Background

Leptin is an adipose tissue-derived hormone that plays a crucial role in body weight, appetite, and behaviour regulation. Leptin controls energy balance as an indicator of adiposity levels and as a modulator of the reward system, which is associated with liking palatable foods. Obesity is characterized by expanded adipose tissue mass and consequently, elevated concentrations of leptin in blood. Leptin's therapeutic potential for most forms of obesity is hampered by leptin resistance and a narrow dose–response window.

Scope of Review

This review describes the current knowledge of the brain regions and intracellular pathways through which leptin promotes negative energy balance and restrains neural circuits affecting food reward. We also describe mechanisms that hinder these biological responses in obesity and highlight potential therapeutic interventions.

Major Conclusions

Additional research is necessary to understand how pathways engaged by leptin in different brain regions are interconnected in the control of energy balance.

Obesity is a major contributor to metabolic and cardiovascular disease. Although senescent cells have been shown to accumulate in adipose tissue, the role of senescence in obesity-induced metabolic disorders and in cardiac dysfunction is not yet clear; therefore, the therapeutic potential of managing senescence in obesity-related metabolic and cardiac disorders remains to be fully defined.

Objective

We investigated the beneficial effects of a senolytic cocktail (dasatinib and quercetin) on senescence and its influence on obesity-related parameters.

Methods and Results

We found that the increase in body weight and adiposity, glucose intolerance, insulin resistance, dyslipidemia, hyperleptinemia, and hepatic disorders which were induced by an obesogenic diet were alleviated by senolytic cocktail treatment in mice. Treatment with senolytic compounds eliminated senescent cells, counteracting the activation of the senescence program and DNA damage in white adipose tissue (WAT) observed with an obesogenic diet. Moreover, the senolytic cocktail prevented the brown adipose tissue (BAT) whitening and increased the expression of the thermogenic gene profile in BAT and pWAT. In the hearts of obese mice, senolytic combination abolished myocardial maladaptation, reducing the senescence-associated secretory phenotype (SASP) and DNA damage, repressing cardiac hypertrophy, and improving diastolic dysfunction. Additionally, we showed that treatment with the senolytic cocktail corrected gene expression programs associated with fatty acid metabolism, oxidative phosphorylation, the P53 pathway, and DNA repair, which were all downregulated in obese mice.

Conclusions

Collectively, these data suggest that a senolytic cocktail can prevent the activation of the senescence program in the heart and WAT and activate the thermogenic program in BAT. Our results suggest that targeting senescent cells may be a novel therapeutic strategy for alleviating obesity-related metabolic and cardiac disorders.

Objective

The molecular mechanisms underlying the maintenance and adaptability of the neuromuscular junction (NMJ) remain poorly understood. This study aimed to investigate the role of AMP-activated protein kinase (AMPK) as a key regulator of NMJ stability and plasticity.

Method

A comprehensive, multifaceted approach was employed, integrating genetic, physiological, and pharmacological methodologies to elucidate the role of skeletal muscle AMPK in modulating the neuromuscular synapse.

Results

Our findings reveal an increased abundance of AMPK transcripts within the NMJ and an age-associated decline in AMPK activity and synapse-specific mitochondrial gene expression. Young mice null for skeletal muscle AMPK displayed a neuromuscular phenotype akin to aged animals. Pharmacological AMPK stimulation facilitated its localization in subsynaptic myonuclei, preceded the induction of several NMJ-related transcripts, and enhanced myotube acetylcholine receptor clustering. Exercise-induced AMPK activation in mouse muscle elicited a broad NMJ-related gene response, consistent with human exercise data.

Conclusions

These findings highlight a critical role for AMPK in the maintenance and remodeling of the NMJ, highlighting its potential as a therapeutic target for age-related and neuromuscular disorders.

Objectives

Susceptibility to obesity in humans is driven by the intricate interplay of genetic, environmental and behavioural factors. Moreover, the mechanisms linking maternal obesity to infertility remain largely understudied. In this study, we investigated how variable susceptibility to obesity in mice affects ovarian steroidogenesis, with a particular focus on the leptin-mediated dysregulation of Nodal signalling pathway in theca cells (TC).

Methods

C56BL/6J (B6) and 129S1/SvlmJ (129) mice, models of maternal obesity (MO), were fed a chow diet (CD) and a high fat diet (HFD) for 16 weeks. To investigate the contrasting effects of leptin on ovarian steroidogenesis, B6 mice pharmacologically treated with leptin for 16 days on CD were used to model hyperleptinemia, while homozygous ob/ob (−/−) mice with genetic leptin deficiency, also on a CD, were used to examine the effects of obesity in the absence of leptin. Following the characterisation of the mouse phenotype, gonadal fat (GON), whole ovaries (WO), ovarian TC and granulosa cell (GC) fractions were collected for mRNA transcription and protein expression analysis. Finally, in vitro treated ovarian explants obtained from B6 mice were used to further elucidate the effects of Nodal on steroidogenesis.

