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Bruce M. Spiegelman – One of the best experts on this subject based on the ideXlab platform.

  • Characterization of the novel brown Adipocyte Cell Line HIB 1B. Adrenergic pathways involved in regulation of uncoupling protein gene expression
    Journal of Cell Science, 1994
    Co-Authors: Susanne Klaus, Lisa Choy, Odette Champigny, Anne Marie Cassard-doulcier, Susan R. Ross, Bruce M. Spiegelman, Daniel Ricquier
    Abstract:

    The HIB 1B Cell Line, derived from a brown fat tumor of a transgenic mouse, is the first established brown Adipocyte Cell Line capable of expressing the brown fat-specific mitochondrial uncoupling protprotein (UCP). UCP gene expression, which was virtually undetectable under basic conditions, was stimulated by acute catecholamine or cyclic AMP treatment to levels comparable to primary cultures of brown Adipocytes. Elevation of UCP mRNA levels following stimulation was very rapid but transient, decreasing after about 4 hours with a half-life between 9 and 13 hours. Immunoblotting showed the presence of UCP in HIB 1B mitochondria, but expression was much lower than observed in BAT or primary cultures of brown Adipocytes. Upon transfection of HIB 1B Cells with a reporter gene containing the UCP promoter, the activity of the transgene was regulatable by cAMP and norepinephrine. Investigation of the possible adrenergic receptors involved in UCP stimulation showed that specific beta 3-adrenergic agonists were much less effective than nonspecific beta-adrenergic agonists and that mRNA levels of the atypical, fat-specific beta 3-adrenoceptor were lower than those observed in brown Adipocytes differentiated in primary culture. From pharmacological evidence we conclude that beta 3-adrenergic receptors account for approximately 30-40% of catecholamine induced UCP gene stimulation, whereas about 60-70% is stimulated via the classical beta 1/2 adrenergic pathway. We conclude that HIB 1B Cells represent a functional system for the study of mechanisms related to brown adipose thermogenesis.

  • Adipocyte specific transcription factor arf6 is a heterodimeric complex of two nuclear hormone receptors ppar7 and rxra
    Nucleic Acids Research, 1994
    Co-Authors: Peter Tontonoz, Reed A. Graves, Adriane I Budavari, Hediye Erdjumentbromage, Mary Lui, Paul Tempst, Bruce M. Spiegelman
    Abstract:

    Previously, we identified a novel transcription factor, ARF6, as a key regulator of the tissue-specific Adipocyte P2 (aP2) enhancer. In order to identify the proteins which comprise the Adipocyte ARF6 complex, we have purified this DNA binding activity from a cultured Adipocyte Cell Line. We have developed a system for growth and differentiation of HIB-1B brown Adipocytes in suspension culture that facilitates the production of large quantities of Adipocyte nuclear extract. ARF6 was purified from HIB-1B nuclear extract by a combination of conventional and sequence-specific DNA affinity chromotography. Chemical sequencing and mass spectral analysis of tryptic peptides derived from the purified polypeptides identifies the ARF6 complex as a heterodimer of the retinoid X receptor alpha (RXR alpha) and the murine peroxisome proliferator activated receptor gamma (PPAR gamma). Of the known PPAR gamma isoforms, PPAR gamma is the predominant form expressed in adipose tissue. These results suggest that PPAR gamma 2 serves a unique function among PPAR family members as an important regulator of Adipocyte-specific gene expression.

  • Hibernoma formation in transgenic mice and isolation of a brown Adipocyte Cell Line expressing the uncoupling protein gene.
    Proceedings of the National Academy of Sciences of the United States of America, 1992
    Co-Authors: Susan R. Ross, Susanne Klaus, Daniel Ricquier, Lisa Choy, Reed A. Graves, Niles Fox, Veronica Solevjeva, Bruce M. Spiegelman
    Abstract:

    Abstract Transgenic mice were produced containing the Adipocyte-specific regulatory region from the Adipocyte P2 (aP2) gene linked to the simian virus 40 transforming genes. Most of the transgenic mice developed brown fat tumors (hibernomas) in their interscapular brown adipose tissue. Hibernoma formation was noticeable in some of the mice as early as 1 day after birth and most of the mice developed very large tumors by 1 month of age. All of the tumor tissue expressed the brown fat-specific uncoupling protprotein (UCP) gene as well as the aP2 gene. Several of the tumors have been used to establish cultured Cell Lines and at least one of these Lines can be induced to differentiate into brown Adipocytes. The cultured Adipocytes express mRNA for UCP upon stimulation with N6,O2′-dibutyryladenosine 3′,5′-cyclic monophosphate, norepinephrine, isoproterenol or D7114, a beta 3 adrenergic agonist. Thus, regulation of the key thermogenic gene UCP can now be studied in an established Cell Line.

