| 1. |
- Bader, Erik, et al.
(författare)
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Identification of proliferative and mature beta-cells in the islets of Langerhans
- 2016
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 535:7612, s. 430-
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Tidskriftsartikel (refereegranskat)abstract
- Insulin-dependent diabetes is a complex multifactorial disorder characterized by loss or dysfunction of beta-cells. Pancreatic beta-cells differ in size, glucose responsiveness, insulin secretion and precursor cell potential(1-5); understanding the mechanisms that underlie this functional heterogeneity might make it possible to develop new regenerative approaches. Here we show that Fltp (also known as Flattop and Cfap126), a Wnt/planar cell polarity (PCP) effector and reporter gene(6), acts as a marker gene that subdivides endocrine cells into two subpopulations and distinguishes proliferation-competent from mature beta-cells with distinct molecular, physiological and ultrastructural features. Genetic lineage tracing revealed that endocrine subpopulations from Fltp-negative and -positive lineages react differently to physiological and pathological changes. The expression of Fltp increases when endocrine cells cluster together to form polarized and mature 3D islet mini-organs(7-9). We show that 3D architecture and Wnt/PCP ligands are sufficient to trigger beta-cell maturation. By contrast, the Wnt/PCP effector Fltp is not necessary for beta-cell development, proliferation or maturation. We conclude that 3D architecture and Wnt/PCP signalling underlie functional beta-cell heterogeneity and induce beta-cell maturation. The identification of Fltp as a marker for endocrine subpopulations sheds light on the molecular underpinnings of islet cell heterogeneity and plasticity and might enable targeting of endocrine subpopulations for the regeneration of functional beta-cell mass in diabetic patients.
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| 2. |
- Fischer, Katrin, et al.
(författare)
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The scaffold protein p62 regulates adaptive thermogenesis through ATF2 nuclear target activation
- 2020
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- During beta -adrenergic stimulation of brown adipose tissue (BAT), p38 phosphorylates the activating transcription factor 2 (ATF2) which then translocates to the nucleus to activate the expression of Ucp1 and Pgc-1 alpha. The mechanisms underlying ATF2 target activation are unknown. Here we demonstrate that p62 (Sqstm1) binds to ATF2 to orchestrate activation of the Ucp1 enhancer and Pgc-1 alpha promoter. P62(Delta 69-251) mice show reduced expression of Ucp1 and Pgc-1 alpha with impaired ATF2 genomic binding. Modulation of Ucp1 and Pgc-1 alpha expression through p62 regulation of ATF2 signaling is demonstrated in vitro and in vivo in p62(Delta 69-251) mice, global p62(-/-) and Ucp1-Cre p62(flx/flx) mice. BAT dysfunction resulting from p62 deficiency is manifest after birth and obesity subsequently develops despite normal food intake, intestinal nutrient absorption and locomotor activity. In summary, our data identify p62 as a master regulator of BAT function in that it controls the Ucp1 pathway through regulation of ATF2 genomic binding. Beta-adrenergic stimulation of brown adipose tissue leads to thermogenesis via the activating transcription factor 2 (ATF2) mediated expression of the thermogenic genes Ucp1 and Pgc-1 alpha. Here, the authors show that the scaffold protein p62 regulates brown adipose tissue function through modifying ATF2 genomic binding and subsequent Ucp1 and Pgc-1 alpha induction.
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| 3. |
- Schmidt, Sebastian, et al.
(författare)
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A reversible state of hypometabolism in a human cellular model of sporadic Parkinson's disease
- 2023
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Ingår i: Nature Communications. - 2041-1723. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Sporadic Parkinson's Disease (sPD) is a progressive neurodegenerative disorder caused by multiple genetic and environmental factors. Mitochondrial dysfunction is one contributing factor, but its role at different stages of disease progression is not fully understood. Here, we showed that neural precursor cells and dopaminergic neurons derived from induced pluripotent stem cells (hiPSCs) from sPD patients exhibited a hypometabolism. Further analysis based on transcriptomics, proteomics, and metabolomics identified the citric acid cycle, specifically the alpha-ketoglutarate dehydrogenase complex (OGDHC), as bottleneck in sPD metabolism. A follow-up study of the patients approximately 10 years after initial biopsy demonstrated a correlation between OGDHC activity in our cellular model and the disease progression. In addition, the alterations in cellular metabolism observed in our cellular model were restored by interfering with the enhanced SHH signal transduction in sPD. Thus, inhibiting overactive SHH signaling may have potential as neuroprotective therapy during early stages of sPD. Mitochondrial dysfunction is a contributing factor in Parkinson's disease. Here the authors carry out a multilayered omics analysis of Parkinson's disease patient-derived neuronal cells, which reveals a reversible hypometabolism mediated by alpha-ketoglutarate dehydrogenase deficiency, which is correlated with disease progression in the donating patients.
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| 4. |
- Ai, Jiaoyu, et al.
(författare)
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Bcl3 Couples Cancer Stem Cell Enrichment With Pancreatic Cancer Molecular Subtypes
- 2021
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Ingår i: Gastroenterology. - : Elsevier BV. - 0016-5085 .- 1528-0012. ; 161:1, s. 318-332
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Tidskriftsartikel (refereegranskat)abstract
- Background & Aims: The existence of different subtypes of pancreatic ductal adenocarcinoma (PDAC) and their correlation with patient outcome have shifted the emphasis on patient classification for better decision-making algorithms and personalized therapy. The contribution of mechanisms regulating the cancer stem cell (CSC) population in different subtypes remains unknown.Methods: Using RNA-seq, we identified B-cell CLL/lymphoma 3 (BCL3), an atypical nf-κb signaling member, as differing in pancreatic CSCs. To determine the biological consequences of BCL3 silencing in vivo and in vitro, we generated bcl3-deficient preclinical mouse models as well as murine cell lines and correlated our findings with human cell lines, PDX models, and 2 independent patient cohorts. We assessed the correlation of bcl3 expression pattern with clinical parameters and subtypes.Results: Bcl3 was significantly down-regulated in human CSCs. Recapitulating this phenotype in preclinical mouse models of PDAC via BCL3 genetic knockout enhanced tumor burden, metastasis, epithelial to mesenchymal transition, and reduced overall survival. Fluorescence-activated cell sorting analyses, together with oxygen consumption, sphere formation, and tumorigenicity assays, all indicated that BCL3 loss resulted in CSC compartment expansion promoting cellular dedifferentiation. Overexpression of BCL3 in human PDXs diminished tumor growth by significantly reducing the CSC population and promoting differentiation. Human PDACs with low BCL3 expression correlated with increased metastasis, and BCL3-negative tumors correlated with lower survival and nonclassical subtypes.Conclusions: We demonstrate that bcl3 impacts pancreatic carcinogenesis by restraining CSC expansion and by curtailing an aggressive and metastatic tumor burden in PDAC across species. Levels of BCL3 expression are a useful stratification marker for predicting subtype characterization in PDAC, thereby allowing for personalized therapeutic approaches.
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| 5. |
- Cavalieri, Riccardo, et al.
(författare)
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Activating ligands of Uncoupling protein 1 identified by rapid membrane protein thermostability shift analysis
- 2022
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Ingår i: Molecular Metabolism. - : Elsevier BV. - 2212-8778. ; 62
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Uncoupling protein 1 (UCP1) catalyses mitochondrial proton leak in brown adipose tissue to facilitate nutrient oxidation for heat production, and may combat metabolic disease if activated in humans. During the adrenergic stimulation of brown adipocytes, free fatty acids generated from lipolysis activate UCP1 via an unclear interaction. Here, we set out to characterise activator binding to purified UCP1 to clarify the activation process, discern novel activators and the potential to target UCP1.Methods: We assessed ligand binding to purified UCP1 by protein thermostability shift analysis, which unlike many conventional approaches can inform on the binding of hydrophobic ligands to membrane proteins. A detailed activator interaction analysis and screening approach was carried out, supported by investigations of UCP1 activity in liposomes, isolated brown fat mitochondria and UCP1 expression-controlled cell lines.Results: We reveal that fatty acids and other activators influence UCP1 through a specific destabilising interaction, behaving as transport substrates that shift the protein to a less stable conformation of a transport cycle. Through the detection of specific stability shifts in screens, we identify novel activators, including the over-the-counter drug ibuprofen, where ligand analysis indicates that UCP1 has a relatively wide structural specificity for interacting molecules. Ibuprofen successfully induced UCP1 activity in liposomes, isolated brown fat mitochondria and UCP1-expressing HEK293 cells but not in cultured brown adipocytes, suggesting drug delivery differs in each cell type.Conclusions: These findings clarify the nature of the activator-UCP1 interaction and demonstrate that the targeting of UCP1 in cells by approved drugs is in principle achievable as a therapeutic avenue, but requires variants with more effective delivery in brown adipocytes.
