent with the known activation of the inflammatory mediator nuclear factor-κB in adipocytes in vitro and in vivo by 10e12z CLA treatment and demonstrate that adipose is an important target tissue of this isomer that impacts other cell types.

  16. Estrogen-related receptor alpha modulates the expression of adipogenesis-related genes during adipocyte differentiation.

    Science.gov (United States)

    Ijichi, Nobuhiro; Ikeda, Kazuhiro; Horie-Inoue, Kuniko; Yagi, Ken; Okazaki, Yasushi; Inoue, Satoshi

    2007-07-06

    Estrogen-related receptor alpha (ERRalpha) 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 ERRalpha in adipogenesis and white adipose tissue development has not been well studied. Here, we show that ERRalpha and ERRalpha-related transcriptional coactivators, peroxisome proliferator-activated receptor gamma (PPARgamma) coactivator-1alpha (PGC-1alpha) and PGC-1beta, 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 ERRalpha-specific siRNA results in mRNA down-regulation of fatty acid binding protein 4, PPARgamma, and PGC-1alpha in 3T3-L1 cells in the adipogenesis medium. ERRalpha and PGC-1beta 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 ERRalpha in 3T3-L1 cells up-regulates adipogenic marker genes and promotes triglyceride accumulation during 3T3-L1 differentiation. These results suggest that ERRalpha may play a critical role in adipocyte differentiation by modulating the expression of various adipogenesis-related genes.

  17. Molecular mechanism of 9-cis-retinoic acid inhibition of adipogenesis in 3T3-L1 cells

    Energy Technology Data Exchange (ETDEWEB)

    Sagara, Chiaki; Takahashi, Katsuhiko [Laboratory of Physiological Chemistry, Institute of Medicinal Chemistry, Hoshi University, Shinagawa, Tokyo 142-8501 (Japan); Kagechika, Hiroyuki [School of Biomedical Science, Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Chiyoda, Tokyo 101-0062 (Japan); Takahashi, Noriko, E-mail: t-noriko@hoshi.ac.jp [Laboratory of Physiological Chemistry, Institute of Medicinal Chemistry, Hoshi University, Shinagawa, Tokyo 142-8501 (Japan)

    2013-03-29

    Highlights: ► We examined the effects of 9-cis-RA on adipogenesis in mouse preadipocyte 3T3-L1. ► 9-cis-RA inhibited lipid accumulation in adipogenetically-induced 3T3-L1 cells. ► A RXR pan-antagonist suppressed the inhibitory effects of 9-cis-RA on adipogenesis. ► This antagonist had no effects on RXRα and PPARγ levels in 9-cis-RA-treated cells. ► 9-cis-RA-induced decrease in both RXRα and PPARγ was independent of RXR activation. -- Abstract: Retinoic acid (RA) signaling is mediated by specific nuclear hormone receptors. Here we examined the effects of 9-cis-RA on adipogenesis in mouse preadipocyte 3T3-L1 cells. 9-cis-RA inhibits the lipid accumulation of adipogenetically induced 3T3-L1 cells. The complex of retinoid X receptor α (RXRα) with peroxisome proliferator-activated receptor γ (PPARγ) is a major transcription factor in the process of adipogenesis, and the levels of these molecules were decreased by 9-cis-RA treatment. A RXR pan-antagonist suppressed 9-cis-RA’s inhibitory effects on adipogenesis, but not on the intracellular levels of both RXRα and PPARγ. These results suggest that 9-cis-RA could inhibit adipogenesis by activating RXR, and decrease both RXR and PPARγs levels in a RXR activation-independent manner.

  18. A Herbal Formula HT048, Citrus unshiu and Crataegus pinnatifida, Prevents Obesity by Inhibiting Adipogenesis and Lipogenesis in 3T3-L1 Preadipocytes and HFD-Induced Obese Rats

    Directory of Open Access Journals (Sweden)

    Yoon Hee Lee

    2015-05-01

    Full Text Available HT048 is a combination composed of Crataegus pinnatifida leaf and Citrus unshiu peel extracts. This study aimed to investigate potential anti-obesity effect of the combination. The 3T3-L1 adipocytes were treated with different doses of HT048 and triglyceride accumulation, glycerol release and adipogenesis-related genes were analyzed. For in vivo study, male Sprague Dawley rats were divided according to experimental diets: the chow diet group, the high-fat diet (HFD group, the HFD supplemented with orlistat group, the HFD supplemented with HT048 group (0.2% or 0.4% for 12 weeks. We measured the body weight, serum lipid levels and the expression of genes involved lipid metabolism. HT048 treatment dose-dependently suppressed adipocyte differentiation and stimulated glycerol release. The expressions of PPARγ and C/EBPα mRNA were decreased by HT048 treatment in adipocytes. HT048 supplementation significantly reduced the body and fat weights in vivo. Serum lipid levels were significantly lower in the HT048 supplemented groups than those of the HFD group. Expression of the hepatic lipogenesis-related genes were decreased and expression of the β-oxidation-related genes were increased in rats fed HT048 compared to that of animals fed HFD. These results suggest that HT048 has a potential benefit in preventing obesity through the inhibition of lipogenesis and adipogenesis.

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

    Directory of Open Access Journals (Sweden)

    Yang Jiao

    2016-02-01

    Full Text Available 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.

  20. The edible red alga, Gracilaria verrucosa, inhibits lipid accumulation and ROS production, but improves glucose uptake in 3T3-L1 cells.

    Science.gov (United States)

    Woo, Mi-Seon; Choi, Hyeon-Son; Lee, Ok-Hwan; Lee, Boo-Yong

    2013-07-01

    Gracilaria verrucosa is a red alga that is widely distributed in seaside areas of many countries. We examined the effect of G. verrucosa extract on adipogenesis, reactive oxygen species (ROS) production, and glucose uptake in 3T3-L1 cells. Oil red O staining and a nitroblue tetrazolium assay showed that G. verrucosa extract inhibited lipid accumulation and ROS production, respectively. mRNA levels of adipogenic transcription factors, peroxisome proliferator-activated receptor gamma and CCAAT/enhancer-binding protein alpha, as well as of their target gene, adipocyte protein 2, were reduced upon treatment with G. verrucosa extract. However, G. verrucosa extract increased glucose uptake, glucose transporter-4 expression, and AMP-activated protein kinaseα (AMPKα) phosphorylation compared to the control. Our results suggest that the anti-adipogenic and insulin-sensitive effects of G. verrucosa extract can be recapitulated to activation of AMPKα.

  1. Characterization of adipocyte differentiation from human mesenchymal stem cells in bone marrow

    Directory of Open Access Journals (Sweden)

    Huang Hai-Yan

    2010-05-01

    Full Text Available Abstract Background Adipocyte hyperplasia is associated with obesity and arises due to adipogenic differentiation of resident multipotent stem cells in the vascular stroma of adipose tissue and remote stem cells of other organs. The mechanistic characterization of adipocyte differentiation has been researched in murine pre-adipocyte models (i.e. 3T3-L1 and 3T3-F442A, revealing that growth-arrest pre-adipocytes undergo mitotic clonal expansion and that regulation of the differentiation process relies on the sequential expression of three key transcription factors (C/EBPβ, C/EBPα and PPARγ. However, the mechanisms underlying adipocyte differentiation from multipotent stem cells, particularly human mesenchymal stem cells (hBMSCs, remain poorly understood. This study investigated cell cycle regulation and the roles of C/EBPβ, C/EBPα and PPARγ during adipocyte differentiation from hBMSCs. Results Utilising a BrdU incorporation assay and manual cell counting it was demonstrated that induction of adipocyte differentiation in culture resulted in 3T3-L1 pre-adipocytes but not hBMSCs undergoing mitotic clonal expansion. Knock-down and over-expression assays revealed that C/EBPβ, C/EBPα and PPARγ were required for adipocyte differentiation from hBMSCs. C/EBPβ and C/EBPα individually induced adipocyte differentiation in the presence of inducers; PPARγ alone initiated adipocyte differentiation but the cells failed to differentiate fully. Therefore, the roles of these transcription factors during human adipocyte differentiation are different from their respective roles in mouse. Conclusions The characteristics of hBMSCs during adipogenic differentiation are different from those of murine cells. These findings could be important in elucidating the mechanisms underlying human obesity further.

  2. Soy pinitol acts partly as an insulin sensitizer or insulin mediator in 3T3-L1 preadipocytes.

    Science.gov (United States)

    Do, Gyeong-Min; Choi, Myung-Sook; Kim, Hye-Jin; Woo, Myung-Nam; Lee, Mi-Kyung; Jeon, Seon-Min

    2008-02-01

    The blood glucose-lowering property of pinitol is mediated via the insulin signaling pathway. This study was carried out to evaluate the effects of soy pinitol on adipogenesis in a 3T3-L1 cell line; 3T3-L1 preadipocytes were treated with pinitol (0-1 mM) together with insulin for 9 days. The regulation of lipid metabolism was assessed by oil-red-O staining of intracellular lipids and real-time PCR of adipogenesis-related factors. The inhibition of cell proliferation was estimated by MTT assay. Pinitol treatment did not inhibit lipid accumulation, nor did it affect expression of adipogenesis-related factors, including ADD1, aP2 and FAS, in a dose-dependent manner. Expression of adiponectin, GLUT4, IRS, C/EBPalpha and PPARgamma mRNAs, however, increased in cells treated with 0.5 mM and/or 1 mM pinitol. Pinitol treatment did not affect the inhibition of cell growth and proliferation in a dose-dependent manner. Accordingly, we suggest that pinitol is nontoxic to this cell line, and that it enhances adipogenesis by acting as an insulin sensitizer or insulin mediator via the upregulation of adiponectin, GLUT4, IRS, C/EBPalpha and PPARgamma in 3T3-L1 preadipocytes.

  3. Pmch-deficiency in rats is associated with normal adipocyte differentiation and lower sympathetic adipose drive.

    Science.gov (United States)

    Mul, Joram D; O'Duibhir, Eoghan; Shrestha, Yogendra B; Koppen, Arjen; Vargoviç, Peter; Toonen, Pim W; Zarebidaki, Eleen; Kvetnansky, Richard; Kalkhoven, Eric; Cuppen, Edwin; Bartness, Timothy J

    2013-01-01

    The orexigenic neuropeptide melanin-concentrating hormone (MCH), a product of Pmch, is an important mediator of energy homeostasis. Pmch-deficient rodents are lean and smaller, characterized by lower food intake, body-, and fat mass. Pmch is expressed in hypothalamic neurons that ultimately are components in the sympathetic nervous system (SNS) drive to white and interscapular brown adipose tissue (WAT, iBAT, respectively). MCH binds to MCH receptor 1 (MCH1R), which is present on adipocytes. Currently it is unknown if Pmch-ablation changes adipocyte differentiation or sympathetic adipose drive. Using Pmch-deficient and wild-type rats on a standard low-fat diet, we analyzed dorsal subcutaneous and perirenal WAT mass and adipocyte morphology (size and number) throughout development, and indices of sympathetic activation in WAT and iBAT during adulthood. Moreover, using an in vitro approach we investigated the ability of MCH to modulate 3T3-L1 adipocyte differentiation. Pmch-deficiency decreased dorsal subcutaneous and perirenal WAT mass by reducing adipocyte size, but not number. In line with this, in vitro 3T3-L1 adipocyte differentiation was unaffected by MCH. Finally, adult Pmch-deficient rats had lower norepinephrine turnover (an index of sympathetic adipose drive) in WAT and iBAT than wild-type rats. Collectively, our data indicate that MCH/MCH1R-pathway does not modify adipocyte differentiation, whereas Pmch-deficiency in laboratory rats lowers adiposity throughout development and sympathetic adipose drive during adulthood.

  4. The orphan nuclear receptor Rev-Erbalpha is a peroxisome proliferator-activated receptor (PPAR) gamma target gene and promotes PPARgamma-induced adipocyte differentiation

    DEFF Research Database (Denmark)

    Fontaine, Coralie; Dubois, Guillaume; Duguay, Yannick;

    2003-01-01

    Rev-Erbalpha (NR1D1) is an orphan nuclear receptor encoded on the opposite strand of the thyroid receptor alpha gene. Rev-Erbalpha mRNA is induced during adipocyte differentiation of 3T3-L1 cells, and its expression is abundant in rat adipose tissue. Peroxisome proliferator-activated receptor gamma...... (PPARgamma) (NR1C3) is a nuclear receptor controlling adipocyte differentiation and insulin sensitivity. Here we show that Rev-Erbalpha expression is induced by PPARgamma activation with rosiglitazone in rat epididymal and perirenal adipose tissues in vivo as well as in 3T3-L1 adipocytes in vitro...... for this nuclear receptor as a promoter of adipocyte differentiation....

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

    Science.gov (United States)

    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.