Results

The significant gain in body weight (BW) and fat mass (FM) in HFD-fed B6 mice (p < 0.05), was associated with increased mRNA transcription of the adipose tissue expansion genes Polymerase I and transcript release factor (Cavin), Secreted frizzled-related protein 5 (Sfrp5) and Mesoderm specific transcript (Mest) in GON (p < 0.05). Furthermore, the HFD-fed B6 mice presented also impaired glucose metabolism and insulin sensitivity (p < 0.05). In contrast, the HFD-fed 129 mice exhibited no changes in BW and FM, maintaining glucose and insulin metabolism. At the ovarian level, decreased protein expression of Steroidogenic Acute Regulatory Protein (StAR) in WO obtained from HFD-fed B6 mice (p = 0.05), was followed by reduced transcription of key steroidogenic genes like Star and Cytochrome P450 17a1 (Cyp17a) in TC (p < 0.05). Furthermore, the transcription of Nodal and its receptors was downregulated (p < 0.05), whereas mRNA levels of Suppressor of cytokine signalling 3 (Socs3) and SMAD family member 7 (Smad7) were upregulated in TC obtained from HFD-fed B6 mice (p < 0.05). No changes were seen in the genes regulating steroidogenesis, Nodal signalling, or Socs3 and Smad7 activity in the ovaries of HFD-fed 129 mice. Importantly, the pharmacological treatment of lean mice with leptin, upregulated the ovarian transcription of Socs3 and Smad7, while downregulating Nodal and its receptors (p < 0.05). Finally, in vitro pharmacological inhibition of Nodal signalling pathway in ovarian explants isolated from CD-fed B6 mice decreased the transcription of Star and Cyp17a in TC (p < 0.05), whereas Nodal treatment of explants obtained from HFD-fed B6 mice restored the transcription of both genes (p < 0.05).

Conclusions

Increased susceptibility to obesity in MO is associated with systemic hyperleptinemia and hypoestrogenism due to compromised ovarian steroidogenesis, largely driven by the inhibitory effects of leptin-Smad7 pathway on Nodal signalling activity in the TC compartment of ovarian follicles.

Objective

B7-H4 (B7S1, B7x, VTCN1) is an important immune checkpoint molecule that maintains immune homeostasis and is also expressed in pancreatic β cells. The polymorphism of B7-H4 influences the prevalence of Type 2 diabetes (T2D), suggesting a potential role of B7-H4 in the physiological function of pancreatic β cells and the pathogenesis of T2D.

Methods

β-cell-specific B7-H4 knockout mice (B7-H4 cKO mice) and their wild-type littermates were used to investigate the in vivo effects of B7-H4 on pancreatic β-cell morphology and function. AAV2/8-ins2-B7H4 and a control virus were infused via the pancreatic intraduct into high-fat diet (HFD)-treated mice to elucidate the therapeutic effect of B7-H4. RNA sequencing was conducted on primary islets. A Luminex assay was used to quantify cytokine changes in B7-H4 cKO mice. Electron microscopy imaging was used to observe insulin secretory vesicles in pancreatic β cells.

Results

Lesion of B7-H4 in β cells results in glucose intolerance due to reduced β-cell mass and deficient insulin secretion, whereas overexpression of B7-H4 in β cells ameliorates glucose intolerance in HFD-fed mice. Mechanistically, B7-H4 deficiency activates signal transducer and activator of transcription 5 (Stat5) signalling, which inhibits the expression of apolipoprotein F (Apof), leading to reduced cholesterol efflux and accumulated cholesterol in β cells, thereby impairing insulin processing and secretion. Overexpression of Apof in β cells or intraperitoneal injection of a Stat5 inhibitor reverses the metabolic phenotype and insulin secretion deficiency in B7-H4 cKO mice.

Conclusion

Our study demonstrated that B7-H4 plays an important role in regulating β-cell mass and insulin secretion, which may shed new light on the development of novel strategies for T2D treatment.

Objective

Dietary protein restriction induces adaptive changes in food preference, increasing protein consumption over carbohydrates or fat. We investigated whether motivation and reward signaling underpin these preferences.

