Publications by authors named "Sebastian Brachs"

25 Publications

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Dietary-challenged mice with Alzheimer-like pathology show increased energy expenditure and reduced adipocyte hypertrophy and steatosis.

Aging (Albany NY) 2021 Apr 16;13. Epub 2021 Apr 16.

Department of Endocrinology and Metabolism, Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin 10117, Germany.

Alzheimer's disease (AD) is frequently accompanied by progressing weight loss, correlating with mortality. Counter-intuitively, weight loss in old age might predict AD onset but obesity in midlife increases AD risk. Furthermore, AD is associated with diabetes-like alterations in glucose metabolism. Here, we investigated metabolic features of amyloid precursor protein overexpressing APP23 female mice modeling AD upon long-term challenge with high-sucrose (HSD) or high-fat diet (HFD). Compared to wild type littermates (WT), APP23 females were less prone to mild HSD-induced and considerable HFD-induced glucose tolerance deterioration, despite unaltered glucose tolerance during normal-control diet. Indirect calorimetry revealed increased energy expenditure and hyperactivity in APP23 females. Dietary interventions, especially HFD, had weaker effects on lean and fat mass gain, steatosis and adipocyte hypertrophy of APP23 than WT mice, as shown by H-magnetic-resonance-spectroscopy, histological and biochemical analyses. Proteome analysis revealed differentially regulated expression of mitochondrial proteins in APP23 livers and brains. In conclusion, hyperactivity, increased metabolic rate, and global mitochondrial dysfunction potentially add up to the development of AD-related body weight changes in APP23 females, becoming especially evident during diet-induced metabolic challenge. These findings emphasize the importance of translating this metabolic phenotyping into human research to decode the metabolic component in AD pathogenesis.
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http://dx.doi.org/10.18632/aging.202978DOI Listing
April 2021

Group 3 Innate Lymphoid Cells Program a Distinct Subset of IL-22BP-Producing Dendritic Cells Demarcating Solitary Intestinal Lymphoid Tissues.

Immunity 2020 11;53(5):1015-1032.e8

Laboratory of Innate Immunity, Department of Microbiology, Infectious Diseases and Immunology, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Hindenburgdamm 30, 12203 Berlin, Germany; Berlin Institute of Health (BIH), Anna-Louisa-Karsch Strasse 2, 10117 Berlin, Germany; Mucosal and Developmental Immunology, Deutsches Rheuma-Forschungszentrum (DRFZ), an institute of the Leibniz Association, 10117 Berlin, Germany. Electronic address:

Solitary intestinal lymphoid tissues such as cryptopatches (CPs) and isolated lymphoid follicles (ILFs) constitute steady-state activation hubs containing group 3 innate lymphoid cells (ILC3) that continuously produce interleukin (IL)-22. The outer surface of CPs and ILFs is demarcated by a poorly characterized population of CD11c cells. Using genome-wide single-cell transcriptional profiling of intestinal mononuclear phagocytes and multidimensional flow cytometry, we found that CP- and ILF-associated CD11c cells were a transcriptionally distinct subset of intestinal cDCs, which we term CIA-DCs. CIA-DCs required programming by CP- and ILF-resident CCR6 ILC3 via lymphotoxin-β receptor signaling in cDCs. CIA-DCs differentially expressed genes associated with immunoregulation and were the major cellular source of IL-22 binding protein (IL-22BP) at steady state. Mice lacking CIA-DC-derived IL-22BP exhibited diminished expression of epithelial lipid transporters, reduced lipid resorption, and changes in body fat homeostasis. Our findings provide insight into the design principles of an immunoregulatory checkpoint controlling nutrient absorption.
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http://dx.doi.org/10.1016/j.immuni.2020.10.012DOI Listing
November 2020

Impact of C-reactive protein on osteo-/chondrogenic transdifferentiation and calcification of vascular smooth muscle cells.

Aging (Albany NY) 2019 08 3;11(15):5445-5462. Epub 2019 Aug 3.

Department of Internal Medicine and Cardiology, Charité - Universitätsmedizin Berlin, Campus Virchow-Klinikum, Berlin 13353, Germany.

Medial vascular calcification occurs during the aging process and is strongly accelerated by chronic kidney disease (CKD). Elevated C-reactive protein (CRP) levels are associated with vascular calcification, cardiovascular events and mortality in CKD patients. CRP is an important promoter of vascular inflammation. Inflammatory processes are critically involved in initiation and progression of vascular calcification. Thus, the present study explored a possible impact of CRP on vascular calcification. We found that CRP promoted osteo-/chondrogenic transdifferentiation and aggravated phosphate-induced osteo-/chondrogenic transdifferentiation and calcification of primary human aortic smooth muscle cells (HAoSMCs). These effects were paralleled by increased cellular oxidative stress and corresponding pro-calcific downstream-signaling. Antioxidants or p38 MAPK inhibition suppressed CRP-induced osteo-/chondrogenic signaling and mineralization. Furthermore, silencing of Fc fragment of IgG receptor IIa (FCGR2A) blunted the pro-calcific effects of CRP. Vascular CRP expression was increased in the klotho-hypomorphic mouse model of aging as well as in HAoSMCs during calcifying conditions. In conclusion, CRP augments osteo-/chondrogenic transdifferentiation of vascular smooth muscle cells through mechanisms involving FCGR2A-dependent induction of oxidative stress. Thus, systemic inflammation may actively contribute to the progression of vascular calcification.
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http://dx.doi.org/10.18632/aging.102130DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6710049PMC
August 2019

Genetic Nicotinamide -Methyltransferase () Deficiency in Male Mice Improves Insulin Sensitivity in Diet-Induced Obesity but Does Not Affect Glucose Tolerance.

Diabetes 2019 03 14;68(3):527-542. Epub 2018 Dec 14.