  6. Hyperosmotic stress inhibits insulin receptor substrate-1 function by distinct mechanisms in 3T3-L1 adipocytes

    DEFF Research Database (Denmark)

    Gual, Philippe; Gonzalez, Teresa; Grémeaux, Thierry;

    2003-01-01

    . Furthermore, the mammalian target of rapamycin (mTOR) inhibitor rapamycin prevented the osmotic shock-induced phosphorylation of IRS-1 on Ser307. The inhibition of mTOR completely reversed the inhibitory effect of hyperosmotic stress on insulin-induced IRS-1 tyrosine phosphorylation and PI 3-kinase activation....... In addition, prolonged osmotic stress enhanced the degradation of IRS proteins through a rapamycin-insensitive pathway and a proteasome-independent process. These data support evidence of new mechanisms involved in osmotic stress-induced cellular insulin resistance. Short-term osmotic stress induces...

  7. NAD(P)H:quinone oxidoreductase 1 activity reduces hypertrophy in 3T3-L1 adipocytes

    Science.gov (United States)

    The nuclear factor E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) pathway responds to oxidative stress via control of the expression of several antioxidant genes. Recent efforts demonstrate that Nrf2 modulates development of adiposity and adipogenesis. However little is kno...

  8. Insulin-Mimetic Action of Rhoifolin and Cosmosiin Isolated from Citrus grandis (L. Osbeck Leaves: Enhanced Adiponectin Secretion and Insulin Receptor Phosphorylation in 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Yerra Koteswara Rao

    2011-01-01

    Full Text Available Citrus grandis (L. Osbeck (red wendun leaves have been used in traditional Chinese medicine to treat several illnesses including diabetes. However, there is no scientific evidence supporting these actions and its active compounds. Two flavone glycosides, rhoifolin and cosmosiin were isolated for the first time from red wendun leaves and, identified these leaves are rich source for rhoifolin (1.1%, w/w. In differentiated 3T3-L1 adipocytes, rhoifolin and cosmosiin showed dose-dependent response in concentration range of o.oo1–5 μM and 1–20 μM, respectively, in biological studies beneficial to diabetes. Particularly, rhoifolin and cosmosiin at 0.5 and 20 μM, respectively showed nearly similar response to that 10 nM of insulin, on adiponectin secretion level. Furthermore, 5 μM of rhoifolin and 20 μM of cosmosiin showed equal potential with 10 nM of insulin to increase the phosphorylation of insulin receptor-β, in addition to their positive effect on GLUT4 translocation. These findings indicate that rhoifolin and cosmosiin from red wendun leaves may be beneficial for diabetic complications through their enhanced adiponectin secretion, tyrosine phosphorylation of insulin receptor-β and GLUT4 translocation.

  9. Metallomics approach to changes in element concentration during differentiation from fibroblasts into adipocytes by element array analysis.

    Science.gov (United States)

    Ogra, Yasumitsu; Nagasaki, Shu; Yawata, Ayako; Anan, Yasumi; Hamada, Koichi; Mizutani, Akihiro

    2016-04-01

    We aimed to establish an element array analysis that involves the simultaneous detection of all elements in cells and the display of changes in element concentration before and after a cellular event. In this study, we demonstrated changes in element concentration during the differentiation of 3T3-L1 mouse fibroblasts into adipocytes. This metallomics approach yielded unique information of cellular response to physiological and toxicological events.

  10. Enhancement of insulin sensitivity in adipocytes by ginger.

    Science.gov (United States)

    Sekiya, Keizo; Ohtani, Atsuko; Kusano, Shuichi

    2004-01-01

    Antidiabetic and hypoglycemic drugs have been reported to enhance adipocyte differentiation of 3T3-L1 preadipocytes. We previously reported that ginseng (active constituents: ginsenosides) enhanced the differentiation [1]. In this experiment, effect of some ginger group food extracts on the adipocyte differentiation was investigated using cultured mouse 3T3-L1 preadipocytes. 3T3-L1 cells were grown as monolayer cultures at 37 degrees C in DMEM supplemented by 10% FBS under the atmosphere of 5% CO(2)-95% air. Ginger extracts were found to enhance the adipocyte differentiation. Active constituent was purified and identified as gingerol. In the gingerol-treated cells, insulin-sensitive glucose uptake was increased. It is expected that ginger enhance the insulin-sensitivity, and improve chronic disease, such as diabetes.

  11. Kaempferol suppresses lipid accumulation by inhibiting early adipogenesis in 3T3-L1 cells and zebrafish.

    Science.gov (United States)

    Lee, Yeon-Joo; Choi, Hyeon-Son; Seo, Min-Jung; Jeon, Hui-Jeon; Kim, Kui-Jin; Lee, Boo-Yong

    2015-08-01

    Kaempferol is a flavonoid present in Kaempferia galanga and Opuntia ficus indica var. saboten. Recent studies have suggested that it has anti-oxidant, anti-inflammatory, anti-cancer, and anti-obesity effects. In this study, we focused on the anti-adipogenic effects of kaempferol during adipocyte differentiation. The results showed that kaempferol inhibits lipid accumulation in adipocytes and zebrafish. Oil Red O and Nile Red staining showed that the number of intracellular lipid droplets decreased in adipocytes and zebrafish treated with kaempferol. LPAATθ (lysophosphatidic acid acyltransferase), lipin1, and DGAT1 (triglyceride synthetic enzymes) and FASN and SREBP-1C (fatty acid synthetic proteins) showed decreased expression levels in the presence of kaempferol. In addition, treatment of kaempferol showed an inhibitory activity on cell cycle progression. Kaempferol delayed cell cycle progression from the S to G2/M phase through the regulation of cyclins in a dose-dependent manner. Kaempferol blocked the phosphorylation of AKT (protein kinase B) and mammalian target of rapamycin (mTOR) signaling pathway during the early stages of adipogenesis. In addition, kaempferol down-regulated pro-early adipogenic factors such as CCAAT-enhancer binding proteins β (C/EBPβ), and Krüppel-like factors (KLFs) 4 and 5, while anti-early adipogenic factors, such as KLF2 and pref-1(preadipocyte factor-1), were upregulated. These kaempferol-mediated regulations of early adipogenic factors resulted in the attenuation of late adipogenic factors such as C/EBPα and peroxisome proliferator-activated receptor γ (PPARγ). These results were supported in zebrafish based on the decrease in lipid accumulation and expression of adipogenic factors. Our results indicated that kaempferol might have an anti-obesity effect by regulating lipid metabolism.

  12. Anti-Adipogenic Effects of Ethanol Extracts Prepared from Selected Medicinal Herbs in 3T3-L1 Cells

    Science.gov (United States)

    Park, Min-Jun; Song, Ji-Hye; Shon, Myung-Soo; Kim, Hae Ok; Kwon, O Jun; Roh, Seong-Soo; Kim, Choon Young; Kim, Gyo-Nam

    2016-01-01

    Obesity is a major risk factor for various metabolic diseases such as cardiovascular disease, hypertension, and type 2 diabetes mellitus. In this study, we prepared ethanol extracts from Agastache rugosa (ARE), Chrysanthemum zawadskii (CZE), Mentha arvensis (MAE), Perilla frutescens (PFE), Leonurus sibiricus (LSE), Gardenia jasminoides (GJE), and Lycopus coreanus (LCE). The anti-oxidant and anti-adipogenic effects were evaluated. The IC50 values for ascorbic acid and LCE against 2,2-diphenyl-1-picrylhydrazyl radicals were 246.2 μg/mL and 166.2 μg/mL, respectively, followed by ARE (186.6 μg/mL), CZE (198.6 μg/mL), MAE (337.1 μg/mL), PFE (415.3 μg/mL), LSE (548.2 μg/mL), and GJE (626.3 μg/mL). In non-toxic concentration ranges, CZE had a strong inhibitory effect against 3T3-L1 adipogenes (84.5%) than those of the other extracts. Furthermore, the anti-adipogenic effect of CZE is largely limited in the early stage of adipogenesis, and we revealed that the inhibitory role of CZE in adipogenesis is required for the activation of Wnt signaling. Our results provide scientific evidence that the anti-adipogenic effect of CZE can be applied as an ingredient for the development of functional foods and nutri-cosmetics for obesity prevention. PMID:27752499

  13. Dietary polyphenols preconditioning protects 3T3-L1 preadipocytes from mitochondrial alterations induced by oxidative stress.

    Science.gov (United States)

    Baret, Pascal; Septembre-Malaterre, Axelle; Rigoulet, Michel; Lefebvre d'Hellencourt, Christian; Priault, Muriel; Gonthier, Marie-Paule; Devin, Anne

    2013-01-01

    Numerous studies indicate that an increase in reactive oxygen species (ROS) significantly affects white adipose tissue biology and leads to an inflammatory profile and insulin resistance, which could contribute to obesity-associated diabetes and cardiovascular diseases. Mitochondria play a key role in adipose tissue energy metabolism and constitute the main source of cellular ROS such as H(2)O(2). Polyphenols constitute the most abundant antioxidants provided by the human diet. Indeed, they are widely distributed in fruits, vegetables and some plant-derived beverages such as coffee and tea. Thus, the biological effects of dietary polyphenols that may increase the antioxidant capacity of the body against obesity-induced oxidative stress are of high interest. Here, we studied the capacity of polyphenols to modulate the impact of oxidative stress on the mitochondria of preadipocytes, which are important cells governing the adipose tissue development for energy homeostasis. Whereas H(2)O(2) treatment induces a proliferation arrest associated with an increase in mitochondrial content in 3T3-L1 preadipocytes, preconditioning with some major dietary polyphenols totally or partially protects the cells against oxidative stress consequences. This article is part of a Directed Issue entitled: Bioenergetic dysfunction, adaptation and therapy.

  14. Dynamic tracking and mobility analysis of single GLUT4 storage vesicle in live 3T3-L1 cells

    Institute of Scientific and Technical Information of China (English)

    Chen Hong LI; Li BAI; Dong Dong LI; Sheng XIA; Tao XU

    2004-01-01

    Glucose transporter 4 (GLUT4) is responsible for insulin-stimulated glucose transporting into the insulin-sensitive fat and muscle cells. The dynamics of GLUT4 storage vesicles (GSVs) remains to be explored and it is unclear how GSVs are arranged based on their mobility. We examined this issue in 3T3-L1 cells via investigating the three-dimensional mobility of single GSV labeled with EGFP-fused GLUT4. A thin layer of cytosol right adjacent to the plasma membrane was illuminated and successively imaged at 5 Hz under a total internal reflection fluorescence microscope with a penetration depth of 136 nm. Employing single particle tracking, the three-dimensional subpixel displacement of single GSV was tracked at a spatial precision of 22 nm. Both the mean square displacement and the diffusion coefficient were calculated for each vesicle. Tracking results revealed that vesicles moved as if restricted within a cage that has a mean radius of 160 nm, suggesting the presence of some intracellular tethering matrix. By constructing the histogram of the diffusion coefficients of GSVs, we observed a smooth distribution instead of the existence of distinct groups. The result indicates that GSVs are dynamically retained in a continuous and wide range of mobility rather than into separate classes.

  15. Novel function of the retinoblastoma protein in fat: regulation of white versus brown adipocyte differentiation

    DEFF Research Database (Denmark)

    Hansen, Jacob B; te Riele, Hein; Kristiansen, Karsten

    2004-01-01

    The differentiation of white and brown fat cells is controlled by a similar set of transcription factors, including PPARgamma and C/EBPalpha. However, despite many similarities between the two types of fat cells, they carry out essentially opposite functions in vivo, with white adipocytes being...... the major energy store and brown adipocytes being potent energy-dissipaters through thermogenesis. Yet, little is known about factors differentially regulating the formation of white and brown fat cells. Members of the retinoblastoma protein family (pRB, p107, p130) have been implicated in the regulation...... of adipocyte differentiation, and expression and phosphorylation of the three retinoblastoma family proteins oscillate in a characteristic manner during differentiation of the white preadipocyte cell line 3T3-L1. We have recently demonstrated a surprising function of the retinoblastoma protein...

  16. Pmch-deficiency in rats is associated with normal adipocyte differentiation and lower sympathetic adipose drive.

    Directory of Open Access Journals (Sweden)

    Joram D Mul

    Full Text Available The orexigenic neuropeptide melanin-concentrating hormone (MCH, a product of Pmch, is an important mediator of energy homeostasis. Pmch-deficient rodents are lean and smaller, characterized by lower food intake, body-, and fat mass. Pmch is expressed in hypothalamic neurons that ultimately are components in the sympathetic nervous system (SNS drive to white and interscapular brown adipose tissue (WAT, iBAT, respectively. MCH binds to MCH receptor 1 (MCH1R, which is present on adipocytes. Currently it is unknown if Pmch-ablation changes adipocyte differentiation or sympathetic adipose drive. Using Pmch-deficient and wild-type rats on a standard low-fat diet, we analyzed dorsal subcutaneous and perirenal WAT mass and adipocyte morphology (size and number throughout development, and indices of sympathetic activation in WAT and iBAT during adulthood. Moreover, using an in vitro approach we investigated the ability of MCH to modulate 3T3-L1 adipocyte differentiation. Pmch-deficiency decreased dorsal subcutaneous and perirenal WAT mass by reducing adipocyte size, but not number. In line with this, in vitro 3T3-L1 adipocyte differentiation was unaffected by MCH. Finally, adult Pmch-deficient rats had lower norepinephrine turnover (an index of sympathetic adipose drive in WAT and iBAT than wild-type rats. Collectively, our data indicate that MCH/MCH1R-pathway does not modify adipocyte differentiation, whereas Pmch-deficiency in laboratory rats lowers adiposity throughout development and sympathetic adipose drive during adulthood.