Methods and Results

In an operant task, protein-restricted male mice responded more for liquid protein rewards, but not carbohydrate, fat, or sweet rewards compared to non-restricted mice. When the number of responses required to access protein reward varied, protein-restricted mice exhibited higher operant responses at moderate to high response requirements. The protein restriction-induced increase in operant responding for protein was absent in Fgf21-KO mice and mice with neuron-specific deletion of the FGF21 co-receptor beta-Klotho (Klb^Cam2ka). Fiber photometry recording of VTA dopamine neurons revealed that oral delivery of maltodextrin triggered a larger dopamine neuron activation than casein in control diet-fed mice, while casein triggered a larger activation in low-protein diet-fed mice. This restriction-induced shift in nutrient-specific VTA dopamine signaling was lost in Fgf21-KO mice.

Conclusion

These data suggest that the increased FGF21 during protein restriction acts in the brain to induce a protein-specific appetite by specifically enhancing the reward value of protein-containing foods and the motivation to consume them.

Background

As the primary source of glucose during fasting, hepatic gluconeogenesis is rigorously regulated to maintain euglycemia. Abnormal gluconeogenesis in the liver can lead to hyperglycemia, a key diagnostic marker and the primary pathological contributor to type 2 diabetes (T2D) and metabolic disorders. Hepatic nuclear factor-4 (HNF4α) is an important regulator of gluconeogenesis. In this study, we identify leupaxin (LPXN) as a novel coactivator for HNF4α. Although previous studies have shown that LPXN is highly correlated with cancer types such as B-cell differentiation and hepatocellular carcinoma progression, the role of LPXN in gluconeogenesis remains unknown.

Methods

We initially used protein pull-down assays, mass spectrometry and luciferase assays to identify the coactivator that interacts with HNF4α in gluconeogenesis. We further leveraged cell cultures and mouse models to validate the functional importance of molecular pathway during gluconeogenesis by using adenovirus-mediated overexpression and adeno-associated virus shRNA–mediated knockdown both in vivo and ex vivo, such as in ob/db/DIO mice, HepG2 and primary hepatocytes. Following, we used CUT&Tag and chip qPCR to identify the LPXN-mediated mechanisms underlying the observed abnormal gluconeogenesis. Additionally, we assessed the translational relevance of our findings using human liver tissues from both healthy donors and patients with obesity/type 2 diabetes.

Results

We found that LPXN interacts with HNF4α to participate in gluconeogenesis. Knockdown of LPXN expression in the liver effectively enhanced glucose metabolism, while its overexpression in the liver effectively inhibited it. Mechanistically, LPXN could translocate into the nucleus and was essential for regulating gluconeogenesis by binding to the PEPCK promoter, which controlled the expression of an enzyme involved in gluconeogenesis, mainly through the Gcg-cAMP-PKA pathway. Additionally, LPXN expression was found to be increased in the livers of patients with steatosis and diabetes, supporting a pathological role of LPXN.

Conclusions

Taken together, our study provides evidence that LPXN plays a critical role in modulating hepatic gluconeogenesis, thereby reinforcing the fact that targeting LPXN may be a potential approach for the treatment of diabetes and metabolic disorders.

Objective

Glucagon is a crucial regulator of glucose and lipid metabolism as well as whole-body energy balance. Thus, modulation of glucagon receptor (GCGR) activity in the context of single-molecule multi-receptor co-agonists has become an emerging therapeutic target against obesity and obesity-associated metabolic dysfunction. To better elucidate the role of GCGR-signaling when paired with incretin receptor signaling or on its own, we developed, LY3324954, a GCGR agonist with improved potency and selectivity as compared to the native glucagon peptide.

Methods

LY3324954 was administered to DIO mice, rats, dogs, and monkeys to evaluate pharmacokinetic (PK) profile. Biweekly treatments were conducted in lean and DIO mice to characterize LY3324954-effects on glucose homeostasis and energy balance. Single dose studies were also conducted in liver Gcgr-deficient mice to establish receptor specificity.

Results

LY3324954 also exhibited extended PK profile in DIO mice, rats, dogs, and monkeys. When administered every 72 h, LY3324954 treatment stimulated transient glucose and insulin excursions in lean mice. In diet-induced obese mice, LY3324954 treatment stimulates energy expenditure, weight loss, and a reduction of adiposity in a dose-dependent manner. Benefit to whole-body lipid homeostasis was likewise observed in these mice.

Conclusions

Taken together, these studies characterize a long-acting and potent GCGR-agonist and its regulation of glucose and lipid metabolism as well as whole-body energy balance following both acute and chronic treatment in mice.