Claude Forest – One of the best experts on this subject based on the ideXlab platform.

  • Isolation and characterization of the mouse cytosolic phosphoenolpyruvate carboxykinase (GTP) gene: evidence for tissue-specific hypersensitive sites.
    Molecular and cellular endocrinology, 1999
    Co-Authors: Christine P. Williams, Catherine Postic, Danielle Robin, Pierre Robin, Joseph Parrinello, Richard L. Printz, Mark A. Magnuson, Daryl K. Granner, K Shelton, Claude Forest
    Abstract:

    A 72 kilobase pair DNA fragment that contains the mouse phosphoenolpyruvate carboxykinase (PEPCK) gene locus, pck1, was isolated from a genomic bacterial artificial chromosome library. The region from approximately -5.5 to +6.6 kilobase pairs relative to the pck1 transcription start site was sequenced and exhibits a high degree of homology to the rat and human genes. Additionally, the chromatin structure of the PEPCK gene in mouse liver resembles that seen in rat. Backcross panel analysis of a microsatellite sequence confirms that the gene is located on chromosome 2. Hypersensitive site analysis was performed on nuclei isolated from the Adipocyte Cell Line 3T3-F442A in the preadipose and adipose states. Several hypersensitive sites are present in the undifferentiated 3T3-F442A Cells, before PEPCK mRNA is detected. The same sites are present after differentiation, however, the sensitivity of mHS 3 increases relative to the others. We conclude that the chromatin is open in 3T3-F442A Cells and that factors are able to bind in the undifferentiated state but that something else is required for transcription.

  • ISOLATION AND CHARACTERIZATION OF THE MOUSE CYTOSOLIC PHOSPHOENOLPYRUVATE CARBOXYKINASE (GTP) GENE : EVIDENCE FOR TISSUE-SPECIFIC HYPERSENSITIVE SITES
    Molecular and Cellular Endocrinology, 1999
    Co-Authors: Christine P. Williams, Catherine Postic, Danielle Robin, Pierre Robin, Joseph Parrinello, Kathy D. Shelton, Richard L. Printz, Mark A. Magnuson, Daryl K. Granner, Claude Forest
    Abstract:

    Abstract A 72 kilobase pair DNA fragment that contains the mouse phosphoenolpyruvate carboxykinase (PEPCK) gene locus, pck1, was isolated from a genomic bacterial artificial chromosome library. The region from ∼−5.5 to +6.6 kilobase pairs relative to the pck1 transcription start site was sequenced and exhibits a high degree of homology to the rat and human genes. Additionally, the chromatin structure of the PEPCK gene in mouse liver resembles that seen in rat. Backcross panel analysis of a microsatellite sequence confirms that the gene is located on chromosome 2. Hypersensitive site analysis was performed on nuclei isolated from the Adipocyte Cell Line 3T3-F442A in the preadipose and adipose states. Several hypersensitive sites are present in the undifferentiated 3T3-F442A Cells, before PEPCK mRNA is detected. The same sites are present after differentiation, however, the sensitivity of mHS 3 increases relative to the others. We conclude that the chromatin is open in 3T3-F442A Cells and that factors are able to bind in the undifferentiated state but that something else is required for transcription.

Lotfi Chouchane – One of the best experts on this subject based on the ideXlab platform.

  • Comprehensive molecular characterization of human Adipocytes reveals a transient brown phenotype
    Journal of Translational Medicine, 2015
    Co-Authors: Andrea Guennoun, Melissa Kazantzis, Remy Thomas, Martin Wabitsch, Daniel Tews, Konduru Seetharama Sastry, Mouaadh Abdelkarim, Vladimir Zilberfarb, Arthur Donny Strosberg, Lotfi Chouchane
    Abstract:

