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Objectives: Dual incretin agonists are among the most effective pharmaceutical treatments for obesity and type 2 diabetes to date. Such therapeutics can target two receptors, such as the glucagon-like peptide-1 (GLP-1) receptor and the glucose-dependent insulinotropic polypeptide (GIP) receptor in the case of tirzepatide, to improve glycemia and reduce body weight. Regarding body weight effects, GIPR signaling is thought to involve at least two relevant mechanisms: the enhancement of food intake reduction and the attenuation of aversive effects caused by GLP-1R agonists. Although it is known that dual GLP-1R-GIPR agonism produces greater weight loss than GLP-1R agonism alone, the precise mechanism is unknown., Methods: To address this question, we used mice lacking GIPR in the whole body, GABAergic neurons, or glutamatergic neurons. These mice were given various combinations of GLP-1R and GIPR agonist drugs with subsequent food intake and conditioned taste aversion measurements., Results: A GIPR knockout in either the whole body or selectively in inhibitory GABAergic neurons protects against diet-induced obesity, whereas a knockout in excitatory glutamatergic neurons had a negligible effect. Furthermore, we found that GIPR in GABAergic neurons is essential for the enhanced weight loss efficacy of dual incretin agonism, yet, surprisingly, its removal enhances the effect of GLP-1R agonism alone. Finally, GIPR knockout in GABAergic neurons prevents the anti-aversive effects of GIPR agonism., Conclusions: Our findings are consistent with GIPR research at large in that both enhancement and removal of GIPR signaling are metabolically beneficial. Notably, however, our findings suggest that future obesity therapies designed to modulate GIPR signaling, whether by agonism or antagonism, would be best targeted towards GABAergic neurons., Competing Interests: Declaration of competing interest R.J.S. has received research support from Novo Nordisk, Fractyl, Astra Zeneca, Congruence Therapeutics, Eli Lilly, Bullfrog AI, Glyscend Therapeutics and Amgen. RJS has served as a paid consultant for Novo Nordisk, Eli Lilly, CinRx, Fractyl, Structure Therapeutics, Crinetics, Amgen and Congruence Therapeutics. R.J.S. has equity in Bullfrog AI and Rewind. R.L., S.W., and C.R. are employees of AstraZeneca. DJD has received remuneration for consulting or speaking from Amgen, AstraZeneca, Boerhinger Ingelheim Kallyope, Merck, Novo Nordisk, Pfizer Inc and Zeleand Pharma and Mt. Sinai Hospital receives investigator-initiated grant support for studies in the Drucker lab from Amgen, Novo Nordisk, Pfizer and Zealand Pharma Inc., (Copyright © 2024. Published by Elsevier GmbH.)

Objectives: Growth differentiation factor 15 (GDF15) acts on the receptor dimer of GDNF family receptor alpha-like (GFRAL) and Rearranged during transfection (RET). While Gfral-expressing cells are known to be present in the area postrema and nucleus of the solitary tract (AP/NTS) located in the brainstem, the presence of Gfral- expressing cells in other sites within the central nervous system and peripheral tissues is not been fully addressed. Our objective was to thoroughly investigate whether GFRAL is expressed in peripheral tissues and in brain sites different from the brainstem., Methods: From Gfral::eGFP mice we collected tissue from 12 different tissues, including brain, and used single molecule in-situ hybridizations to identify cells within those tissues expressing Gfral. We then contrasted the results with human Gfral-expression by analyzing publicly available single-cell RNA sequencing data., Results: In mice we found readably detectable Gfral mRNA within the AP/NTS but not within other brain sites. Within peripheral tissues, we failed to detect any Gfral-labelled cells in the vast majority of examined tissues and when present, were extremely rare. Single cell sequencing of human tissues confirmed GFRAL-expressing cells are detectable in some sites outside the AP/NTS in an extremely sparse manner. Importantly, across the utilized methodologies, smFISH, genetic Gfral reporter mice and scRNA-Seq, we failed to detect Gfral-labelled cells with all three., Conclusions: Through highly sensitive and selective technologies we show Gfral expression is overwhelmingly restricted to the brainstem and expect that GDF15 and GFRAL-based therapies in development for cancer cachexia will specifically target AP/NTS cells., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

The combination of decreasing food intake and increasing energy expenditure represents a powerful strategy for counteracting cardiometabolic diseases such as obesity and type 2 diabetes 1 . Yet current pharmacological approaches require conjugation of multiple receptor agonists to achieve both effects 2-4 , and so far, no safe energy-expending option has reached the clinic. Here we show that activation of neurokinin 2 receptor (NK2R) is sufficient to suppress appetite centrally and increase energy expenditure peripherally. We focused on NK2R after revealing its genetic links to obesity and glucose control. However, therapeutically exploiting NK2R signalling has previously been unattainable because its endogenous ligand, neurokinin A, is short-lived and lacks receptor specificity 5,6 . Therefore, we developed selective, long-acting NK2R agonists with potential for once-weekly administration in humans. In mice, these agonists elicit weight loss by inducing energy expenditure and non-aversive appetite suppression that circumvents canonical leptin signalling. Additionally, a hyperinsulinaemic-euglycaemic clamp reveals that NK2R agonism acutely enhances insulin sensitization. In diabetic, obese macaques, NK2R activation significantly decreases body weight, blood glucose, triglycerides and cholesterol, and ameliorates insulin resistance. These findings identify a single receptor target that leverages both energy-expending and appetite-suppressing programmes to improve energy homeostasis and reverse cardiometabolic dysfunction across species., Competing Interests: Competing interests: T.M., J.H.E., J.B.H., D.P.C., M.B.F.G., M.T.-C., T.W.S. and Z.G.-H. work or have worked in some capacity for Embark Laboratories ApS, a company developing therapeutics for the treatment of T2D and obesity. P.K. is a consultant for Embark Laboratories ApS. The use of and chemical composition for NK2R agonists are patented by the University of Copenhagen and Embark Laboratories ApS, respectively. R.J.S. has received research support from Novo Nordisk, Fractyl, AstraZeneca, Congruence Therapeutics, Eli Lilly, Bullfrog AI, Glycsend Therapeutics and Amgen. R.J.S. has served as a paid consultant for Novo Nordisk, Eli Lilly, CinRx, Fractyl, Structure Therapeutics, Crinetics and Congruence Therapeutics. R.J.S. has equity in Calibrate, Rewind and Levator Therapeutics. The other authors declare no competing interests., (© 2024. The Author(s).)