Stress-induced behaviours are driven by complex neural circuits and some neuronal populations concurrently modulate diverse behavioural and physiological responses to stress. Glucagon-like peptide-1 (GLP-1)-producing preproglucagon (PPG) neurons within the lower brainstem caudal nucleus of the solitary tract (cNTS) are particularly sensitive to stressful stimuli and are implicated in multiple physiological and behavioural responses to interoceptive and psychogenic threats. However, the afferent inputs driving stress-induced activation of PPG neurons are largely unknown, and the role of PPG neurons in anxiety-like behaviour is controversial. Through chemogenetic manipulations we reveal that cNTS PPG neurons have the ability to moderately increase anxiety-like behaviours in mice in a sex-dependent manner. Using an intersectional approach, we show that input from the paraventricular nucleus of the hypothalamus (PVN) drives activation of both the cNTS as a whole and PPG neurons in particular in response to acute restraint stress, but that while this input is rich in corticotropin-releasing hormone (CRH), PPG neurons do not express significant levels of receptors for CRH and are not activated following lateral ventricle delivery of CRH. Finally, we demonstrate that cNTS-projecting PVN neurons are necessary for the ability of restraint stress to suppress food intake in male mice. Our findings reveal sex differences in behavioural responses to PPG neural activation and highlight a hypothalamic-brainstem pathway in stress-induced hypophagia.

Objective

The control of energy balance involves neural circuits in the central nervous system, including AGRP neurons in the arcuate nucleus of the hypothalamus (ARC). AGRP neurons are crucial for energy balance and their increased activity during fasting is critical to promote feeding behavior. The activity of these neurons is influenced by multiple signals including those acting on G-protein coupled receptors (GPCR) activating different intracellular signaling pathways. We sought to determine whether discrete G-protein mediated signaling in AGRP neurons, promotes differential regulation of feeding and whole-body glucose homeostasis.

Methods

To test the contribution of Gαq/11 or Gαs signaling, we developed congenital mouse lines expressing the different DREADD receptors (i.e., hM3q and rM3s), in AGRP neurons. Then we elicited chemogenetic activation of AGRP neurons in these mice during the postprandial state to determine the impact on feeding and glucose homeostasis.

Results

Activation of AGRP neurons via hM3q and rM3s promoted hyperphagia. In contrast, only hM3q activation of AGRP neurons of the hypothalamic arcuate nucleus during the postprandial state enhanced whole-body glucose disposal by reducing sympathetic nervous system activity to the pancreas and liver, promoting glucose-stimulated insulin secretion, glycogen deposition and improving glucose tolerance.

Conclusions

These data indicate that AGRP neurons regulate food intake and glucose homeostasis through distinct GPCR-dependent signaling pathways and suggest that the transient increase in AGRP neuron activity may contribute to the beneficial effects of fasting on glycemic control.

Inorganic polyphosphate (polyP) is an evolutionary and ancient polymer composed by orthophosphate units linked by phosphoanhydride bonds. In mammalian cells, polyP shows a high localization in mammalian mitochondria, and its regulatory role in various aspects of bioenergetics has already been demonstrated, via molecular mechanism(s) yet to be fully elucidated. In recent years, a role for polyP in signal transduction, from brain physiology to the bloodstream, has also emerged.

Objective

In this manuscript, we explored the intriguing possibility that the effects of polyP on signal transduction could be mechanistically linked to those exerted on bioenergetics.

Methods

To conduct our studies, we used a combination of cellular and animal models.

Results

Our findings demonstrate for the first time the intimate crosstalk between the levels of polyP and the activation status of the AMPK signaling pathway, via a mechanism involving free phosphate homeostasis. AMPK is a key player in mammalian cell signaling, and a crucial regulator of cellular and mitochondrial homeostasis. Our results show that the depletion of mitochondrial polyP in mammalian cells downregulates the activity of AMPK. Moreover, increased levels of polyP activate AMPK. Accordingly, the genetic downregulation of AMPKF0611 impairs polyP levels in both SH-SY5Y cells and in the brains of female mice.

Conclusions

This manuscript sheds new light on the regulation of AMPK and positions polyP as a potent regulator of mammalian cell physiology beyond mere bioenergetics, paving the road for using its metabolism as an innovative pharmacological target in pathologies characterized by dysregulated bioenergetics.

Objectives

Arginine vasopressin (AVP), known as an antidiuretic hormone, is also crucial in metabolic homeostasis. Although AVP receptor-deficient mice exhibit various abnormalities in glucose and lipid metabolism, the mechanism underlying these symptoms remains unclear. This study aimed to explore the involvement of the gut hormones including glucagon-like peptide-1 (GLP-1) and microbiota as essential mediators.

