Objective

Obesity increases deposition of extracellular matrix (ECM) components of cardiac tissue. Since obesity aggregates with insulin resistance and heart disease, it is imperative to determine whether the increased ECM deposition contributes to this disease cluster. The hypotheses tested in this study were that in cardiac tissue of obese mice i) increased deposition of ECM components (collagens and hyaluronan) contributes to cardiac insulin resistance and that a reduction in these components improves cardiac insulin action and ii) reducing excess collagens and hyaluronan mitigates obesity-associated cardiac dysfunction.

Methods

Genetic and pharmacological approaches that manipulated collagen and hyaluronan contents were employed in obese C57BL/6 mice fed a high fat (HF) diet. Cardiac insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp and cardiac function was measured by pressure-volume loop analysis in vivo.

Results

We demonstrated a tight association between increased ECM deposition with cardiac insulin resistance. Increased collagen deposition by genetic deletion of matrix metalloproteinase 9 (MMP9) exacerbated cardiac insulin resistance and pirfenidone, a clinically available anti-fibrotic medication which inhibits collagen expression, improved cardiac insulin resistance in obese mice. Furthermore, decreased hyaluronan deposition by treatment with PEGylated human recombinant hyaluronidase PH20 (PEGPH20) improved cardiac insulin resistance in obese mice. These relationships corresponded to functional changes in the heart. Both PEGPH20 and pirfenidone treatment in obese mice ameliorated HF diet-induced abnormal myocardial remodelling.

Conclusion

Our results provide important new insights into the role of ECM deposition in the pathogenesis of cardiac insulin resistance and associated dysfunction in obesity of distinct mouse models. These findings support the novel therapeutic potential of targeting early cardiac ECM abnormalities in the prevention and treatment of obesity-related cardiovascular complications.

Objectives

A high proportion of women with advanced epithelial ovarian cancer (EOC) experience weakness and cachexia. This relationship is associated with increased morbidity and mortality. EOC is the most lethal gynecological cancer, yet no preclinical cachexia model has demonstrated the combined hallmark features of metastasis, ascites development, muscle loss and weakness in adult immunocompetent mice.

Methods

Here, we evaluated a new model of ovarian cancer-induced cachexia with the advantages of inducing cancer in adult immunocompetent C57BL/6J mice through orthotopic injections of EOC cells in the ovarian bursa. We characterized the development of metastasis, ascites, muscle atrophy, muscle weakness, markers of inflammation, and mitochondrial stress in the tibialis anterior (TA) and diaphragm ∼45, ∼75 and ∼90 days after EOC injection.

Results

Primary ovarian tumour sizes were progressively larger at each time point while severe metastasis, ascites development, and reductions in body, fat and muscle weights occurred by 90 Days. There were no changes in certain inflammatory (TNFα), atrogene (MURF1 and Atrogin) or GDF15 markers within both muscles whereas IL-6 was increased at 45 and 90 Day groups in the diaphragm. TA weakness in 45 Day preceded atrophy and metastasis that were observed later (75 and 90 Day, respectively). The diaphragm demonstrated both weakness and atrophy in 45 Day. In both muscles, this pre-severe-metastatic muscle weakness corresponded with considerable reprogramming of gene pathways related to mitochondrial bioenergetics as well as reduced functional measures of mitochondrial pyruvate oxidation and creatine-dependent ADP/ATP cycling as well as increased reactive oxygen species emission (hydrogen peroxide). Remarkably, muscle force per unit mass at 90 days was partially restored in the TA despite the presence of atrophy and severe metastasis. In contrast, the diaphragm demonstrated progressive weakness. At this advanced stage, mitochondrial pyruvate oxidation in both muscles exceeded control mice suggesting an apparent metabolic super-compensation corresponding with restored indices of creatine-dependent adenylate cycling.

Conclusions

This mouse model demonstrates the concurrent development of cachexia and metastasis that occurs in women with EOC. The model provides physiologically relevant advantages of inducing tumour development within the ovarian bursa in immunocompetent adult mice. Moreover, the model reveals that muscle weakness in both TA and diaphragm precedes severe metastasis while weakness also precedes atrophy in the TA. An underlying mitochondrial bioenergetic stress corresponded with this early weakness. Collectively, these discoveries can direct new research towards the development of therapies that target pre-atrophy and pre-severe-metastatic weakness during EOC in addition to therapies targeting cachexia.