Objective: Optimal weight loss involves decreasing adipose tissue while preserving lean muscle mass. Identifying molecular mediators that preserve lean muscle mass is therefore a clinically important goal. We have shown that circulating, postprandial FGF19 levels are lower in patients with obesity and decrease further with comorbidities such as type 2 diabetes and MASLD. Preclinical studies have shown that FGF15 (mouse ortholog of human FGF19) is necessary to protect against lean muscle mass loss following metabolic surgery-induced weight loss in a mouse model of diet-induced obesity. We evaluated if non-surgical weight loss interventions also lead to increased systemic levels of FGF19 and whether FGF19 levels are predictive of lean muscle mass following rapid weight loss in human subjects with obesity., Research Design and Methods: Weight loss was induced in 176 subjects with obesity via a very low-energy diet, VLED (800 kcal/d) in the form of total liquid meal replacement for 3-4 months. We measured plasma FGF19 levels at baseline and following VLED-induced weight loss. Multiple linear regression was performed to assess if FGF19 levels were predictive of lean mass at baseline (obesity) and following VLED., Results: Postprandial levels of FGF19 increased significantly following VLED-weight loss. Multiple linear regression analysis showed that baseline (obesity) FGF19 levels, but not post VLED FGF19 levels, significantly predicted the percent of lean muscle mass after VLED-induced weight loss, while controlling for age, sex, and the baseline percent lean mass., Conclusion: These data identify gut-muscle communication and FGF19 as a potentially important mediator of the preservation of lean muscle mass during rapid weight loss., Competing Interests: Declaration of competing interest RJS has received research support from Novo Nordisk, Fractyl, Astra Zeneca, Congruence Therapeutics, Eli Lilly, Bullfrog AI, Glycsend Therapeutics and Amgen. RJS has served as a paid consultant for Novo Nordisk, Eli Lilly, CinRx, Fractyl, Structure Therapeutics, Crinetics and Congruence Therapeutics. RJS has equity in Calibrate, Rewind and Levator Therapeutics. AER receives salary support from Rewind. JHS is a paid employee of Amgen Inc.The remaining authors declare no competing interests., (Copyright © 2024. Published by Elsevier Masson SAS.)

Classic hereditary hemochromatosis (HH) is an autosomal recessive iron-overload disorder resulting from loss-of-function mutations of the HFE gene. Patients with HH exhibit excessive hepatic iron accumulation that predisposes these patients to liver disease, including the risk for developing liver cancer. Chronic iron overload also poses a risk for the development of metabolic disorders such as obesity, type 2 diabetes, and insulin resistance. We hypothesized that liraglutide, GLP1 receptor agonist, alters iron metabolism while also reducing body weight and glucose tolerance in a mouse model of HH (global HFE knockout, HFE KO) and diet-induced obesity and glucose intolerance. The total body HFE KO and wild-type control mice were fed high-fat diet for 8 weeks. Mice were subdivided into liraglutide and vehicle-treated groups and received daily subcutaneous administration of the respective treatment once daily for 18 weeks. Liraglutide improved glucose tolerance and hepatic lipid markers and reduced body weight in a mouse model of HH, the HFE KO mouse, similar to wild-type controls. Importantly, our data show that liraglutide alters iron metabolism in HFE KO mice, leading to decreased circulating and stored iron levels in HFE KO mice. These observations highlight the potential that GLP1 receptor agonist could be used to reduce iron overload in addition to reducing body weight and improving glucose regulation in HH patients., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com. See the journal About page for additional terms.)

Fibroblast growth factor 15/19 (FGF15/19, mouse/human ortholog) is expressed in the ileal enterocytes of the small intestine and released postprandially in response to bile acid absorption. Previous reports of FGF15-/- mice have limited our understanding of gut-specific FGF15's role in metabolism. Therefore, we studied the role of endogenous gut-derived FGF15 in bile acid, cholesterol, glucose, and energy balance. We found that circulating levels of FGF19 were reduced in individuals with obesity and comorbidities, such as type 2 diabetes and metabolic dysfunction-associated fatty liver disease. Gene expression analysis of ileal FGF15-positive cells revealed differential expression during the obesogenic state. We fed standard chow or a high-fat metabolic dysfunction-associated steatohepatitis-inducing diet to control and intestine-derived FGF15-knockout (FGF15INT-KO) mice. Control and FGF15INT-KO mice gained similar body weight and adiposity and did not show genotype-specific differences in glucose, mixed meal, pyruvate, and glycerol tolerance. FGF15INT-KO mice had increased systemic bile acid levels but decreased cholesterol levels, pointing to a primary role for gut-derived FGF15 in regulating bile acid and cholesterol metabolism when exposed to obesogenic diet. These studies show that intestinal FGF15 plays a specific role in bile acid and cholesterol metabolism regulation but is not essential for energy and glucose balance.