Methods

We used the mouse GLP-1-secreting cell line, GLUTag, and performed live cell imaging to examine the contribution of V1a and V1b vasopressin receptors (V1aR and V1bR, respectively) to GLP-1 secretion. We next investigated the hormone dynamics of V1aR-deficient mice (V1aR^−/− mice), V1bR-deficient mice (V1bR^−/− mice), and V1aR/V1bR-double deficient mice (V1aR^−/− V1bR^−/−mice).

Results

AVP induced the increase in intracellular Ca²⁺ levels and GLP-1 secretion from GLUTag cells in a V1aR and V1bR-dependent manner. AVP receptor-deficient mice, particularly V1aR^−/−V1bR^−/− mice, demonstrated impaired secretion of GLP-1 and peptide YY secreted by enteroendocrine L cells. V1aR^−/−V1bR^−/−mice also exhibited abnormal lipid accumulation in the brown adipose tissue and skeletal muscle. We discovered that V1aR^−/− V1bR^−/− mice showed increased Paneth cell-related gene expression in the small intestine, which was attributed to increased fecal butyric acid levels. Exposure to butyric acid reduced GLP-1 secretion in L cell line. Additionally, human Paneth cell-related gene expressions negatively correlated with that of V1 receptor genes.

Conclusions

The deficiency in V1 receptor genes may increase gut butyric acid levels and impair the function of L cells, thus dysregulating lipid homeostasis in the brown adipose tissue and skeletal muscle. This study highlights the importance of appropriate control of the gut microbiota and its metabolites, including butyric acid, for the optimum functioning of enteroendocrine cells.

Objective

Infiltration of adipocytes into the pancreatic parenchyma has been linked to impaired insulin secretion in individuals with increased genetic risk of T2D and prediabetic conditions. However, the study of this ectopic fat depot has been limited by the lack of suitable in vitro models.

Methods

Here, we developed a novel 3D model of functionally mature human pancreatic adipose tissue organoids by aggregating human pancreatic adipose tissue-derived stromal vascular fraction (SVF) cells into organoids and differentiating them over 19 days.

Results

These organoids carry biological properties of the in situ pancreatic fat, presenting levels of adipogenic markers comparable to native pancreatic adipocytes and improved lipolytic and anti-lipolytic response compared to conventional 2D cultures. The organoids harbour a small population of immune cells, mimicking in vivo adipose environment. Furthermore, they express GIPR, allowing investigation of incretin effects in pancreatic fat. In accordance, GIP and the dual GLP1R/GIPR agonist tirzepatide stimulate lipolysis but had distinct effects on the expression of proinflammatory cytokines.

Conclusions

This novel adipose organoid model is a valuable tool to study the metabolic impact of incretin signalling in pancreatic adipose tissue, revealing potential therapeutic targets of incretins beyond islets. The donor-specific metabolic memory of these organoids enables examination of the pancreatic fat-islet crosstalk in a donor-related metabolic context.

Objective

Steroidogenic factor-1 (SF1) neurons of the ventromedial hypothalamus play key roles in the regulation of food intake, body weight and glucose metabolism. The bile acid receptor Takeda G protein-coupled receptor 5 (TGR5) is expressed in the hypothalamus, where it determines some of the actions of bile acids on food intake and body weight through still poorly defined neuronal mechanisms. Here, we examined the role of TGR5 in SF1 neurons in the regulation of energy balance and glucose metabolism.

Methods

We used a genetic approach combined with metabolic phenotyping and molecular analyses to establish the effect of TGR5 deletion in SF1 neurons on meal pattern, body weight, body composition, energy expenditure and use of energy substrates as well as on possible changes in glucose handling and insulin sensitivity.

Results

Our findings reveal that TGR5 in SF1 neurons does not play a major role in the regulation of food intake or body weight under standard chow, but it is involved in the adaptive feeding response to the acute exposure to cold or to a hypercaloric, high-fat diet, without changes in energy expenditure. Notably, TGR5 in SF1 neurons hinder glucose metabolism, since deletion of the receptor improves whole-body glucose uptake through heightened insulin signaling in the hypothalamus and in the brown adipose tissue.

Conclusions

TGR5 in SF1 neurons favours satiety by differently modifying the meal pattern in response to specific metabolic cues. These studies also reveal a novel key function for TGR5 in SF1 neurons in the regulation of whole-body insulin sensitivity, providing new insight into the role played by neuronal TGR5 in the regulation of metabolism.

Objective

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.

