BACKGROUND: Sodium-glucose co-transporter 2 inhibitors (SGLT2i), when combined with metformin (COMBI), offer multi-organ protective effects in patients with type 2 diabetes (T2D), particularly those at high risk of cardiovascular or renal complications. However, the underlying molecular mechanisms remain poorly understood. METHODS: We profiled 303 targeted serum metabolites in 1494 participants of the KORA study, including T2D patients treated with COMBI therapy, metformin monotherapy, or no glucose-lowering medication. Additionally, metabolomic profiling was quantified on seven tissues (plasma, liver, adrenal glands, adipose tissue, testis, lung, and cerebellum), and related hepatic transcripts were evaluated in 40 mice. Multivariable linear regression analyses, adjusted for age, sex, BMI, lifestyle, glycemic, and cardiovascular risk factors, were applied to human data; tissue-specific regression analyses were conducted for murine samples. Identified metabolites were further investigated using biochemical pathway analyses and literature review. RESULTS: COMBI therapy was associated with significant changes in metabolite profiles. In humans, 10 metabolites were significantly altered compared to metformin monotherapy. In mice, 82 altered metabolites were identified in plasma, 52 in liver, 30 in adrenal glands, 12 in adipose tissue, seven in testis, seven in lung, and six in cerebellum. COMBI therapy lowered threonine concentrations in both human serum and murine plasma but raised threonine, glycine, and urea cycle metabolites (citrulline, asymmetric dimethyl arginine (ADMA), and ornithine) in murine liver. This was accompanied by enhanced hepatic expression of Slc38a2, a threonine transporter gene. In humans, urea cycle metabolites correlated strongly with the fibrosis-4 index, a marker of liver fibrosis. Additionally, COMBI therapy elevated ketone body markers, such as hydroxybutyrylcarnitine, across murine liver, plasma, adrenal glands, adipose tissue, and testis. CONCLUSIONS: COMBI therapy modulates amino acid metabolism, the urea cycle, and ketone body production, suggesting potential mechanisms underlying its protective effects against liver fibrosis and male subfertility. These findings provide novel insights into the systemic metabolic actions of COMBI therapy and highlight its translational potential to improve clinical outcomes in T2D patients.

The prevalence of metabolic diseases, including obesity and type 2 diabetes, continues to rise. Although GLP-1 receptor agonists (GLP-1RAs) now provide the first effective treatment options for patients with obesity, many still fail to reach their target weight. Interleukin (IL)-22 has emerged as a promising therapeutic due to its ability to modulate key metabolic factors. This study evaluated the potential of a novel long-acting lipidated IL-22 analogue as a complementary treatment to GLP-1RAs in mouse models. Lipidated IL-22 induced up to 20% weight reduction as a monotherapy and up to 40% in combination with GLP-1RAs, demonstrating additive efficacy. Lipidated IL-22 preserved lean body mass with less than half the lean mass loss of GLP-1RAs or caloric restriction. Glycemic control was also enhanced, with lipidated IL-22 normalizing blood glucose, and improving insulin sensitivity independent of food intake. Mechanistically, lipidated IL-22 enhanced intestinal secretion of anorectic factors like PYY and doubled fecal energy loss through reduced intestinal calorie absorption. These findings demonstrate the novel intestinal mechanisms of action of lipidated IL-22 and its additive potential to GLP-1RA treatment. Therefore, lipidated IL-22 is strongly positioned as a novel anti-obesity treatment that can address critical unmet needs in the treatment of metabolic diseases.

The FAIR principles guide data stewardship towards maximizing the value of scientific data while offering a high level of flexibility to accommodate differences in standards and scientific practices. Research communities have developed and implemented domain-specific workflows to make their data FAIR. This work compares the implementation of two externally developed structured FAIRification workflows-a generic workflow and a domain-specific workflow- using the example of metadata captured in diabetes research in Germany and applying the FAIR data maturity model developed by the Research Data Alliance. Interestingly, the implementation of both workflows required similar resources and led us to achieve the same FAIRness rating. We therefore conclude that the adaptations made in the FAIRification workflow for health research data improve efficiency but do not necessarily lead to higher FAIRness scores when applied to core data sets. Based on the results of our workflow comparison, we identified a list of requirements that should be met for the FAIRification of a core data set regardless of the workflow employed. In the future, FAIR data strategies and infrastructure should be planned and implemented as early as possible in the FAIRification journey. It is anticipated that this comparative analysis will help establish standard operating procedures for the FAIRification of core data sets for health studies.