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| 6. |
- Divakaruni, Ajit S., et al.
(författare)
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A practical guide for the analysis, standardization and interpretation of oxygen consumption measurements
- 2022
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Ingår i: Nature Metabolism. - : Springer Science and Business Media LLC. - 2522-5812. ; 4:8, s. 978-994
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Tidskriftsartikel (refereegranskat)abstract
- Measurement of oxygen consumption is a powerful and uniquely informative experimental technique. It can help identify mitochondrial mechanisms of action following pharmacologic and genetic interventions, and characterize energy metabolism in physiology and disease. The conceptual and practical benefits of respirometry have made it a frontline technique to understand how mitochondrial function can interface with—and in some cases control—cell physiology. Nonetheless, an appreciation of the complexity and challenges involved with such measurements is required to avoid common experimental and analytical pitfalls. Here we provide a practical guide to oxygen consumption measurements covering the selection of experimental models and instrumentation, as well as recommendations for the collection, interpretation and normalization of data. These guidelines are provided with the intention of aiding experimental design and enhancing the overall reputability, transparency and reliability of oxygen consumption measurements.
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| 7. |
- Dludla, Phiwayinkosi V., et al.
(författare)
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Uncoupling proteins as a therapeutic target to protect the diabetic heart
- 2018
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Ingår i: Pharmacological Research. - : Elsevier BV. - 1043-6618 .- 1096-1186. ; 137, s. 11-24
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Forskningsöversikt (refereegranskat)abstract
- Myocardial remodeling and dysfunction caused by accelerated oxidative damage is a widely reported phenomenon within a diabetic state. Altered myocardial substrate preference appears to be the major cause of enhanced oxidative stress-mediated cell injury within a diabetic heart. During this process, exacerbated free fatty acid flux causes an abnormal increase in mitochondrial membrane potential leading to the overproduction of free radical species and subsequent cell damage. Uncoupling proteins (UCPs) are expressed within the myocardium and can protect against free radical damage by modulating mitochondrial respiration, leading to reduced production of reactive oxygen species. Moreover, transgenic animals lacking UCPs have been shown to be more susceptible to oxidative damage and display reduced cardiac function when compared to wild type animals. This suggests that tight regulation of UCPs is necessary for normal cardiac function and in the prevention of diabetes-induced oxidative damage. This review aims to enhance our understanding of the pathophysiological mechanisms relating to the role of UCPs in a diabetic heart, and further discuss known pharmacological compounds and hormones that can protect a diabetic heart through the modulation of UCPs.
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| 8. |
- Elhag, Sara, et al.
(författare)
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Differences in Cell-Intrinsic Inflammatory Programs of Yolk Sac and Bone Marrow Macrophages
- 2021
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Ingår i: Cells. - : MDPI AG. - 2073-4409. ; 10:12
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Tidskriftsartikel (refereegranskat)abstract
- Background: Tissue-resident macrophages have mixed developmental origins. They derive in variable extent from yolk sac (YS) hematopoiesis during embryonic development. Bone marrow (BM) hematopoietic progenitors give rise to tissue macrophages in postnatal life, and their contribution increases upon organ injury. Since the phenotype and functions of macrophages are modulated by the tissue of residence, the impact of their origin and developmental paths has remained incompletely understood. Methods: In order to decipher cell-intrinsic macrophage programs, we immortalized hematopoietic progenitors from YS and BM using conditional HoxB8, and carried out an in-depth functional and molecular analysis of differentiated macrophages. Results: While YS and BM macrophages demonstrate close similarities in terms of cellular growth, differentiation, cell death susceptibility and phagocytic properties, they display differences in cell metabolism, expression of inflammatory markers and inflammasome activation. Reduced abundance of PYCARD (ASC) and CASPASE-1 proteins in YS macrophages abrogated interleukin-1 beta production in response to canonical and non-canonical inflammasome activation. Conclusions: Macrophage ontogeny is associated with distinct cellular programs and immune response. Our findings contribute to the understanding of the regulation and programming of macrophage functions.
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| 9. |
- Fischer, Katrin, et al.
(författare)
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Alternatively activated macrophages do not synthesize catecholamines or contribute to adipose tissue adaptive thermogenesis
- 2017
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Ingår i: Nature Medicine. - : Springer Science and Business Media LLC. - 1078-8956 .- 1546-170X. ; 23:5, s. 623-630
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Tidskriftsartikel (refereegranskat)abstract
- Adaptive thermogenesis is the process of heat generation in response to cold stimulation. It is under the control of the sympathetic nervous system, whose chief effector is the catecholamine norepinephrine (NE). NE enhances thermogenesis through beta 3-adrenergic receptors to activate brown adipose tissue and by 'browning' white adipose tissue. Recent studies have reported that alternative activation of macrophages in response to interleukin (IL)-4 stimulation induces the expression of tyrosine hydroxylase (TH), a key enzyme in the catecholamine synthesis pathway, and that this activation provides an alternative source of locally produced catecholamines during the thermogenic process. Here we report that the deletion of Th in hematopoietic cells of adult mice neither alters energy expenditure upon cold exposure nor reduces browning in inguinal adipose tissue. Bone marrow-derived macrophages did not release NE in response to stimulation with IL-4, and conditioned media from IL-4-stimulated macrophages failed to induce expression of thermogenic genes, such as uncoupling protein 1 (Ucp1), in adipocytes cultured with the conditioned media. Furthermore, chronic treatment with IL-4 failed to increase energy expenditure in wild-type, Ucp1(-/-) and interleukin-4 receptor-alpha double-negative (Il4ra(-/-)) mice. In agreement with these findings, adipose-tissue-resident macrophages did not express TH. Thus, we conclude that alternatively activated macrophages do not synthesize relevant amounts of catecholamines, and hence, are not likely to have a direct role in adipocyte metabolism or adaptive thermogenesis.
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| 10. |
- Fyda, Thomas Jacob, et al.
(författare)
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Disruption of thermogenic UCP1 predated the divergence of pigs and peccaries
- 2020
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Ingår i: Journal of Experimental Biology. - : The Company of Biologists. - 0022-0949 .- 1477-9145. ; 223:15
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Tidskriftsartikel (refereegranskat)abstract
- Uncoupling protein 1 (UCP1) governs non-shivering thermogenesis in brown adipose tissue. It has been estimated that pigs lost UCP1 similar to 20 million years ago (MYA), dictating cold intolerance among piglets. Our current understanding of the root causes of UCP1 loss are, however, incomplete. Thus, examination of additional species can shed light on these fundamental evolutionary questions. Here, we investigated UCP1 in the Chacoan peccary (Catagonus wagneri), a member of the Tayassuid lineage that diverged from pigs during the late Eocene-mid Oligocene. Exons 1 and 2 have been deleted in peccary UCP1 and the remaining exons display additional inactivating mutations. A common nonsense mutation in exon 6 revealed that UCP1 was pseudogenized in a shared ancestor of pigs and peccaries. Our selection pressure analyses indicate that the inactivation occurred 36.2-44.3 MYA during the mid-late Eocene, which is much earlier than previously thought. Importantly, pseudogenized UCP1 provides the molecular rationale for cold sensitivity and current tropical biogeography of extant peccaries.