Sanofi Research and Development, Frankfurt am Main, Germany

Antisense oligonucleotide knockdown (ASO-KD) of nicotinamide -methyltransferase (NNMT) in high-fat diet (HFD)-fed mice has been reported to reduce weight gain and plasma insulin levels and to improve glucose tolerance. Using NNMT-ASO-KD or NNMT knockout mice (NNMT), we tested the hypothesis that deletion protects against diet-induced obesity and its metabolic consequences in males and females on obesity-inducing diets. We also examined samples from a human weight reduction (WR) study for adipose (a) expression and plasma 1-methylnicotinamide (MNAM) levels. In Western diet (WD)-fed female mice, NNMT-ASO-KD reduced body weight, fat mass, and insulin level and improved glucose tolerance. Although NNMT mice fed a standard diet had no obvious phenotype, NNMT males fed an HFD showed strongly improved insulin sensitivity (IS). Furthermore, NNMT females fed a WD showed reduced weight gain, less fat, and lower insulin levels. However, no improved glucose tolerance was observed in NNMT mice. Although expression in human fat biopsy samples increased during WR, corresponding plasma MNAM levels significantly declined, suggesting that other mechanisms besides a expression modulate circulating MNAM levels during WR. In summary, upon NNMT deletion or knockdown in males and females fed different obesity-inducing diets, we observed sex- and diet-specific differences in body composition, weight, and glucose tolerance and estimates of IS.
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http://dx.doi.org/10.2337/db18-0780DOI Listing
March 2019

Distinct Housing Conditions Reveal a Major Impact of Adaptive Immunity on the Course of Obesity-Induced Type 2 Diabetes.

Front Immunol 2018 28;9:1069. Epub 2018 May 28.

Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité - Universitätsmedizin Berlin, Freie Universität Berlin, Humboldt-Universität zu Berlin, Berlin, Germany.

Obesity is associated with adipose tissue inflammation, insulin resistance, and the development of type 2 diabetes (T2D). However, our knowledge is mostly based on conventional murine models and promising preclinical studies rarely translated into successful therapies. There is a growing awareness of the limitations of studies in laboratory mice, housed in abnormally hygienic specific pathogen-free (SPF) conditions, as relevant aspects of the human immune system remain unappreciated. Here, we assessed the impact of housing conditions on adaptive immunity and metabolic disease processes during high-fat diet (HFD). We therefore compared diet-induced obesity in SPF mice with those housed in non-SPF, so-called "antigen exposed" (AE) conditions. Surprisingly, AE mice fed a HFD maintained increased insulin levels to compensate for insulin resistance, which was reflected in islet hyperplasia and improved glucose tolerance compared to SPF mice. By contrast, we observed higher proportions of effector/memory T cell subsets in blood and liver of HFD AE mice accompanied by the development of non-alcoholic steatohepatitis-like liver pathology. Thus, our data demonstrate the impact of housing conditions on metabolic alterations. Studies in AE mice, in which physiological microbial exposure was restored, could provide a tool for revealing therapeutic targets for immune-based interventions for T2D patients.
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http://dx.doi.org/10.3389/fimmu.2018.01069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5985496PMC
July 2019

Inflammatory Pathways Regulated by Tumor Necrosis Receptor-Associated Factor 1 Protect From Metabolic Consequences in Diet-Induced Obesity.

Circ Res 2018 03 22;122(5):693-700. Epub 2018 Jan 22.

From the Cardiology and Angiology I, University Heart Center and Medical Center (N.A.M., C.C., B.D., N.H., K.P., T.M., F.W., P.S., I.H., T.H., C.v.z.M., C.B., A.Z., D.W.), Faculty of Biology (N.A.M.), Department of Radiology, Medical Physics, Medical Center (D.v.E.), Hematology and Oncology (D.P.), and Department of Urology (R.S.), University of Freiburg, Germany; Inflammation Biology, La Jolla Institute for Allergy and Immunology, CA (K.B., A.B.P., E.E., K.L., D.W.); Neurosurgery, University of Erlangen, Germany (B.S.); Center for Cardiovascular Research (U.K.) and Department of Endocrinology & Metabolism, Center for Cardiovascular Research (CCR), Germany (S.B.), Charité-Universitätsmedizin Berlin, Germany; and Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK), Partner Site Berlin, Germany (S.B.).

Rationale: The coincidence of inflammation and metabolic derangements in obese adipose tissue has sparked the concept of met-inflammation. Previous observations, however, suggest that inflammatory pathways may not ultimately cause dysmetabolism.

Objective: We have revisited the relationship between inflammation and metabolism by testing the role of TRAF (tumor necrosis receptor-associated factor)-1, an inhibitory adapter of inflammatory signaling of TNF (tumor necrosis factor)-α, IL (interleukin)-1β, and TLRs (toll-like receptors).

Methods And Results: Mice deficient for TRAF-1, which is expressed in obese adipocytes and adipose tissue lymphocytes, caused an expected hyperinflammatory phenotype in adipose tissue with enhanced adipokine and chemokine expression, increased leukocyte accumulation, and potentiated proinflammatory signaling in macrophages and adipocytes in a mouse model of diet-induced obesity. Unexpectedly, TRAF-1 mice were protected from metabolic derangements and adipocyte growth, failed to gain weight, and showed improved insulin resistance-an effect caused by increased lipid breakdown in adipocytes and UCP (uncoupling protein)-1-enabled thermogenesis. TRAF-1-dependent catabolic and proinflammatory cues were synergistically driven by β3-adrenergic and inflammatory signaling and required the presence of both TRAF-1-deficient adipocytes and macrophages. In human obesity, TRAF-1-dependent genes were upregulated.