  17. Hypochlorous acid via peroxynitrite activates protein kinase Cθ and insulin resistance in adipocytes

    OpenAIRE

    Zhou, Jun; Wang, Qilong; Ding, Ye; Zou, Ming-hui

    2015-01-01

    We recently reported that genetic deletion of myeloperoxidase (MPO) alleviates obesity-related insulin resistance in mice in vivo. How MPO impairs insulin sensitivity in adipocytes is poorly characterized. As hypochlorous acid (HOCl) is a principal oxidant product generated by MPO, we evaluated the effects of HOCl on insulin signaling in adipocytes differentiated from 3T3-L1 cells. Exposure of 3T3-L1 adipocytes to exogenous HOCl (200 μmol/l) attenuated insulin-stimulated 2-deoxyglucose uptake...

  18. The role of Smad Ubiquitin Regulatory Factor-1: SMURF1 in the differentiation 3T3-L1

    OpenAIRE

    Muñiz Diego, María del Carmen

    2013-01-01

    The Smad Ubiquitin Regulatory Factor-1: SMURF1 is an E3 ligase. This E3 ligase has been linked to several important biological pathways, including the bone morphogenetic protein pathway, the non-canonical Wnt pathway and the mitogen-activated protein kinase pathway. Multiple functions of Smurf1 have been discovered in cell growth and morphogenesis, cell migration, cell polarity and autophagy. Previous studies, in 2003, have demonstrated that overexpression of Smurf1 induces proteasomal degrad...

  19. γ-Oryzanol Enhances Adipocyte Differentiation and Glucose Uptake

    Directory of Open Access Journals (Sweden)

    Chang Hwa Jung

    2015-06-01

    Full Text Available Recent studies show that brown rice improves glucose intolerance and potentially the risk of diabetes, although the underlying molecular mechanisms remain unclear. One of the phytochemicals found in high concentration in brown rice is γ-oryzanol (Orz, a group of ferulic acid esters of phytosterols and triterpene alcohols. Here, we found that Orz stimulated differentiation of 3T3-L1 preadipocytes and increased the protein expression of adipogenic marker genes such as peroxisome proliferator-activated receptor gamma (PPAR-γ and CCAAT/enhanced binding protein alpha (C/EBPα. Moreover, Orz significantly increased the glucose uptake in insulin-resistant cells and translocation of glucose transporter type 4 (GLUT4 from the cytosol to the cell surface. To investigate the mechanism by which Orz stimulated cell differentiation, we examined its effects on cellular signaling of the mammalian target of rapamycin complex 1 (mTORC1, a central mediator of cellular growth and proliferation. The Orz treatment increased mTORC1 kinase activity based on phosphorylation of 70-kDa ribosomal S6 kinase 1 (S6K1. The effect of Orz on adipocyte differentiation was dependent on mTORC1 activity because rapamycin blocks cell differentiation in Orz-treated cells. Collectively, our results indicate that Orz stimulates adipocyte differentiation, enhances glucose uptake, and may be associated with cellular signaling mediated by PPAR-γ and mTORC1.

  20. Annexin A3 as a negative regulator of adipocyte differentiation.

    Science.gov (United States)

    Watanabe, Takenori; Ito, Yoshimasa; Sato, Asuka; Hosono, Takashi; Niimi, Shingo; Ariga, Toyohiko; Seki, Taiichiro

    2012-10-01

    Annexin A3 is a protein belonging to the annexin family, and it is mainly present in cellular membranes as a phospholipid-binding protein that binds via the calcium ion. However, its physiological function remains to be clarified. We examined the expression of annexin A3 in mouse tissues and found for the first time that annexin A3 mRNA and its protein were expressed more strongly in adipose tissues than in other tissues. In adipose tissues, annexin A3-expressing cells were present in the stromal vascular fraction, and precisely identical to Pref-1-positive preadipocytes, Pref-1 being an epidermal growth factor repeat-containing transmembrane protein that inhibits adipogenesis. In 3T3-L1 cells, used as a model of adipogenesis, annexin A3 was down-regulated at an early phase of adipocyte differentiation, and this pattern paralleled that of Pref-1. Suppression of annexin A3 in these cells with siRNA caused elevation of the PPARγ2 mRNA level and lipid droplet accumulation. In conclusion, our data suggest that annexin A3 is a negative regulator of adipocyte differentiation.

  1. Differentiation of Pre-Adipocytes in Modelled Microgravity

    Science.gov (United States)

    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. Role of extrathyroidal TSHR expression in adipocyte differentiation and its association with obesity

    Directory of Open Access Journals (Sweden)

    Lu Sumei

    2012-01-01

    Full Text Available Abstract Background Obesity is known to be associated with higher risks of cardiovascular disease, metabolic syndrome, and diabetes mellitus. Thyroid-stimulating hormone (TSHR is the receptor for thyroid-stimulating hormone (TSH, or thyrotropin, the key regulator of thyroid functions. The expression of TSHR, once considered to be limited to thyrocytes, has been so far detected in many extrathyroidal tissues including liver and fat. Previous studies have shown that TSHR expression is upregulated when preadipocytes differentiate into mature adipocytes, suggestive of a possible role of TSHR in adipogenesis. However, it remains unclear whether TSHR expression in adipocytes is implicated in the pathogenesis of obesity. Methods In the present study, TSHR expression in adipose tissues from both mice and human was analyzed, and its association with obesity was evaluated. Results We here showed that TSHR expression was increased at both mRNA and protein levels when 3T3-L1 preadipocytes were induced to differentiate. Knockdown of TSHR blocked the adipocyte differentiation of 3T3-L1 preadipocytes as evaluated by Oil-red-O staining for lipid accumulation and by RT-PCR analyses of PPAR-γ and ALBP mRNA expression. We generated obesity mice (C57/BL6 by high-fat diet feeding and found that the TSHR protein expression in visceral adipose tissues from obesity mice was significantly higher in comparison with the non-obesity control mice (P Conclusion Taken together, these results suggested that TSHR is an important regulator of adipocyte differentiation. Dysregulated expression of TSHR in adipose tissues is associated with obesity, which may involve a mechanism of excess adipogenesis.

  3. Regulation of Autophagy-Related Protein and Cell Differentiation by High Mobility Group Box 1 Protein in Adipocytes

    Directory of Open Access Journals (Sweden)

    Huanhuan Feng

    2016-01-01

    Full Text Available High mobility group box 1 protein (HMGB1 is a molecule related to the development of inflammation. Autophagy is vital to maintain cellular homeostasis and protect against inflammation of adipocyte injury. Our recent work focused on the relationship of HMGB1 and autophagy in 3T3-L1 cells. In vivo experimental results showed that, compared with the normal-diet group, the high-fat diet mice displayed an increase in adipocyte size in the epididymal adipose tissues. The expression levels of HMGB1 and LC3II also increased in epididymal adipose tissues in high-fat diet group compared to the normal-diet mice. The in vitro results indicated that HMGB1 protein treatment increased LC3II formation in 3T3-L1 preadipocytes in contrast to that in the control group. Furthermore, LC3II formation was inhibited through HMGB1 knockdown by siRNA. Treatment with the HMGB1 protein enhanced LC3II expression after 2 and 4 days but decreased the expression after 8 and 10 days among various differentiation stages of adipocytes. By contrast, FABP4 expression decreased on the fourth day and increased on the eighth day. Hence, the HMGB1 protein modulated autophagy-related proteins and lipid-metabolism-related genes in adipocytes and could be a new target for treatment of obesity and related metabolic diseases.

  4. Expression of Caveolin 1 is enhanced by DNA demethylation during adipocyte differentiation. status of insulin signaling.

    Science.gov (United States)

    Palacios-Ortega, Sara; Varela-Guruceaga, Maider; Milagro, Fermín Ignacio; Martínez, José Alfredo; de Miguel, Carlos

    2014-01-01

    Caveolin 1 (Cav-1) is an essential constituent of adipocyte caveolae which binds the beta subunit of the insulin receptor (IR) and is implicated in the regulation of insulin signaling. We have found that, during adipocyte differentiation of 3T3-L1 cells the promoter, exon 1 and first intron of the Cav-1 gene undergo a demethylation process that is accompanied by a strong induction of Cav-1 expression, indicating that epigenetic mechanisms must have a pivotal role in this differentiation process. Furthermore, IR, PKB-Akt and Glut-4 expression are also increased during the differentiation process suggesting a coordinated regulation with Cav-1. Activation of Cav-1 protein by phosphorylation arises during the differentiation process, yet in fully mature adipocytes insulin is no longer able to significantly increase Cav-1 phosphorylation. However, these long-term differentiated cells are still able to respond adequately to insulin, increasing IR and PKB-Akt phosphorylation and glucose uptake. The activation of Cav-1 during the adipocyte differentiation process could facilitate the maintenance of insulin sensitivity by these fully mature adipocytes isolated from additional external stimuli. However, under the influence of physiological conditions associated to obesity, such as chronic inflammation and hypoxia, insulin sensitivity would finally be compromised.

  5. Activation of liver X receptors prevents statin-induced death of 3T3-L1 preadipocytes

    DEFF Research Database (Denmark)

    Madsen, Lise; Petersen, Rasmus K; Steffensen, Knut R;

    2008-01-01

    to differentiate by standard isobutylmethylxanthine/dexamethasone/insulin treatment in the presence of statins, they failed to differentiate and underwent massive apoptosis. The simultaneous addition of selective LXR agonists prevented the statin-induced apoptosis. By using mouse embryo fibroblasts from wild...... of LXRalpha, we demonstrate that the response to LXR agonists is LXR-dependent. Interestingly, LXR-mediated rescue of statin-induced apoptosis was not related to up-regulation of genes previously shown to be involved in the antiapoptotic action of LXR. Furthermore, forced expression of Bcl-2 did not prevent...... of IkappaBalpha prevented LXR agonist-dependent rescue of statin-induced apoptosis. Thus, the results presented in this paper provide novel insight into the action of statins on and LXR-dependent inhibition of apoptosis....

  6. 黄芩水提液对3T3-L1脂肪细胞增殖、诱导分化及脂联素启动子荧光素酶活性的影响%The Effect of Scutellaria Baicalensis Water Extract on Proliferation, Cytokines mRNA Expressions and Promoter Activity of 3T3-L1 Cells

    Institute of Scientific and Technical Information of China (English)

    崔琳; 路玲玲; 李强; 宰军华; 刘卫红; 王小晓

    2015-01-01

    目的:本研究旨在观察黄芩水提液(Scutellaria BaicalensisWater Extract,SBWE)对3T3-L1前体细胞增殖、分化,对脂肪细胞因子脂联素表达以及脂联素(Adiponectin,ADP)启动子荧光素酶活性的影响,从分子生物学角度阐述SBWE降脂作用的可能机理.方法:通过体外培养3T3-L1细胞,采用MTT法检测SBWE对3T3-L1细胞增殖能力的影响;通过诱导脂肪细胞分化成为成熟脂肪细胞,观察SBWE对脂肪形成的影响;化学发光法检测脂联素启动子双荧光素酶报告基因活性;荧光定量PCR法检测脂联素mRNA(Adipoq)表达.结果:与正常组相比,给予3T3-L1细胞0.01、0.1、1 mg?mL-1浓度的SBWE 24 h,可显著抑制细胞的增殖活性(P<0.05);0.1、1 mg?mL-1浓度的SBWE能够降低3T3-L1细胞分化为脂肪细胞的数量,并减少细胞内脂滴聚集,但无明显剂量依赖性;0.01、0.1 mg?mL-1浓度SBWE能显著提高脂联素基因启动子荧光素酶活性,与空载体比较差异有统计学意义(P<0.05);与正常组相比,给予3T3-L1细胞0.1 mg?mL-1SBWE 24 h,诱导前后的脂肪细胞Adipoq表达均明显增加(P<0.05).结论:SBWE可有效抑制3T3-L1脂肪细胞的增殖、分化,同时增加脂联素基因表达,这可能是通过增强脂联素基因启动子荧光素酶活性实现,这些为黄芩水提液减肥的作用机制提供一定的基础.%The study was designed to measure the effect of S.baicalensiswater extract (SBWE) on 3T3-L1 cells and its adiponectin (ADP) mRNA (Adipoq) and promoter luciferase activity.Cell survival rate was determined by MTT assay.The expression of Adipoq was measured by real-time PCR,while the luciferase report systems of Adipoq were used to transfer 3T3-L1 cells.The luciferase activities of the transferred cells were compared by luciferase assay.It was found that the mRNA expression of Adipoq was decreased in comparison with the control group.The luciferase activity showed a stronger ADP promoter activity in 3T3-L1 cells in

  7. ATF3 inhibits PPARγ-stimulated transactivation in adipocyte cells

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Min-Kyung; Jung, Myeong Ho, E-mail: jung0603@pusan.ac.kr

    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.