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| 11. |
- Gaudry, Michael James, 1990-
(författare)
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Comparative analysis of the thermogenic protein UCP1 across the mammalian phylogeny
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Uncoupling protein 1 (UCP1) drives mitochondrial inefficiency to produce heat in mammalian brown adipose tissue (BAT). Many mammalian species rely on this form of adaptive non-shivering thermogenesis (NST) to defend high body temperatures in the cold. Little is known regarding how evolution may have shaped UCP1 function to reflect distinct thermoregulatory requirements of various lineages. This thesis merges genetic and functional data while using a comparative approach to gain insights into the evolutionary rise of thermogenic UCP1, as well as its repeated loss of function among several eutherian lineages. UCP1 structure-function relationships and mechanistic insights are gained by examining natural mutations among the orthologues of different species. In Paper I, we reveal that, like eutherian mammals, marsupial UCP1 is expressed in adipose tissue of developing young gray short-tailed opossums (Monodelphis domestica) and coincides with the onset of thermoregulatory competence. Transcriptomic analyses reveal partial browning signatures in adipose tissue of young opossums, resembling eutherian beige adipose tissue. Overexpression of marsupial UCP1 in a mammalian test system (HEK293 cells), however, reveals its lack of thermogenic functionality. I then performed ancestral reconstruction of UCP1 and demonstrate that the thermogenic function arose in the stem eutherian ancestor. In Paper II, I uncover that UCP1 not only became pseudogenized in pigs (e.g. Sus spp.), but in a common ancestor of both pigs and peccaries (e.g. Catagonus wagneri) as indicated from a shared inactivating mutation, re-calibrating the timeline of this inactivation and our understanding of how it may limit the geographic distribution of modern peccaries. In Paper III, I uncover a novel UCP1 pseudogene unique to the largest seals, elephant seals (Mirounga spp.), showing that UCP1 is retained within most members of the seal lineage for neonatal defense of body temperatures, but its loss coincides with the extreme body sizes attained by elephant seals. In Paper IV, we functionally verify that N-terminal truncation or frameshift mutation repair cannot rescue the thermogenic function of elephant seal UCP1. By contrast, we verify the thermogenic capacity of UCP1 of the small-bodied harbor seal (Phoca vitulina), matching that of terrestrial eutherians.In Paper V, we examine UCP1 in the naked mole-rat (Heterocephalus glaber), a species that displays a natural mutation to the histidine pair motif that has been previously deemed crucial for UCP1 function. We hypothesized that this may underlie the poor thermoregulatory abilities of the species. Our assessment of UCP1 mutants, however, reveal that the naked mole-rat retains UCP1 function and that the histidine pair motif is unnecessary for the GDP-sensitive thermogenic function of the protein, providing important structure-function information of UCP1 and questioning a proposed mechanistic model. In summary, this thesis utilizes UCP1 as a biomarker to trace the evolution of mammalian NST and thermoregulation. Insights gained provide clues to the various factors influencing mammalian endothermy and hints of structure-function relationships in this thermogenic protein.
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| 12. |
- Gaudry, Michael J., et al.
(författare)
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Comparative functional analyses of UCP1 to unravel evolution, ecophysiology and mechanisms of mammalian thermogenesis
- 2021
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Ingår i: Comparative Biochemistry and Physiology - Part B. - : Elsevier BV. - 1096-4959 .- 1879-1107. ; 255
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Tidskriftsartikel (refereegranskat)abstract
- Brown adipose tissue (BAT), present in many placental mammals, provides adaptive nonshivering thermogenesis (NST) for body temperature regulation and has facilitated survival in diverse thermal niches on our planet. Intriguingly, several key details on the molecular mechanisms of NST and their potential ecophysiological adaptations are still unknown. Comparative studies at the whole animal level are unpragmatic, due to the diversity and complexity of thermoregulation among different species. We propose that the molecular evolution of mitochondrial uncoupling protein 1 (UCP1), a central component for BAT thermogenesis, represents a powerful opportunity to unravel key questions of mammalian thermoregulation. Comparative analysis of UCP1 may elucidate how its thermogenic function arose, how environmental selection has shaped protein function to support ecophysiological requirements, and how the enigmatic molecular mechanism of proton leak is governed. Several approaches for the assessment of UCP1 function in vitro have been introduced over the years. For comparative characterization of UCP1, we put forward the overexpression of UCP1 orthologues and mutated variants in a mammalian cell system as a primary strategy and discuss advantageous aspects in contrast to other experimental systems. In turn, we suggest how remaining experimental caveats can be solved by complimentary test systems before physiological consolidation in the animal model. Furthermore, we highlight the appropriate bioenergetic techniques to perform the functional analyses on UCP1. The comparative characterizations of diverse UCP1 variants may enable key insights into open questions surrounding the molecular basis of NST.
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| 13. |
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| 14. |
- Gaudry, Michael J., et al.
(författare)
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Molecular evolution of thermogenic uncoupling protein 1 and implications for medical intervention of human disease
- 2019
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Ingår i: Molecular Aspects of Medicine. - : Elsevier BV. - 0098-2997 .- 1872-9452. ; 68, s. 6-17
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Forskningsöversikt (refereegranskat)abstract
- In eutherian mammals, brown adipose tissue (BAT) permits non-shivering thermogenesis (NST) through high metabolic rates catalyzed by the unique mitochondrial uncoupling protein 1 (UCP1). The tissue has recently gained remarkable attention due to its discovery in adult humans. Approaching BAT and UCP1 as therapeutic targets to combust surplus energy bares high potential to combat the epidemic of the metabolic syndrome that has precipitated in our society as a result of our modern lifestyles. Our understanding of the physiological and molecular control of BAT may benefit tremendously from consideration of its evolution that basically outlines the blueprint of how to construct a fat burning tissue. Here, we discuss the evolutionary history of UCP1 and BAT, from its origins and emergence to its downfall in several mammalian lineages. Additionally, we delineate the annotation of UCPs in vertebrates by analyzing genomic organization and summarize the phylogeny of UCP1 within the closest relatives of humans, the great apes. Outlining whether the molecular networks controlling thermogenesis in adipose tissue (commonly known as the browning potential) pre-dated the classical thermogenic function of BAT and UCP1, and whether the evolutionary inactivation of UCP1 enhanced compensatory thermogenic mechanisms, should be of major interest to those who aim to access adipose tissue thermogenesis in a biomedical context.
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| 15. |
- Gaudry, Michael J., et al.
(författare)
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Molecular evolution of uncoupling proteins and implications for brain function
- 2019
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Ingår i: Neuroscience Letters. - : Elsevier BV. - 0304-3940 .- 1872-7972. ; 696, s. 140-145
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Forskningsöversikt (refereegranskat)abstract
- Uncoupling proteins (UCPs) belong to the mitochondrial anion carrier superfamily and catalyze important metabolic functions at the mitochondrial inner membrane. While the thermogenic role of UCP1 in brown fat of eutherian mammals is well established, the molecular functions of UCP1 in ectothermic vertebrates and of other UCP paralogs remain less clear. Here, we critically discuss the existence of brain UCPs and their potential roles. Applying phylogenetic classification of novel UCPs, we summarize the evidence for brain UCP1 among vertebrates, the role of UCP2 in specific brain areas, and the existence of brain-specific UCPs. The phylogenetic analyses and discussion on functional data should alert the scientific community that the molecular function of so-called UCP1 homologues is by far not clarified and possibly relates to neither thermogenesis nor mitochondrial uncoupling.
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| 16. |
- Gaudry, Michael J., 1990-, et al.
(författare)
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Terrestrial Birth and Body Size Tune UCP1 Functionality in Seals
- 2024
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Ingår i: Molecular biology and evolution. - 0737-4038 .- 1537-1719. ; 41:4
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Tidskriftsartikel (refereegranskat)abstract
- The molecular evolution of the mammalian heater protein UCP1 is a powerful biomarker to understand thermoregulatory strategies during species radiation into extreme climates, such as aquatic life with high thermal conductivity. While fully aquatic mammals lost UCP1, most semiaquatic seals display intact UCP1 genes, apart from large elephant seals. Here, we show that UCP1 thermogenic activity of the small-bodied harbor seal is equally potent compared to terrestrial orthologs, emphasizing its importance for neonatal survival on land. In contrast, elephant seal UCP1 does not display thermogenic activity, not even when translating a repaired or a recently highlighted truncated version. Thus, the thermogenic benefits for neonatal survival during terrestrial birth in semiaquatic pinnipeds maintained evolutionary selection pressure on UCP1 function and were only outweighed by extreme body sizes among elephant seals, fully eliminating UCP1-dependent thermogenesis.