Conclusions: Enhancing TRAF-1-dependent inflammatory pathways in a gain-of-function approach protected from metabolic derangements in diet-induced obesity. These findings identify TRAF-1 as a regulator of dysmetabolism in mice and humans and question the pathogenic role of chronic inflammation in metabolism.
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http://dx.doi.org/10.1161/CIRCRESAHA.117.312055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5834385PMC
March 2018

Impact of Swiprosin-1/Efhd2 on Adult Hippocampal Neurogenesis.

Stem Cell Reports 2018 02 11;10(2):347-355. Epub 2018 Jan 11.

Department of Stem Cell Biology, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Glueckstrasse 6, Erlangen 91054, Germany; IZKF Junior Research Group III and BMBF Research Group Neuroscience, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU), Erlangen 91054, Germany. Electronic address:

Swiprosin-1/Efhd2 (Efhd2) is highly expressed in the CNS during development and in the adult. EFHD2 is regulated by Ca binding, stabilizes F-actin, and promotes neurite extension. Previous studies indicated a dysregulation of EFHD2 in human Alzheimer's disease brains. We hypothesized a detrimental effect of genetic ablation of Efhd2 on hippocampal integrity and specifically investigated adult hippocampal neurogenesis. Efhd2 was expressed throughout adult neuronal development and in mature neurons. We observed a severe reduction of the survival of adult newborn neurons in Efhd2 knockouts, starting at the early neuroblast stage. Spine formation and dendrite growth of newborn neurons were compromised in full Efhd2 knockouts, but not upon cell-autonomous Efhd2 deletion. Together with our finding of severe hippocampal tauopathy in Efhd2 knockout mice, these data connect Efhd2 to impaired synaptic plasticity as present in Alzheimer's disease and identify a role of Efhd2 in neuronal survival and synaptic integration in the adult hippocampus.
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http://dx.doi.org/10.1016/j.stemcr.2017.12.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5830914PMC
February 2018

Regulation of body weight and energy homeostasis by neuronal cell adhesion molecule 1.

Nat Neurosci 2017 Aug 19;20(8):1096-1103. Epub 2017 Jun 19.

Max Delbrück Center for Molecular Medicine, Berlin, Germany.

Susceptibility to obesity is linked to genes regulating neurotransmission, pancreatic beta-cell function and energy homeostasis. Genome-wide association studies have identified associations between body mass index and two loci near cell adhesion molecule 1 (CADM1) and cell adhesion molecule 2 (CADM2), which encode membrane proteins that mediate synaptic assembly. We found that these respective risk variants associate with increased CADM1 and CADM2 expression in the hypothalamus of human subjects. Expression of both genes was elevated in obese mice, and induction of Cadm1 in excitatory neurons facilitated weight gain while exacerbating energy expenditure. Loss of Cadm1 protected mice from obesity, and tract-tracing analysis revealed Cadm1-positive innervation of POMC neurons via afferent projections originating from beyond the arcuate nucleus. Reducing Cadm1 expression in the hypothalamus and hippocampus promoted a negative energy balance and weight loss. These data identify essential roles for Cadm1-mediated neuronal input in weight regulation and provide insight into the central pathways contributing to human obesity.
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http://dx.doi.org/10.1038/nn.4590DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5533218PMC
August 2017

Renal function is independently associated with circulating betatrophin.

PLoS One 2017 3;12(3):e0173197. Epub 2017 Mar 3.

Dept. of Endocrinology, Diabetes and Nutrition, Charité-Universitätsmedizin Berlin, Berlin, Germany.

Objective: Betatrophin has been identified as a marker linking liver with beta cell function and lipid metabolism in murine models. Until now, the regulation of circulating betatrophin in humans is not entirely clear. We here analyzed the relation of betatrophin levels to phenotypes of the metabolic syndrome and speculated that renal function might influence circulating betatrophin levels and explain age-dependent changes of betatrophin.

Subjects: We analyzed blood samples from 535 individuals participating in the Metabolic Syndrome Berlin Potsdam study.

Results: In a crude analysis we found a positive correlation between betatrophin levels and HbA1c (r = 0.24; p < 0.001), fasting glucose (r = 0.20; p < 0.001) and triglycerides (r = 0.12; p = 0.007). Furthermore betatrophin was positively correlated with age (r = 0.47; p <0.001), systolic blood pressure (r = 0.17; p < 0.001), intima media thickness (r = 0.26; p < 0.001) and negatively correlated with CKD-EPI eGFR (r = -0.33; p < 0.001) as an estimate of renal function. Notably, eGFR remained highly associated with betatrophin after adjustment for age, waist circumference, gender, HbA1c and lipid parameters in a multivariate linear regression model (β = -0.197, p< 0.001).

Conclusions: Our data suggest that circulating levels of betatrophin depend on age, gender, waist circumference, total/HDL cholesterol ratio and renal function. Especially the association to eGFR highlights the importance for future studies to address renal function as possible influence on betatrophin regulation and consider eGFR as potential confounder when analyzing the role of betatrophin in humans.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0173197PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5336269PMC
August 2017

Inhibition of citrate cotransporter Slc13a5/mINDY by RNAi improves hepatic insulin sensitivity and prevents diet-induced non-alcoholic fatty liver disease in mice.

Mol Metab 2016 11 13;5(11):1072-1082. Epub 2016 Aug 13.

Department of Endocrinology, Diabetes and Nutrition, Center for Cardiovascular Research, Charité - University School of Medicine, Berlin, 10117, Germany; DZHK (German Center for Cardiovascular Research), Partner Site, Berlin, Germany. Electronic address:

Objective: Non-alcoholic fatty liver disease is a world-wide health concern and risk factor for cardio-metabolic diseases. Citrate uptake modifies intracellular hepatic energy metabolism and is controlled by the conserved sodium-dicarboxylate cotransporter solute carrier family 13 member 5 (SLC13A5, mammalian homolog of INDY: mINDY). In er and INDY reduction decreased whole-body lipid accumulation. Genetic deletion of in mice protected from diet-induced adiposity and insulin resistance. We hypothesized that inducible hepatic mINDY inhibition should prevent the development of fatty liver and hepatic insulin resistance.