Glucagon-like peptide-1 receptor agonists (GLP-1RAs) exert anti-inflammatory effects relevant to the chronic complications of type 2 diabetes. Although GLP-1RAs attenuate T cell-mediated gut and systemic inflammation directly through the gut intraepithelial lymphocyte GLP-1R, how GLP-1RAs inhibit systemic inflammation in the absence of widespread immune expression of the GLP-1R remains uncertain. Here, we show that GLP-1R activation attenuates the induction of plasma tumor necrosis factor alpha (TNF-α) by multiple Toll-like receptor agonists. These actions are not mediated by hematopoietic or endothelial GLP-1Rs but require central neuronal GLP-1Rs. In a cecal slurry model of polymicrobial sepsis, GLP-1RAs similarly require neuronal GLP-1Rs to attenuate detrimental responses associated with sepsis, including sickness, hypothermia, systemic inflammation, and lung injury. Mechanistically, GLP-1R activation leads to reduced TNF-α via α 1 -adrenergic, δ-opioid, and κ-opioid receptor signaling. These data extend emerging concepts of brain-immune networks and posit a new gut-brain GLP-1R axis for suppression of peripheral inflammation., Competing Interests: Declaration of interests D.J.D. has consulted for Altimmune, Amgen, Kallyope, Merck, Novo Nordisk, Pfizer Inc., and Sanofi Inc. Mt. Sinai Hospital receives operating grant support from Amgen, Novo Nordisk, and Pfizer Inc. for studies in the Drucker lab. R.J.S. has consulted for Novo Nordisk, Scohia, CinRx, Fractyl, Structure Therapeutics, Congruenc Therapeutics, Calibrate, and Rewind and receives operating grant support from Novo Nordisk, Astra Zeneca, Fractyl, and Eli Lilly Inc., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Obesity is a chronic disease that affects more than 650 million adults worldwide. Obesity not only is a significant health concern on its own, but predisposes to cardiometabolic comorbidities, including coronary heart disease, dyslipidemia, hypertension, type 2 diabetes, and some cancers. Lifestyle interventions effectively promote weight loss of 5% to 10%, and pharmacological and surgical interventions even more, with some novel approved drugs inducing up to an average of 25% weight loss. Yet, maintaining weight loss over the long-term remains extremely challenging, and subsequent weight gain is typical. The mechanisms underlying weight regain remain to be fully elucidated. The purpose of this Pennington Biomedical Scientific Symposium was to review and highlight the complex interplay between the physiological, behavioral, and environmental systems controlling energy intake and expenditure. Each of these contributions were further discussed in the context of weight-loss maintenance, and systems-level viewpoints were highlighted to interpret gaps in current approaches. The invited speakers built upon the science of obesity and weight loss to collectively propose future research directions that will aid in revealing the complicated mechanisms involved in the weight-reduced state., (© 2023 The Obesity Society.)

We previously reported that mice lacking the protein optic atrophy 1 (OPA1 BKO) in brown adipose tissue (BAT) display induction of the activating transcription factor 4 (ATF4), which promotes fibroblast growth factor 21 (FGF21) secretion as a batokine. FGF21 increases metabolic rates under baseline conditions but is dispensable for the resistance to diet-induced obesity (DIO) reported in OPA1 BKO mice (Pereira et al., 2021). To determine alternative mediators of this phenotype, we performed transcriptome analysis, which revealed increased levels of growth differentiation factor 15 (GDF15), along with increased protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) levels in BAT. To investigate whether ATF4 induction was mediated by PERK and evaluate the contribution of GDF15 to the resistance to DIO, we selectively deleted PERK or GDF15 in OPA1 BKO mice. Mice with reduced OPA1 and PERK levels in BAT had preserved ISR activation. Importantly, simultaneous deletion of OPA1 and GDF15 partially reversed the resistance to DIO and abrogated the improvements in glucose tolerance. Furthermore, GDF15 was required to improve cold-induced thermogenesis in OPA1 BKO mice. Taken together, our data indicate that PERK is dispensable to induce the ISR, but GDF15 contributes to the resistance to DIO, and is required for glucose homeostasis and thermoregulation in OPA1 BKO mice by increasing energy expenditure., Competing Interests: JJ, LG, EW, AM, SB, KK, JK, JC, EA, RP No competing interests declared, RS R.J.S. has received research support from Fractyl, Novo Nordisk, AstraZeneca, and Eli Lilly. R.J.S. has served on scientific advisory boards for Novo Nordisk, CinRX, Scohia, Fractyl and Structure Therapeutics. R.J.S. is a stakeholder of Calibrate and Rewind, (© 2023, Jena, García-Peña et al.)

The adipose-derived hormone leptin acts via its receptor (LepRb) in the brain to control energy balance. A potentially unidentified population of GABAergic hypothalamic LepRb neurons plays key roles in the restraint of food intake and body weight by leptin. To identify markers for candidate populations of LepRb neurons in an unbiased manner, we performed single-nucleus RNA-Seq of enriched mouse hypothalamic LepRb cells, identifying several previously unrecognized populations of hypothalamic LepRb neurons. Many of these populations displayed strong conservation across species, including GABAergic Glp1r-expressing LepRb (LepRbGlp1r) neurons, which expressed more Lepr than other LepRb cell populations. Ablating Lepr from LepRbGlp1r cells provoked hyperphagic obesity without impairing energy expenditure. Similarly, improvements in energy balance caused by Lepr reactivation in GABA neurons of otherwise Lepr-null mice required Lepr expression in GABAergic Glp1r-expressing neurons. Furthermore, restoration of Glp1r expression in LepRbGlp1r neurons in otherwise Glp1r-null mice enabled food intake suppression by the GLP1R agonist, liraglutide. Thus, the conserved GABAergic LepRbGlp1r neuron population plays crucial roles in the suppression of food intake by leptin and GLP1R agonists.