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| 17. |
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| 18. |
- Grandoch, Maria, et al.
(författare)
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4-Methylumbelliferone improves the thermogenic capacity of brown adipose tissue
- 2019
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Ingår i: Nature Metabolism. - : Springer Science and Business Media LLC. - 2522-5812. ; 1:5, s. 546-559
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Tidskriftsartikel (refereegranskat)abstract
- Therapeutic increase in brown adipose tissue (BAT) thermogenesis is of great interest, as BAT activation counteracts obesity and insulin resistance. Hyaluronan (HA) is a glycosaminoglycan, found in the extracellular matrix, that is synthesized by HA synthases (HAS1, HAS2, and HAS3) from sugar precursors and accumulates in diabetic conditions. Its synthesis can be inhibited by the small molecule 4-methylumbelliferone (4-MU). Here we show that inhibition of HA synthesis by 4-MU or genetic deletion of Has2 and Has3 improves the thermogenic capacity of BAT, reduces body-weight gain, and improves glucose homeostasis independently of adrenergic stimulation in mice on a diabetogenic diet. In this context, we validated a novel magnetic resonce T2 mapping approach for in vivo visualization of BAT activation. Inhibition of HA synthesis increases glycolysis, BAT respiration, and uncoupling protein 1 (UCP1) expression. In addition, we show that 4-MU increases BAT capacity without inducing chronic stimulation and propose that 4-MU, a clinically approved, prescription-free drug, could be repurposed to treat obesity and diabetes.
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| 19. |
- Herrnhold, Marie, et al.
(författare)
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Adverse bioenergetic effects of N-acyl amino acids in human adipocytes overshadow beneficial mitochondrial uncoupling
- 2023
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Ingår i: Redox Biology. - 2213-2317. ; 66
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Enhancing energy turnover via uncoupled mitochondrial respiration in adipose tissue has great potential to improve human obesity and other metabolic complications. However, the amount of human brown adipose tissue and its uncoupling protein 1 (UCP1) is low in obese patients. Recently, a class of endogenous molecules, N-acyl amino acids (NAAs), was identified as mitochondrial uncouplers in murine adipocytes, presumably acting via the adenine nucleotide translocator (ANT). Given the translational potential, we investigated the bioenergetic effects of NAAs in human adipocytes, characterizing beneficial and adverse effects, dose ranges, amino acid derivatives and underlying mechanisms.Method: NAAs with neutral (phenylalanine, leucine, isoleucine) and polar (lysine) residues were synthetized and assessed in intact and permeabilized human adipocytes using plate-based respirometry. The Seahorse technology was applied to measure bioenergetic parameters, dose-dependency, interference with UCP1 and adenine nucleotide translocase (ANT) activity, as well as differences to the established chemical uncouplers niclosamide ethanolamine (NEN) and 2,4-dinitrophenol (DNP).Result: NAAs with neutral amino acid residues potently induce uncoupled respiration in human adipocytes in a dose-dependent manner, even in the presence of the UCP1-inhibitor guanosine diphosphate (GDP) and the ANT-inhibitor carboxyatractylate (CAT). However, neutral NAAs significantly reduce maximal oxidation rates, mitochondrial ATP-production, coupling efficiency and reduce adipocyte viability at concentrations above 25 μM. The in vitro therapeutic index (using induced proton leak and viability as determinants) of NAAs is lower than that of NEN and DNP.Conclusion: NAAs are potent mitochondrial uncouplers in human adipocytes, independent of UCP1 and ANT. However, previously unnoticed adverse effects harm adipocyte functionality, reduce the therapeutic index of NAAs in vitro and therefore question their suitability as anti-obesity agents without further chemical modifications.
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| 20. |
- Holthaus, Lisa, et al.
(författare)
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CD4(+) T cell activation, function, and metabolism are inhibited by low concentrations of DMSO
- 2018
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Ingår i: JIM - Journal of Immunological Methods. - : Elsevier BV. - 0022-1759 .- 1872-7905. ; 463, s. 54-60
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Tidskriftsartikel (refereegranskat)abstract
- Dimethyl sulfoxide (DMSO) is a polar organic solvent used in a wide range of biological applications. DMSO is routinely used as a cryoprotectant for long-term cell freezing as well as to dissolve peptides and drugs for immune cell functional assays. Here, human CD4(+) T cell activation, cytokine production, proliferation, and metabolism were investigated after stimulation in the presence of 0.01% to 1%, DMSO, representing concentrations commonly used in vitro. Surface expression of the activation markers CD69, CD25 and CD154 after polyclonal activation of CD4(+) T cells was inhibited by 0.25% or higher concentrations of DMSO. The frequencies of IL-21(+), IL-4(+), and IL-22(+) CD4(+) T cells, following polyclonal activation were variably inhibited by DMSO at concentrations ranging from 0.25% to 1%, whereas IFN gamma(+) cells were unaffected. CD4(+) T cell proliferation after anti-CD3 or antigen stimulation was inhibited by 0.5% DMSO and abolished by 1% DMSO. After polyclonal stimulation, glucose uptake was inhibited in the presence of 1% DMSO, but only minor effects on CD4+ T cell respiration were observed. Consistent with the immune effects, the gene expression of early signaling and activation pathways were inhibited in CD4+ T cells in the presence of 1% DMSO. Our study revealed that DMSO at concentrations generally used for in vitro studies of T cells impacts multiple features of T cell function. Therefore, we urge care when adding DMSO-containing preparations to T cell cultures.
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| 21. |
- Hytönen, Jarkko, et al.
(författare)
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Activation of Peroxisome Proliferator-Activated Receptor-δ as Novel Therapeutic Strategy to Prevent In-Stent Restenosis and Stent Thrombosis
- 2016
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Ingår i: Arteriosclerosis, Thrombosis and Vascular Biology. - 1079-5642 .- 1524-4636. ; 36:8, s. 1534-1548
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Tidskriftsartikel (refereegranskat)abstract
- Objective Drug-eluting coronary stents reduce restenosis rate and late lumen loss compared with bare-metal stents; however, drug-eluting coronary stents may delay vascular healing and increase late stent thrombosis. The peroxisome proliferator-activated receptor-delta (PPAR) exhibits actions that could favorably influence outcomes after drug-eluting coronary stents placement. Approach and Results Here, we report that PPAR ligand-coated stents strongly reduce the development of neointima and luminal narrowing in a rabbit model of experimental atherosclerosis. Inhibition of inflammatory gene expression and vascular smooth muscle cell (VSMC) proliferation and migration, prevention of thrombocyte activation and aggregation, and proproliferative effects on endothelial cells were identified as key mechanisms for the prevention of restenosis. Using normal and PPAR-depleted VSMCs, we show that the observed effects of PPAR ligand GW0742 on VSMCs and thrombocytes are PPAR receptor dependent. PPAR ligand treatment induces expression of pyruvate dehydrogenase kinase isozyme 4 and downregulates the glucose transporter 1 in VSMCs, thus impairing the ability of VSMCs to provide the increased energy demands required for growth factor-stimulated proliferation and migration. Conclusions In contrast to commonly used drugs for stent coating, PPAR ligands not only inhibit inflammatory response and proliferation of VSMCs but also prevent thrombocyte activation and support vessel re-endothelialization. Thus, pharmacological PPAR activation could be a promising novel strategy to improve drug-eluting coronary stents outcomes.
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| 22. |
- Jastroch, Martin, et al.