Objective

The lactational period is associated with profound hyperphagia to accommodate the energy demands of nursing. These changes are important for the long-term metabolic health of the mother and children as altered feeding during lactation increases the risk of mothers and offspring developing metabolic disorders later in life. However, the specific behavioral mechanisms and neural circuitry mediating the hyperphagia of lactation are incompletely understood.

Methods

Here, we utilized home cage feeding devices to characterize the dynamics of feeding behavior in lactating mice. A combination of pharmacological and behavioral assays were utilized to determine how lactation alters meal structure, circadian aspects of feeding, hedonic feeding, and sensitivity to hunger and satiety signals in lactating mice. Finally, we utilized chemogenetic, immunohistochemical, and in vivo imaging approaches to characterize the role of hypothalamic agouti-related peptide (AgRP) neurons in lactational-hyperphagia.

Results

The lactational period is associated with increased meal size, altered circadian patterns of feeding, reduced sensitivity to gut-brain satiety signals, and enhanced sensitivity to negative energy balance. Hypothalamic AgRP neurons display increased sensitivity to negative energy balance and altered in vivo activity during the lactational state. Further, using in vivo imaging approaches we demonstrate that AgRP neurons are directly activated by lactation. Chemogenetic inhibition of AgRP neurons acutely reduces feeding in lactating mice, demonstrating an important role for these neurons in lactational-hyperphagia.

Conclusions

Together, these results show that lactation collectively alters multiple components of feeding behavior and position AgRP neurons as an important cellular substrate mediating the hyperphagia of lactation.

Objectives

Cachexia is a metabolic disorder and comorbidity with cancer and heart failure. The syndrome impacts more than thirty million people worldwide, accounting for 20% of all cancer deaths. In acute myeloid leukemia, somatic mutations of the metabolic enzyme isocitrate dehydrogenase 1 and 2 cause the production of the oncometabolite D2-hydroxyglutarate (D2-HG). Increased production of D2-HG is associated with heart and skeletal muscle atrophy, but the mechanistic links between metabolic and proteomic remodeling remain poorly understood. Therefore, we assessed how oncometabolic stress by D2-HG activates autophagy and drives skeletal muscle loss.

Methods

We quantified genomic, metabolomic, and proteomic changes in cultured skeletal muscle cells and mouse models of IDH-mutant leukemia using RNA sequencing, mass spectrometry, and computational modeling.

Results

D2-HG impairs NADH redox homeostasis in myotubes. Increased NAD+ levels drive activation of nuclear deacetylase Sirt1, which causes deacetylation and activation of LC3, a key regulator of autophagy. Using LC3 mutants, we confirm that deacetylation of LC3 by Sirt1 shifts its distribution from the nucleus into the cytosol, where it can undergo lipidation at pre-autophagic membranes. Sirt1 silencing or p300 overexpression attenuated autophagy activation in myotubes. In vivo, we identified increased muscle atrophy and reduced grip strength in response to D2-HG in male vs. female mice. In male mice, glycolytic intermediates accumulated, and protein expression of oxidative phosphorylation machinery was reduced. In contrast, female animals upregulated the same proteins, attenuating the phenotype in vivo. Network modeling and machine learning algorithms allowed us to identify candidate proteins essential for regulating oncometabolic adaptation in mouse skeletal muscle.

Conclusions

Our multi-omics approach exposes new metabolic vulnerabilities in response to D2-HG in skeletal muscle and provides a conceptual framework for identifying therapeutic targets in cachexia.

Objective

The prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) continues to rise with the increasing obesity epidemic. Rezdiffra as an activator of a thyroid hormone receptor-beta is the only Food and Drug Administration approved therapy. As such, there is a critical need to improve our understanding of gene expression regulation and signaling transduction in MASLD to develop new therapies. Matrin-3 is a DNA- and RNA-binding protein involved in the pathogenesis of human diseases. Here we examined its previously uncharacterized role in limiting hepatic steatosis and stress response via the constitutive androstane receptor (CAR).