Obesity promotes triple-negative breast cancer (TNBC), and effective interventions are urgently needed to break the obesity-TNBC link. Epidemiologic studies indicate that bariatric surgery reduces TNBC risk, while evidence is limited or conflicted for weight loss via low-fat diet (LFD) or calorie restriction (CR). Using a murine model of obesity-driven TNBC, we compared the antitumor effects of vertical sleeve gastrectomy (VSG) with LFD, chronic CR, and intermittent CR. Each intervention generated weight and fat loss and suppressed tumor growth relative to obese mice (greatest suppression with CR). VSG and CR regimens exerted both similar and unique effects, as assessed using multiomics approaches, in reversing obesity-associated transcript, epigenetics, secretome, and microbiota changes and restoring antitumor immunity. Thus, in a murine model of TNBC, bariatric surgery and CR each reverse obesity-driven tumor growth via shared and distinct antitumor mechanisms, and CR is superior to VSG in reversing obesity's procancer effects.

Growth differentiation factor 15 (GDF15) induces weight loss and increases insulin action in obese rodents. Whether and how GDF15 improves insulin action without weight loss is unknown. Obese rats were treated with GDF15 and displayed increased insulin tolerance 5 h later. Lean and obese female and male mice were treated with GDF15 on days 1, 3, and 5 without weight loss and displayed increased insulin sensitivity during a euglycemic hyperinsulinemic clamp on day 6 due to enhanced suppression of endogenous glucose production and increased glucose uptake in WAT and BAT. GDF15 also reduced glucagon levels during clamp independently of the GFRAL receptor. The insulin-sensitizing effect of GDF15 was completely abrogated in GFRAL KO mice and also by treatment with the β-adrenergic antagonist propranolol and in β1,β2-adrenergic receptor KO mice. GDF15 activation of the GFRAL receptor increases β-adrenergic signaling, in turn, improving insulin action in the liver and white and brown adipose tissue., Competing Interests: Declaration of interests S.B.J. is an employee of Novo Nordisk, a pharmaceutical company producing and selling medicine for the treatment of diabetes and obesity. R.J.S. has received research support from Novo Nordisk, Fractyl, Astra Zeneca, and Eli Lilly. R.J.S. has served as a paid consultant for Novo Nordisk, Eli Lilly, Scohia, CinRx, Fractyl, Structure Therapeutics, and Congruence Therapeutics. R.J.S. has equity in Calibrate and Rewind., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Regenerating family member gamma, Reg3γ (the mouse homolog of human REG3A), belonging to the antimicrobial peptides (AMPs), functions as a part of the host immune system to maintain spatial segregation between the gut bacteria and the host in the intestine via bactericidal activity. There is emerging evidence that gut manipulations such as bariatric surgery, dietary supplementation or drug treatment to produce metabolic benefits alter the gut microbiome. In addition to changes in a wide range of gut hormones, these gut manipulations also induce the expression of Reg3γ in the intestine. Studies over the past decades have revealed that Reg3γ not only plays a role in the gut lumen but can also contribute to host physiology through interaction with the gut microbiota. Herein, we discuss the current knowledge regarding the biology of Reg3γ, its role in various metabolic functions, and new opportunities for therapeutic strategies to treat metabolic disorders., (© 2023. The Author(s).)

Exercise is a first-line treatment for type 2 diabetes and preserves β-cell function by hitherto unknown mechanisms. We postulated that proteins from contracting skeletal muscle may act as cellular signals to regulate pancreatic β-cell function. We used electric pulse stimulation (EPS) to induce contraction in C2C12 myotubes and found that treatment of β-cells with EPS-conditioned medium enhanced glucose-stimulated insulin secretion (GSIS). Transcriptomics and subsequent targeted validation revealed growth differentiation factor 15 (GDF15) as a central component of the skeletal muscle secretome. Exposure to recombinant GDF15 enhanced GSIS in cells, islets, and mice. GDF15 enhanced GSIS by upregulating the insulin secretion pathway in β-cells, which was abrogated in the presence of a GDF15 neutralizing antibody. The effect of GDF15 on GSIS was also observed in islets from GFRAL-deficient mice. Circulating GDF15 was incrementally elevated in patients with pre- and type 2 diabetes and positively associated with C-peptide in humans with overweight or obesity. Six weeks of high-intensity exercise training increased circulating GDF15 concentrations, which positively correlated with improvements in β-cell function in patients with type 2 diabetes. Taken together, GDF15 can function as a contraction-induced protein that enhances GSIS through activating the canonical signaling pathway in a GFRAL-independent manner., Article Highlights: Exercise improves glucose-stimulated insulin secretion through direct interorgan communication. Contracting skeletal muscle releases growth differentiation factor 15 (GDF15), which is required to synergistically enhance glucose-stimulated insulin secretion. GDF15 enhances glucose-stimulated insulin secretion by activating the canonical insulin release pathway. Increased levels of circulating GDF15 after exercise training are related to improvements in β-cell function in patients with type 2 diabetes., (© 2023 by the American Diabetes Association.)