(författare)
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Importance of adipocyte browning in the evolution of endothermy
- 2020
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Ingår i: Philosophical Transactions of the Royal Society of London. Biological Sciences. - : The Royal Society. - 0962-8436 .- 1471-2970. ; 375:1793
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Forskningsöversikt (refereegranskat)abstract
- Endothermy changes the relationship between organisms and their environment fundamentally, and it is therefore of major ecological and evolutionary significance. Endothermy is characterized by non-shivering thermogenesis, that is metabolic heat production in the absence of muscular activity. In many eutherian mammals, brown adipose tissue (BAT) is an evolutionary innovation that facilitates non-shivering heat production in mitochondria by uncoupling food-derived substrate oxidation from chemical energy (ATP) production. Consequently, energy turnover is accelerated resulting in increased heat release. The defining characteristics of BAT are high contents of mitochondria and vascularization, and the presence of uncoupling protein 1. Recent insights, however, reveal that a range of stimuli such as exercise, diet and the immune system can cause the browning of white adipocytes, thereby increasing energy expenditure and heat production even in the absence of BAT. Here, we review the molecular mechanisms that cause browning of white adipose tissue, and their potential contribution to thermoregulation. The significance for palaeophysiology lies in the presence of adipose tissue and the mechanisms that cause its browning and uncoupling in all amniotes. Hence, adipocytes may have played a role in the evolution of endothermy beyond the more specific evolution of BAT in eutherians. This article is part of the theme issue 'Vertebrate palaeophysiology'.
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| 23. |
- Jastroch, Martin, et al.
(författare)
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Pros and cons for the evidence of adaptive non-shivering thermogenesis in marsupials
- 2021
-
Ingår i: Journal of Comparative Physiology. B, Biochemical, Systemic, and Environmental Physiology. - : Springer Science and Business Media LLC. - 0174-1578 .- 1432-136X. ; 191:6, s. 1085-1095
-
Forskningsöversikt (refereegranskat)abstract
- The thermogenic mechanisms supporting endothermy are still not fully understood in all major mammalian subgroups. In placental mammals, brown adipose tissue currently represents the most accepted source of adaptive non-shivering thermogenesis. Its mitochondrial protein UCP1 (uncoupling protein 1) catalyzes heat production, but the conservation of this mechanism is unclear in non-placental mammals and lost in some placentals. Here, we review the evidence for and against adaptive non-shivering thermogenesis in marsupials, which diverged from placentals about 120-160 million years ago. We critically discuss potential mechanisms that may be involved in the heat-generating process among marsupials.
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| 24. |
- Jastroch, Martin, et al.
(författare)
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When pigs fly, UCP1 makes heat
- 2015
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Ingår i: MOLECULAR METABOLISM. - : Elsevier BV. - 2212-8778. ; 4:5, s. 359-362
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Brown and beige adipose tissue may represent important therapeutic targets for the treatment of diabetes and obesity as these organs dissipate nutrient energy as heat through the thermogenic uncoupling protein 1 (UCP1). While mice are commonly used to mimic the potential effects of brown/beige adipose tissue that may act in human metabolism, new animal models are edging into the market for translational medicine. Pigs reflect human metabolism better than mice in multiple parameters such as obesity-induced hyperglycemia, cholesterol profiles and energy metabolism. Recently, it was reported that energy expenditure and body temperature in pigs is induced by the hormone leptin, and that leptin's action is mediated by UCP1 in adipose tissue. Given the tremendous importance of identifying molecular mechanisms for targeting therapeutics, we critically examine the evidence supporting the presence of UCP1 in pigs and conclude that methodological shortcomings prevent an unequivocal claim for the presence of UCP1 in pigs. Despite this, we believe that leptin's effects on energy expenditure in pigs are potentially more transformative to human medicine in the absence of UCP1, as adult and obese humans possess only minor amounts of UCP1. In general, we propose that the biology of new animal models requires attention to comparative studies with humans given the increasing amount of genomic information for various animal species.
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| 25. |
- Kabra, Uma D, et al.
(författare)
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Direct Substrate Delivery into Mitochondrial-Fission Deficient Pancreatic Islets Rescues Insulin Secretion
- 2017
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 66:5, s. 1247-1257
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Tidskriftsartikel (refereegranskat)abstract
- In pancreatic beta cells, mitochondrial bioenergetics control glucose-stimulated insulin secretion (GSIS). Mitochondrial dynamics are generally associated with quality control, maintaining the functionality of bioenergetics. By acute pharmacological inhibition of mitochondrial fission protein Drp1, we here demonstrate that mitochondrial fission is necessary for GSIS in mouse and human islets. We confirm that genetic silencing of Drp1 increases mitochondrial proton leak in MIN6 cells. However, our comprehensive analysis of pancreatic islet bioenergetics reveals that Drp1 does not control insulin secretion via its effect on proton leak but instead via modulation of glucose-fuelled respiration. Notably, pyruvate fully rescues the impaired insulin secretion of fission-deficient beta cells, demonstrating that defective mitochondrial dynamics solely impact substrate supply upstream of oxidative phosphorylation. The present findings provide novel insights in how mitochondrial dysfunction may cause pancreatic beta cell failure. In addition, the results will stimulate new thinking in the intersecting fields of mitochondrial dynamics and bioenergetics, as treatment of defective dynamics in mitochondrial diseases appears to be possible by improving metabolism upstream of mitochondria.
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| 26. |
- Kabra, Uma D., et al.
(författare)
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Drp1 Overexpression Decreases Insulin Content in Pancreatic MIN6 Cells
- 2022
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Ingår i: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 23:20
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Tidskriftsartikel (refereegranskat)abstract
- Mitochondrial dynamics and bioenergetics are central to glucose-stimulated insulin secretion by pancreatic beta cells. Previously, we demonstrated that a disturbance in glucose-invoked fission impairs insulin secretion by compromising glucose catabolism. Here, we investigated whether the overexpression of mitochondrial fission regulator Drp1 in MIN6 cells can improve or rescue insulin secretion. Although Drp1 overexpression slightly improves the triggering mechanism of insulin secretion of the Drp1-knockdown cells and has no adverse effects on mitochondrial metabolism in wildtype MIN6 cells, the constitutive presence of Drp1 unexpectedly impairs insulin content, which leads to a reduction in the absolute values of secreted insulin. Coherent with previous studies in Drp1-overexpressing muscle cells, we found that the upregulation of ER stress-related genes (BiP, Chop, and Hsp60) possibly impacts insulin production in MIN6 cells. Collectively, we confirm the important role of Drp1 for the energy-coupling of insulin secretion but unravel off-targets effects by Drp1 overexpression on insulin content that warrant caution when manipulating Drp1 in disease therapy.
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| 27. |
- Kabra, Uma D., et al.
(författare)
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Mitochondrial Dynamics and Insulin Secretion
- 2023
-
Ingår i: International Journal of Molecular Sciences. - 1661-6596 .- 1422-0067. ; 24:18
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Forskningsöversikt (refereegranskat)abstract
- Mitochondria are involved in the regulation of cellular energy metabolism, calcium homeostasis, and apoptosis. For mitochondrial quality control, dynamic processes, such as mitochondrial fission and fusion, are necessary to maintain shape and function. Disturbances of mitochondrial dynamics lead to dysfunctional mitochondria, which contribute to the development and progression of numerous diseases, including Type 2 Diabetes (T2D). Compelling evidence has been put forward that mitochondrial dynamics play a significant role in the metabolism-secretion coupling of pancreatic β cells. The disruption of mitochondrial dynamics is linked to defects in energy production and increased apoptosis, ultimately impairing insulin secretion and β cell death. This review provides an overview of molecular mechanisms controlling mitochondrial dynamics, their dysfunction in pancreatic β cells, and pharmaceutical agents targeting mitochondrial dynamic proteins, such as mitochondrial division inhibitor-1 (mdivi-1), dynasore, P110, and 15-oxospiramilactone (S3).
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| 28. |
- Kabra, Uma D., et al.