  8. Piperine, a component of black pepper, decreases eugenol-induced cAMP and calcium levels in non-chemosensory 3T3-L1 cells.

    Science.gov (United States)

    Yoon, Yeo Cho; Kim, Sung-Hee; Kim, Min Jung; Yang, Hye Jeong; Rhyu, Mee-Ra; Park, Jae-Ho

    2015-01-01

    This study investigated the effects of an ethanol extract of black pepper and its constituent, piperine, on odorant-induced signal transduction in non-chemosensory cells. An ethanol extract of black pepper decreased eugenol-induced cAMP and calcium levels in preadipocyte 3T3-L1 cells with no toxicity. Phosphorylation of CREB (cAMP response element-binding protein) was down-regulated by the black pepper extract. The concentration (133.8 mg/g) and retention time (5.5 min) of piperine in the ethanol extract were quantified using UPLC-MS/MS. Pretreatment with piperine decreased eugenol-induced cAMP and calcium levels in 3T3-L1 cells. Piperine also decreased the phosphorylation of CREB, which is up-regulated by eugenol. These results suggest that piperine inhibits the eugenol-induced signal transduction pathway through modulation of cAMP and calcium levels and phosphorylation of CREB in non-chemosensory cells.

  9. Effect of TNF-Alpha on Caveolin-1 Expression and Insulin Signaling During Adipocyte Differentiation and in Mature Adipocytes

    Directory of Open Access Journals (Sweden)

    Sara Palacios-Ortega

    2015-07-01

    Full Text Available Background/Aims: Tumor necrosis factor-α (TNF-α-mediated chronic low-grade inflammation of adipose tissue is associated with obesity and insulin resistance. Caveolin-1 (Cav-1 is the central component of adipocyte caveolae and has an essential role in the regulation of insulin signaling. The effects of TNF-α on Cav-1 expression and insulin signaling during adipocyte differentiation and in mature adipocytes were studied. Methods: 3T3-L1 cells were differentiated (21 days in the presence TNF-α (10 ng/mL and mature adipocytes were also treated with TNF-α for 48 hours. Cav-1 and insulin receptor (IR gene methylation were determined as well as Cav-1, IR, PKB/AKT-2 and Glut-4 expression and activation by real time RT-PCR and western blot. Baseline and insulin-induced glucose uptake was measured by the 2-[C14]-deoxyglucose uptake assay. Results: TNF-α slowed down the differentiation program, hindering the expression of some insulin signaling intermediates without fully eliminating insulin-mediated glucose uptake. In mature adipocytes, TNF-α did not compromise lipid-storage capacity, but downregulated the expression of the insulin signaling intermediates, totally blocking insulin-mediated glucose uptake. Insulin sensitivity correlated with the level of activated phospho-Cav-1 in both situations, strongly suggesting the direct contribution of Cav-1 to the maintenance of this physiological response. Conclusion: Cav-1 activation by phosphorylation seems to be essential for the maintenance of an active and insulin-sensitive glucose uptake.

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

    Energy Technology Data Exchange (ETDEWEB)

    Hao, Chan-Juan; Cheng, Xue-Jia; Xia, Hong-Fei, E-mail: hongfeixia@yahoo.com.cn; 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.

  11. Inhibition of protein kinase CbetaII increases glucose uptake in 3T3-L1 adipocytes through elevated expression of glucose transporter 1 at the plasma membrane.

    NARCIS (Netherlands)

    Bosch, R.R.; Bazuine, M.; Wake, M.M.; Span, P.N.; Olthaar, A.J.; Schurmann, A.; Maassen, J.A.; Hermus, A.R.M.M.; Willems, P.H.G.M.; Sweep, C.G.J.

    2003-01-01

    The mechanism via which diacylglycerol-sensitive protein kinase Cs (PKCs) stimulate glucose transport in insulin-sensitive tissues is poorly defined. Phorbol esters, such as phorbol-12-myristate-13-acetate (PMA), are potent activators of conventional and novel PKCs. Addition of PMA increases the rat

  12. Glutamine, insulin and glucocorticoids regulate glutamine synthetase expression in C2C12 myotubes, Hep G2 hepatoma cells and 3T3 L1 adipocytes

    OpenAIRE

    Wang, Yanxin; Watford, Malcolm

    2006-01-01

    The cell-specific regulation of glutamine synthetase expression was studied in three cell lines. In C2C12 myotubes, glucocorticoids increased the abundance of both glutamine synthetase protein and mRNA. Culture in the absence of glutamine also resulted in very high glutamine synthetase protein abundance but mRNA levels were unchanged. Glucocorticoids also increased the abundance of glutamine synthetase mRNA in Hep G2 hepatoma cells but this was not reflected in changes in protein abundance. C...

  13. Insulin elevates leptin secretion and mRNA levels via cyclic AMP in 3T3-L1 adipocytes deprived of glucose

    Directory of Open Access Journals (Sweden)

    Tomomi Tsubai

    2016-11-01

    Conclusion: Insulin alone stimulates leptin secretion and elevates leptin mRNA levels via cAMP under the lack of glucose metabolism, while glucose is a significant and ambivalent effector on the insulin effects of leptin.

  14. Adrenergic receptor stimulation attenuates insulin-stimulated glucose uptake in 3T3-L1 adipocytes by inhibiting GLUT4 translocation

    NARCIS (Netherlands)

    Mulder, A.; Tack, C.J.J.; Olthaar, A.J.; Smits, P.; Sweep, C.G.J.; Bosch, R.R.

    2005-01-01

    Activation of the sympathetic nervous system inhibits insulin-stimulated glucose uptake. However, the underlying mechanisms are incompletely understood. Therefore, we studied the effects of catecholamines on insulin-stimulated glucose uptake and insulin-stimulated translocation of GLUT4 to the plasm

  15. Adipocyte differentiation and leptin expression

    DEFF Research Database (Denmark)

    Hwang, C S; Loftus, T M; Mandrup, S

    1997-01-01

    , most notably those of the C/EBP and PPAR families, which combine to regulate each other and to control the expression of adipocyte-specific genes. One such gene, i.e. the obese gene, was recently identified and found to encode a hormone, referred to as leptin, that plays a major role in the regulation...... of energy intake and expenditure. The hormonal and transcriptional control of adipocyte differentiation is discussed, as is the role of leptin and other factors secreted by the adipocyte that participate in the regulation of adipose homeostasis....

  16. Effects of lysophosphatidic acid on the in vitro proliferation and differentiation of a novel porcine preadipocyte cell line.

    Science.gov (United States)

    Nobusue, Hiroyuki; Kondo, Daisuke; Yamamoto, Makiko; Kano, Koichiro

    2010-12-01

    We examined the effects of lysophosphatidic acid (LPA) on in vitro proliferation and differentiation of a porcine preadipocyte cell line, DFAT-P, and a mouse preadipocyte cell line, 3T3-L1. During the proliferation and differentiation phases, DFAT-P and 3T3-L1 cells expressed only the endothelial differentiation gene (EDG)-2 receptor and not EDG-4 and EDG-7 receptors. LPA promoted the proliferation of DFAT-P cells more extensively than that of 3T3-L1 cells. After adipogenic induction, LPA inhibited glycerol-3-phosphate dehydrogenase activity and lipid droplet accumulation, and suppressed peroxisome proliferator-activated receptor γ (PPARγ) protein expression, this inhibitory effect in DFAT-P cells was twice as high as that in 3T3-L1 cells. Furthermore, treatments with low LPA concentrations significantly inhibited adipocyte differentiation in DFAT-P cells but not in 3T3-L1 cells. We conclude that LPA promotes the proliferation of porcine preadipocytes through the EDG-2 receptor but inhibits their differentiation, and these effects depend on the down-regulation of PPARγ expression via the EDG-2 receptor. Furthermore, DFAT-P cells are more sensitive to LPA than 3T3-L1 cells. These findings in a porcine model will contribute to the understanding of LPA action mechanisms on in vitro proliferation and differentiation of preadipocytes in domestic animals and/or humans.

  17. Nonivamide enhances miRNA let-7d expression and decreases adipogenesis PPARγ expression in 3T3-L1 cells.

    Science.gov (United States)

    Rohm, Barbara; Holik, Ann-Katrin; Kretschy, Nicole; Somoza, Mark M; Ley, Jakob P; Widder, Sabine; Krammer, Gerhard E; Marko, Doris; Somoza, Veronika

    2015-06-01

    Red pepper and its major pungent principle, capsaicin (CAP), have been shown to be effective anti-obesity agents by reducing energy intake, enhancing energy metabolism, decreasing serum triacylglycerol content, and inhibiting adipogenesis via activation of the transient receptor potential cation channel subfamily V member 1 (TRPV1). However, the binding of CAP to the TRPV1 receptor is also responsible for its pungent sensation, strongly limiting its dietary intake. Here, the effects of a less pungent structural CAP-analog, nonivamide, on adipogenesis and underlying mechanisms in 3T3-L1 cells were studied. Nonivamide was found to reduce mean lipid accumulation, a marker of adipogenesis, to a similar extent as CAP, up to 10.4% (P < 0.001). Blockage of the TRPV1 receptor with the specific inhibitor trans-tert-butylcyclohexanol revealed that the anti-adipogenic activity of nonivamide depends, as with CAP, on TRPV1 receptor activation. In addition, in cells treated with nonivamide during adipogenesis, protein levels of the pro-adipogenic transcription factor peroxisome-proliferator activated receptor γ (PPARγ) decreased. Results from miRNA microarrays and digital droplet PCR analysis demonstrated an increase in the expression of the miRNA mmu-let-7d-5p, which has been associated with decreased PPARγ levels.

  18. Effects of Yerba maté, a Plant Extract Formulation (“YGD” and Resveratrol in 3T3-L1 Adipogenesis

    Directory of Open Access Journals (Sweden)

    Juliana C. Santos

    2014-10-01

    Full Text Available We aimed to evaluate the in vitro effects of yerba maté, YGD (a herbal preparation containing yerba maté, guarana and damiana, and resveratrol on adipogenesis. The anti-adipogenic effects of yerba mate, YGD, resveratrol and YGD + resveratrol and yerba mate + resveratrol combinations were evaluated in 3T3-L1 cells by Oil Red staining, cellular triglyceride content, and PCR quantitative array. The results demonstrated that all of the tested compounds inhibited adipogenesis. Yerba maté extract significantly down-regulated the expression of genes that play an important role in regulating adipogenesis, such as Adig, Axin, Cebpa, Fgf10, Lep, Lpl, and Pparγ2. In addition, these genes, YGD also repressed Bmp2, Ccnd1, Fasn, and Srebf1. Resveratrol also modulated the expression of Adig, Bmp2, Ccnd1, C/EBPα, Fasn, Fgf10, Lep, Lpl, and Pparγ2. Moreover, resveratrol repressed Cebpb, Cdk4, Fgf2, and Klf15. The yerba maté extract and YGD up-regulated the expression of genes involved in inhibiting adipogenesis, such as Dlk-1, Klf2, and Ucp1. Resveratrol also induced the expression of Klf2 and Ucp1. In addition resveratrol modulated the Ddit3, Foxo1, Sirt1, and Sirt2. The combined effects of these compounds on gene expression showed similar results observed from individual treatments. Our data indicates that the synergy between the compounds favors the inhibition of adipogenesis.

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

    Directory of Open Access Journals (Sweden)

    Hansen Jacob B

    2011-05-01

    Full Text Available Abstract 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

  20. 三种植物甾醇调控SRE元件和3T3-L1细胞脂肪化活性的作用%Effects of Three Plant Sterols on Activity of Sterol Regulatory Element and Adipogenesis of 3T3L-1

    Institute of Scientific and Technical Information of China (English)

    陈彦光; 张弦; 刘健

    2013-01-01

    [目的]为了研究植物甾醇对脂质代谢的调控作用.[方法]利用所构建的SRE荧光素酶活性测定系统,体外考察了豆甾醇、β-谷甾醇和菜油甾醇3种常见的植物甾醇对SRE元件的调控活性.[结果]植物甾醇能够增强SRE元件活性.植物甾醇也可以增加3T3-L1细胞的脂肪化作用.[结论]植物甾醇可能正调控脂质的合成.%[Objective] The research aimed to study the regulating effect of plant sterol on lipid metabolism.[Method] Luciferase reporter vector pGL3-basic that contained one or three sterol regulatory element (SRE) was transfected into HepG2,and luciferase activity of three plant sterols (stigmasterol,β-sitosterol and campesterol) was analyzed.[Result] The plant sterols had positive regulator of SRE.And plant sterols could increase adipogenesis of 3T3L-1.[Conclusion] Plant sterols could increase adipogenesis by increasing SRE activity.