Objective: Glucagon-like peptide-1 receptor (GLP-1R) agonists (GLP-1RA) and fibroblast growth factor-21 (FGF21) confer similar metabolic benefits. GLP-1RA induce FGF21, leading us to investigate mechanisms engaged by the GLP-1RA liraglutide to increase FGF21 levels and the metabolic relevance of liraglutide-induced FGF21., Methods: Circulating FGF21 levels were measured in fasted male C57BL/6J, neuronal GLP-1R knockout, β-cell GLP-1R knockout, and liver peroxisome proliferator-activated receptor alpha knockout mice treated acutely with liraglutide. To test the metabolic relevance of liver FGF21 in response to liraglutide, chow-fed control and liver Fgf21 knockout (Liv Fgf21-/- ) mice were treated with vehicle or liraglutide in metabolic chambers. Body weight and composition, food intake, and energy expenditure were measured. Since FGF21 reduces carbohydrate intake, we measured body weight in mice fed matched diets with low- (LC) or high-carbohydrate (HC) content and in mice fed a high-fat, high-sugar (HFHS) diet. This was done in control and Liv Fgf21-/- mice and in mice lacking neuronal β-klotho (Klb) expression to disrupt brain FGF21 signaling., Results: Liraglutide increases FGF21 levels independently of decreased food intake via neuronal GLP-1R activation. Lack of liver Fgf21 expression confers resistance to liraglutide-induced weight loss due to attenuated reduction of food intake in chow-fed mice. Liraglutide-induced weight loss was impaired in Liv Fgf21-/- mice when fed HC and HFHS diets but not when fed a LC diet. Loss of neuronal Klb also attenuated liraglutide-induced weight loss in mice fed HC or HFHS diets., Conclusions: Our findings support a novel role for a GLP-1R-FGF21 axis in regulating body weight in a dietary carbohydrate-dependent manner., (Copyright © 2023 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Vertical sleeve gastrectomy (VSG), the most utilized bariatric procedure in clinical practice, greatly reduces body weight and improves a variety of metabolic disorders. However, one of its long-term complications is bone loss and increased risk of fracture. Elevated circulating sclerostin (SOST) and granulocyte-colony stimulating factor (G-CSF) concentrations have been considered as potential contributors to VSG-associated bone loss. To test these possibilities, we administrated antibodies to SOST or G-CSF receptor and investigated alterations to bone and marrow niche following VSG. Neutralizing either SOST or G-CSF receptor did not alter beneficial effects of VSG on adiposity and hepatic steatosis, and anti-SOST treatment provided a further improvement to glucose tolerance. SOST antibodies partially reduced trabecular and cortical bone loss following VSG by increasing bone formation, whereas G-CSF receptor antibodies had no effects on bone mass. The expansion in myeloid cellularity and reductions in bone marrow adiposity seen with VSG were partially eliminated by treatment with Anti-G-CSF receptor. Taken together, these experiments demonstrate that antibodies to SOST or G-CSF receptor may act through independent mechanisms to partially block effects of VSG on bone loss or marrow niche cells, respectively., Competing Interests: Declaration of competing interest RJS has received research support from Novo Nordisk, AstraZeneca and Fractyl. RJS has served as a paid consultant to Novo Nordisk, Scohia, CinRx, Structure Therapeutics and has equity in Calibrate and Rewind. SJH and ANE hold stock in Regeneron. OAM has received grant support from Regeneron Pharmaceuticals, Inc., CombiGene AB, and Rejuvenate Bio. All other authors have declared that no conflicts of interest exist., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Deoxynivalenol (DON), a type B trichothecene mycotoxin contaminating grains, promotes nausea, emesis and anorexia. With DON exposure, circulating levels of intestinally derived satiation hormones, including glucagon-like peptide 1 (GLP-1) are elevated. To directly test whether GLP-1 signaling mediates the effects of DON, we examined the response of GLP-1 or GLP-1R-deficient mice to DON injection. We found comparable anorectic and conditioned taste avoidance learning responses in GLP-1/GLP-1R deficient mice compared to control littermates, suggesting that GLP-1 is not necessary for the effects of DON on food intake and visceral illness. We then used our previously published data from translating ribosome affinity purification with RNA sequencing (TRAP-seq) analysis of area postrema neurons that express the receptor for the circulating cytokine growth differentiation factor (GDF15), growth differentiation factor a-like (GFRAL). Interestingly, this analysis showed that a cell surface receptor for DON, calcium sensing receptor (CaSR), is heavily enriched in GFRAL neurons. Given that GDF15 potently reduces food intake and can cause visceral illness by signaling through GFRAL neurons, we hypothesized that DON may also signal by activating CaSR on GFRAL neurons. Indeed, circulating GDF15 levels are elevated after DON administration but both GFRAL knockout and GFRAL neuron-ablated mice exhibited similar anorectic and conditioned taste avoidance responses compared to WT littermates. Thus, GLP-1 signaling and GFRAL signaling and neurons are not required for DON-induced visceral illness or anorexia.