In recent decades, obesity and diabetes have reached pandemic levels, with obesity now recognised as a major health risk factor. Evidence shows that metabolic diseases are more pronounced in the offspring of malnourished parents, suggesting that predisposition can be inherited via epigenetic information in gametes. This has sparked growing interest in small regulatory RNAs in sperm as carriers of epigenetic inheritance. However, the functional annotation of dysregulated sperm microRNAs (miRNAs) in obesity and diabetes remains limited. This work addresses this gap by analysing publicly available datasets of diet-regulated sperm miRNAs and linking them to genes functionally associated with obesity and diabetes. We systematically identified diet-responsive sperm miRNAs and overlapped their predicted targets with genes associated with metabolic phenotypes, as catalogued by the International Mouse Phenotyping Consortium (IMPC). First, in a sequence-based approach, we uncovered 11,272 and 6528 potential target genes for miRNAs regulated by the acute and chronic HFD interventions, respectively. Second, by overlapping these predicted target genes of sperm miRNAs with our IMPC-derived list of 889 genes associated with obesity and diabetes, we identified 805 acute- and 546 chronic-HFD predicted response genes. This approach thus associates function with regulated miRNAs and revealed distinct miRNA-gene networks in acute versus chronic HFD models, including shared nodes in pathways related to insulin signalling, lipid metabolism, and β-cell function. To support further research, we provide the field with the ShinyFatSperm App (https://reproproteomics.shinyapps.io/ShinyFatSperm/), which facilitates the functional interpretation of diet-regulated sperm miRNAs and enables users to explore their roles in the intergenerational transmission of metabolic disease risk. Taken together, our findings reinforce the concept that paternal dietary exposures can influence offspring health through epididymal- and sperm-borne miRNAs, and related epigenetic mechanisms. This work provides a roadmap for hypothesis-driven investigation into the intergenerational inheritance of metabolic diseases and highlights the urgent need for translational strategies to interrupt this cycle.

Ferroptosis, driven by uncontrolled peroxidation of membrane phospholipids, is distinct from other cell death modalities because it lacks an initiating signal and is surveilled by endogenous antioxidant defenses. Glutathione peroxidase 4 (GPX4) is the guardian of ferroptosis, although its membrane-protective function remains poorly understood. Here, structural and functional analyses of a missense mutation in GPX4 (p.R152H), which causes early-onset neurodegeneration, revealed that this variant disrupts membrane anchoring without considerably impairing its catalytic activity. Spatiotemporal Gpx4 deletion or neuron-specific GPX4R152H expression in mice induced degeneration of cortical and cerebellar neurons, accompanied by progressive neuroinflammation. Patient induced pluripotent stem cell (iPSC)-derived cortical neurons and forebrain organoids displayed increased ferroptotic vulnerability, mirroring key pathological features, and were sensitive to ferroptosis inhibition. Neuroproteomics revealed Alzheimer's-like signatures in affected brains. These findings highlight the necessity of proper GPX4 membrane anchoring, establish ferroptosis as a key driver of neurodegeneration, and provide the rationale for targeting ferroptosis as a therapeutic strategy in neurodegenerative disease.

Heart weight (HW) is a critical parameter in cardiology and mouse research, commonly normalized to body weight (BW) or tibia length (TL) to account for size differences. Ratio-based normalization, however, assumes strict proportionality between variables, an assumption that is rarely tested and may bias group comparisons. We analysed HW, BW, and TL measurements from over 25,000 C57BL/6N wildtype mice generated by the International Mouse Phenotyping Consortium. Sex- and age-stratified analyses were combined with simulation-based modelling to evaluate empirical scaling relationships and the statistical behaviour of ratio-based normalization. Across all age and sex groups, correlations between HW, BW, and TL were negligible to weak, indicating substantial deviations from proportionality. Simulations demonstrated that ratio-based normalization can generate misleading results, including spurious or reversed group differences, when proportionality assumptions are violated. Ratios were consistent with linear and allometric models only under strictly proportional conditions, characterized by regression lines passing through the origin. Linear models with covariate adjustment and allometric scaling provide more robust and biologically meaningful frameworks for organ weight analysis. Ratio-based normalization should be avoided unless key mathematical assumptions are met.