  1. Transcriptional networks controlling adipocyte differentiation

    DEFF Research Database (Denmark)

    Siersbæk, R; Mandrup, Susanne

    2011-01-01

    Adipocyte differentiation is regulated by a complex cascade of signals that drive the transcriptional reprogramming of the fibroblastic precursors. Genome-wide analyses of chromatin accessibility and binding of adipogenic transcription factors make it possible to generate "snapshots" of the trans...

  2. Suppressive effects of quercetin-3-O-(6″-Feruloyl)-β-D-galactopyranoside on adipogenesis in 3T3-L1 preadipocytes through down-regulation of PPARγ and C/EBPα expression.

    Science.gov (United States)

    Yang, Lei; Li, Xiao-Fan; Gao, Lei; Zhang, Ya-Ou; Cai, Guo-Ping

    2012-03-01

    Obesity is a chronic, costly disease, and flavonoids such as quercetin have been proven to play protective roles against it. This study investigated the suppressive effect of quercetin-3-O-(6″-feruloyl)-β-D-galactopyranoside (QFG) on adipogenesis of 3T3-L1 preadipocytes. Quercetin-3-O-(6″-feruloyl)-β-D-galactopyranoside and quercetin were both extracted from Psidium guajava (Myrtaceae, commonly known as guava) leaves and were evaluated for their suppressive effect on adipogenesis by means of oil red O staining and triglyceride assay. It was shown that QFG inhibited adipogenesis in a dose- and time-dependent manner, and it exerted a stronger effect than did quercetin at the same concentration. Quantitative real-time polymerase chain reaction and western blotting were conducted to further examine the differentiation expression of marker genes and transcriptional factors. Both mRNA and protein expression of the key adipogenic transcriptional factors, peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT (cytidine-cytidine-adenosine-adenosine-thymidine)/enhancer-binding protein alpha (C/EBPα), were inhibited by QFG. Moreover, the mRNA expression patterns of key participants in the Wnt-β-catenin pathway were not altered during the QFG-induced adipogenesis inhibition. These results suggest that QFG effectively suppresses adipogenesis and that it exerts its role mainly through the significant down-regulation of PPARγ and C/EBPα and, probably, via a Wnt-β-catenin independent pathway.

  3. Genome-wide profiling of peroxisome proliferator-activated receptor γ in primary epididymal, inguinal, and brown adipocytes reveals depot-selective binding correlated with gene expression

    DEFF Research Database (Denmark)

    Siersbæk, Majken; Loft, Anne; Jørgensen, Mads Malik Aagaard;

    2012-01-01

    epididymal, inguinal, and brown adipose tissues. While these PPARγ binding profiles are overall similar, there are clear depot-selective binding sites. Most PPARγ binding sites previously mapped in 3T3-L1 adipocytes can also be detected in primary adipocytes, but there are a large number of PPARγ binding...... sites that are specific to the primary cells, and these tend to be located in closed chromatin regions in 3T3-L1 adipocytes. The depot-selective binding of PPARγ is associated with highly depot-specific gene expression. This indicates that PPARγ plays a role in the induction of genes characteristic...... of different adipocyte lineages and that preadipocytes from different depots are differentially preprogrammed to permit PPARγ lineage-specific recruitment even when differentiated in vitro....

  4. 7-Chloroarctinone-b as a new selective PPARγ antagonist potently blocks adipocyte differentiation

    Institute of Scientific and Technical Information of China (English)

    Yong-tao LI; Li LI; Jing CHEN; Tian-cen HU; Jin HUANG; Yue-wei GUO; Hua-liang JIANG; Xu SHEN

    2009-01-01

    Aim: Peroxisome proliferator-activated receptor gamma (PPARy) is a therapeutic target for obesity, cancer and diabetes mellitus. In order to develop potent lead compounds for obesity treatment, we screened a natural product library for novel PPARy antagonists with inhibitory effects on adipocyte differentiation. Methods: Surface plasmon resonance (SPR) technology and cell-based transactivation assay were used to screen for PPARy antago-nists. To investigate the antagonistic mechanism of the active compound, we measured its effect on PPARy/RXRα heterodimerization and PPARy co-activator recruitment using yeast two-hybrid assay, Gal4/UAS cell-based assay and SPR based assay. The 3T3-L1 cell differentiation assay was used to evaluate the effect of the active compound on adipocyte differentiation. Results: A new thiophene-acetylene type of natural product, 7-chloroarctinone-b (CAB), isolated from the roots of Rhaponticum uniflo-rum, was discovered as a novel PPARγ antagonist capable of inhibiting rosiglitazone-induced PPARγ transcriptional activity. SPR analy-sis suggested that CAB bound tightly to PPARγ and considerably antagonized the potent PPARy agonist rosigtitazone-stimulated PPARγ-LBD/RXRα-LBD binding. Gal4/UAS and yeast two-hybrid assays were used to evaluate the antagonistic activity of CAB on rosiglitazone-induced recruitment of the coactivator for PPARy. CAB could efficiently antagonize both hormone and rosiglitazone-induced adipocyte differentiation in cell culture. Conclusion: CAB shows antagonistic activity to PPARγ and can block the adipocyte differentiation, indicating it may be of potential use as a lead therapeutic compound for obesity.

  5. 3T3-L1脂肪细胞膜FGF-21结合蛋白的初步鉴定%Identification of Binding Partners of Fibroblast Growth Factor-21 in Cell Membrane of 3T3-L1 Cells

    Institute of Scientific and Technical Information of China (English)

    王文飞; 任桂萍; 侯玉婷; 李德山

    2008-01-01

    成纤维细胞生长因子(fibroblast growth factor,FGF)-21是最近发现的1个可以独立调节血糖的细胞因子,有望成为治疗2型糖尿病的备选药物,但是,FGF-21调解血糖的机理尚不十分清楚,为探讨该因子功能受体,应用偶联方法,以313-L1脂肪细胞为靶标,以FGF-21为诱饵,在3T3-L1脂肪细胞膜上寻找结合蛋白,结果表明,生物素标记的FGF-21可与脂肪细胞膜蛋白形成分子质量大小约300 kD以上两组复合物,竞争试验显示,非标记的FGF-21可与生物素标记的FGF-21竞争、抑制标记的FGF-21参入复合物;应用非标记FGF-21剂量越大,抑制后者参入复合物的程度越强,结果提示,该复合物是FGF-21特异性的,此外,随着生物素标记的FGF-21剂量增加,观察到的标记复合物越多;但是,当FGF-21剂量达12.5 mg/L以上时,观察到的复合物数量不再增加,实验结果提示,复合物形成与FGF-21剂量相关;FGF-21特异结合的蛋白质结合位点饱和后,复合物形成量最大,同时,采用FGF受体特异性抑制剂SU5402可特异性抑制FGF-21在3T3-L1脂肪细胞中的促进葡萄糖吸收作用,提示本实验所观察到的FGF-21-膜蛋白复合物可能就是FGF-21-FGF受体.

  6. JAZF1 can regulate the expression of lipid metabolic genes and inhibit lipid accumulation in adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Ming, Guang-feng [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Changsha 410078, Hunan (China); Department of Critical Care Medicine, Xiangya Hospital, Central South University, Changsha 410008, Hunan (China); Xiao, Di; Gong, Wei-jing [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Changsha 410078, Hunan (China); Liu, Hui-xia; Liu, Jun [Department of Geriatrics, Xiangya Hospital, Central South University, Changsha 410008, Hunan (China); Zhou, Hong-hao [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Changsha 410078, Hunan (China); Liu, Zhao-qian, E-mail: liuzhaoqian63@126.com [Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, Central South University, Changsha 410078, Hunan (China)

    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.

  7. Global mapping of cell type-specific open chromatin by FAIRE-seq reveals the regulatory role of the NFI family in adipocyte differentiation.

    Directory of Open Access Journals (Sweden)

    Hironori Waki

    2011-10-01

    Full Text Available Identification of regulatory elements within the genome is crucial for understanding the mechanisms that govern cell type-specific gene expression. We generated genome-wide maps of open chromatin sites in 3T3-L1 adipocytes (on day 0 and day 8 of differentiation and NIH-3T3 fibroblasts using formaldehyde-assisted isolation of regulatory elements coupled with high-throughput sequencing (FAIRE-seq. FAIRE peaks at the promoter were associated with active transcription and histone modifications of H3K4me3 and H3K27ac. Non-promoter FAIRE peaks were characterized by H3K4me1+/me3-, the signature of enhancers, and were largely located in distal regions. The non-promoter FAIRE peaks showed dynamic change during differentiation, while the promoter FAIRE peaks were relatively constant. Functionally, the adipocyte- and preadipocyte-specific non-promoter FAIRE peaks were, respectively, associated with genes up-regulated and down-regulated by differentiation. Genes highly up-regulated during differentiation were associated with multiple clustered adipocyte-specific FAIRE peaks. Among the adipocyte-specific FAIRE peaks, 45.3% and 11.7% overlapped binding sites for, respectively, PPARγ and C/EBPα, the master regulators of adipocyte differentiation. Computational motif analyses of the adipocyte-specific FAIRE peaks revealed enrichment of a binding motif for nuclear family I (NFI transcription factors. Indeed, ChIP assay showed that NFI occupy the adipocyte-specific FAIRE peaks and/or the PPARγ binding sites near PPARγ, C/EBPα, and aP2 genes. Overexpression of NFIA in 3T3-L1 cells resulted in robust induction of these genes and lipid droplet formation without differentiation stimulus. Overexpression of dominant-negative NFIA or siRNA-mediated knockdown of NFIA or NFIB significantly suppressed both induction of genes and lipid accumulation during differentiation, suggesting a physiological function of these factors in the adipogenic program. Together, our

  8. SIRT1 Limits Adipocyte Hyperplasia through c-Myc Inhibition.

    Science.gov (United States)

    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.

  9. Effects of xanthohumol-rich hop extract on the differentiation of preadipocytes.

    Science.gov (United States)

    Kiyofuji, Ayane; Yui, Kazuki; Takahashi, Koki; Osada, Kyoichi

    2014-01-01

    Xanthohumol is a major prenylated, hydrophobic flavonoid found in the female inflorescences of the hop plant (Humulus lupulus L.). In this study, we examined the effects of xanthohumol-rich hop extract containing 17.8% xanthohumol and 12.4% isoxanthohumol on the differentiation and adipogenesis of 3T3L1 cells. We observed that the extract inhibited the differentiation of 3T3L1 cells and intracellular fat droplets via the regulation of adipogenic factors such as the peroxisome proliferator-activated receptor gamma, CCAAT/enhancer binding protein alpha, and adipocyte fatty acid-binding protein.

  10. PPARgamma in adipocyte differentiation and metabolism

    DEFF Research Database (Denmark)

    Siersbaek, Rasmus; Nielsen, Ronni; Mandrup, Susanne

    2010-01-01

    Adipocyte differentiation is controlled by a tightly regulated transcriptional cascade in which PPARgamma and members of the C/EBP family are key players. Here we review the roles of PPARgamma and C/EBPs in adipocyte differentiation with emphasis on the recently published genome-wide binding prof...

  11. Free fatty acids and IL-6 induce adipocyte galectin-3 which is increased in white and brown adipose tissues of obese mice.

    Science.gov (United States)

    Krautbauer, Sabrina; Eisinger, Kristina; Hader, Yvonne; Buechler, Christa

    2014-10-01

    Galectin-3 regulates immune cell function and clearance of advanced glycation end products. Galectin-3 is increased in serum of obese humans and mice and most studies suggest that this protein protects from inflammation in metabolic diseases. Current data show that galectin-3 is markedly elevated in the liver, subcutaneous and intra-abdominal fat depots of mice fed a high fat diet and ob/ob mice. Galectin-3 is also increased in brown adipose tissues of these animals and immunohistochemistry confirms higher levels in adipocytes. Raised galectin-3 in obese white adipocytes has been described in the literature and regulation of adipocyte galectin-3 by metabolites with a role in obesity has been analyzed. Galectin-3 is expressed in 3T3-L1 fibroblasts and human preadipocytes and is modestly induced in mature adipocytes. In 3T3-L1 adipocytes galectin-3 is localized in the cytoplasm and is also detected in cell supernatants. Glucose does not alter soluble galectin-3. Lipopolysaccharide has no effect while TNF reduces and IL-6 raises this lectin in cell supernatants. Palmitate and oleate modestly elevate soluble galectin-3. Differentiation of 3T3-L1 cells in the presence of 100 μM and 200 μM linoleate induces soluble galectin-3 and cellular levels are upregulated by the higher concentration. Current data suggest that free fatty acids and IL-6 increase galectin-3 in adipocytes and thereby may contribute to higher levels in obesity.

  12. 6-Gingerol Suppresses Adipocyte-Derived Mediators of Inflammation In Vitro and in High-Fat Diet-Induced Obese Zebra Fish.