Objective: To compare the proteomic composition of follicular fluid from women with normal weight vs. women with obesity but without a history of polycystic ovary syndrome or known ovarian dysfunction undergoing in vitro fertilization., Design: Cross-sectional., Setting: Academic medical center., Patient(s): Eight women with normal weight and 8 women with obesity undergoing in vitro fertilization and without a history of polycystic ovary syndrome, ovulatory dysfunction, diminished ovarian reserve, or known endometriosis were included in the analysis., Intervention(s): Not applicable., Main Outcome Measure(s): Proteomic assessment using liquid chromatography-mass spectrometry analysis., Result(s): The mean age of women with normal weight was similar to that of women with obesity (32.9 vs. 32.6 years, not significant). The mean body mass index of women with normal weight was 21.2 kg/m 2 compared with a body mass index of 37.1 kg/m 2 in women with obesity. A total of 1,174 proteins were identified with ≥2 peptides present. Twenty-five proteins were found to be significantly altered in the follicular fluid from women with obesity. Of these 25 proteins, 19 were up-regulated and 6 were down-regulated. Notably, C-reactive protein was 11-fold higher in the follicular fluid from women with obesity than in the follicular fluid from women with normal weight., Conclusion(s): Obesity is associated with dysregulation at the level of the follicle, including alterations in proteins related to inflammation and metabolism. These include proteins with emerging roles in energy homeostasis and follicular regulation., (Copyright © 2022 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Changing composition of the gut microbiome is an important component of the gut adaptation to various environments, which have been implicated in various metabolic diseases including obesity and type 2 diabetes, but the mechanisms by which the microbiota influence host physiology remain contentious. Here we find that both diets high in the fermentable fiber inulin and vertical sleeve gastrectomy increase intestinal expression and circulating levels of the anti-microbial peptide Reg3g. Moreover, a number of beneficial effects of these manipulations on gut function, energy balance, and glucose regulation are absent in Reg3g knockout mice. Peripheral administration of various preparations of Reg3g improves glucose tolerance, and this effect is dependent on the putative receptor Extl3 in the pancreas. These data suggest Reg3g acts both within the lumen and as a gut hormone to link the intestinal microbiome to various aspects of host physiology that may be leveraged for novel treatment strategies., Competing Interests: Declaration of interests R.J.S. has received research support from Novo Nordisk, Astra Zeneca, Pfizer, and Fractyl. R.J.S has served as a paid consultant and/or scientific advisory board member to Novo Nordisk, Scohia, Fractyle and ShouTi Pharma. R.J.S. has equity or option positions in Rewind and Calibrate Health. S.L.-A. is a paid employee of Novo Nordisk., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Gut intraepithelial lymphocytes (IELs) are thought to calibrate glucagon-like peptide 1 (GLP-1) bioavailability, thereby regulating systemic glucose and lipid metabolism. Here, we show that the gut IEL GLP-1 receptor (GLP-1R) is not required for enteroendocrine L cell GLP-1 secretion and glucose homeostasis nor for the metabolic benefits of GLP-1R agonists (GLP-1RAs). Instead, the gut IEL GLP-1R is essential for the full effects of GLP-1RAs on gut microbiota. Moreover, independent of glucose control or weight loss, the anti-inflammatory actions of GLP-1RAs require the gut IEL GLP-1R to selectively restrain local and systemic T cell-induced, but not lipopolysaccharide-induced, inflammation. Such effects are mediated by the suppression of gut IEL effector functions linked to the dampening of proximal T cell receptor signaling in a protein-kinase-A-dependent manner. These data reposition key roles of the L cell-gut IEL GLP-1R axis, revealing mechanisms linking GLP-1R activation in gut IELs to modulation of microbiota composition and control of intestinal and systemic inflammation., Competing Interests: Declaration of interests D.J.D. has served as a speaker or consultant for Altimmune, Amgen, AMT Inc., Eli Lilly Inc., Kallyope Inc., Merck Inc., Novo Nordisk Inc., and Pfizer Inc. R.J.S. serves as a paid consultant for Novo Nordisk, Scohia, Fractyl, and ShouTi. R.J.S. has equity positions in Calibrate and Rewind Health., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Metformin is a blood-glucose-lowering medication with physiological effects that extend beyond its anti-diabetic indication. Recently, it was reported that metformin lowers body weight via induction of growth differentiation factor 15 (GDF15), which suppresses food intake by binding to the GDNF family receptor α-like (GFRAL) in the hindbrain. Here, we corroborate that metformin increases circulating GDF15 in mice and humans, but we fail to confirm previous reports that the GDF15-GFRAL pathway is necessary for the weight-lowering effects of metformin. Instead, our studies in wild-type, GDF15 knockout, and GFRAL knockout mice suggest that the GDF15-GFRAL pathway is dispensable for the effects of metformin on energy balance. The data presented here question whether metformin is a sufficiently strong stimulator of GDF15 to drive anorexia and weight loss and emphasize that additional work is needed to untangle the relationship among metformin, GDF15, and energy balance., Competing Interests: Declaration of interests A.B.K. and C.C. are co-founders of Ousia Pharma ApS, a biotech company developing therapeutics for obesity. J.S.Q. and K.F. have received research funding from Novo Nordisk. R.J.S. has received research support from Novo Nordisk and Astra Zeneca. R.J.S. has served as a paid consultant for Novo Nordisk, Scohia, Fractyl, and ShouTi Pharma. R.J.S. has equity positions in Calibrate and Rewind., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Body weight and adiposity represent biologically controlled parameters that are influenced by a combination of genetic, developmental and environmental variables. Although the hypothalamus plays a crucial role in matching caloric intake with energy expenditure to achieve a stable body weight, it is now recognized that neuronal circuits in the hindbrain not only serve to produce nausea and to terminate feeding in response to food consumption or during pathological states, but also contribute to the long-term control of body weight. Additionally, recent work has identified hindbrain neurons that are capable of suppressing food intake without producing aversive responses like those associated with nausea. Here we review recent advances in our understanding of the hindbrain neurons that control feeding, particularly those located in the area postrema and the nucleus tractus solitarius. We frame this information in the context of new atlases of hindbrain neuronal populations and develop a model of the hindbrain circuits that control food intake and energy balance, suggesting important areas for additional research., (© 2022. Springer Nature Limited.)