Objective

Exercise plays a crucial role in maintaining and improving human health. However, the precise molecular mechanisms that govern the body’s response to exercise or/compared to periods of inactivity remain elusive. Current evidence appears to suggest that exercise exerts a seemingly dual influence on macrophage polarization states, inducing both pro-immune response M1 activation and cell-repair-focused M2 activation. To reconcile this apparent paradox, we leveraged a comprehensive meta-analysis of 75 diverse exercise and immobilization published datasets (7000+ samples), encompassing various exercise modalities, sampling techniques, and species.

Methods

75 exercise and immobilization expression datasets were identified and processed for analysis. The data was analyzed using boolean relationships which uses binary gene expression relationships in order to increase the signal to noise achieved from the data, allowing for the use of comparison across such a diverse set of datasets. We utilized a boolean relationship-aided macrophage gene model [1], to model the macrophage polarization state in pre and post exercise samples in both immediate exercise and long term training.

Results

Our modeling uncovered a key temporal dynamic: exercise triggers an immediate M1 surge, while long term training transitions to sustained M2 activation. These patterns were consistent across different species (human vs mouse), sampling methods (blood vs muscle biopsy), and exercise type (resistance vs endurance), and routinely showed statistically significant results. Immobilization was shown to have the opposite effect of exercise by triggering an immediate M2 activation. Individual characteristics like gender, exercise intensity and age were found to impact the degree of polarization without changing the overall patterns. To model macrophages within the specific context of muscle tissue, we identified a focused gene set signature of muscle resident macrophage polarization, allowing for the precise measurement of macrophage activity in response to exercise within the muscle.

Conclusions

These consistent patterns across all 75 examined studies suggest that the long term health benefits of exercise stem from its ability to orchestrate a balanced and temporally-regulated interplay between pro-immune response (M1) and reparative macrophage activity (M2). Similarly, it suggests that an imbalance between pro-immune and cell repair responses could facilitate disease development. Our findings shed light on the intricate molecular choreography behind exercise-induced health benefits with a particular insight on its effect on the macrophages within the muscle.

Objective

The early consumption of calorie-rich diet disrupts circadian rhythms and has adverse effects on memory, yet the effects of time-restricted feeding (TRF) and the underlying molecular mechanisms are unknown. Here, we set out to identify the behavioral and molecular circadian rhythms disruptions generated by juvenile obesogenic diet consumption and their restoration by TRF in male mice.

Methods

Metabolic rhythms were measured by indirect calorimetry and memory performances by behavioral tasks. Hippocampal translatome (pS6_TRAP), enrichment and co-regulated gene network analyses were conducted to identify the molecular pathways involved in memory impairments and their restoration by TRF. Differential exon usage analyses, mass spectrometry and pharmacological intervention were used to confirm thyroid hormone signaling involvement.

Results

We show that four weeks of TRF restore the rhythmicity of metabolic parameters and prevents memory impairments in mice fed a high fat-high sucrose (HFS) diet since weaning, independently of body fat levels. Hippocampal translatome and differential exon usage analyses indicate that impaired memory of mice under ad libitum HFS diet is accompanied by reduced thyroid hormone signaling and altered expression of astrocytic genes regulating glutamate neurotransmission. TRF restored the diurnal expression variation of part of these genes and intra-hippocampal infusion of T3, the active form of thyroid hormone, rescues memory performances and astrocytic gene expression of ad libitum HFS diet-fed mice.

Conclusions

Thus, thyroid hormones contribute to the TRF positive effects on both metabolism and memory in mice fed an obesogenic diet, highlighting this nutritional approach as a powerful tool in addressing obesity brain comorbidities and paving the way for further mechanistic studies on hippocampal thyroid signaling.

Objectives

Deficiency in the transcription factor (TF) GLI-Similar 3 (GLIS3) in humans and mice leads to the development of polycystic kidney disease (PKD). In this study, we investigate the role of GLIS3 in the regulation of energy metabolism and mitochondrial functions in relation to its role in normal kidney and metabolic reprogramming in PKD pathogenesis.

Methods

Transcriptomics, cistromics, and metabolomics were used to obtain insights into the role of GLIS3 in the regulation of energy homeostasis and mitochondrial metabolism in normal kidney and PKD pathogenesis using GLIS3-deficient mice.