Methods: Adult C57BL/6J mice were fed a Western diet (60% kcal from fat, 21% kcal from carbohydrate) ad libitum. Knockdown of mINDY was induced by weekly injection of a chemically modified, liver-selective siRNA for 8 weeks. Mice were metabolically characterized and the effect of mINDY suppression on glucose tolerance as well as insulin sensitivity was assessed with an ipGTT and a hyperinsulinemic-euglycemic clamp. Hepatic lipid accumulation was determined by biochemical measurements and histochemistry.

Results: Within the 8 week intervention, hepatic mINDY expression was suppressed by a liver-selective siRNA by over 60%. mINDY knockdown improved hepatic insulin sensitivity (i.e. insulin-induced suppression of endogenous glucose production) of C57BL/6J mice in the hyperinsulinemic-euglycemic clamp. Moreover, the siRNA-mediated mINDY inhibition prevented neutral lipid storage and triglyceride accumulation in the liver, while we found no effect on body weight.

Conclusions: We show that inducible mINDY inhibition improved hepatic insulin sensitivity and prevented diet-induced non-alcoholic fatty liver disease in adult C57BL6/J mice. These effects did not depend on changes of body weight or body composition.
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http://dx.doi.org/10.1016/j.molmet.2016.08.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5081411PMC
November 2016

Chronic Activation of Hepatic Nrf2 Has No Major Effect on Fatty Acid and Glucose Metabolism in Adult Mice.

PLoS One 2016 4;11(11):e0166110. Epub 2016 Nov 4.

Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, Germany.

The transcription factor NF-E2-related factor 2 (Nrf2) induces cytoprotective genes, but has also been linked to the regulation of hepatic energy metabolism. In order to assess the pharmacological potential of hepatic Nrf2 activation in metabolic disease, Nrf2 was activated over 7 weeks in mice on Western diet using two different siRNAs against kelch-like ECH-associated protein 1 (Keap1), the inhibitory protein of Nrf2. Whole genome expression analysis followed by pathway analysis demonstrated successful knock-down of Keap1 expression and induction of Nrf2-dependent genes involved in anti-oxidative stress defense and biotransformation, proving the activation of Nrf2 by the siRNAs against Keap1. Neither the expression of fatty acid- nor carbohydrate-handling proteins was regulated by Keap1 knock-down. Metabolic profiling of the animals did also not show effects on plasma and hepatic lipids, energy expenditure or glucose tolerance. The data indicate that hepatic Keap1/Nrf2 is not a major regulator of glucose or lipid metabolism in mice.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0166110PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5096693PMC
June 2017

Weight Loss Partially Restores Glucose-Driven Betatrophin Response in Humans.

J Clin Endocrinol Metab 2016 Nov 26;101(11):4014-4020. Epub 2016 Jul 26.

Medical Clinic of Endocrinology, Diabetes, and Nutrition (L.M., S.B.,A.-M.D., M.B., V.L., R.J.v.S., T.B., J.S., K.M.) and Department of Pediatric Endocrinology and Diabetology (A.E., H.K.), Charité-Universitaetsmedizin Berlin, 10117 Berlin, Germany, Charité-Center for Cardiovascular Research (L.M., S.B., A.-M.D., M.B., V.L., R.J.v.S., T.B., J.S., K.M.), 10117 Berlin, Germany, and Clinical Research Unit (A.-M.D., K.M.), Berlin Institute of Health 10117 Berlin, Germany; and Experimental and Clinical Research Center (S.B., M.B., R.J.v.S., J.S., K.M.), 13125 Berlin, Germany.

Context: Recently a potential role of betatrophin was shown in the postprandial switch from lipid to glucose metabolism.

Objective: The objective of the study was to analyze whether obesity is associated with altered postprandial betatrophin response and whether this could be restored by weight loss. Design, Setting, Participants, and Intervention: Oral glucose load was performed in 12 lean individuals at baseline as well as in 20 obese subjects before and after a 12-week structured weight-loss program at an endocrinology research center. Euglycemic hyperinsulinemic clamps were performed in the obese cohort. The effect of insulin and different glucose concentrations on betatrophin expression were analyzed in 3T3-L1 adipocytes.

Main Outcome Measure: Circulating betatrophin levels during a glucose challenge were measured.

Results: The betatrophin level decreases after an oral glucose intake (P < .001). This correlates with the increase of glucose levels (r = -0.396; P < .05). Hyperinsulinemia results in an increase of betatrophin. In vitro experiments in 3T3-L1 adipocytes confirmed that insulin and low glucose concentration increases betatrophin expression, whereas a further elevation of glucose levels blunts this effect. Obese subjects are characterized by lower fasting betatrophin (600.6 ± 364.4 vs 759.5 ± 197.9 pg/mL; P < .05) and a more pronounced betatrophin suppression during the glucose challenge. The impaired betatrophin response in obese subjects is restored after weight loss and is comparable with lean individuals.

Conclusions: Obesity is associated with increased betatrophin suppression after an oral glucose load, which is driven by increased hyperglycemia. Given the metabolic properties of betatrophin, this may indicate that betatrophin is tightly linked to obesity-associated metabolic disturbances. In line with such an assumption, weight loss almost completely eliminated this phenomenon.
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http://dx.doi.org/10.1210/jc.2016-1788DOI Listing
November 2016

Adipocyte-specific blockade of gamma-secretase, but not inhibition of Notch activity, reduces adipose insulin sensitivity.