Glucagon-like peptide-1 (GLP-1) regulates energy homeostasis via activation of the GLP-1 receptors (GLP-1Rs) in the central nervous system. However, the mechanism by which the central GLP-1 signal controls blood glucose levels, especially in different nutrient states, remains unclear. Here, we defined a population of glucose-sensing GLP-1R neurons in the dorsomedial hypothalamic nucleus (DMH), by which endogenous GLP-1 decreases glucose levels via the cross-talk between the hypothalamus and pancreas. Specifically, we illustrated the sufficiency and necessity of DMH GLP-1R in glucose regulation. The activation of the DMH GLP-1R neurons is mediated by a cAMP-PKA-dependent inhibition of a delayed rectifier potassium current. We also dissected a descending control of DMH GLP-1R -dorsal motor nucleus of the vagus nerve (DMV)-pancreas activity that can regulate glucose levels by increasing insulin release. Thus, our results illustrate how central GLP-1 action in the DMH can induce a nutrient state-dependent reduction in blood glucose level.

Vertical sleeve gastrectomy (VSG) results in an increase in the number of hormone-secreting enteroendocrine cells (EECs) in the intestinal epithelium; however, the mechanism remains unclear. Notably, the beneficial effects of VSG are lost in a mouse model lacking the nuclear bile acid receptor farnesoid X receptor (FXR). FXR is a nuclear transcription factor that has been shown to regulate intestinal stem cell (ISC) function in cancer models. Therefore, we hypothesized that the VSG-induced increase in EECs is due to changes in intestinal differentiation driven by an increase in bile acid signaling through FXR. To test this, we performed VSG in mice that express EGFP in ISC/progenitor cells and performed RNA-Seq on GFP-positive cells sorted from the intestinal epithelia. We also assessed changes in EEC number (marked by glucagon-like peptide-1, GLP-1) in mouse intestinal organoids following treatment with bile acids, an FXR agonist, and an FXR antagonist. RNA-Seq of ISCs revealed that bile acid receptors are expressed in ISCs and that VSG explicitly alters expression of several genes that regulate EEC differentiation. Mouse intestinal organoids treated with bile acids and 2 different FXR agonists increased GLP-1-positive cell numbers, and administration of an FXR antagonist blocked these effects. Taken together, these data indicate that VSG drives ISC fate toward EEC differentiation through bile acid signaling.

Objective: Vertical Sleeve Gastrectomy (VSG) is one of the most efficacious treatments for obesity and its comorbidities. Although a range of evidence suggests that alterations of the microbiota in the distal gut following VSG are pivotal to these metabolic improvements, the effect of surgery to alter the microbiota of the proximal intestine and its effect on host physiology remain largely unknown. As the main bacteria in the upper small intestine, Lactobacillus subspecies have been appreciated as important regulators of gut function. These bacteria also regulate intestinal Hypoxia- Inducible Factor 2α (HIF2α) signaling that plays an integral role in gut physiology and iron absorption. In the present study, we sought to determine the impact of VSG on Lactobacillus spp. in the small intestine and potential downstream impacts of Lactobacillus spp. on HIF2α, specifically in the duodenum., Methods: To determine the effects of VSG on the microbiota and HIF2α signaling in the duodenum, VSG surgeries were performed on diet-induced obese mice. To further probe the relationship between Lactobacillus spp. and HIF2α signaling in the duodenum, we applied a customized high-fat but iron-deficient diet on mice to increase duodenal HIF2α signaling and determined alterations of gut bacteria. To explore the causal role of Lactobacillus spp. in duodenal HIF2α signaling activation, we chronically administered probiotics containing Lactobacillus spp. to high-fat-fed obese mice. Lastly, we studied the effect of lactate, the major metabolite of Lactobacilli, on HIF2α in ex vivo duodenal organoids., Results: There were pronounced increases in the abundance of Lactobacillus spp. in samples isolated from duodenal epithelium in VSG-operated mice as compared to sham-operated mice. This was accompanied by an increase in the expression of genes that are targets of HIF2α in the duodenum of VSG-treated mice. Activating HIF2α signaling with a high-fat but iron-deficient diet resulted in weight loss, improvements in glucose regulation, and increased Lactobacillus spp. richness in the duodenum as compared to mice on an iron-replete diet. Chronic administration of probiotics containing Lactobacillus spp. not only increased HIF2α signaling in the duodenum such as occurs after VSG but also resulted in reduced weight gain and improved glucose tolerance in high-fat-fed mice. Furthermore, lactate was able to activate HIF2α in ex vivo duodenal organoids., Conclusions: These results support a model whereby VSG increases duodenal Lactobacillus richness and potentially stimulates intestinal HIF2α signaling via increased lactate production., (Copyright © 2022 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