Results

Transcriptome analysis showed that many genes critical for mitochondrial biogenesis, oxidative phosphorylation (OXPHOS), fatty acid oxidation (FAO), and the tricarboxylic acid (TCA) cycle, including Tfam, Tfb1m, Tfb2m, Ppargc1a, Ppargc1b, Atp5j2, Hadha, and Sdha, are significantly suppressed in kidneys from both ubiquitous and tissue-specific Glis3-deficient mice. ChIP-Seq analysis demonstrated that GLIS3 is associated with the regulatory region of many of these genes, indicating that their transcription is directly regulated by GLIS3. Cistrome analyses revealed that GLIS3 binding loci frequently located near those of hepatocyte nuclear factor 1-Beta (HNF1B) and nuclear respiratory factor 1 (NRF1) suggesting GLIS3 regulates transcription of many metabolic and mitochondrial function-related genes in coordination with these TFs. Seahorse analysis and untargeted metabolomics corroborated that mitochondrial OXPHOS utilization is suppressed in GLIS3-deficient kidneys and showed that key metabolites in glycolysis, TCA cycle, and glutamine pathways were altered indicating increased reliance on aerobic glycolysis and glutamine anaplerosis. These features of metabolic reprogramming may contribute to a bioenergetic environment that supports renal cyst formation and progression in Glis3-deficient mice kidneys.

Conclusions

We identify GLIS3 as a novel positive regulator of the transition from aerobic glycolysis to OXPHOS in normal early postnatal kidney development by directly regulating the transcription of mitochondrial metabolic genes. Loss of GLIS3 induces several features of renal cell metabolic reprogramming. Our study identifies GLIS3 as a new participant in an interconnected transcription regulatory network, that includes HNF1B and NRF1, critical in the regulation of mitochondrial-related gene expression and energy metabolism in normal postnatal kidneys and PKD pathogenesis in Glis3-deficient mice.

Objective

Thyroid hormone (TH) action is mediated by thyroid hormone receptor (THR) isoforms. While THRβ1 is likely the main isoform expressed in liver, its role in human hepatocytes is not fully understood.

Methods

To elucidate the role of THRβ1 action in human hepatocytes we used CRISPR/Cas9 editing to knock out THRβ1 in induced pluripotent stem cells (iPSC). Following directed differentiation to the hepatic lineage, iPSC-derived hepatocytes were then interrogated to determine the role of THRβ1 in ligand-independent and -dependent functions.

Results

We found that the loss of THRβ1 promoted alterations in proliferation rate and metabolic pathways regulated by T3, including gluconeogenesis, lipid oxidation, fatty acid synthesis, and fatty acid uptake. We observed that key genes involved in liver metabolism are regulated through both T3 ligand-dependent and -independent THRβ1 signaling mechanisms. Finally, we demonstrate that following THRβ1 knockout, several key metabolic genes remain T3 responsive suggesting they are THRα targets.

Conclusions

These results highlight that iPSC-derived hepatocytes are an effective platform to study mechanisms regulating TH signaling in human hepatocytes.

Objective

The study aimed to investigate the effects of glucagon on metabolic pathways in mouse models of obesity, fatty liver disease, and type 2 diabetes (T2D) to determine the extent and variability of hepatic glucagon resistance in these conditions.

Methods

We investigated glucagon's effects in mouse models of fatty liver disease, obesity, and type 2 diabetes (T2D), including male BKS-db/db, high-fat diet-fed, and western diet-fed C57Bl/6 mice. Glucagon tolerance tests were performed using the selective glucagon receptor agonist acyl-glucagon (IUB288). Blood glucose, serum and liver metabolites include lipids and amino acids were measured. Additionally, liver protein expression related to glucagon signalling and a comprehensive liver metabolomics were performed.

Results

Western diet-fed mice displayed impaired glucagon response, with reduced blood glucose and PKA activation. In contrast, high-fat diet-fed and db/db mice maintained normal glucagon sensitivity, showing significant elevations in blood glucose and phospho-PKA motif protein expression. Acyl-glucagon treatment also lowered liver alanine and histidine levels in high-fat diet-fed mice, but not in western diet-fed mice. Additionally, some amino acids, such as methionine, were increased by acyl-glucagon only in chow diet control mice. Despite normal glucagon sensitivity in PKA signalling, db/db mice had a distinct metabolomic response, with acyl-glucagon significantly altering 90 metabolites in db/+ mice but only 42 in db/db mice, and classic glucagon-regulated metabolites, such as cyclic adenosine monophosphate (cAMP), being less responsive in db/db mice.

Conclusions

The study reveals that hepatic glucagon resistance in obesity and T2D is complex and not uniform across metabolic pathways, underscoring the complexity of glucagon action in these conditions.

Objectives

Obesity, diabetes and high-calorie diets are associated with defective sperm function and lowered male fertility. Mature spermatozoa primarily use fructose and glucose, and glucose and glycerol metabolism are important for sperm function. We recently discovered a novel mammalian enzyme, glycerol-3-phosphate (Gro3P) phosphatase (G3PP), and showed that it operates the glycerol shunt by hydrolyzing Gro3P to glycerol, and regulates glucose, lipid and energy metabolism in pancreatic β-cells and liver. We now observed that G3PP expression is the highest in the testis and spermatozoa, and investigated its role in male fertility.