Methods

Matrin-3 floxed and liver-specific knockout mice were fed either a chow diet or 60 kcal% high-fat diet (HFD) for up to 16 weeks. The mice were euthanized for different analysis including liver histology, lipid levels, and gene expression. Bulk RNA-seq, bulk ATAC-seq, and single-nucleus Multiome were used to examine changes of transcriptome and chromatin accessibility in the liver. Integrative bioinformatics analysis of our data and publicly available datasets and different biochemical assays were performed to identify underlying the molecular mechanisms mediating matrin-3's effects. Liver-tropic adeno-associated virus was used to restore the expression of CAR for lipid, acute phase genes, and histological analysis.

Results

Matrin-3 expression is induced in the steatotic livers of mice. Liver-specific matrin-3 deletion exacerbated HFD-induced steatosis, acute phase response, and inflammation in the liver of female mice. The transcriptome and chromatin accessibility were re-programmed in the liver of these mice with signatures indicating that CAR signaling is dysregulated. Mechanistically, matrin-3 interacts with CAR mRNA, and matrin-3 deficiency promotes CAR mRNA degradation. Consequently, matrin-3 deletion impaired CAR signaling by reducing CAR expression. Matrin-3 levels positively correlate with CAR expression in human livers. Ces2a and Il1r1 were identified as new target genes of CAR. Interestingly, we found that CAR discords with the expression of its target genes including Cyp2b10 and Ces2a in response to HFD, indicating CAR signaling is dysregulated by HFD despite increased CAR expression. Dysregulated CAR signaling upon matrin-3 deficiency reduced Ces2a and de-repressed Il1r1 expression. CAR restoration partially abrogated the dysregulated gene expression, exacerbated hepatic steatosis, acute phase response, and inflammation in liver-specific matrin-3 knockout mice fed a HFD.

Conclusions

Our findings demonstrate that matrin-3 is a key upstream regulator maintaining CAR signaling upon metabolic stress, and the matrin-3-CAR axis limits hepatic steatosis and stress response signaling that may give insights for therapeutic intervention.

Objective

In this investigation, we addressed the contribution of the core circadian clock factor, BMAL1, in skeletal muscle to both acute transcriptional responses to exercise and transcriptional remodeling in response to exercise training. Additionally, we adopted a systems biology approach to investigate how loss of skeletal muscle BMAL1 altered peripheral tissue homeostasis as well as exercise training adaptations in iWAT, liver, heart, and lung of male mice.

Methods

Combining inducible skeletal muscle specific BMAL1 knockout mice, physiological testing and standardized exercise protocols, we performed a multi-omic analysis (transcriptomics, chromatin accessibility and metabolomics) to explore loss of muscle BMAL1 on muscle and peripheral tissue responses to exercise.

Results

Muscle-specific BMAL1 knockout mice demonstrated a blunted transcriptional response to acute exercise, characterized by the lack of upregulation of well-established exercise responsive transcription factors including Nr4a3 and Ppargc1a. Six weeks of exercise training in muscle-specific BMAL1 knockout mice induced significantly greater and divergent transcriptomic and metabolomic changes in muscle. Surprisingly, liver, lung, inguinal white adipose and heart showed divergent exercise training transcriptomes with less than 5% of ‘exercise-training’ responsive genes shared for each tissue between genotypes.

Conclusions

Our investigation has uncovered the critical role that BMAL1 plays in skeletal muscle as a key regulator of gene expression programs for both acute exercise and training adaptations. In addition, our work has uncovered the significant impact that altered exercise response in muscle and its likely impact on the system plays in the peripheral tissue adaptations to exercise training. Our work also demonstrates that if the muscle adaptations diverge to a more maladaptive state this is linked to increased gene expression signatures of inflammation across many tissues. Understanding the molecular targets and pathways contributing to health vs. maladaptive exercise adaptations will be critical for the next stage of therapeutic design for exercise mimetics.

Objective

Aberrant glucolipid metabolism in the heart is a characteristic factor in diabetic cardiomyopathy (DbCM). Super-enhancers-driven noncoding RNAs (seRNAs) are emerging as powerful regulators in the progression of cardiac diseases. However, the functions of seRNAs in DbCM have not been fully elucidated.