    Science.gov (United States)

    Choi, Jia; Kim, Kui-Jin; Kim, Byung-Hak; Koh, Eun-Jeong; Seo, Min-Jung; Lee, Boo-Yong

    2017-02-01

    The present study was performed to investigate the molecular mechanism of 6-gingerol on adipocyte-mediated systemic inflammation in vitro and in high-fat diet-induced obese zebra fish. 6-Gingerol decreased adipogenesis due to the suppression of adipocyte differentiation markers, including peroxisome proliferator-activated receptor gamma, CCAATT enhancer binding protein α, and adipocyte protein 2, and triglyceride synthesis enzymes, including sterol regulatory element-binding protein-1, fatty acid synthase, lysophosphatidic acid acyltransferase, and acyl-coA : diacylglycerol acyltransferase 1, in 3T3-L1. A coculture insert system using 3T3-L1 with RAW 264.7 (coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages) revealed that 6-gingerol increased anti-inflammatory cytokine interleukin-10. The expression of TNFα, monocyte chemotactic protein-1, interleukin-1β, and interleukin-6 were decreased in the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages treated with 6-gingerol. Moreover, the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages treated with 6-gingerol inhibited the protein expression of TNFα and monocyte chemotactic protein-1 in RAW 264.7. 6-Gingerol decreased c-JUN N-terminal kinase and I kappa B kinase beta and its downstream target AP-1 expression in the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages. Furthermore, 6-gingerol decreased the expression of inducible nitric oxide synthase stimulated by the coculture insert system using fully differentiated 3T3-L1 cells with RAW 264.7 macrophages in RAW 264.7 and attenuated nitric oxide production in diet-induced obese zebra fish. Our results suggest that 6-gingerol suppresses inflammation through the regulation of the c-JUN N-terminal kinase-I kappa B kinase beta and its downstream targets.

  13. Role of adipocyte-derived lipoprotein lipase in adipocyte hypertrophy

    Directory of Open Access Journals (Sweden)

    Orlando Robert A

    2007-10-01

    Full Text Available Abstract Background A major portion of available fatty acids for adipocyte uptake is derived from lipoprotein lipase (LPL-mediated hydrolysis of circulating lipoprotein particles. In vivo studies aimed at identifying the precise role of adipocyte-derived LPL in fat storage function of adipose tissue have been unable to provide conclusive evidence due to compensatory mechanisms that activate endogenous fatty acid synthesis. To address this gap in knowledge, we have measured the effect of reducing adipocyte LPL expression on intracellular lipid accumulation using a well-established cultured model of adipocyte differentiation. Methods siRNA specific for mouse LPL was transfected into 3T3-L1 adipocytes. Expression of LPL was measured by quantitative real-time PCR and cell surface-associated LPL enzymatic activity was measured by colorimetric detection following substrate (p-nitrophenyl butyrate hydrolysis. Apolipoprotein CII and CIII expression ratios were also measured by qRT-PCR. Intracellular lipid accumulation was quantified by Nile Red staining. Results During differentiation of 3T3-L1 pre-adipocytes, LPL mRNA expression increases 6-fold resulting in a 2-fold increase in cell surface-associated LPL enzymatic activity. Parallel to this increase in LPL expression, we found that intracellular lipids increased ~10-fold demonstrating a direct correlation between adipocyte-derived LPL expression and lipid storage. We next reduced LPL expression in adipocytes using siRNA transfections to directly quantify the contributions of adipocyte-derived LPL to lipid storage, This treatment reduced LPL mRNA expression and cell surface-associated LPL enzymatic activity to ~50% of non-treated controls while intracellular lipid levels were reduced by 80%. Exogenous addition of purified LPL (to restore extracellular lipolytic activity or palmitate (as a source of free fatty acids to siRNA-treated cells restored intracellular lipid levels to those measured for non

  14. Berberine regulates peroxisome proliferator-activated receptors and positive transcription elongation factor b expression in diabetic adipocytes.

    Science.gov (United States)

    Zhou, Jiyin; Zhou, Shiwen

    2010-12-15

    Berberine has hypoglycemic and hypolipidemic effects on diabetic rats. This study investigated the relationship between hypoglycemic and hypolipidemic effects of berberine and peroxisome proliferator-activated receptors (PPARs) and positive transcription elongation factor b (P-TEFb) (including cyclin-dependent kinase 9 (CDK9) and cyclin T1) in white adipose tissue of diabetic rats and RNA interference-treated 3T3-L1 cells. Berberine promoted differentiation and inhibited lipid accumulation of 3T3-L1 cells, further decreased PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression and decreased tumor necrosis factor α content in supernatants of both control and RNA interference-treated 3T3-L1 cells. After a 16-week induction with 35 mg/kg streptozotocin (i.p.) and high-carbohydrate/high-fat diet, diabetic rats were treated with 75, 150 and 300 mg/kg berberine and 100 mg/kg fenofibrate or 4 mg/kg rosiglitazone for another 16 weeks. Berberine decreased white adipose tissue to body weight ratio and adipocyte size and increased adipocyte number. Berberine upregulated PPARα/δ/γ, CDK9 and cyclin T1 mRNA and protein expression in adipose tissue, decreased tumor necrosis factor α and free fatty acid content and increased lipoprotein lipase activity in serum and adipose tissue. Berberine modulated metabolic related PPARs expression and differentiation related P-TEFb expression in adipocytes, which are associated with its hypoglycemic and hypolipidemic effects.

  15. DNA microarray analysis of genes differentially expressed in adipocyte differentiation

    Indian Academy of Sciences (India)

    Chunyan Yin; Yanfeng Xiao; Wei Zhang; Erdi Xu; Weihua Liu; Xiaoqing Yi; Ming Chang

    2014-06-01

    In the present study, the human liposarcoma cell line SW872 was used to identify global changes in gene expression profiles occurring during adipogenesis. We further explored some of the genes expressed during the late phase of adipocyte differentiation. These genes may play a major role in promoting excessive proliferation and accumulation of lipid droplets, which contribute to the development of obesity. By using microarray-based technology, we examined differential gene expression in early differentiated adipocytes and late differentiated adipocytes. Validated genes exhibited a ≥ 10-fold increase in the late phase of adipocyte differentiation by polymerase chain reaction (RT-PCR). Compared with undifferentiated preadipocytes, we found that 763 genes were increased in early differentiated adipocytes, and 667 genes were increased in later differentiated adipocytes. Furthermore, 21 genes were found being expressed 10-fold higher in the late phase of adipocyte differentiation. The results were in accordance with the RT-PCR test, which validated 11 genes, namely, CIDEC, PID1, LYRM1, ADD1, PPAR2, ANGPTL4, ADIPOQ, ACOX1, FIP1L1, MAP3K2 and PEX14. Most of these genes were found being expressed in the later phase of adipocyte differentiation involved in obesity-related diseases. The findings may help to better understand the mechanism of obesity and related diseases.

  16. Effect of clenbuterol on apoptosis, adipogenesis, and lipolysis in adipocytes.

    Science.gov (United States)

    Kim, Hye-Kyeong; Della-Fera, Mary Anne; Hausman, Dorothy B; Baile, Clifton A

    2010-09-01

    Clenbuterol, a beta(2)-adrenergic receptor (beta(2)-AR) selective agonist, has been shown to decrease body fat in animals and can induce apoptosis in adipose tissue in mice. We hypothesized that direct actions of a beta-adrenergic receptor agonist on adipocytes could trigger the observed apoptotic effect. The hypothesis was inspected by investigating the direct effect of clenbuterol on apoptosis, adipogenesis, and lipolysis in vitro using the 3T3-L1 cell line and rat primary adipocytes. Cells were treated with 10(-9) to 10(-5) M clenbuterol depending on the experiments. There was no apoptotic effect of clenbuterol both in 3T3-L1 cells and rat primary adipocytes. Adipogenesis monitored by Oil Red O staining and AdipoRed assay was modestly decreased by clenbuterol treatment (p < 0.05). In fully differentiated primary adipocytes, clenbuterol increased basal lipolysis compared with the control (p < 0.01). In summary, direct stimulation of beta(2)-AR by clenbuterol does not cause apoptosis in adipocytes, despite a direct lipolytic stimulation and attenuation of adipogenesis.

  17. 非诺贝特通过Sirt1-NF-κB途径调控肿瘤坏死因子α诱导的3T3-L1脂肪细胞炎症反应

    Institute of Scientific and Technical Information of China (English)

    林琴琴; 林蓉; 张继业; 王维蓉; 杨莉娜; 张凯帆

    2011-01-01

    目的探讨非诺贝特对肿瘤坏死因子α(TNF-α)诱导3T3-L1成熟脂肪细胞中炎症相关蛋白CD40L/CD40表达的影响及其是否通过Sirt1-NF-κB途径调控TNF-α诱导的3T3-L1脂肪细胞炎症反应。方法 3T3-L1前体脂肪细胞诱导分化成为成熟脂肪细胞后,免疫荧光检测Sirt1及CD40L/CD40的表达定位。TNF-α刺激细胞以及加入非诺贝特、Sirt1特异性激动剂和抑制剂和NF-κB信号通路阻断剂后,采用免疫印迹法检测细胞内Sirt1、CD40和NF-κB蛋白表达水平,采用RT-Q-PCR检测Sirt1和CD40 mRNA表达水平。结果油红O染色结果表明成功建立诱导分化体系。Sirt1在3T3-L1成熟脂肪中有表达且主要定位于细胞核。免疫荧光和逆转录PCR结果表明,3T3-L1成熟脂肪细胞表达CD40且主要定位于细胞膜上,但不表达CD40L。非诺贝特剂量依赖性下调TNF-α(10μg/L)诱导的成熟脂肪细胞中CD40蛋白和mRNA的表达,上调TNF-α诱导的成熟脂肪细胞中Sirt1蛋白和mRNA的表达。Sirt1特异性激动剂白藜芦醇增强非诺贝特对TNF-α诱导的成熟脂肪细胞中CD40蛋白和mRNA表达的抑制作用,组蛋白去乙酰化酶抑制剂尼克酰胺和Sirt1特异性抑制剂Sirtinol减弱非诺贝特的抑制作用。

  18. Adipocyte induced arterial calcification is prevented with sodium thiosulfate

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Neal X., E-mail: xuechen@iupui.edu [Divison of Nephrology, Indiana University School of Medicine, Indianapolis, IN (United States); O’Neill, Kalisha; Akl, Nader Kassis [Divison of Nephrology, Indiana University School of Medicine, Indianapolis, IN (United States); Moe, Sharon M. [Divison of Nephrology, Indiana University School of Medicine, Indianapolis, IN (United States); Roudebush VA Medical Center, Indianapolis, IN (United States)

    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

  19. Establishment of a preadipocyte cell line derived from mature adipocytes of GFP transgenic mice and formation of adipose tissue.

    Science.gov (United States)

    Nobusue, Hiroyuki; Endo, Tsuyoshi; Kano, Koichiro

    2008-06-01

    We established a preadipocyte cell line from mature adipocytes obtained from subcutaneous fat tissue of green fluorescent protein (GFP) transgenic mice. The floating top layer, containing mature adipocytes, was isolated from subcutaneous fat tissue by collagenase digestion and filtration. Fluorescence-activated cell sorting and microscopic analysis revealed that the floating cell fraction comprised a highly homogeneous adipocyte population with no adipose stromal-vascular cells. Isolated mature adipocytes dedifferentiated into fibroblast-like cells and actively proliferated in ceiling culture. In vitro studies showed that the cells could redifferentiate into mature adipocytes in an identical way to 3T3-L1 preadipocytes. No changes in the differentiation pattern were observed during the propagation of our cells. They were successfully maintained and differentiated for at least 22 passages. We named these cells dedifferentiated fat (DFAT-GFP) cells. When DFAT-GFP cells were implanted subcutaneously into C57BL/6N mice, they developed highly vascularized fat pads that morphologically resembled normal subcutaneous adipose tissue and consisted of GFP-positive cells; however, implanted 3T3-L1 cells did not have such an effect on the mice. We conclude that DFAT-GFP cells provide a model that should enable us to study the mechanisms of adipocyte differentiation and adipose tissue formation in vivo and in vitro.

  20. Modulation of chromatin access during adipocyte differentiation

    DEFF Research Database (Denmark)

    Mandrup, Susanne; Hager, Gordon L

    2012-01-01

    Cellular development requires reprogramming of the genome to modulate the gene program of the undifferentiated cell and allow expression of the gene program unique to differentiated cells. A number of key transcription factors involved in this reprogramming of preadipocytes to adipocytes have bee...