(författare)
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Respiratory Parameters for the Classification of Dysfunctional Insulin Secretion by Pancreatic Islets
- 2021
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Ingår i: Metabolites. - : MDPI AG. - 2218-1989. ; 11:6
-
Tidskriftsartikel (refereegranskat)abstract
- The development of obesity and type 2 diabetes (T2D) has been associated with impaired mitochondrial function. In pancreatic beta (beta) cells, mitochondrial energy metabolism plays a central role in triggering and controlling glucose-stimulated insulin secretion (GSIS). Here, we have explored whether mitochondrial bioenergetic parameters assessed with Seahorse extracellular flux technology can quantitatively predict insulin secretion. We metabolically stressed male C57BL/6 mice by high-fat feeding (HFD) and measured the glucose sensitivity of islet respiration and insulin secretion. The diet-induced obese (DIO) mice developed hyperinsulinemia, but no pathological secretory differences were apparent between isolated DIO and chow islets. Real-time extracellular flux analysis, however, revealed a lower respiratory sensitivity to glucose in DIO islets. Correlation of insulin secretion with respiratory parameters uncovers compromised insulin secretion in DIO islets by oxidative power. Normalization to increased insulin contents during DIO improves the quantitative relation between GSIS and respiration, allowing to classify dysfunctional properties of pancreatic insulin secretion, and thereby serving as valuable biomarker for pancreatic islet glucose responsiveness and health.
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| 29. |
- Keipert, Susanne, et al.
(författare)
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Endogenous FGF21-signaling controls paradoxical obesity resistance of UCP1-deficient mice
- 2020
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- Brown adipose thermogenesis increases energy expenditure and relies on uncoupling protein 1 (UCP1), however, UCP1 knock-out mice show resistance to diet-induced obesity at room temperature. Here, the authors show that this resistance relies on FGF21-signaling, inducing the browning of white adipose tissue. Uncoupling protein 1 (UCP1) executes thermogenesis in brown adipose tissue, which is a major focus of human obesity research. Although the UCP1-knockout (UCP1 KO) mouse represents the most frequently applied animal model to judge the anti-obesity effects of UCP1, the assessment is confounded by unknown anti-obesity factors causing paradoxical obesity resistance below thermoneutral temperatures. Here we identify the enigmatic factor as endogenous FGF21, which is primarily mediating obesity resistance. The generation of UCP1/FGF21 double-knockout mice (dKO) fully reverses obesity resistance. Within mild differences in energy metabolism, urine metabolomics uncover increased secretion of acyl-carnitines in UCP1 KOs, suggesting metabolic reprogramming. Strikingly, transcriptomics of metabolically important organs reveal enhanced lipid and oxidative metabolism in specifically white adipose tissue that is fully reversed in dKO mice. Collectively, this study characterizes the effects of endogenous FGF21 that acts as master regulator to protect from diet-induced obesity in the absence of UCP1.
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| 30. |
- Keipert, Susanne, 1980-, et al.
(författare)
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Two-stage evolution of mammalian adipose tissue thermogenesis
- 2024
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Ingår i: Science. - 0036-8075 .- 1095-9203. ; 384:6700, s. 1111-1117
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Tidskriftsartikel (refereegranskat)abstract
- Brown adipose tissue (BAT) is a heater organ that expresses thermogenic uncoupling protein 1 (UCP1) to maintain high body temperatures during cold stress. BAT thermogenesis is considered an overarching mammalian trait, but its evolutionary origin is unknown. We show that adipose tissue of marsupials, which diverged from eutherian mammals ~150 million years ago, expresses a nonthermogenic UCP1 variant governed by a partial transcriptomic BAT signature similar to that found in eutherian beige adipose tissue. We found that the reconstructed UCP1 sequence of the common eutherian ancestor displayed typical thermogenic activity, whereas therian ancestor UCP1 is nonthermogenic. Thus, mammalian adipose tissue thermogenesis may have evolved in two distinct stages, with a prethermogenic stage in the common therian ancestor linking UCP1 expression to adipose tissue and thermal stress. We propose that in a second stage, UCP1 acquired its thermogenic function specifically in eutherians, such that the onset of mammalian BAT thermogenesis occurred only after the divergence from marsupials.
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| 31. |
- Keuper, Michaela, et al.
(författare)
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The good and the BAT of metabolic sex differences in thermogenic human adipose tissue
- 2021
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Ingår i: Molecular and Cellular Endocrinology. - : Elsevier BV. - 0303-7207 .- 1872-8057. ; 533
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Tidskriftsartikel (refereegranskat)abstract
- Thermogenic adipose tissue, which comprises classical brown and beige adipose tissue, has the ability to improve systemic metabolism. Its identification in adult humans has fostered extensive investigations on the therapeutic value to counteract obesity and metabolic disorders. Sex and gender differences of human thermogenic adipose tissue, however, are still understudied despite their importance for personalized treatment options. Here, we review studies reporting human sex differences of thermogenic adipose tissue and related potential improvements of systemic energy metabolism. An increasing body of evidence suggests higher prevalence, mass and activity of thermogenic adipose tissue in women, but the consequences for metabolic disease progression and mechanisms are largely unknown. Therefore, we also discuss observations on sex-specific adipose metabolism in experimental mouse and rat studies that may assist to establish molecular mechanisms and instruct future investigations in humans.
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| 32. |
- Khani, Sajjad, et al.
(författare)
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Cold-induced expression of a truncated adenylyl cyclase 3 acts as rheostat to brown fat function
- 2024
-
Ingår i: Nature Metabolism. - 2522-5812. ; 6:6, s. 1053-1075
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Tidskriftsartikel (refereegranskat)abstract
- Promoting brown adipose tissue (BAT) activity innovatively targets obesity and metabolic disease. While thermogenic activation of BAT is well understood, the rheostatic regulation of BAT to avoid excessive energy dissipation remains ill-defined. Here, we demonstrate that adenylyl cyclase 3 (AC3) is key for BAT function. We identified a cold-inducible promoter that generates a 5′ truncated AC3 mRNA isoform (Adcy3-at), whose expression is driven by a cold-induced, truncated isoform of PPARGC1A (PPARGC1A-AT). Male mice lacking Adcy3-at display increased energy expenditure and are resistant to obesity and ensuing metabolic imbalances. Mouse and human AC3-AT are retained in the endoplasmic reticulum, unable to translocate to the plasma membrane and lack enzymatic activity. AC3-AT interacts with AC3 and sequesters it in the endoplasmic reticulum, reducing the pool of adenylyl cyclases available for G-protein-mediated cAMP synthesis. Thus, AC3-AT acts as a cold-induced rheostat in BAT, limiting adverse consequences of cAMP activity during chronic BAT activation.
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| 33. |
- Liskiewicz, D., et al.
(författare)
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Neuronal loss of TRPM8 leads to obesity and glucose intolerance in male mice
- 2023
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Ingår i: Molecular Metabolism. - : Elsevier BV. - 2212-8778. ; 72
-
Tidskriftsartikel (refereegranskat)abstract
- Objective: Mice with global deletion of the transient receptor potential channel melastatin family member 8 (TRPM8) are obese, and treatment of diet-induced obese (DIO) mice with TRPM8 agonists decrease body weight. Whether TRPM8 signaling regulates energy metabolism via central or peripheral effects is unknow. Here we assessed the metabolic phenotype of mice with either Nestin Cre-mediated neuronal loss of TRPM8, or with deletion of TRPM8 in Advillin Cre positive sensory neurons of the peripheral nervous system (PNS).Methods: Nestin Cre- and Advillin Cre-Trpm8 knock-out (KO) mice were metabolically phenotyped under chronic exposure to either chow or high-fat diet (HFD), followed by assessment of energy and glucose metabolism.Results: At room temperature, chow-fed neuronal Trpm8 KO are obese and show decreased energy expenditure when acutely treated with the TRPM8 selective agonist icilin. But body weight of neuronal Trpm8 KO mice is indistinguishable from wildtype controls at thermoneutrality, or when mice are chronically exposed to HFD-feeding. In contrast to previous studies, we show that the TRPM8 agonist icilin has no direct effect on brown adipocytes, but that icilin stimulates energy expenditure, at least in part, via neuronal TRPM8 signaling. We further show that lack of TRPM8 in sensory neurons of the PNS does not lead to a metabolically relevant phenotype.Conclusions: Our data indicate that obesity in TRPM8-deficient mice is centrally mediated and likely originates from alterations in energy expenditure and/or thermal conductance, but does not depend on TRPM8 signaling in brown adipocytes or sensory neurons of the PVN.