Methods

Super enhancers and their associated seRNAs were screened and identified by H3K27ac ChIP-seq data in the Encyclopedia of DNA Elements (ENCODE) dataset. A dual-luciferase reporter assay was performed to analyze the function of super-enhancers on the transcription of peroxisome proliferator-activated receptor α-related seRNA (PPARα-seRNA). A DbCM mouse model was established using db/db leptin receptor-deficient mice. Adeno-associated virus serotype 9-seRNA (AAV9-seRNA) was injected via the tail vein to evaluate the role of seRNA in DbCM. The underlying mechanism was explored through RNA pull-down, RNA and chromatin immunoprecipitation, and chromatin isolation by RNA purification.

Results

PPARα-seRNA was regulated by super-enhancers and its levels were increased in response to high glucose and palmitic acid stimulation in cardiomyocytes. Functionally, PPARα-seRNA overexpression aggravated lipid deposition, reduced glucose uptake, and repressed energy production. In contrast, PPARα-seRNA knockdown ameliorated metabolic disorder in vitro. In vivo, overexpression of PPARα-seRNA exacerbated cardiac metabolic disorder and deteriorated cardiac dysfunction, myocardial fibrosis, and hypertrophy in DbCM. Mechanistically, PPARα-seRNA bound to the histone demethylase KDM4B (Lysine-specific demethylase 4B) and decreased H3K9me3 levels in the promoter region of PPARα, ultimately enhancing its transcription.

Conclusions

Our study revealed the pivotal function of a super-enhancer-driven long noncoding RNA (lncRNA), PPARα-seRNA, in the deterioration of cardiac function and the exacerbation of metabolic abnormalities in diabetic cardiomyopathy, which recruited KDM4B to the promoter region of PPARα and repression of its transcription. This suggests a promising therapeutic strategy for the treatment of DbCM.

Mitochondria facilitate thousands of biochemical reactions, covering a broad spectrum of anabolic and catabolic processes. Here we demonstrate that the adipocyte mitochondrial proteome is markedly altered across multiple models of insulin resistance and reveal a consistent decrease in the level of the mitochondrial processing peptidase miPEP.

Objective

To determine the role of miPEP in insulin resistance.

Methods

To experimentally test this observation, we generated adipocyte-specific miPEP knockout mice to interrogate its role in the aetiology of insulin resistance.

Results

We observed a strong phenotype characterised by enhanced insulin sensitivity and reduced adiposity, despite normal food intake and physical activity. Strikingly, these phenotypes vanished when mice were housed at thermoneutrality, suggesting that metabolic protection conferred by miPEP deletion hinges upon a thermoregulatory process. Tissue specific analysis of miPEP deficient mice revealed an increment in muscle metabolism, and upregulation of the protein FBP2 that is involved in ATP hydrolysis in the gluconeogenic pathway.

Conclusion

These findings suggest that miPEP deletion initiates a compensatory increase in skeletal muscle metabolism acting as a protective mechanism against diet-induced obesity and insulin resistance.

Objective

Bariatric surgery is an effective treatment to obesity, leading to weight loss and improvement in glycemia, that is characterized by hypersecretion of gastrointestinal hormones. However, weight regain and relapse of hyperglycemia are not uncommon. We set to identify mechanisms that can enhance gastrointestinal hormonal secretion following surgery to sustain weight loss.

Methods

We investigated the effect of somatostatin (Sst) inhibition on the outcomes of bariatric surgery using a mouse model of sleeve gastrectomy (SG).

Results

Sst knockout (sst-ko) mice fed with a calorie-rich diet gained weight normally and had a mild favorable metabolic phenotype compared to heterozygous sibling controls, including elevated plasma levels of GLP-1. Mathematical modeling of the feedback inhibition between Sst and GLP-1 showed that Sst exerts its maximal effect on GLP-1 under conditions of high hormonal stimulation, such as following SG. Obese sst-ko mice that underwent SG had higher levels of GLP-1 compared with heterozygous SG-operated controls. The SG-sst-ko mice regained less weight than controls and maintained lower glycemia months after surgery. Obese wild-type mice that underwent SG and were treated daily with a Sst receptor inhibitor for two months had higher GLP-1 levels, regained less weight, and improved metabolic profile compared to saline-treated SG-operated controls, and compared to inhibitor or saline-treated sham-operated obese mice.