  1. Aldose reductases influence prostaglandin F2α levels and adipocyte differentiation in male mouse and human species.

    Science.gov (United States)

    Pastel, Emilie; Pointud, Jean-Christophe; Loubeau, Gaëlle; Dani, Christian; Slim, Karem; Martin, Gwenaëlle; Volat, Fanny; Sahut-Barnola, Isabelle; Val, Pierre; Martinez, Antoine; Lefrançois-Martinez, Anne-Marie

    2015-05-01

    Aldose reductases (AKR1B) are widely expressed oxidoreductases whose physiological function remains elusive. Some isoforms are genuine prostaglandin F2α (PGF2α) synthases, suggesting they might influence adipose homeostasis because PGF2α inhibits adipogenesis. This was shown by Akr1b7 gene ablation in the mouse, which resulted in increased adiposity related to a lower PGF2α content in fat. Yet humans have no ortholog gene for Akr1b7, so the role of aldose reductases in human adipose homeostasis remains to be explored. We analyzed expression of genes encoding human and mouse aldose reductase isoforms in adipose tissues and differentiating adipocytes to assess conserved mechanisms regulating PGF2α synthesis and adipogenesis. The Akr1b3 gene encoded the most abundant isoform in mouse adipose tissue, whereas Akr1b7 encoded the only isoform enriched in the stromal vascular fraction. Most mouse aldose reductase gene expression peaked in early adipogenesis of 3T3-L1 cells and diminished with differentiation. In contrast with its mouse ortholog Akr1b3, AKR1B1 expression increased throughout differentiation of human multipotent adipose-derived stem cells, paralleling PGF2α release, whereas PGF2α receptor (FP) levels collapsed in early differentiation. Pharmacological inhibition of aldose reductase using Statil altered PGF2α production and enhanced human multipotent adipose-derived stem adipocyte differentiation. As expected, the adipogenic effects of Statil were counteracted by an FP agonist (cloprostenol). Thus, in both species aldose reductase-dependent PGF2α production could be important in early differentiation to restrict adipogenesis. PGF2α antiadipogenic signaling could then be toned down through the FP receptor or aldose reductases down-regulation in human and mouse cells, respectively. Our data suggest that aldose reductase inhibitors could have obesogenic potential.

  2. An Model to Probe the Regulation of Adipocyte Differentiation under Hyperglycemia

    Directory of Open Access Journals (Sweden)

    Kusampudi Shilpa

    2013-06-01

    Full Text Available BackgroundThe aim of this study was an in vitro investigation of the effect of high glucose concentration on adipogenesis, as prolonged hyperglycemia alters adipocyte differentiation.Methods3T3-L1 preadipocytes differentiated in the presence of varying concentrations of glucose (25, 45, 65, 85, and 105 mM were assessed for adipogenesis using AdipoRed (Lonza assay. Cell viability and proliferation were measured using MTT reduction and [3H] thymidine incorporation assay. The extent of glucose uptake and glycogen synthesis were measured using radiolabelled 2-deoxy-D-[1-3H] glucose and [14C]-UDP-glucose. The gene level expression was evaluated using reverse transcription-polymerase chain reaction and protein expression was studied using Western blot analysis.ResultsGlucose at 105 mM concentration was observed to inhibit adipogenesis through inhibition of CCAAT-enhancer-binding proteins, sterol regulatory element-binding protein, peroxisome proliferator-activated receptor and adiponectin. High concentration of glucose induced stress by increasing levels of toll-like receptor 4, nuclear factor κB and tumor necrosis factor α thereby generating activated preadipocytes. These cells entered the state of hyperplasia through inhibition of p27 and proliferation was found to increase through activation of protein kinase B via phosphoinositide 3 kinase dependent pathway. This condition inhibited insulin signaling through decrease in insulin receptor β. Although the glucose transporter 4 (GLUT4 protein remained unaltered with the glycogen synthesis inhibited, the cells were found to exhibit an increase in glucose uptake via GLUT1.ConclusionAdipogenesis in the presence of 105 mM glucose leads to an uncontrolled proliferation of activated preadipocytes providing an insight towards understanding obesity.

  3. Signal transducer and activator of transcription 5B (STAT5B) modulates adipocyte differentiation via MOF.

    Science.gov (United States)

    Gao, Peng; Zhang, Yuchao; Liu, Yuantao; Chen, Jicui; Zong, Chen; Yu, Cong; Cui, Shang; Gao, Weina; Qin, Dandan; Sun, Wenchuan; Li, Xia; Wang, Xiangdong

    2015-12-01

    The role and mechanism of signal transducer and activator of transcription 5B (STAT5B) in adipogenesis remain unclear. In this study, our data showed that Males absent on the first (MOF) protein expression was increased during 3 T3-L1 preadipocytes differentiation accompanied with STAT5B expression increasing. Over-expression STAT5B enhanced MOF promoter trans-activation in HeLa cells. Mutagenesis assay and ChIP analysis exhibited that STAT5B was able to bind MOF promoter. Knocking-down STAT5B in 3 T3-L1 preadipocytes led to decreased expression of MOF, but resulted in increased expression of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα) and fatty acid-binding protein 4 (Fabp4), which were important factors or enzymes for adipogenesis. We also found that knocking-down MOF in 3 T3-L1 preadipocytes resulted in increased expression of PPARγ, C/EBPα and Fabp4, which was in the same trend as STAT5B knocking-down. Over-expression MOF resulted in reduced promoter trans-activation activity of C/EBPα. These results suggest that STAT5B and MOF work as negative regulators in adipogenesis, and STAT5B modulates preadipocytes differentiation partially by regulating MOF expression.

  4. Translocator protein (18 kDa) as a pharmacological target in adipocytes to regulate glucose homeostasis.

    Science.gov (United States)

    Li, Jiehan; Papadopoulos, Vassilios

    2015-09-01

    As a major regulator in obesity and its associated metabolic complications, the proper functioning of adipocytes is crucial for health maintenance, thus serving as an important target for the development of anti-obese and anti-diabetic therapies. There is increasing evidence that mitochondrial malfunction is a pivotal event in disturbing adipocyte cell homeostasis. Among major mitochondrial structure components, the high-affinity drug- and cholesterol-binding outer mitochondrial membrane translocator protein (18 kDa; TSPO) has shown importance across a broad spectrum of mitochondrial functions. Recent studies demonstrated the presence of TSPO in white adipocyte mitochondria of mice, and administration of TSPO drug ligands to obese mice reduced weight gain and lowered glucose level. Therefore, it is of great interest to assess whether TSPO in adipocytes could serve as a drug target to regulate adipocyte activities with potential influence on weight control and glucose metabolism. Two structurally distinct TSPO drug ligands, PK 11195 and FGIN-1-27, improved the intracellular dynamics of 3T3-L1 adipocytes, such as the production and release of adipokines, glucose uptake, and adipogenesis. TSPO knockdown in either differentiated adipocytes or preadipocytes impaired these functions. Findings from 3T3-L1 cells were related to human primary cells, where TSPO expression was tightly associated with the metabolic state of primary adipocytes and the differentiation of primary preadipocytes. These results suggest that TSPO expression is essential to safeguard healthy adipocyte functions, and that TSPO activation in adipocytes improves their metabolic status in regulating glucose homeostasis. Adipocyte TSPO may serve as a pharmacologic target for the treatment of obesity and diabetes.

  5. The dietary fatty acid 10E12Z-CLA induces epiregulin expression through COX-2 dependent PGF(2α) synthesis in adipocytes.

    Science.gov (United States)

    Belda, Benjamin J; Thompson, Jerry T; Sinha, Raghu; Prabhu, K Sandeep; Vanden Heuvel, John P

    2012-10-01

    Conjugated linoleic acids (CLAs) are a group of dietary fatty acids that are widely marketed as weight loss supplements. The isomer responsible for this effect is the trans-10, cis-12 CLA (10E12Z-CLA) isomer. 10E12Z-CLA treatment during differentiation of 3T3-L1 adipocytes induces expression of prostaglandin-endoperoxide synthase-2 (Cyclooxygenase-2; COX-2). This work demonstrates that COX-2 is also induced in fully differentiated 3T3-L1 adipocytes after a single treatment of 10E12Z-CLA at both the mRNA (20-40 fold) and protein level (7 fold). Furthermore, prostaglandin (PG)F(2α), but not PGE(2), is significantly increased 10 fold. In female BALB/c mice fed 0.5% 10E12Z-CLA for 10 days, COX-2 was induced in uterine adipose (2 fold). In vitro, pharmacological COX-2 inhibition did not block the effect of 10E12Z-CLA on adipocyte-specific gene expression although PGF(2α) was dose-dependently decreased. These studies demonstrate that PGF(2α) was not by itself responsible for the reduction in adipocyte character due to 10E12Z-CLA treatment. However, PGF(2α), either exogenously or endogenously in response to 10E12Z-CLA, increased the expression of the potent mitogen and epidermal growth factor (EGF) receptor (EGFR) ligand epiregulin in 3T3-L1 adipocytes. Blocking PGF(2α) signaling with the PGF(2α) receptor (FP) antagonist AL-8810 returned epiregulin mRNA levels back to baseline. Although this pathway is not directly responsible for adipocyte dependent gene expression, these results suggest that this signaling pathway may still have broad effect on the adipocyte and surrounding cells.

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

    Energy Technology Data Exchange (ETDEWEB)

    Lasrich, Dorothee; Bartelt, Alexander [Department of Biochemistry and Molecular Cell Biology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246 Hamburg (Germany); Grewal, Thomas, E-mail: thomas.grewal@sydney.edu.au [Faculty of Pharmacy A15, The University of Sydney, Sydney, NSW 2006 (Australia); Heeren, Joerg, E-mail: heeren@uke.de [Department of Biochemistry and Molecular Cell Biology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246 Hamburg (Germany)

    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.

  7. Limonin, a Component of Dictamni Radicis Cortex, Inhibits Eugenol-Induced Calcium and cAMP Levels and PKA/CREB Signaling Pathway in Non-Neuronal 3T3-L1 Cells.

    Science.gov (United States)

    Yoon, Yeo Cho; Kim, Sung-Hee; Kim, Min Jung; Yang, Hye Jeong; Rhyu, Mee-Ra; Park, Jae-Ho

    2015-12-10

    Limonin, one of the major components in dictamni radicis cortex (DRC), has been shown to play various biological roles in cancer, inflammation, and obesity in many different cell types and tissues. Recently, the odorant-induced signal transduction pathway (OST) has gained attention not only because of its function in the perception of smell but also because of its numerous physiological functions in non-neuronal cells. However, little is known about the effects of limonin and DRC on the OST pathway in non-neuronal cells. We investigated odorant-stimulated increases in Ca(2+) and cAMP, major second messengers in the OST pathway, in non-neuronal 3T3-L1 cells pretreated with limonin and ethanol extracts of DRC. Limonin and the extracts significantly decreased eugenol-induced Ca(2+) and cAMP levels and upregulated phosphorylation of CREB and PKA. Our results demonstrated that limonin and DRC extract inhibit the OST pathway in non-neuronal cells by modulating Ca(2+) and cAMP levels and phosphorylation of CREB.

  8. Curcuma longa polyphenols improve insulin-mediated lipid accumulation and attenuate proinflammatory response of 3T3-L1 adipose cells during oxidative stress through regulation of key adipokines and antioxidant enzymes.

    Science.gov (United States)

    Septembre-Malaterre, Axelle; Le Sage, Fanny; Hatia, Sarah; Catan, Aurélie; Janci, Laurent; Gonthier, Marie-Paule

    2016-07-08

    Plant polyphenols may exert beneficial action against obesity-related oxidative stress and inflammation which promote insulin resistance. This study evaluated the effect of polyphenols extracted from French Curcuma longa on 3T3-L1 adipose cells exposed to H2 O2 -mediated oxidative stress. We found that Curcuma longa extract exhibited high amounts of curcuminoids identified as curcumin, demethoxycurcumin, and bisdemethoxycurcumin, which exerted free radical-scavenging activities. Curcuma longa polyphenols improved insulin-mediated lipid accumulation and upregulated peroxisome proliferator-activated receptor-gamma gene expression and adiponectin secretion which decreased in H2 O2 -treated cells. Curcuminoids attenuated H2 O2 -enhanced production of pro-inflammatory molecules such as interleukin-6, tumor necrosis factor-alpha, monocyte chemoattractant protein-1, and nuclear factor κappa B. Moreover, they reduced intracellular levels of reactive oxygen species elevated by H2 O2 and modulated the expression of genes encoding superoxide dismutase and catalase antioxidant enzymes. Collectively, these findings highlight that Curcuma longa polyphenols protect adipose cells against oxidative stress and may improve obesity-related metabolic disorders. © 2016 BioFactors, 42(4):418-430, 2016.

  9. Gelidium elegans, an edible red seaweed, and hesperidin inhibit lipid accumulation and production of reactive oxygen species and reactive nitrogen species in 3T3-L1 and RAW264.7 cells.

    Science.gov (United States)

    Jeon, Hui-Jeon; Seo, Min-Jung; Choi, Hyeon-Son; Lee, Ok-Hwan; Lee, Boo-Yong

    2014-11-01

    Gelidium elegans is an edible red alga native to the intertidal area of northeastern Asia. We investigated the effect of G. elegans extract and its main flavonoids, rutin and hesperidin, on lipid accumulation and the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in 3T3-L1 and RAW264.7 cells. Our data show that G. elegans extract decreased lipid accumulation and ROS/RNS production in a dose-dependent manner. The extract also inhibited the mRNA expression of adipogenic transcription factors, such as peroxisome proliferator-activated receptor gamma and CCAAT/enhancer-binding protein alpha, while enhancing the protein expression of the antioxidant enzymes superoxide dismutases 1 and 2, glutathione peroxidase, and glutathione reductase compared with controls. In addition, lipopolysaccharide-induced nitric oxide production was significantly reduced in G. elegans extract-treated RAW264.7 cells. In analysis of the effects of G. elegans flavonoids on lipid accumulation and ROS/RNS production, only hesperidin showed an inhibitory effect on lipid accumulation and ROS production; rutin did not affect adipogenesis and ROS status. The antiadipogenic effect of hesperidin was evidenced by the downregulation of peroxisome proliferator-activated receptor gamma, CCAAT/enhancer-binding protein alpha, and fatty acid binding protein 4 gene expression. Collectively, our data suggest that G. elegans is a potential food source containing antiobesity and antioxidant constituents.