    AbstractBackgroundFunctional brown adipose tissue (BAT), involved in energy expenditure, has recently been detected in substantial amounts in adults. Formerly overlooked BAT has now become an attractive anti-obesity target.Methods and resultsMolecular characterization of human brown and white Adipocytes, using a myriad of techniques including high-throughput RNA sequencing and functional assays, showed that PAZ6 and SW872 Cells exhibit classical molecular and phenotypic markers of brown and white Adipocytes, respectively. However, the pre-Adipocyte Cell Line SGBS presents a versatile phenotype. A transit expression of classical brown markers such as UCP1 and PPARγ peaked and decLined at day 28 post-differentiation initiation. Conversely, white Adipocyte markers, including Tcf21, showed reciprocal behavior. Interestingly, leptin levels peaked at day 28 whereas the highest adiponectin mRNA levels were detected at day 14 of differentiation. Phenotypic analysis of the abundance and shape of lipid droplets were consistent with the molecular patterns. Accordingly, the oxidative capacity of SGBS Adipocytes peaked on differentiation day 14 and decLined progressively towards differentiation day 28.ConclusionsOur studies have unveiled a new phenotype of human Adipocytes, providing a tool to identify molecular gene expression patterns and pathways involved in the conversion between white and brown Adipocytes.

  • Comprehensive molecular characterization of human Adipocytes reveals a transient brown phenotype
    Journal of translational medicine, 2015
    Co-Authors: Andrea Guennoun, Melissa Kazantzis, Remy Thomas, Martin Wabitsch, Daniel Tews, Mouaadh Abdelkarim, Vladimir Zilberfarb, Arthur Donny Strosberg, Konduru S. Sastry, Lotfi Chouchane
    Abstract:

    Functional brown adipose tissue (BAT), involved in energy expenditure, has recently been detected in substantial amounts in adults. Formerly overlooked BAT has now become an attractive anti-obesity target. Molecular characterization of human brown and white Adipocytes, using a myriad of techniques including high-throughput RNA sequencing and functional assays, showed that PAZ6 and SW872 Cells exhibit classical molecular and phenotypic markers of brown and white Adipocytes, respectively. However, the pre-Adipocyte Cell Line SGBS presents a versatile phenotype. A transit expression of classical brown markers such as UCP1 and PPARγ peaked and decLined at day 28 post-differentiation initiation. Conversely, white Adipocyte markers, including Tcf21, showed reciprocal behavior. Interestingly, leptin levels peaked at day 28 whereas the highest adiponectin mRNA levels were detected at day 14 of differentiation. Phenotypic analysis of the abundance and shape of lipid droplets were consistent with the molecular patterns. Accordingly, the oxidative capacity of SGBS Adipocytes peaked on differentiation day 14 and decLined progressively towards differentiation day 28. Our studies have unveiled a new phenotype of human Adipocytes, providing a tool to identify molecular gene expression patterns and pathways involved in the conversion between white and brown Adipocytes.

Christine P. Williams – One of the best experts on this subject based on the ideXlab platform.

  • Isolation and characterization of the mouse cytosolic phosphoenolpyruvate carboxykinase (GTP) gene: evidence for tissue-specific hypersensitive sites.
    Molecular and cellular endocrinology, 1999
    Co-Authors: Christine P. Williams, Catherine Postic, Danielle Robin, Pierre Robin, Joseph Parrinello, Richard L. Printz, Mark A. Magnuson, Daryl K. Granner, K Shelton, Claude Forest
    Abstract:

    A 72 kilobase pair DNA fragment that contains the mouse phosphoenolpyruvate carboxykinase (PEPCK) gene locus, pck1, was isolated from a genomic bacterial artificial chromosome library. The region from approximately -5.5 to +6.6 kilobase pairs relative to the pck1 transcription start site was sequenced and exhibits a high degree of homology to the rat and human genes. Additionally, the chromatin structure of the PEPCK gene in mouse liver resembles that seen in rat. Backcross panel analysis of a microsatellite sequence confirms that the gene is located on chromosome 2. Hypersensitive site analysis was performed on nuclei isolated from the Adipocyte Cell Line 3T3-F442A in the preadipose and adipose states. Several hypersensitive sites are present in the undifferentiated 3T3-F442A Cells, before PEPCK mRNA is detected. The same sites are present after differentiation, however, the sensitivity of mHS 3 increases relative to the others. We conclude that the chromatin is open in 3T3-F442A Cells and that factors are able to bind in the undifferentiated state but that something else is required for transcription.