Mol Metab 2016 Feb 2;5(2):113-121. Epub 2015 Dec 2.

Department of Medicine, Columbia University, New York, NY 10032, USA. Electronic address:

Objective: As the obesity pandemic continues to expand, novel molecular targets to reduce obesity-related insulin resistance and Type 2 Diabetes (T2D) continue to be needed. We have recently shown that obesity is associated with reactivated liver Notch signaling, which, in turn, increases hepatic insulin resistance, opening up therapeutic avenues for Notch inhibitors to be repurposed for T2D. Herein, we tested the systemic effects of γ-secretase inhibitors (GSIs), which prevent endogenous Notch activation, and confirmed these effects through creation and characterization of two different adipocyte-specific Notch loss-of-function mouse models through genetic ablation of the Notch transcriptional effector Rbp-Jk (A-Rbpj) and the obligate γ-secretase component Nicastrin (A-Nicastrin).

Methods: Glucose homeostasis and both local adipose and systemic insulin sensitivity were examined in GSI-treated, A-Rbpj and A-Nicastrin mice, as well as vehicle-treated or control littermates, with complementary in vitro studies in primary hepatocytes and 3T3-L1 adipocytes.

Results: GSI-treatment increases hepatic insulin sensitivity in obese mice but leads to reciprocal lowering of adipose glucose disposal. While A-Rbpj mice show normal body weight, adipose development and mass and unchanged adipose insulin sensitivity as control littermates, A-Nicastrin mice are relatively insulin-resistant, mirroring the GSI effect on adipose insulin action.

Conclusions: Notch signaling is dispensable for normal adipocyte function, but adipocyte-specific γ-secretase blockade reduces adipose insulin sensitivity, suggesting that specific Notch inhibitors would be preferable to GSIs for application in T2D.
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http://dx.doi.org/10.1016/j.molmet.2015.11.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4735659PMC
February 2016

Enhanced insulin signaling in density-enhanced phosphatase-1 (DEP-1) knockout mice.

Mol Metab 2015 Apr 12;4(4):325-36. Epub 2015 Feb 12.

Center for Cardiovascular Research/CCR, Institute of Laboratory Medicine, Clinical Chemistry and Pathobiochemistry, Hessische Str. 3-4, 10115 Berlin, Charité - Universitätsmedizin Berlin, Germany.

Objective: Insulin resistance can be triggered by enhanced dephosphorylation of the insulin receptor or downstream components in the insulin signaling cascade through protein tyrosine phosphatases (PTPs). Downregulating density-enhanced phosphatase-1 (DEP-1) resulted in an improved metabolic status in previous analyses. This phenotype was primarily caused by hepatic DEP-1 reduction.

Methods: Here we further elucidated the role of DEP-1 in glucose homeostasis by employing a conventional knockout model to explore the specific contribution of DEP-1 in metabolic tissues. Ptprj (-/-) (DEP-1 deficient) and wild-type C57BL/6 mice were fed a low-fat or high-fat diet. Metabolic phenotyping was combined with analyses of phosphorylation patterns of insulin signaling components. Additionally, experiments with skeletal muscle cells and muscle tissue were performed to assess the role of DEP-1 for glucose uptake.

Results: High-fat diet fed-Ptprj (-/-) mice displayed enhanced insulin sensitivity and improved glucose tolerance. Furthermore, leptin levels and blood pressure were reduced in Ptprj (-/-) mice. DEP-1 deficiency resulted in increased phosphorylation of components of the insulin signaling cascade in liver, skeletal muscle and adipose tissue after insulin challenge. The beneficial effect on glucose homeostasis in vivo was corroborated by increased glucose uptake in skeletal muscle cells in which DEP-1 was downregulated, and in skeletal muscle of Ptprj (-/-) mice.

Conclusion: Together, these data establish DEP-1 as novel negative regulator of insulin signaling.
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http://dx.doi.org/10.1016/j.molmet.2015.02.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4354926PMC
April 2015

The AP-1 transcription factor Fra1 inhibits follicular B cell differentiation into plasma cells.

J Exp Med 2014 Oct 6;211(11):2199-212. Epub 2014 Oct 6.

Division of Molecular Immunology, Nikolaus Fiebiger Center, Department of Internal Medicine III, Department of Radiation Oncology, Division of Ophthalmology, Department Kopfklinik, University of Erlangen-Nuremberg, D91054 Erlangen, Germany Institute for Osteology and Biomechanics, University Medical Center Hamburg-Eppendorf, D20246 Hamburg, Germany

The cornerstone of humoral immunity is the differentiation of B cells into antibody-secreting plasma cells. This process is tightly controlled by a regulatory gene network centered on the transcriptional repressor B lymphocyte-induced maturation protein 1 (Blimp1). Proliferation of activated B cells is required to foster Blimp1 expression but needs to be terminated to avoid overshooting immune reactions. Activator protein 1 (AP-1) transcription factors become quickly up-regulated upon B cell activation. We demonstrate that Fra1, a Fos member of AP-1, enhances activation-induced cell death upon induction in activated B cells. Moreover, mice with B cell-specific deletion of Fra1 show enhanced plasma cell differentiation and exacerbated antibody responses. In contrast, transgenic overexpression of Fra1 blocks plasma cell differentiation and immunoglobulin production, which cannot be rescued by Bcl2. On the molecular level, Fra1 represses Blimp1 expression and interferes with binding of the activating AP-1 member c-Fos to the Blimp1 promoter. Conversely, overexpression of c-Fos in Fra1 transgenic B cells releases Blimp1 repression. As Fra1 lacks transcriptional transactivation domains, we propose that Fra1 inhibits Blimp1 expression and negatively controls plasma cell differentiation through binding to the Blimp1 promoter. In summary, we demonstrate that Fra1 negatively controls plasma cell differentiation by repressing Blimp1 expression.
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http://dx.doi.org/10.1084/jem.20130795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4203943PMC
October 2014

The adaptor protein Swiprosin-1/EFhd2 is dispensable for platelet function in mice.