Methods

We examined G3PP expression during spermatogenesis in mouse and assessed male fertility and spermatozoon function in conditional germ cell specific G3PP-KO (cG3PP-KO) mice and tamoxifen-inducible conditional germ cell G3PP-KO (icG3PP-KO) mice. We also determined the structural and metabolic parameters and oxidative stress in the spermatozoa from icG3PP-KO and control mice.

Results

G3PP expression in mouse spermatocytes and spermatids markedly increases during spermatogenesis. Male cG3PP-KO mice, in which germ cell G3PP is deleted from embryonic stage, are infertile due to dysfunctional sperm with reduced motility and capacitation, and elevated spontaneous acrosomal reaction and oxidative stress. However, icG3PP-KO male mice do not have altered fertility, due to the presence of ∼10% normal spermatozoa. icG3PP-KO spermatozoa display significantly reduced functionality and morphological and ultrastructural alterations. The icG3PP-KO spermatozoa show reduced glycerol production, elevated levels of Gro3P and reactive oxygen species (ROS), and oxidative stress that is associated with increased mitochondrial membrane potential.

Conclusions

Germ cell G3PP deletion leads to the generation of spermatozoa that are functionally and structurally abnormal, likely due to the build-up of Gro3P that increases mitochondrial membrane potential, ROS, and oxidative stress and alters spermatozoa function. Overall, the results indicate that G3PP and the glycerol shunt are essential for normal spermatozoa function and male fertility.

Objective

Dysregulated glucagon secretion and inadequate functional beta cell mass are hallmark features of diabetes. While glucagon receptor (GCGR) antagonism ameliorates hyperglycemia and elicits beta cell regeneration in pre-clinical models of diabetes, it also promotes alpha and delta cell hyperplasia. We sought to investigate the mechanism by which loss of glucagon action impacts pancreatic islet non-alpha cells, and the relevance of these observations in a human islet context.

Methods

We used zebrafish, rodents, and transplanted human islets comprising six different models of interrupted glucagon signaling to examine their impact on delta and beta cell proliferation and mass. We also used models with global deficiency of the cationic amino acid transporter, SLC7A2, and mTORC1 inhibition via rapamycin, to determine whether amino acid-dependent nutrient sensing was required for islet non-alpha cell growth.

Results

Inhibition of glucagon signaling stimulated delta cell proliferation in mouse and transplanted human islets, and in mouse islets. This was rapamycin-sensitive and required SLC7A2. Likewise, gcgr deficiency augmented beta cell proliferation via SLC7A2- and mTORC1-dependent mechanisms in zebrafish and promoted cell cycle engagement in rodent beta cells but was insufficient to drive a significant increase in beta cell mass in mice.

Conclusions

Our findings demonstrate that interruption of glucagon signaling augments islet non-alpha cell proliferation in zebrafish, rodents, and transplanted human islets in a manner requiring SLC7A2 and mTORC1 activation. An increase in delta cell mass may be leveraged for future beta cell regeneration therapies relying upon delta cell reprogramming.

Objectives

Histone acetylation is an important epigenetic modification that regulates various biological processes and cell homeostasis. Acetyl-CoA, a hub molecule of metabolism, is the substrate for histone acetylation, thus linking metabolism with epigenetic regulation. However, still relatively little is known about the dynamics of histone acetylation and its dependence on metabolic processes, due to the lack of integrated methods that can capture site-specific histone acetylation and deacetylation reactions together with the dynamics of acetyl-CoA synthesis.

Methods

In this study, we present a novel proteo-metabo-flux approach that combines mass spectrometry-based metabolic flux analysis of acetyl-CoA and histone acetylation with computational modelling. We developed a mathematical model to describe metabolic label incorporation into acetyl-CoA and histone acetylation based on experimentally measured relative abundances.

Results

We demonstrate that our approach is able to determine acetyl-CoA synthesis dynamics and site-specific histone acetylation and deacetylation reaction rate constants, and that consideration of the metabolically labelled acetyl-CoA fraction is essential for accurate determination of histone acetylation dynamics. Furthermore, we show that without correction, changes in metabolic fluxes would be misinterpreted as changes in histone acetylation dynamics, whereas our proteo-metabo-flux approach allows to distinguish between the two processes.

Conclusions

Our proteo-metabo-flux approach expands the repertoire of metabolic flux analysis and cross-omics and represents a valuable approach to study the regulatory interplay between metabolism and epigenetic regulation by histone acetylation.