  • ISOLATION AND CHARACTERIZATION OF THE MOUSE CYTOSOLIC PHOSPHOENOLPYRUVATE CARBOXYKINASE (GTP) GENE : EVIDENCE FOR TISSUE-SPECIFIC HYPERSENSITIVE SITES
    Molecular and Cellular Endocrinology, 1999
    Co-Authors: Christine P. Williams, Catherine Postic, Danielle Robin, Pierre Robin, Joseph Parrinello, Kathy D. Shelton, Richard L. Printz, Mark A. Magnuson, Daryl K. Granner, Claude Forest
    Abstract:

    Abstract A 72 kilobase pair DNA fragment that contains the mouse phosphoenolpyruvate carboxykinase (PEPCK) gene locus, pck1, was isolated from a genomic bacterial artificial chromosome library. The region from ∼−5.5 to +6.6 kilobase pairs relative to the pck1 transcription start site was sequenced and exhibits a high degree of homology to the rat and human genes. Additionally, the chromatin structure of the PEPCK gene in mouse liver resembles that seen in rat. Backcross panel analysis of a microsatellite sequence confirms that the gene is located on chromosome 2. Hypersensitive site analysis was performed on nuclei isolated from the Adipocyte Cell Line 3T3-F442A in the preadipose and adipose states. Several hypersensitive sites are present in the undifferentiated 3T3-F442A Cells, before PEPCK mRNA is detected. The same sites are present after differentiation, however, the sensitivity of mHS 3 increases relative to the others. We conclude that the chromatin is open in 3T3-F442A Cells and that factors are able to bind in the undifferentiated state but that something else is required for transcription.

Daniel Ricquier – One of the best experts on this subject based on the ideXlab platform.

  • Characterization of the novel brown Adipocyte Cell Line HIB 1B. Adrenergic pathways involved in regulation of uncoupling protein gene expression
    Journal of Cell Science, 1994
    Co-Authors: Susanne Klaus, Lisa Choy, Odette Champigny, Anne Marie Cassard-doulcier, Susan R. Ross, Bruce M. Spiegelman, Daniel Ricquier
    Abstract:

    The HIB 1B Cell Line, derived from a brown fat tumor of a transgenic mouse, is the first established brown Adipocyte Cell Line capable of expressing the brown fat-specific mitochondrial uncoupling protein (UCP). UCP gene expression, which was virtually undetectable under basic conditions, was stimulated by acute catecholamine or cyclic AMP treatment to levels comparable to primary cultures of brown Adipocytes. Elevation of UCP mRNA levels following stimulation was very rapid but transient, decreasing after about 4 hours with a half-life between 9 and 13 hours. Immunoblotting showed the presence of UCP in HIB 1B mitochondria, but expression was much lower than observed in BAT or primary cultures of brown Adipocytes. Upon transfection of HIB 1B Cells with a reporter gene containing the UCP promoter, the activity of the transgene was regulatable by cAMP and norepinephrine. Investigation of the possible adrenergic receptors involved in UCP stimulation showed that specific beta 3-adrenergic agonists were much less effective than nonspecific beta-adrenergic agonists and that mRNA levels of the atypical, fat-specific beta 3-adrenoceptor were lower than those observed in brown Adipocytes differentiated in primary culture. From pharmacological evidence we conclude that beta 3-adrenergic receptors account for approximately 30-40% of catecholamine induced UCP gene stimulation, whereas about 60-70% is stimulated via the classical beta 1/2 adrenergic pathway. We conclude that HIB 1B Cells represent a functional system for the study of mechanisms related to brown adipose thermogenesis.

  • Hibernoma formation in transgenic mice and isolation of a brown Adipocyte Cell Line expressing the uncoupling protein gene.
    Proceedings of the National Academy of Sciences of the United States of America, 1992
    Co-Authors: Susan R. Ross, Susanne Klaus, Daniel Ricquier, Lisa Choy, Reed A. Graves, Niles Fox, Veronica Solevjeva, Bruce M. Spiegelman
    Abstract:

    Abstract Transgenic mice were produced containing the Adipocyte-specific regulatory region from the Adipocyte P2 (aP2) gene linked to the simian virus 40 transforming genes. Most of the transgenic mice developed brown fat tumors (hibernomas) in their interscapular brown adipose tissue. Hibernoma formation was noticeable in some of the mice as early as 1 day after birth and most of the mice developed very large tumors by 1 month of age. All of the tumor tissue expressed the brown fat-specific uncoupling protein (UCP) gene as well as the aP2 gene. Several of the tumors have been used to establish cultured Cell Lines and at least one of these Lines can be induced to differentiate into brown Adipocytes. The cultured Adipocytes express mRNA for UCP upon stimulation with N6,O2′-dibutyryladenosine 3′,5′-cyclic monophosphate, norepinephrine, isoproterenol or D7114, a beta 3 adrenergic agonist. Thus, regulation of the key thermogenic gene UCP can now be studied in an established Cell Line.