PLoS One 2014 22;9(9):e107139. Epub 2014 Sep 22.

University of Würzburg, Department of Experimental Biomedicine, University Hospital and Rudolf Virchow Center for Experimental Biomedicine, Würzburg, Germany.

Background: Platelets are anuclear cell fragments derived from bone marrow megakaryocytes that safeguard vascular integrity, but may also cause pathological vessel occlusion. Reorganizations of the platelet cytoskeleton and agonist-induced intracellular Ca2+-mobilization are crucial for platelet hemostatic function. EF-hand domain containing 2 (EFhd2, Swiprosin-1) is a Ca2+-binding cytoskeletal adaptor protein involved in actin remodeling in different cell types, but its function in platelets is unknown.

Objective: Based on the described functions of EFhd2 in immune cells, we tested the hypothesis that EFhd2 is a crucial adaptor protein for platelet function acting as a regulator of Ca2+-mobilization and cytoskeletal rearrangements.

Methods And Results: We generated EFhd2-deficient mice and analyzed their platelets in vitro and in vivo. Efhd2-/- mice displayed normal platelet count and size, exhibited an unaltered in vivo life span and showed normal Ca2+-mobilization and activation/aggregation responses to classic agonists. Interestingly, upon stimulation of the immunoreceptor tyrosine-based activation motif-coupled receptor glycoprotein (GP) VI, Efhd2-/- platelets showed a slightly increased coagulant activity. Furthermore, absence of EFhd2 had no significant impact on integrin-mediated clot retraction, actomyosin rearrangements and spreading of activated platelets on fibrinogen. In vivo EFhd2-deficiency resulted in unaltered hemostatic function and unaffected arterial thrombus formation.

Conclusion: These results show that EFhd2 is not essential for platelet function in mice indicating that other cytoskeletal adaptors may functionally compensate its loss.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107139PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4170979PMC
June 2015

The Ca2+ sensor protein swiprosin-1/EFhd2 is present in neurites and involved in kinesin-mediated transport in neurons.

PLoS One 2014 18;9(8):e103976. Epub 2014 Aug 18.

Division of Molecular Immunology, Dept. of Internal Medicine III, Universitätsklinikum Erlangen, and Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.

Swiprosin-1/EFhd2 (EFhd2) is a cytoskeletal Ca2+ sensor protein strongly expressed in the brain. It has been shown to interact with mutant tau, which can promote neurodegeneration, but nothing is known about the physiological function of EFhd2 in the nervous system. To elucidate this question, we analyzed EFhd2-/-/lacZ reporter mice and showed that lacZ was strongly expressed in the cortex, the dentate gyrus, the CA1 and CA2 regions of the hippocampus, the thalamus, and the olfactory bulb. Immunohistochemistry and western blotting confirmed this pattern and revealed expression of EFhd2 during neuronal maturation. In cortical neurons, EFhd2 was detected in neurites marked by MAP2 and co-localized with pre- and post-synaptic markers. Approximately one third of EFhd2 associated with a biochemically isolated synaptosome preparation. There, EFhd2 was mostly confined to the cytosolic and plasma membrane fractions. Both synaptic endocytosis and exocytosis in primary hippocampal EFhd2-/- neurons were unaltered but transport of synaptophysin-GFP containing vesicles was enhanced in EFhd2-/- primary hippocampal neurons, and notably, EFhd2 inhibited kinesin mediated microtubule gliding. Therefore, we found that EFhd2 is a neuronal protein that interferes with kinesin-mediated transport.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0103976PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136728PMC
May 2015

Swiprosin-1/EFhd2 limits germinal center responses and humoral type 2 immunity.

Eur J Immunol 2014 Nov 16;44(11):3206-19. Epub 2014 Sep 16.

Division of Molecular Immunology, Department of Internal Medicine III, Nikolaus Fiebiger Center, Universitätsklinikum Erlangen and Friedrich-Alexander University Erlangen-Nuremberg, Nuremberg, Germany.

Activated B cells are selected for in germinal centers by regulation of their apoptosis. The Ca2+ -binding cytoskeletal adaptor protein Swiprosin-1/EFhd2 (EFhd2) can promote apoptosis in activated B cells. We therefore hypothesized that EFhd2 might limit humoral immunity by repressing both the germinal center reaction and the expected enhancement of immune responses in the absence of EFhd2. Here, we established EFhd2(-/-) mice on a C57BL/6 background, which revealed normal B- and T-cell development, basal Ab levels, and T-cell independent type 1, and T-cell independent type 2 responses. However, T cell-dependent immunization with sheep red blood cells and infection with the helminth Nippostrongylus brasiliensis (N.b) increased production of antibodies of multiple isotypes, as well as germinal center formation in EFhd2(-/-) mice. In addition, serum IgE levels and numbers of IgE+ plasma cells were strongly increased in EFhd2(-/-) mice, both after primary as well as after secondary N.b infection. Finally, mixed bone marrow chimeras unraveled an EFhd2-dependent B cell-intrinsic contribution to increased IgE plasma cell numbers in N.b-infected mice. Hence, we established a role for EFhd2 as a negative regulator of germinal center-dependent humoral type 2 immunity, with implications for the generation of IgE.
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http://dx.doi.org/10.1002/eji.201444479DOI Listing
November 2014

The mammalian INDY homolog is induced by CREB in a rat model of type 2 diabetes.

Diabetes 2014 Mar 12;63(3):1048-57. Epub 2013 Nov 12.

University of Potsdam, Institute of Nutritional Science, Nutritional Biochemistry, Potsdam, Germany.