Glutaric aciduria type 1 is caused by inherited deficiency of glutaryl-CoA dehydrogenase and subsequent accumulation of neurotoxic metabolites. Clinically, the disease is characterized by striatal damage and dystonic movement disorder in untreated infants. Despite newborn screening and pre-symptomatic therapy start, about one-third of patients still develop neurological symptoms. Furthermore, progressive white matter changes and chronic kidney disease highlights the need for improved therapies. To elucidate the potential of substrate reduction therapy for GA1 we investigated whether aminoadipate-semialdehyde synthetase, the first enzyme of the lysine oxidation pathway, could serve as therapeutic target. Therefore, we studied whether Gcdh knockout (KO) mice, a known animal model for GA1, were rescued by additional knockout of Aass. Gcdh/Aass KO mice were clinically indistinguishable from wild-type mice and showed a marked reduction of glutaric acid in brain (20.9 µg/mg protein vs. 59.2 µg/mg protein; p = 0.001), liver (23.5 µg/mg protein vs. 104.8 µg/mg protein; p = 0.001), and urine (11.9 mol/mol creatinine vs. 166.5 mol/mol creatinine; p = 0.001). The effect was less pronounced for 3-hydroxyglutaric acid. Unlike Gcdh KO mice, Gcdh/Aass KO mice did not develop a severe phenotype under high-lysine diet. In conclusion, knockout of Aass partially rescues the severe phenotype of Gcdh KO mice, providing a potential therapeutic target.

BACKGROUND & AIMS: Obesity and type 2 diabetes are global health challenges driven by genetic and environmental factors, including diet. While intermittent fasting improves metabolic health, the hepatic mechanisms linking feeding transitions to systemic metabolic regulation remain unclear. We investigated whether Indian Hedgehog (Ihh), a liver-derived hepatokine, coordinates metabolic responses to nutritional transitions. METHODS: We employed genetic and epigenetic tools, including liver-specific deletion of the PRC2 component Eed, to study Ihh regulation. In vivo metabolic phenotyping, thermogenic gene profiling, and Ihh immunoneutralization assessed its function. VLDL-associated Ihh levels were measured and their correlations with metabolic traits were analyzed in humans. RESULTS: Ihh is induced upon feeding and promotes adipose thermogenesis, enhancing metabolic flexibility. The Ihh locus in hepatocytes resides in a bivalent chromatin state; hepatic Eed deletion derepresses Ihh, conferring resistance to diet-induced obesity and insulin resistance. Immunoneutralization of Ihh reverses this protection, confirming its necessity. Ihh circulates in complex with VLDL. Human Ihh-VLDL levels decline with age and correlate with improved metabolic parameters, including insulin sensitivity, HDL/LDL ratio, and reduced adiposity. CONCLUSIONS & IMPLICATIONS: Ihh is a liver-derived, epigenetically regulated hepatokine that links nutrient timing to systemic metabolic control by stimulating thermogenesis and promoting glucose homeostasis. These findings identify Ihh as a key inter-organ signal coupling hepatic chromatin dynamics to energy balance. The age-related decline in circulating Ihh and its strong association with metabolic health suggest that enhancing Ihh signaling may represent a novel therapeutic avenue for obesity and type 2 diabetes.

The epididymis is a highly specialised organ essential for promoting the post-testicular functional maturation of spermatozoa, a process underpinning male fertility. This review examines the latest proteomics advances that have been used to unravel the complex molecular landscape of the epididymis, revealing the dynamic protein networks that shape sperm function beyond their genomic and transcriptomic blueprints. Here, we highlight how high-resolution mass spectrometry has helped to map the proteomic signatures of epididymal tissue, luminal extracellular vesicles (epididymosomes), and spermatozoa at different maturation stages, pinpointing key regulators of motility, capacitation, fertilisation competence, and immune regulation. However, critical knowledge gaps remain, including deep protein characterisation of the cytoplasmic droplet, epididymal fluid, and relatively underexplored anatomical tissue segments such as the corpus and cauda epididymis. We discuss how integrating global proteomic insights with complementary omics, single cell proteomics and advanced imaging is poised to reveal the spatial and temporal refinement of the sperm proteome, providing insights into how its disruption may contribute to idiopathic infertility. To promote data accessibility and accelerate discovery in epididymal biology, we introduce ShinyEpididymis (https://reproproteomics.shinyapps.io/ShinyEpididymis/), an interactive, web-based resource integrating publicly available proteomic datasets from spermatozoa, epididymosomes, and epididymal tissue. This platform enables researchers to rapidly query proteins of interest, explore spatial patterns of expression, and identify potential biomarkers or therapeutic targets. By consolidating current knowledge and defining future priorities, this review positions proteomics at the forefront of understanding epididymal biology, emphasising its clinical relevance and untapped potential for diagnosing and treating male infertility.

Understanding the complex factors influencing mammalian metabolism and body weight homeostasis is a long-standing challenge requiring knowledge of energy intake, absorption and expenditure. Using measurements of respiratory gas exchange, indirect calorimetry can provide non-invasive estimates of whole-body energy expenditure. However, inconsistent measurement units and flawed data normalization methods have slowed progress in this field. This guide aims to establish consensus standards to unify indirect calorimetry experiments and their analysis for more consistent, meaningful and reproducible results. By establishing community-driven standards, we hope to facilitate data comparison across research datasets. This advance will allow the creation of an in-depth, machine-readable data repository built on shared standards. This overdue initiative stands to markedly improve the accuracy and depth of efforts to interrogate mammalian metabolism. Data sharing according to established best practices will also accelerate the translation of basic findings into clinical applications for metabolic diseases afflicting global populations.