DNA methylation is dynamically regulated in metabolic diseases, but it remains unclear whether the changes are causal or consequential. Therefore, we used a longitudinal approach to refine the onset of metabolic and DNA methylation changes at high temporal resolution. Male C57BL/6N mice were fed with 60 % high-fat diet (HFD) for up to 12 weeks and metabolically characterized weekly. Liver was collected after 1, 2, 4, 5, 6, 7, 8, and 12 weeks and hepatic DNA methylation and gene expression were analyzed. A subset of obese mice underwent vertical sleeve gastrectomy (VSG) or metformin treatment and livers were studied. Distinct hepatic gene expression patterns developed upon feeding HFD, with genes from the fatty acid metabolism pathway being predominantly altered. When comparing metabolic data with gene expression and DNA methylation, in particular Fgf21 DNA methylation decreased before the onset of increased Fgf21 expression and metabolic changes. Neither weight loss induced by VSG nor improved glucose tolerance by metformin treatment could revert hepatic Fgf21 DNA methylation or expression. Our data emphasize the dynamic induction of DNA methylation upon metabolic stimuli. Reduced Fgf21 DNA methylation established before massive overexpression of Fgf21, which is likely an adaptive effort of the liver to maintain glucose homeostasis despite the developing insulin resistance and steatosis. Fgf21 DNA methylation resisted reversion by intervention strategies, illustrating the long-term effects of unhealthy lifestyle. Our data provide a temporal roadmap to the development of hepatic insulin resistance, comprehensively linking DNA methylation with gene expression and metabolic data., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Growth differentiation factor 15 (GDF15), a TGFβ superfamily cytokine, acts through its receptor, cell line-derived neurotrophic factorfamily receptor α-like (GFRAL), to suppress food intake and promote nausea. GDF15 is broadly expressed at low levels but increases in states of disease such as cancer, cachexia, and sepsis. Whether GDF15 is necessary for inducing sepsis-associated anorexia and body weight loss is currently unclear. To test this we used a model of moderate systemic infection in GDF15KO and GFRALKO mice with lipopolysaccharide (LPS) treatment to define the role of GDF15 signaling in infection-mediated physiologic responses. Since physiological responses to LPS depend on housing temperature, we tested the effects of subthermoneutral and thermoneutral conditions on eliciting anorexia and inducing GDF15. Our data demonstrate a conserved LPS-mediated increase in circulating GDF15 levels in mouse, rat, and human. However, we did not detect differences in LPS-induced anorexia between WT and GDF15KO or GFRALKO mice. Furthermore, there were no differences in anorexia or circulating GDF15 levels at either thermoneutral or subthermoneutral housing conditions in LPS-treated mice. These data demonstrate that GDF15 is not necessary to drive food intake suppression in response to moderate doses of LPS. NEW & NOTEWORTHY Although many responses to LPS depend on housing temperature, the anorexic response to LPS does not. LPS results in a potent and rapid increase in circulating levels of GDF15 in mice, rats, and humans. Nevertheless, GDF15 and its receptor (GFRAL) are not required for the anorexic response to systemic LPS administration. The anorexic response to LPS likely involves a myriad of complex physiological alterations.

Gastric bypass and vertical sleeve gastrectomy (VSG) remain the most potent and durable treatments for obesity and type 2 diabetes but are also associated with iron deficiency. The transcription factor HIF2α, which regulates iron absorption in the duodenum, increases following these surgeries. Increasing iron levels by means of dietary supplementation or hepatic hepcidin knockdown does not undermine the effects of VSG, indicating that metabolic improvements following VSG are not secondary to lower iron levels. Gut-specific deletion of Vhl results in increased constitutive duodenal HIF2α signaling and produces a profound lean, glucose-tolerant phenotype that mimics key effects of VSG. Interestingly, intestinal Vhl deletion also results in increased intestinal secretion of GLP-1, which is essential for these metabolic benefits. These data demonstrate a role for increased duodenal HIF2α signaling in regulating crosstalk between iron-regulatory systems and other aspects of systemic physiology important for metabolic regulation., Competing Interests: Declaration of interests R.J.S. has received research support from Novo Nordisk, Astra Zeneca, Pfizer, Energesis, Kintai, and Ionis. R.J.S. has served as a paid consultant for Novo Nordisk, Scohia, Kintai, Eli Lilly, and Ionis. R.J.S. has equity positions in Calibrate and Rewind., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Compelling evidence shows that glucagon-like peptide-1 (GLP-1) has a profound effect in restoring normoglycemia in type 2 diabetic patients by increasing pancreatic insulin secretion. Although L-cells are the primary source of circulating GLP-1, the current therapies do not target L-cells to increase GLP-1 levels. Our study aimed to determine the molecular underpinnings of GLP-1 secretion as an impetus to identify new interventions to target endogenous L-cells., Methods: We used genetic mouse models of intestine-specific overexpression of hypoxia-inducible factor (HIF)-1α and HIF-2α (Vhl ΔIE ), conditional overexpression of intestinal HIF-2α (Hif-2α LSL;Vilin-Cre/ERT2 ), and intestine-specific HIF-2α knockout mice (Hif-2α ΔIE ) to show that HIF signaling, especially HIF-2α, regulates GLP-1 secretion., Results: Our data show that intestinal HIF signaling improved glucose homeostasis in a GLP-1-dependent manner. Intestinal HIF potentiated GLP-1 secretion via the lipid sensor G-protein-coupled receptor (GPR)40 enriched in L-cells. We show that HIF-2α regulates GPR40 in L-cells and potentiates fatty acid-induced GLP-1 secretion via extracellular regulated kinase (ERK). Using a genetic model of intestine-specific overexpression of HIF-2α, we show that HIF-2α is sufficient to increase GLP-1 levels and attenuate diet-induced metabolic perturbations such as visceral adiposity, glucose intolerance, and hepatic steatosis. Lastly, we show that intestinal HIF-2α signaling acts as a priming mechanism crucial for postprandial lipid-mediated GLP-1 secretion. Thus, disruption of intestinal HIF-2α decreases GLP-1 secretion., Conclusions: In summary, we show that intestinal HIF signaling, particularly HIF-2α, regulates the lipid sensor GPR40, which is crucial for the lipid-mediated GLP-1 secretion, and suggest that HIF-2α is a potential target to induce endogenous GLP-1 secretion., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)