Reduced expression of the INDY (I'm not dead yet) tricarboxylate carrier increased the life span in different species by mechanisms akin to caloric restriction. Mammalian INDY homolog (mIndy, SLC13A5) gene expression seems to be regulated by hormonal and/or nutritional factors. The underlying mechanisms are still unknown. The current study revealed that mIndy expression and [(14)C]-citrate uptake was induced by physiological concentrations of glucagon via a cAMP-dependent and cAMP-responsive element-binding protein (CREB)-dependent mechanism in primary rat hepatocytes. The promoter sequence of mIndy located upstream of the most frequent transcription start site was determined by 5'-rapid amplification of cDNA ends. In silico analysis identified a CREB-binding site within this promoter fragment of mIndy. Functional relevance for the CREB-binding site was demonstrated with reporter gene constructs that were induced by CREB activation when under the control of a fragment of a wild-type promoter, whereas promoter activity was lost after site-directed mutagenesis of the CREB-binding site. Moreover, CREB binding to this promoter element was confirmed by chromatin immunoprecipitation in rat liver. In vivo studies revealed that mIndy was induced in livers of fasted as well as in high-fat-diet-streptozotocin diabetic rats, in which CREB is constitutively activated. mIndy induction was completely prevented when CREB was depleted in these rats by antisense oligonucleotides. Together, these data suggest that mIndy is a CREB-dependent glucagon target gene that is induced in fasting and in type 2 diabetes. Increased mIndy expression might contribute to the metabolic consequences of diabetes in the liver.
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http://dx.doi.org/10.2337/db13-0749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3968437PMC
March 2014

Monoclonal antibodies to discriminate the EF hand containing calcium binding adaptor proteins EFhd1 and EFhd2.

Monoclon Antib Immunodiagn Immunother 2013 Aug;32(4):237-45

Division of Molecular Immunology, Department of Medicine III, University of Erlangen-Nürnberg, Nikolaus Fiebiger Center, Erlangen, Germany.

Small Ca(2+) binding adaptor proteins of the EF hand family play important roles in neuronal and immune cell Ca(2+) signaling. Swiprosin-1/EFhd2 (EFhd2) and Swiprosin-2/EFhd1 (EFhd1) are conserved and very homologous Ca(2+) binding adaptor proteins of the EF hand family, with possibly redundant functions. In particular, EFhd2 has been proposed to be involved in B cell signaling and neuropathological disorders. Little is known thus far about the expression of EFhd2 on the single cell level in tissue sections or blood cells. Here we describe the generation of four specific anti-EFhd2 monoclonal antibodies. These recognize murine and human EFhd2, but not murine EFhd1, and their binding site maps to a region in the N-terminal part of EFhd2, where EFhd2 and EFhd1 differ most. Moreover, to detect EFhd1 specifically, we also generated anti-EFhd1 polyclonal antibodies, making use of a singular peptide of the N-terminal part of the protein. Using anti-EFhd2 MAb, we reveal two EFhd2 pools in B cells, one at the membrane and one cytoplasmic pool. Staining of human peripheral blood mononuclear cells shows EFhd2 expression in B cells but a ∼5 fold higher expression in monocytes. Taken together, EFhd2 monoclonal antibodies will be valuable to assess the real subcellular localization and expression level of EFhd2 in healthy and diseased primary cells and tissues.
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http://dx.doi.org/10.1089/mab.2013.0014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3733322PMC
August 2013

The B cell receptor-induced calcium flux involves a calcium mediated positive feedback loop.

Cell Calcium 2012 May 10;51(5):411-7. Epub 2012 Feb 10.

Division of Molecular Immunology, Department of Internal Medicine III, University of Erlangen-Nürnberg, Nikolaus Fiebiger Center, Glückstr. 6, 91054 Erlangen, Germany.

The B cell receptor (BCR)-elicited calcium flux results in activation of mature B cells. We have recently shown that the adaptor protein Swiprosin-1/EFhd2 (EFhd2) amplifies the BCR-induced calcium flux in B cell lines. EFhd2 is a calcium binding adaptor protein with two predicted EF-hands. Here we asked whether these domains are functional and control its function. Using a blot-overlay assay with radioactive calcium we show that both EF-hands of EFhd2 have an intrinsic capacity to bind calcium. Equilibrium centrifugation confirmed that EFhd2 binds 2 calcium ions, with an apparent Kd of 110 μM. Point mutations revealed that the conserved residues E116 and E152, which reside in the canonical calcium binding loop in EF-hands 1 and 2, are essential for calcium binding by EFhd2. These mutations as well as deletion of the EF-hands, in particular EF-hand 1, abolished the ability of EFhd2 to restore BCR-induced calcium signaling in EFhd2-deficient WEHI231 cells. N-terminal deletions, but not C-terminal deletions, acted similarly. Thus, the N-terminal part of EFhd2 as well as calcium binding to its EF-hands control the intracellular calcium concentration in response to BCR stimulation in WEHI231 cells. Hence, EFhd2 regulates the BCR-elicited calcium flux through a calcium-dependent positive feedback mechanism in WEHI231 cells.
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http://dx.doi.org/10.1016/j.ceca.2012.01.004DOI Listing
May 2012

Fraternal twins: Swiprosin-1/EFhd2 and Swiprosin-2/EFhd1, two homologous EF-hand containing calcium binding adaptor proteins with distinct functions.

Cell Commun Signal 2011 Jan 18;9. Epub 2011 Jan 18.

Division of Molecular Immunology, Department of Medicine III, Nikolaus Fiebiger Center, University of Erlangen-Nürnberg, 91054 Erlangen, Germany.

