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Modulators of UCP1-dependent thermogenesis: Glucocorticoids, diet and novel research models
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The activation and recruitment of brown adipose tissue (BAT) thermogenesis has been put forward as a promising strategy to reduce the disease burden of obesity and obesity-related diseases. Heat production by BAT can be attributed to the tissue-specific mitochondrial uncoupling protein 1 (UCP1). Upon activation, UCP1 uncouples substrate oxidation from ATP production, thereby dissipating energy solely as heat and thus facilitating the ‘wasting’ of energy. To date, cold exposure is the strongest known BAT activator. However, to harness the energy wasting potential of BAT as a weight-reducing agent, the search for alternative factors that alter the activation or recruitment state of BAT is ongoing. The goal of this thesis is to obtain a better understanding of compounds and processes that modulate UCP1-dependent thermogenesis. 

We investigate glucocorticoids for their potential to alter the UCP1-dependent thermogenic capacity of mice. We provide the novel insight that glucocorticoid supplementation reduces total BAT UCP1 protein levels, but only in mice housed at thermoneutrality. This reduction occurs at the transcriptional level by direct binding of the liganded glucocorticoid receptor to Ucp1regulatory regions. We also demonstrate that the glucocorticoid-induced reduction in BAT thermogenesis does not contribute to the development of glucocorticoid-induced obesity.

Further, we show that high-fat diet- and cafeteria diet-feeding induces the activation and recruitment of BAT UCP1 protein in the obesity-resistant 129S mouse strain. We demonstrate the importance of this diet-induced modulation of BAT thermogenic capacity by reporting an increased metabolic efficiency in UCP1-ablated mice compared to wild-type mice. 

We finally present two novel models that can be used for the identification of novel modulators of BAT thermogenesis, namely a brown adipocyte clonal cell line derived from adult human BAT, and a UCP1-luciferase reporter mouse which facilitates real-time tracking of endogenous Ucp1expression. Using these models, we identify the genes Mtus1and Kcnk3, and the compound WWL113, as novel modulators of UCP1-dependent thermogenesis. 

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University , 2019.
Keywords [en]
brown adipose tissue, UCP1, glucocorticoids, diet-induced thermogenesis, obesity, physiology
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Bioscience
Identifiers
URN: urn:nbn:se:su:diva-163376ISBN: 978-91-7797-580-9 (print)ISBN: 978-91-7797-581-6 (electronic)OAI: oai:DiVA.org:su-163376DiVA, id: diva2:1277541
Public defence
2019-03-01, Vivi Täckholmsalen (Q-salen), NPQ-huset, Svante Arrhenius Väg 20, Stockholm, 10:00 (English)
Opponent
Supervisors
Funder
The Royal Swedish Academy of Sciences
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 1: Manuscript. Paper 2: Manuscript. Paper 5: Manuscript.

Available from: 2019-02-06 Created: 2019-01-10 Last updated: 2019-01-31Bibliographically approved
List of papers
1. Glucocorticoid-induced obesity develops independently of UCP1
Open this publication in new window or tab >>Glucocorticoid-induced obesity develops independently of UCP1
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

An excess of glucocorticoids is associated with the development of obesity, as is evident from the accumulation of visceral fat in patients suffering from Cushing’s Syndrome. Activated brown adipose tissue (BAT) reduces metabolic efficiency; correspondingly an inactivation of BAT has been proposed to cause glucocorticoid-induced obesity. Here we determine the extent to which changes in BAT function as a result of glucocorticoid treatment contribute to the simultaneous development of obesity. In mice housed at 21 °C and treated with corticosterone for 2 weeks (CORT), we unexpectedly found no change in total BAT uncoupling protein 1 (UCP1) protein levels or in non-shivering thermogenic capacity. In mice housed at thermoneutrality, a humanized condition, we did observe a reduction in total UCP1 protein levels in BAT in response to CORT, which was reflected in reduced brown adipocyte cellular and mitochondrial UCP1-dependent respiration. However, glucocorticoid-induced obesity developed to the same extent in animals housed at 21 °C and 30 °C, while total BAT UCP1 protein levels differed 100-fold between the two groups. In wild-type and UCP1 knock-out mice housed at 30 °C and treated with CORT, obesity also developed to the same extent. Thus, contrary to what has previously been suggested, our results show that the development of glucocorticoid-induced obesity is unrelated to the presence of UCP1.