  10. Limonin, a Component of Dictamni Radicis Cortex, Inhibits Eugenol-Induced Calcium and cAMP Levels and PKA/CREB Signaling Pathway in Non-Neuronal 3T3-L1 Cells

    Directory of Open Access Journals (Sweden)

    Yeo Cho Yoon

    2015-12-01

    Full Text Available Limonin, one of the major components in dictamni radicis cortex (DRC, has been shown to play various biological roles in cancer, inflammation, and obesity in many different cell types and tissues. Recently, the odorant-induced signal transduction pathway (OST has gained attention not only because of its function in the perception of smell but also because of its numerous physiological functions in non-neuronal cells. However, little is known about the effects of limonin and DRC on the OST pathway in non-neuronal cells. We investigated odorant-stimulated increases in Ca2+ and cAMP, major second messengers in the OST pathway, in non-neuronal 3T3-L1 cells pretreated with limonin and ethanol extracts of DRC. Limonin and the extracts significantly decreased eugenol-induced Ca2+ and cAMP levels and upregulated phosphorylation of CREB and PKA. Our results demonstrated that limonin and DRC extract inhibit the OST pathway in non-neuronal cells by modulating Ca2+ and cAMP levels and phosphorylation of CREB.

  11. Gene expression of the zinc transporter ZIP14 (SLC39a14) is affected by weight loss and metabolic status and associates with PPARγ in human adipose tissue and 3T3-L1 pre-adipocytes

    DEFF Research Database (Denmark)

    Juul, Trine Maxel; Smidt, Kamille; Larsen, Agnete

    2015-01-01

    BACKGROUND: The expansion and function of adipose tissue are important during the development of insulin resistance and inflammation in obesity. Zinc dyshomeostasis is common in obese individuals. In the liver, zinc influx transporter ZIP14, affects proliferation and glucose metabolism but the role...... and during adipogenesis. However, in silico analysis revealed that the ZIP14 promoter does not contain PPARγ-binding motifs. CONCLUSIONS: We hypothesize that ZIP14-mediated zinc influx might directly influence PPARγ activity and that ZIP14 may regulate expansion and function of adipose tissue and serve...

  12. 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

    Directory of Open Access Journals (Sweden)

    Chia-Hua Liang

    2013-01-01

    Full Text Available 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.

  13. 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.

    Science.gov (United States)

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

    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 (PPAR γ ), 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.

  14. 鼠PVRL-2慢病毒载体的构建及其在3T3-L1细胞中的表达%Potential role of mouse PVRL-2 gene in the fatty acid metabolism

    Institute of Scientific and Technical Information of China (English)

    马静; 刘晓萌; 张传海; 郑宗基; 赵倩伟; 杨鸣琦; 张雷

    2013-01-01

    Excess fat and cholesterol in food such as meat,eggs or milk could lead to hyperlipoidemia in human.Currently,to explore genes expression and their mechanisms associated with lipid metabolism has been a major focus in veterinary science.Growing bodies of evidence indicated that molecular functions of fatty acid metabolism related genes such as ApoE,ApoC1 and Tomm40 were very well characterized; however,function of their chromosomal neighbor such as PVRL-2 gene in the fatty acid metabolism remains unclear.Present study was aim to investigate potential role of mouse PVRL-2 gene in regulation of fatty acid related gene expression using preadipogenic 3T3-L1 cells.The cells were infected by Lentiviral particles which was produced by lentiviral plasmid containing Pvrl2 gene,and RNA were extracted 48h post viral infection.Quantitative real-time PCR analysis confirmed that PVRL-2 overexpressed more than 100 folds upon PVRL-2 virus transformation compared to the control.Notably,the expression of PPARα gene which is a key player in the fatty acid oxidation was strongly induced (4.5 fold increase) post PVRL-2 viral infection,but not other genes that related to the fatty acid metabolism such as CPT1A,FASN,COX7A,PGC1B,ASADM showed similar changes.Furthermore,bioinformatics analyses revealed that Nectin-2,coded by PVRL-2,should be a transmembrane protein with a signal peptide.In conclusion,the present study demonstrated that overexpression of PVRL-2 induce the expression of PPARα,which highlight the potential roles of PVRL-2 gene in fatty acid metabolism.Future studies are needed to determine detailed molecular function of PVRL-2 gene in fatty acid metabolism.%过多的脂肪和胆固醇随着肉蛋奶被人体摄入是导致人类高血脂等各种疾病诱发的原因之一,而探索脂代谢通路相关基因的表达变化及其调控机制已经成为分子生物学技术在兽医学领域中的研究热点.与高血脂有关的ApoE、ApoC1和Tomm40等基因研究较多,

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

    Science.gov (United States)

    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.

  16. Berberine Suppresses Adipocyte Differentiation via Decreasing CREB Transcriptional Activity.

    Directory of Open Access Journals (Sweden)

    Juan Zhang

    Full Text Available Berberine, one of the major constituents of Chinese herb Rhizoma coptidis, has been demonstrated to lower blood glucose, blood lipid, and body weight in patients with type 2 diabetes mellitus. The anti-obesity effect of berberine has been attributed to its anti-adipogenic activity. However, the underlying molecular mechanism remains largely unknown. In the present study, we found that berberine significantly suppressed the expressions of CCAAT/enhancer-binding protein (C/EBPα, peroxisome proliferators-activated receptor γ2 (PPARγ2, and other adipogenic genes in the process of adipogenesis. Berberine decreased cAMP-response element-binding protein (CREB phosphorylation and C/EBPβ expression at the early stage of 3T3-L1 preadipocyte differentiation. In addition, CREB phosphorylation and C/EBPβ expression induced by 3-isobutyl-1-methylxanthine (IBMX and forskolin were also attenuated by berberine. The binding activities of cAMP responsive element (CRE stimulated by IBMX and forskolin were inhibited by berberine. The binding of phosphorylated CREB to the promoter of C/EBPβ was abrogated by berberine after the induction of preadipocyte differentiation. These results suggest that berberine blocks adipogenesis mainly via suppressing CREB activity, which leads to a decrease in C/EBPβ-triggered transcriptional cascades.

  17. Berberine Suppresses Adipocyte Differentiation via Decreasing CREB Transcriptional Activity.

    Science.gov (United States)

    Zhang, Juan; Tang, Hongju; Deng, Ruyuan; Wang, Ning; Zhang, Yuqing; Wang, Yao; Liu, Yun; Li, Fengying; Wang, Xiao; Zhou, Libin

    2015-01-01

    Berberine, one of the major constituents of Chinese herb Rhizoma coptidis, has been demonstrated to lower blood glucose, blood lipid, and body weight in patients with type 2 diabetes mellitus. The anti-obesity effect of berberine has been attributed to its anti-adipogenic activity. However, the underlying molecular mechanism remains largely unknown. In the present study, we found that berberine significantly suppressed the expressions of CCAAT/enhancer-binding protein (C/EBP)α, peroxisome proliferators-activated receptor γ2 (PPARγ2), and other adipogenic genes in the process of adipogenesis. Berberine decreased cAMP-response element-binding protein (CREB) phosphorylation and C/EBPβ expression at the early stage of 3T3-L1 preadipocyte differentiation. In addition, CREB phosphorylation and C/EBPβ expression induced by 3-isobutyl-1-methylxanthine (IBMX) and forskolin were also attenuated by berberine. The binding activities of cAMP responsive element (CRE) stimulated by IBMX and forskolin were inhibited by berberine. The binding of phosphorylated CREB to the promoter of C/EBPβ was abrogated by berberine after the induction of preadipocyte differentiation. These results suggest that berberine blocks adipogenesis mainly via suppressing CREB activity, which leads to a decrease in C/EBPβ-triggered transcriptional cascades.

  18. Effects of fatty acid regulation on visfatin gene expression in adipocytes

    Institute of Scientific and Technical Information of China (English)

    WEN Yu; WANG Hong-wei; WU Jing; LU Hui-ling; HU Xiu-fen; Katherine Cianflone

    2006-01-01

    .5 mmol/L oleate or 1.0 mmol/L palmitate. Visfatin mRNA expression increased during differentiation more than 1.5-fold. Bovine serum albumin (BSA) did not influence visfatin mRNA expression compared with the control group. Dose-response studies demonstrated that addition of 0.125 mmol/L oleate and palmitate to 3T3-L1 adipocytes decreased visfatin mRNA expression significantly (78%, 77%, respectively,relative to untreated control, P<0.05), and further to 65% (relative to untreated control, P<0.05) and 55% (relative to untreated control, P<0.01) at 1.0 mmol/L FFA. Furthermore, the suppression on preadipocytes was similar to that of adipocytes, which reached a maximal reduction of 44% (oleate, P<0.05) and 47% (palmitate, P<0.05) at 1.0 mmol/L FFA.Conclusions Oleic acid and palmitic acid may induce insulin resistance in 3T3-L1 adipocytes and preadipocytes. Downregulation of visfatin mRNA may contribute to impair insulin sensitivity caused by oleate and palmitate.

  19. Genistein inhibits differentiation of primary human adipocytes.

    Science.gov (United States)

    Park, Hea Jin; Della-Fera, Mary Anne; Hausman, Dorothy B; Rayalam, Srujana; Ambati, Suresh; Baile, Clifton A

    2009-02-01

    Genistein, a major soy isoflavone, has been reported to exhibit antiadipogenic and proapoptotic potential in vivo and in vitro. It is also a phytoestrogen which has high affinity to estrogen receptor beta. In this study, we determined the effect of genistein on adipogenesis and estrogen receptor (ER) alpha and beta expression during differentiation in primary human preadipocytes. Genistein inhibited lipid accumulation in a dose-dependent manner at concentrations of 6.25 microM and higher, with 50 microM genistein inhibiting lipid accumulation almost completely. Low concentrations of genistein (3.25 microM) increased cell viability and higher concentrations (25 and 50 microM) decreased it by 16.48+/-1.35% (P<.0001) and 50.68+/-1.34% (P<.0001). Oil Red O staining was used to confirm the effects on lipid accumulation. The inhibition of lipid accumulation was associated with inhibition of glycerol-3-phosphate dehydrogenase activity and down-regulation of expression of adipocyte-specific genes, including peroxisome proliferator-activated receptor gamma, CCAAT/enhancer binding protein alpha, glycerol-3-phosphate dehydrogenase, adipocyte fatty acid binding protein, fatty acid synthase, sterol regulatory element-binding protein 1, perilipin, leptin, lipoprotein lipase and hormone-sensitive lipase. These effects of genistein during the differentiation period were associated with down-regulation of ERalpha and ERbeta expression. This study adds to the elucidation of the molecular pathways involved in the inhibition of adipogenesis by phytoestrogens.

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

    Science.gov (United States)

    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.

  1. Integrator complex plays an essential role in adipose differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Otani, Yuichiro; Nakatsu, Yusuke [Department of Medical Chemistry, Division of Molecular Medical Science, Graduate School of Biomedical Sciences, Hiroshima University (Japan); Sakoda, Hideyuki [Department of Internal Medicine, Graduate School of Medicine, University of Tokyo, Tokyo (Japan); Fukushima, Toshiaki [Department of Medical Chemistry, Division of Molecular Medical Science, Graduate School of Biomedical Sciences, Hiroshima University (Japan); Fujishiro, Midori [Department of Internal Medicine, Graduate School of Medicine, University of Tokyo, Tokyo (Japan); Kushiyama, Akifumi [Department of Internal Medicine, The Institute for Adult Diseases, Asahi Life Foundation, Tokyo (Japan); Okubo, Hirofumi; Tsuchiya, Yoshihiro; Ohno, Haruya [Department of Medical Chemistry, Division of Molecular Medical Science, Graduate School of Biomedical Sciences, Hiroshima University (Japan); Takahashi, Shin-Ichiro [Graduate School of Agriculture and Life Sciences, University of Tokyo, Tokyo (Japan); Nishimura, Fusanori [Department of Dental Science for Health Promotion, Division of Cervico-Gnathostomatology, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima (Japan); Kamata, Hideaki [Department of Medical Chemistry, Division of Molecular Medical Science, Graduate School of Biomedical Sciences, Hiroshima University (Japan); Katagiri, Hideki [Division of Molecular Metabolism and Diabetes, Tohoku University Graduate School of Medicine, Sendai (Japan); Asano, Tomoichiro, E-mail: tasano@hiroshima-u.ac.jp [Department of Medical Chemistry, Divisio