Changes in the intracellular calcium concentration govern cytoskeletal rearrangement, mitosis, apoptosis, transcriptional regulation or synaptic transmission, thereby, regulating cellular effector and organ functions. Calcium binding proteins respond to changes in the intracellular calcium concentration with structural changes, triggering enzymatic activation and association with downstream proteins. One type of calcium binding proteins are EF-hand super family proteins. Here, we describe two recently discovered homologous EF-hand containing adaptor proteins, Swiprosin-1/EF-hand domain containing 2 (EFhd2) and Swiprosin-2/EF-hand domain containing 1 (EFhd1), which are related to allograft inflammatory factor-1 (AIF-1). For reasons of simplicity and concision we propose to name Swiprosin-1/EFhd2 and Swiprosin-2/EFhd1 from now on EFhd2 and EFhd1, according to their respective gene symbols. AIF-1 and Swiprosin-1/EFhd2 are already present in Bilateria, for instance in Drosophila melanogaster and Caenhorhabditis elegans. Swiprosin-2/EFhd1 arose later from gene duplication in the tetrapodal lineage. Secondary structure prediction of AIF-1 reveals disordered regions and one functional EF-hand. Swiprosin-1/EFhd2 and Swiprosin-2/EFhd1 exhibit a disordered region at the N-terminus, followed by two EF-hands and a coiled-coil domain. Whereas both proteins are similar in their predicted overall structure they differ in a non-homologous stretch of 60 amino acids just in front of the EF-hands. AIF-1 controls calcium-dependent cytoskeletal rearrangement in innate immune cells by means of its functional EF-hand. We propose that Swiprosin-1/EFhd2 as well is a cytoskeleton associated adaptor protein involved in immune and brain cell function. Pro-inflammatory conditions are likely to modulate expression and function of Swiprosin-1/EFhd2. Swiprosin-2/EFhd1, on the other hand, modulates apoptosis and differentiation of neuronal and muscle precursor cells, probably through an association with mitochondria. We suggest furthermore that Swiprosin-2/EFhd1 is part of a cellular response to oxidative stress, which could explain its pro-survival activity in neuronal, muscle and perhaps some malignant tissues.
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http://dx.doi.org/10.1186/1478-811X-9-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3036668PMC
January 2011

Swiprosin-1/EFhd2 controls B cell receptor signaling through the assembly of the B cell receptor, Syk, and phospholipase C gamma2 in membrane rafts.

J Immunol 2010 Apr 1;184(7):3665-76. Epub 2010 Mar 1.

Division of Molecular Immunology, Department of Medicine III, Nikolaus Fiebiger Center, University of Erlangen-Nürnberg, Erlangen, Germany.

Compartmentalization of the BCR in membrane rafts is important for its signaling capacity. Swiprosin-1/EFhd2 (Swip-1) is an EF-hand and coiled-coil-containing adaptor protein with predicted Src homology 3 (SH3) binding sites that we identified in membrane rafts. We showed previously that Swip-1 amplifies BCR-induced apoptosis; however, the mechanism of this amplification was unknown. To address this question, we overexpressed Swip-1 and found that Swip-1 amplified the BCR-induced calcium flux in WEHI231, B62.1, and Bal17 cells. Conversely, the BCR-elicited calcium flux was strongly attenuated in Swip-1-silenced WEHI231 cells, and this was due to a decreased calcium mobilization from intracellular stores. Complementation of Swip-1 expression in Swip-1-silenced WEHI231 cells restored the BCR-induced calcium flux and enhanced spleen tyrosine kinase (Syk) tyrosine phosphorylation and activity as well as SLP65/BLNK/BASH and phospholipase C gamma2 (PLCgamma2) tyrosine phosphorylation. Furthermore, Swip-1 induced the constitutive association of the BCR itself, Syk, and PLCgamma2 with membrane rafts. Concomitantly, Swip-1 stabilized the association of BCR with tyrosine-phosphorylated proteins, specifically Syk and PLCgamma2, and enhanced the constitutive interaction of Syk and PLCgamma2 with Lyn. Interestingly, Swip-1 bound to the rSH3 domains of the Src kinases Lyn and Fgr, as well as to that of PLCgamma. Deletion of the predicted SH3-binding region in Swip-1 diminished its association and that of Syk and PLCgamma2 with membrane rafts, reduced its interaction with the SH3 domain of PLCgamma, and diminished the BCR-induced calcium flux. Hence, Swip-1 provides a membrane scaffold that is required for the Syk-, SLP-65-, and PLCgamma2-dependent BCR-induced calcium flux.
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http://dx.doi.org/10.4049/jimmunol.0903642DOI Listing
April 2010

Swimming behaviour of Schistosoma mansoni cercariae: responses to irradiance changes and skin attractants.

Parasitol Res 2008 Mar 22;102(4):685-90. Epub 2007 Dec 22.

Institute for Biology, University Erlangen-Nuernberg, Staudtstr. 5, 91058 Erlangen, Germany.

The swimming behaviour of many cercarial species is governed by sensitive responses to light and dark stimuli. We studied the effect of irradiance changes on swimming behaviour of Schistosoma mansoni cercariae and found only insignificant responses. Decreasing light intensity results in a weak tendency of the cercariae to start swimming movements, and increasing light intensity tends to inhibit the start of swimming. These responses seem not suitable to increase the transmission success. Whether the cercariae show chemo-orientation towards human skin was studied by video-tracking their swimming movements around agar containing human-skin-surface extracts and when immersed into skin extracts. They showed no directed chemotactic orientation, as they did not correct their swimming paths in direction towards the skin-extract substrates, also not when shifting between forward and backward swimming. However, the cercariae shifted more between backward and forward swimming and therewith increased their rate of change of direction. This response may support an accumulation around the skin substrates and could guide the cercariae towards the host's skin surface when they are already in close proximity to it.
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http://dx.doi.org/10.1007/s00436-007-0812-4DOI Listing
March 2008