Keywords
uncoupling protein 1, corticosterone, obesity, thermogenesis, energy balance, brown adipose tissue, thermoneutrality, cold exposure
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Bioscience
Identifiers
urn:nbn:se:su:diva-163421 (URN)
Funder
Swedish Research Council
Available from: 2019-01-02 Created: 2019-01-02 Last updated: 2019-01-10Bibliographically approved
2. Ucp1 transcription is regulated by glucocorticoid receptor binding at Ucp1 regulatory regions
Open this publication in new window or tab >>Ucp1 transcription is regulated by glucocorticoid receptor binding at Ucp1 regulatory regions
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Glucocorticoids suppress Ucp1gene expression in brown adipocytes (BA) and brite/beige adipocytes (WA) in a cell-autonomous manner. The intracellular mechanism through which this suppression is mediated is not known. In in vitrodifferentiated primary cultures of murine brown and brite pre-adipocytes, the presence of dexamethasone (DEX) significantly reduced adrenergically-induced Ucp1mRNA levels, but upregulated the expression of other adrenergically-regulated genes. Additionally, DEX treatment increased NE-induced cAMP accumulation in BA. We thus conclude that the glucocorticoid-induced downregulation of Ucp1transcription occurs independently of a glucocorticoid-induced upregulation of the adrenergic signaling pathway. Using chromatin immunoprecipitation in combination with DNA sequencing in brown adipose tissue isolated from C57Bl/6 and BALB/c mice, we identified 6 glucocorticoid receptor binding sites in Ucp1active enhancer regions marked by H3K27ac. Luciferase reporter gene assays in the BA WT1 cell line indicated that the liganded GR binds to a regulatory region 0-4 kb upstream of the Ucp1transcription start site and thereby suppresses transcription. Thus, our study has identified a direct regulation of Ucp1transcription by the glucocorticoid receptor that occurs independently of the effects of glucocorticoids on adrenergic signaling. 

Keywords
Glucocorticoids, UCP1, transcription, adrenergic signaling, brown adipocyte, glucocorticoid response element
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:su:diva-163988 (URN)
Funder
The Royal Swedish Academy of Sciences
Available from: 2019-01-10 Created: 2019-01-10 Last updated: 2019-01-24Bibliographically approved
3. Genetic and functional characterization of clonally derived adult human brown adipocytes
Open this publication in new window or tab >>Genetic and functional characterization of clonally derived adult human brown adipocytes
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2015 (English)In: Nature Medicine, ISSN 1078-8956, E-ISSN 1546-170X, Vol. 21, no 4, p. 389-394Article in journal (Refereed) Published
Abstract [en]

Brown adipose tissue (BAT) acts in mammals as a natural defense system against hypothermia, and its activation to a state of increased energy expenditure is believed to protect against the development of obesity. Even though the existence of BAT in adult humans has been widely appreciated(1-8), its cellular origin and molecular identity remain elusive largely because of high cellular heterogeneity within various adipose tissue depots. To understand the nature of adult human brown adipocytes at single cell resolution, we isolated clonally derived adipocytes from stromal vascular fractions of adult human BAT from two individuals and globally analyzed their molecular signatures. We used RNA sequencing followed by unbiased genome-wide expression analyses and found that a population of uncoupling protein 1 (UCP1)-positive human adipocytes possessed molecular signatures resembling those of a recruitable form of thermogenic adipocytes (that is, beige adipocytes). In addition, we identified molecular markers that were highly enriched in UCP1-positive human adipocytes, a set that included potassium channel K3 (KCNK3) and mitochondrial tumor suppressor 1 (MTUS1). Further, we functionally characterized these two markers using a loss-of-function approach and found that KCNK3 and MTUS1 were required for beige adipocyte differentiation and thermogenic function. The results of this study present new opportunities for human BAT research, such as facilitating cell-based disease modeling and unbiased screens for thermogenic regulators.