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| 34. |
- Pfab, Alexander, et al.
(författare)
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Inhibition of mitochondrial transcription by the neurotoxin MPP
- 2023
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 425:1
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Tidskriftsartikel (refereegranskat)abstract
- The neurotoxin MPP+ triggers cell death of dopamine neurons and induces Parkinson's disease symptoms in mice and men, but the immediate transcriptional response to this neurotoxin has not been studied. We therefore treated human SH-SY5Y cells with a low dose (0.1 mM) of MPP+ and measured the effect on nascent transcription by precision run-on sequencing (PRO-seq). We found that transcription of the mitochondrial genome was significantly reduced already after 30 min, whereas nuclear gene transcription was unaffected. Inhibition of respiratory complex I by MPP+ led to reduced ATP production, that may explain the diminished activity of mitochondrial RNA polymerase. Our results show that MPP+ has a direct effect on mitochondrial function and transcription, and that other gene expression or epigenetic changes induced by this neurotoxin are secondary effects that reflect a cellular adaptation program.
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| 35. |
- Rollwitz, Erik, et al.
(författare)
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Plate-Based Respirometry to Assess Thermal Sensitivity of Zebrafish Embryo Bioenergetics in situ
- 2021
-
Ingår i: Frontiers in Physiology. - : Frontiers Media SA. - 1664-042X. ; 12
-
Tidskriftsartikel (refereegranskat)abstract
- Oxygen consumption allows measuring the metabolic activity of organisms. Here, we adopted the multi-well plate-based respirometry of the extracellular flux analyzer (Seahorse XF96) to investigate the effect of temperature on the bioenergetics of zebrafish embryos (Danio rerio) in situ. We show that the removal of the embryonic chorion is beneficial for oxygen consumption rates (OCR) and penetration of various mitochondrial inhibitors, and confirm that sedation reduces the variability of OCR. At 48h post-fertilization, embryos (maintained at a routine temperature of 28°C) were exposed to different medium temperatures ranging from 18°C to 37°C for 20h prior OCR measurement. Measurement temperatures from 18°C to 45°C in the XF96 were achieved by lowering the room temperature and active in-built heating. At 18°C assay temperature, basal OCR was low due to decreased ATP-linked respiration, which was not limited by mitochondrial power, as seen in substantial spare respiratory capacity. Basal OCR of the embryos increased with assay temperature and were stable up to 37°C assay temperature, with pre-exposure of 37°C resulting in more thermo-resistant basal OCR measured at 41°C. Adverse effects of the mitochondrial inhibitor oligomycin were seen at 37°C and chemical uncouplers disrupted substrate oxidation gradually with increasing assay temperature. Proton leak respiration increased at assay temperatures above 28°C and compromised the efficiency of ATP production, calculated as coupling efficiency. Thus, temperature impacts mitochondrial respiration by reduced cellular ATP turnover at lower temperatures and by increased proton leak at higher temperatures. This conclusion is coherent with the assessment of heart rate, an independent indicator of systemic metabolic rate, which increased with exposure temperature, peaking at 28°C, and decreased at higher temperatures. Collectively, plate-based respirometry allows assessing distinct parts of mitochondrial energy transduction in zebrafish embryos and investigating the effect of temperature and temperature acclimation on mitochondrial bioenergetics in situ.
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| 36. |
- Russo, Gianluca L., et al.
(författare)
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CRISPR-Mediated Induction of Neuron-Enriched Mitochondrial Proteins Boosts Direct Glia-to-Neuron Conversion
- 2021
-
Ingår i: Cell Stem Cell. - : Elsevier BV. - 1934-5909 .- 1875-9777. ; 28:3, s. 524-534
-
Tidskriftsartikel (refereegranskat)abstract
- Astrocyte-to-neuron conversion is a promising avenue for neuronal replacement therapy. Neurons are particularly dependent on mitochondrial function, but how well mitochondria adapt to the new fate is unknown. Here, we determined the comprehensive mitochondrial proteome of cortical astrocytes and neurons, identifying about 150 significantly enriched mitochondrial proteins for each cell type, including transporters, metabolic enzymes, and cell-type-specific antioxidants. Monitoring their transition during reprogramming revealed late and only partial adaptation to the neuronal identity. Early dCas9-mediated activation of genes encoding mitochondrial proteins significantly improved conversion efficiency, particularly for neuron-enriched but not astrocyte-enriched antioxidant proteins. For example, Sod1 not only improves the survival of the converted neurons but also elicits a faster conversion pace, indicating that mitochondrial proteins act as enablers and drivers in this process. Transcriptional engineering of mitochondrial proteins with other functions improved reprogramming as well, demonstrating a broader role of mitochondrial proteins during fate conversion.
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| 37. |
- Tattikota, Sudhir G., et al.
(författare)
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miR-184 Regulates Pancreatic beta-Cell Function According to Glucose Metabolism
- 2015
-
Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 290:33, s. 20284-20294
-
Tidskriftsartikel (refereegranskat)abstract
- In response to fasting or hyperglycemia, the pancreatic beta-cell alters its output of secreted insulin; however, the pathways governing this adaptive response are not entirely established. Although the precise role of microRNAs (miRNAs) is also unclear, a recurring theme emphasizes their function in cellular stress responses. We recently showed that miR-184, an abundant miRNA in the beta-cell, regulates compensatory proliferation and secretion during insulin resistance. Consistent with previous studies showing miR-184 suppresses insulin release, expression of this miRNA was increased in islets after fasting, demonstrating an active role in the beta-cell as glucose levels lower and the insulin demand ceases. Additionally, miR-184 was negatively regulated upon the administration of a sucrose-rich diet in Drosophila, demonstrating strong conservation of this pathway through evolution. Furthermore, miR-184 and its target Argonaute2 remained inversely correlated as concentrations of extracellular glucose increased, underlining a functional relationship between this miRNA and its targets. Lastly, restoration of Argonaute2 in the presence of miR-184 rescued suppression of miR-375-targeted genes, suggesting these genes act in a coordinated manner during changes in the metabolic context. Together, these results highlight the adaptive role of miR-184 according to glucose metabolism and suggest the regulatory role of this miRNA in energy homeostasis is highly conserved.
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| 38. |
- Tattikota, Sudhir G, et al.
(författare)
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MiR-184 regulates pancreatic β-cell function according to glucose metabolism.
- 2015
-
Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 290:33, s. 20284-20294
-
Tidskriftsartikel (refereegranskat)abstract
- In response to fasting or hyperglycemia, the pancreatic β-cell alters its output of secreted insulin; however the pathways governing this adaptive response are not entirely established. While the precise role of microRNAs (miRNAs) is also unclear, a recurring theme emphasizes their function in cellular stress responses. We recently showed that miR-184, an abundant miRNA in the β-cell, regulates compensatory proliferation and secretion during insulin resistance. Consistent with previous studies showing miR-184 suppresses insulin release, expression of this miRNA was increased in islets after fasting, demonstrating an active role in the β-cell as glucose levels lower and the insulin demand ceases. Additionally, miR-184 was negatively regulated upon administration of a sucrose-rich diet in Drosophila demonstrating strong conservation of this pathway through evolution. Furthermore, miR-184 and its target Argonaute2 (Ago2) remained inversely correlated as concentrations of extracellular glucose increased, underlining a functional relationship between this miRNA and its targets. Lastly, restoration of Ago2 in the presence of miR-184 rescued suppression of miR-375-targeted genes suggesting these genes act in a coordinated manner during changes in the metabolic context. Together, these results highlight the adaptive role of miR-184 according to glucose metabolism and suggest the regulatory role of this miRNA in energy homeostasis is highly conserved.
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| 39. |
- Tews, D., et al.