Conclusions

Our results suggest that inhibition of Sst signaling enhances the long-term favorable metabolic outcomes of bariatric surgery.

Background

Bioenergetic remodeling of core energy metabolism is essential to the initiation, survival, and progression of cancer cells through exergonic supply of adenosine triphosphate (ATP) and metabolic intermediates, as well as control of redox homeostasis. Mitochondria are evolutionarily conserved organelles that mediate cell survival by conferring energetic plasticity and adaptive potential. Mitochondrial ATP synthesis is coupled to the oxidation of a variety of substrates generated through diverse metabolic pathways. As such, inhibition of the mitochondrial bioenergetic system by restricting metabolite availability, direct inhibition of the respiratory Complexes, altering organelle structure, or coupling efficiency may restrict carcinogenic potential and cancer progression.

Scope of Review

Here, we review the role of bioenergetics as the principal conductor of energetic functions and carcinogenesis while highlighting the therapeutic potential of targeting mitochondrial functions.

Major Conclusions

Mitochondrial bioenergetics significantly contribute to cancer initiation and survival. As a result, therapies designed to limit oxidative efficiency may reduce tumor burden and enhance the efficacy of currently available antineoplastic agents.

Background

Type 1 diabetes (T1D) is a complex multi-system disease which arises from both environmental and genetic factors, resulting in the destruction of insulin-producing pancreatic beta cells. Over the past two decades, human genetic studies have provided new insight into the etiology of T1D, including an appreciation for the role of beta cells in their own demise.

Scope of Review

Here, we outline models supported by human genetic data for the role of beta cell dysfunction and death in T1D. We highlight the importance of strong evidence linking T1D genetic associations to bona fide candidate genes for mechanistic and therapeutic consideration. To guide rigorous interpretation of genetic associations, we describe molecular profiling approaches, genomic resources, and disease models that may be used to construct variant-to-gene links and to investigate candidate genes and their role in T1D.

Major Conclusions

We profile advances in understanding the genetic causes of beta cell dysfunction and death at individual T1D risk loci. We discuss how genetic risk prediction models can be used to address disease heterogeneity. Further, we present areas where investment will be critical for the future use of genetics to address open questions in the development of new treatment and prevention strategies for T1D.

Objective

In response to bacterial inflammation, anorexia of acute illness is protective and is associated with the induction of fasting metabolic programs such as ketogenesis. Forced feeding during the anorectic period induced by bacterial inflammation is associated with suppressed ketogenesis and increased mortality. As ketogenesis is considered essential in fasting adaptation, we sought to determine the role of ketogenesis in illness-induced anorexia.

Methods

A mouse model of inducible hepatic specific deletion of the rate limiting enzyme for ketogenesis (HMG-CoA synthase 2, Hmgcs2) was used to investigate the role of ketogenesis in endotoxemia, a model of bacterial inflammation, and in prolonged starvation.

Results

Mice deficient of hepatic Hmgcs2 failed to develop ketosis during endotoxemia and during prolonged fasting. Surprisingly, hepatic HMGCS2 deficiency and the lack of ketosis did not affect survival, glycemia, or body temperature in response to endotoxemia. Mice with hepatic ketogenic deficiency also did not exhibit any defects in starvation adaptation and were able to maintain blood glucose, body temperature, and lean mass compared to littermate wild-type controls. Mice with hepatic HMGCS2 deficiency exhibited higher levels of plasma acetate levels in response to fasting.

Conclusions

Circulating hepatic-derived ketones do not provide protection against endotoxemia, suggesting that alternative mechanisms drive the increased mortality from forced feeding during illness-induced anorexia. Hepatic ketones are also dispensable for surviving prolonged starvation in the absence of inflammation. Our study challenges the notion that hepatic ketogenesis is required to maintain blood glucose and preserve lean mass during starvation, raising the possibility of extrahepatic ketogenesis and use of alternative fuels as potential means of metabolic compensation.