National Category
Biological Sciences
Research subject
Molecular Bioscience
Identifiers
urn:nbn:se:su:diva-116982 (URN)10.1038/nm.3819 (DOI)000352493600021 ()25774848 (PubMedID)
Note

AuthorCount:13;

Available from: 2015-05-15 Created: 2015-05-05 Last updated: 2019-01-21Bibliographically approved
4. ThermoMouse: An In Vivo Model to Identify Modulators of UCP1 Expression in Brown Adipose Tissue
Open this publication in new window or tab >>ThermoMouse: An In Vivo Model to Identify Modulators of UCP1 Expression in Brown Adipose Tissue
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2014 (English)In: Cell reports, ISSN 2211-1247, E-ISSN 2211-1247, Vol. 9, no 5, p. 1584-1593Article in journal (Refereed) Published
Abstract [en]

Obesity develops when energy intake chronically exceeds energy expenditure. Because brown adipose tissue (BAT) dissipates energy in the form of heat, increasing energy expenditure by augmenting BAT-mediated thermogenesis may represent an approach to counter obesity and its complications. The ability of BAT to dissipate energy is dependent on expression of mitochondrial uncoupling protein 1 (UCP1). To facilitate the identification of pharmacological modulators of BAT UCP1 levels, which may have potential as antiobesity medications, we developed a transgenic model in which luciferase activity faithfully mimics endogenous UCP1 expression and its response to physiologic stimuli. Phenotypic screening of a library using cells derived from this model yielded a small molecule that increases UCP1 expression in brown fat cells and mice. Upon adrenergic stimulation, compound-treated mice showed increased energy expenditure. These tools offer an opportunity to identify pharmacologic modulators of UCP1 expression and uncover regulatory pathways that impact BAT-mediated thermogenesis.

Keywords
UCP1, brown adipose tissue, mouse model, screening, Luciferase
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Cellbiology
Identifiers
urn:nbn:se:su:diva-163375 (URN)10.1016/j.celrep.2014.10.066 (DOI)
Funder
Swedish Research Council
Available from: 2018-12-28 Created: 2018-12-28 Last updated: 2019-01-21Bibliographically approved
5. In the absence of UCP1-mediated diet-induced thermogenesis, obesity is augmented even in the obesity-resistant 129S mouse strain
Open this publication in new window or tab >>In the absence of UCP1-mediated diet-induced thermogenesis, obesity is augmented even in the obesity-resistant 129S mouse strain
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

The attractive tenet that recruitment and activation of brown adipose tissue (BAT) and Uncoupling Protein 1 (UCP1) could counteract the development of obesity and its comorbidities in humans has been experimentally corroborated mainly by experiments demonstrating that UCP1-ablated mice on a C57Bl/6 background (housed exempt from thermal stress) become more obese when fed a high-fat diet. However, concerns may be raised that this outcome of UCP1 ablation is restricted to this very special inbred and particularly obesity-prone mouse strain. We have therefore examined to which degree UCP1 ablation has similar metabolic effects in a mouse strain known tobe obesity resistant: the 129S strain. For this, male 129S2/sv or 129SV/Pas mice and corresponding UCP1-KO mice were fed chow, or a high-fat or a cafeteria diet for 4 weeks. The absence of UCP1 augmented obesity (weight gain, body fat mass, % body fat, fat depot size) in high-fat diet- and cafeteria-fed mice, with a similar or lower food intake, indicating that, when present, UCP1 indeed decreases metabolic efficiency. The increased obesity was due to a decrease in energy expenditure. The consumption of a high-fat or cafeteria diet increased total BAT UCP1 protein levels in wild-type mice, and correspondingly, high-fat diet and cafeteria diet-fed mice demonstrated increased norepinephrine-induced oxygen consumption. There was a positive correlation between body fat and total BAT UCP1 protein content. No evidence for diet-induced adrenergic thermogenesis was found in UCP1-ablated mice. Thus, the obesity-reducing effect of UCP1 is not restricted to a particular, and perhaps not representative, mouse strain. 

National Category
Biochemistry and Molecular Biology
Research subject
Molecular Bioscience
Identifiers
urn:nbn:se:su:diva-163422 (URN)
Funder
The Royal Swedish Academy of Sciences
Available from: 2019-01-09 Created: 2019-01-09 Last updated: 2019-01-24Bibliographically approved

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