(författare)
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Elevated UCP1 levels are sufficient to improve glucose uptake in human white adipocytes
- 2019
-
Ingår i: Redox Biology. - : Elsevier BV. - 2213-2317. ; 26
-
Tidskriftsartikel (refereegranskat)abstract
- Brown adipose tissue (BAT) has been considered beneficial for metabolic health by participating in the regulation of glucose homoeostasis. The browning factors that improve glucose uptake beyond normal levels are still unknown but glucose uptake is not affected in UCP1 knockout mice. Here, we demonstrate in human white adipocytes that basal/resting glucose uptake is improved by solely elevating UCP1 protein levels. Generating human white Simpson-Golabi-Behmel syndrome (SGBS) adipocytes with a stable knockout and overexpression of UCP1, we discovered that UCP1 overexpressing adipocytes significantly improve glucose uptake by 40%. Mechanistically, this is caused by higher glycolytic flux, seen as increased oxygen consumption, extracellular acidification and lactate secretion rates. The improvements in glucose handling are comparable to white-to-brown transitions, as judged by, for the first time, directly comparing in vitro differentiated mouse brown vs white adipocytes. Although no adipogenic, metabolic and mitochondrial gene expressions were significantly altered in SGBS cells, pharmacological inhibition of GLUT1 completely abrogated differences between UCP1 + and control cells, thereby uncovering GLUT1-mediated uptake as permissive gatekeeper. Collectively, our data demonstrate that elevating UCP1 levels is sufficient to improve human white adipocytes as a glucose sink without adverse cellular effects, thus not requiring the adrenergic controlled, complex network of browning which usually hampers translational efforts.
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| 40. |
- Treberg, Jason R., et al.
(författare)
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Comparing Electron Leak in Vertebrate Muscle Mitochondria
- 2018
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Ingår i: Integrative and Comparative Biology. - : Oxford University Press (OUP). - 1540-7063 .- 1557-7023. ; 58:3, s. 495-505
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Tidskriftsartikel (refereegranskat)abstract
- Mitochondrial electron transfer for oxidative ATP regeneration is linked to reactive oxygen species (ROS) production in aerobic eukaryotic cells. Because they can contribute to signaling as well as oxidative damage in cells, these ROS have profound impact for the physiology and survival of the organism. Although mitochondria have been recognized as a potential source for ROS for about 50 years, the mechanistic understanding on molecular sites and processes has advanced recently. Most experimental approaches neglect thermal variability among species although temperature impacts mitochondrial processes significantly. Here we delineate the importance of temperature by comparing muscle mitochondrial ROS formation across species. Measuring the thermal sensitivity of respiration, electron leak rate (ROS formation), and the antioxidant capacity (measured as H2O2 consumption) in intact mitochondria of representative ectothermic and endothermic vertebrate species, our results suggest that using a common assay temperature is inappropriate for comparisons of organisms with differing body temperatures. Moreover, we propose that measuring electron leak relative to the mitochondrial antioxidant capacity (the oxidant ratio) may be superior to normalizing relative to respiration rates or mitochondrial protein for comparisons of mitochondrial metabolism of ROS across species of varying mitochondrial respiratory capacities.
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| 41. |
- Welmanh, Shaun, et al.
(författare)
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Obligatory homeothermy of mesic habitat-adapted African striped mice, Rhabdomys pumilio, is governed by seasonal basal metabolism and year-round 'thermogenic readiness' of brown adipose tissue
- 2022
-
Ingår i: Journal of Experimental Biology. - : The Company of Biologists. - 0022-0949 .- 1477-9145. ; 225:13
-
Tidskriftsartikel (refereegranskat)abstract
- Small mammals undergo thermoregulatory adjustments in response to changing environmental conditions. Whereas small heterothermic mammals can employ torpor to save energy in the cold, homeothermic species must increase heat production to defend normothermia through the recruitment of brown adipose tissue (BAT). Here, we studied thermoregulatory adaptation in an obligate homeotherm, the African striped mouse (Rhabdomys pumilio), captured from a subpopulation living in a mesic, temperate climate with marked seasonal differences. Basal metabolic rate (BMR), nonshivering thermogenesis (NST) and summit metabolic rate (M-sum) increased from summer to winter, with NST and M-sum already reaching maximal rates in autumn, suggesting seasonal preparation for the cold. Typical of rodents, cold-induced metabolic rates were positively correlated with BAT mass. Analysis of cytochrome c oxidase (COX) activity and UCP1 content, however, demonstrated that thermogenic capacity declined with BAT mass. This resulted in seasonal differences in NST being driven by changes in BMR. The increase in BMR was supported by a comprehensive anatomical analysis of metabolically active organs, revealing increased mass proportions in the cold season. The thermoregulatory response of R. pumilio was associated with the maintenance of body mass throughout the year (48.3 +/- 1.4 g), contrasting large summer-winter mass reductions often observed in Holarctic rodents. Collectively. bioenergetic adaptation of this Afrotropical rodent involves seasonal organ adjustments influencing BMR, combined with a constant thermogenic capacity dictated by trade-offs in the thermogenic properties of BAT. Arguably, this high degree of plasticity was a response to unpredictable cold spells throughout the year. Consequently, the reliance on such a resource-intensive thermoregulatory strategy may expose more energetic vulnerability in changing environments of food scarcity and extreme weather conditions due to climate change, with major ramifications for survival of the species.
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| 42. |
- Zhang, Qian, et al.
(författare)
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The glucose-dependent insulinotropic polypeptide (GIP) regulates body weight and food intake via CNS-GIPR signaling
- 2021
-
Ingår i: Cell Metabolism. - : Elsevier BV. - 1550-4131 .- 1932-7420. ; 33:4, s. 833-844
-
Tidskriftsartikel (refereegranskat)abstract
- Uncertainty exists as to whether the glucose-dependent insulinotropic polypeptide receptor (GIPR) should be activated or inhibited for the treatment of obesity. Gipr was recently demonstrated in hypothalamic feeding centers, but the physiological relevance of CNS Gipr remains unknown. Here we show that HFD-fed CNS-Gipr KO mice and humanized (h)GIPR knockin mice with CNS-hGIPR deletion show decreased body weight and improved glucose metabolism. In DIO mice, acute central and peripheral administration of acyl-GIP increases cFos neuronal activity in hypothalamic feeding centers, and this coincides with decreased body weight and food intake and improved glucose handling. Chronic central and peripheral administration of acyl-GIP lowers body weight and food intake in wild-type mice, but shows blunted/absent efficacy in CNS-Gipr KO mice. Also, the superior metabolic effect of GLP-1/GIP co-agonism relative to GLP-1 is extinguished in CNS-Gipr KO mice. Our data hence establish a key role of CNS Gipr for control of energy metabolism.
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| 43. |
- Ziqubu, Khanyisani, et al.
(författare)
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Brown adipose tissue-derived metabolites and their role in regulating metabolism
- 2024
-
Ingår i: Metabolism. - 0026-0495 .- 1532-8600. ; 150
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Forskningsöversikt (refereegranskat)abstract
- The discovery and rejuvenation of metabolically active brown adipose tissue (BAT) in adult humans have offered a new approach to treat obesity and metabolic diseases. Beyond its accomplished role in adaptive thermogenesis, BAT secretes signaling molecules known as “batokines”, which are instrumental in regulating whole-body metabolism via autocrine, paracrine, and endocrine action. In addition to the intrinsic BAT metabolite-oxidizing activity, the endocrine functions of these molecules may help to explain the association between BAT activity and a healthy systemic metabolic profile. Herein, we review the evidence that underscores the significance of BAT-derived metabolites, especially highlighting their role in controlling physiological and metabolic processes involving thermogenesis, substrate metabolism, and other essential biological processes. The conversation extends to their capacity to enhance energy expenditure and mitigate features of obesity and its related metabolic complications. Thus, metabolites derived from BAT may provide new avenues for the discovery of metabolic health-promoting drugs with far-reaching impacts. This review aims to dissect the complexities of the secretory role of BAT in modulating local and systemic metabolism in metabolic health and disease.
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