The transcriptional coactivator PGC-1α has been implicated in the regulation of multiple metabolic processes. However, the previously reported metabolic phenotypes of mice deficient in PGC-1α have been inconsistent. PGC-1α exists as multiple isoforms, including variants transcribed from an alternative first exon. We show here that alternative PGC-1α variants are the main entity that increases PGC-1α during exercise. These variants, unlike the canonical isoform of PGC-1α, are robustly upregulated in human skeletal muscle after exercise. Furthermore, the extent of this upregulation correlates with oxygen consumption. Mice lacking these variants manifest impaired energy expenditure during exercise, leading to the development of obesity and hyperinsulinemia. The alternative variants are also upregulated in brown adipose tissue in response to cold exposure, and mice lacking these variants are intolerant of a cold environment. Our findings thus indicate that an increase in PGC-1α expression, attributable mostly to upregulation of alternative variants, is pivotal for adaptive enhancement of energy expenditure and heat production and thereby essential for the regulation of whole-body energy metabolism.

Objective

Interleukin (IL)-22 is a potential therapeutic protein for the treatment of metabolic diseases such as obesity, type 2 diabetes, and metabolic dysfunction-associated steatotic liver disease due to its involvement in multiple cellular pathways and observed hepatoprotective effects. The short serum half-life of IL-22 has previously limited its use in clinical applications; however, the development of mRNA-lipid nanoparticle (LNP) technology offers a novel therapeutic approach that uses a host-generated IL-22 fusion protein. In the present study, the effects of administration of an mRNA-LNP encoding IL-22 on metabolic disease parameters was investigated in various mouse models.

Methods

C57BL/6NCrl mice were used to confirm mouse serum albumin (MSA)-IL-22 protein expression prior to assessments in C57BL/6NTac and CETP/ApoB transgenic mouse models of metabolic disease. Mice were fed either regular chow or a modified amylin liver nonalcoholic steatohepatitis–inducing diet prior to receiving either LNP-encapsulated MSA-IL-22 or MSA mRNA via intravenous or intramuscular injection. Metabolic markers were monitored for the duration of the experiments, and postmortem histology assessment and analysis of metabolic gene expression pathways were performed.

Results

MSA-IL-22 was detectable for ≥8 days following administration. Improvements in body weight, lipid metabolism, glucose metabolism, and lipogenic and fibrotic marker gene expression in the liver were observed in the MSA-IL-22–treated mice, and these effects were shown to be durable.

Conclusions

These results support the application of mRNA-encoded IL-22 as a promising treatment strategy for metabolic syndrome and associated comorbidities in human populations.

Objective

Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality, and its incidence is increasing due to endemic obesity. HCC is sexually dimorphic in both humans and rodents with higher incidence in males, although the mechanisms contributing to these correlations remain unclear. Here, we examined the role of sphingosine kinase 2 (SphK2), the enzyme that regulates the balance of bioactive sphingolipid metabolites, sphingosine-1-phosphate (S1P) and ceramide, in gender specific MASH-driven HCC.

Methods

Male and female mice were fed a high fat diet with sugar water, a clinically relevant model that recapitulates MASH-driven HCC in humans followed by physiological, biochemical cellular and molecular analyses. In addition, correlations with increased risk of HCC recurrence were determined in patients.

Results

Here, we report that deletion of SphK2 protects both male and female mice from Western diet-induced weight gain and metabolic dysfunction without affecting hepatic lipid accumulation or fibrosis. However, SphK2 deficiency decreases chronic diet-induced hepatocyte proliferation in males but increases it in females. Remarkably, SphK2 deficiency reverses the sexual dimorphism of HCC, as SphK2^−/− male mice are protected whereas the females develop liver cancer. Only in male mice, chronic western diet induced accumulation of the autophagy receptor p62 and its downstream mediators, the antioxidant response target NQO1, and the oncogene c-Myc. SphK2 deletion repressed these known drivers of HCC development. Moreover, high p62 expression correlates with poor survival in male HCC patients but not in females. In hepatocytes, lipotoxicity-induced p62 accumulation is regulated by sex hormones and prevented by SphK2 deletion. Importantly, high SphK2 expression in male but not female HCC patients is associated with a more aggressive HCC differentiation status and increased risk of cancer recurrence.

Conclusions

This work identifies SphK2 as a potential regulator of HCC sexual dimorphism and suggests SphK2 inhibitors now in clinical trials could have opposing, gender-specific effects in patients.