Mutations in the gene encoding Tectonic β-propeller repeat-containing repeat protein 2 (TECPR2) cause hereditary sensory and autonomic neuropathy subtype 9 (HSAN9) which is a fatal neurodevelopmental and neurodegenerative disorder involving the sensory and peripheral nervous system. TECPR2 is ubiquitously expressed and linked to trafficking and sorting within the cell, however, its functional role remains poorly defined. Moreover, molecular insights into pathogenic mechanisms underlying HSAN9 are lacking. Here, we report a novel mouse model which harbors a HSAN9-associated nonsense mutation that causes loss of TECPR2 expression. Mice show altered gait, highly region-specific axonal dystrophy, and extensive local gliosis. The affected medulla area prominently features swollen axons filled with amorphous protein aggregates, glycogen granules, single and double membrane vesicles as well as aberrant organelles including ER and mitochondria whose proteome is distinctly altered. Despite the locally restricted pathology the neuronal demise is detectable in the cerebrospinal fluid and responded to by damage-associated microglia. However, their capacity to clear neuronal debris seems attenuated. Overall, neuronal and microglia phenotypes point to a dysfunctional endolysosomal system when TECPR2 is missing. This was confirmed in TECPR2 knockout cells and linked to TECPR2's interaction with the homotypic fusion and protein sorting (HOPS)-tethering complex. Collectively, we uncovered a role of TECPR2 in endolysosome maintenance which seems relevant for healthy neurons in a particular brain region.

PURPOSE: This study investigates genes contributing to late-adult corneal dystrophies (LACDs) in aged mice, with potential implications for late-onset corneal dystrophies (CDs) in humans. METHODS: The International Mouse Phenotyping Consortium (IMPC) database, containing data from 8901 knockout mouse lines, was filtered to include late-adult mice (49+ weeks) with significant (P < 0.0001) CD phenotypes. Candidate genes were mapped to human orthologs using the Mouse Genome Informatics group, with expression analyzed via PLAE and a literature review for prior CD associations. Comparative analyses of LACD genes from IMPC and established human CD genes from IC3D included protein interactions (STRING), biological processes (PANTHER), and molecular pathways (KEGG). RESULTS: Analysis identified 14 genes linked to late-adult abnormal corneal phenotypes. Of these, 2 genes were previously associated with CDs in humans, while 12 were novel. Seven of the 14 genes (50%) were expressed in the human cornea based on single-cell transcriptomics. Protein-protein interactions via STRING showed several significant interactions with known human CD genes. PANTHER analysis identified six biological processes shared with established human CD genes. Two genes (Rgs2 and Galnt9) were involved in pathways related to human corneal diseases, including cGMP-PKG signaling, mucin-type O-glycan biosynthesis, and oxytocin signaling. Other candidates were implicated in pathways such as pluripotency of stem cells, MAPK signaling, WNT signaling, actin cytoskeleton regulation, and cellular senescence. CONCLUSIONS: This study identified 14 genes linked to LACD in knockout mice, 12 of which are novel in corneal biology. These genes may serve as potential therapeutic targets for treating corneal diseases in aging human populations.

Endurance exercise reduces the risk of metabolic diseases by improving skeletal muscle metabolism, particularly in individuals with overweight and obesity. As biological sex impacts glucose and fatty acid handling in skeletal muscle, we hypothesized sex differences at the transcriptomic and proteomic level in the acute response to exercise and after an 8-week exercise intervention. We analyzed skeletal muscle biopsies from 25 sedentary subjects (16f/9m) with overweight and obesity using transcriptomics and proteomics at baseline, after acute exercise, and following an 8-week endurance training program. Regulation of sex-specific differences was studied in primary myotubes from the donors. At baseline, differentially methylated CpG-sites potentially explain up to 59% of transcriptomic and 67% of proteomic sex differences. Differences were dominated by higher abundance of fast-twitch fiber type proteins, and transcripts and proteins regulating glycogen degradation and glycolysis in males. Females showed higher abundance of proteins regulating fatty acid uptake and storage. Acute exercise induced stress-responsive transcripts and serum myoglobin predominantly in males. Both sexes adapted to 8-week endurance training by upregulating mitochondrial proteins involved in TCA cycle, oxidative phosphorylation, and β-oxidation. Training equalized fast-twitch fiber type protein levels, mainly by reducing them in males. In vivo sex differences in autosomal genes were poorly retained in myotubes but partially restored by sex hormone treatment. In conclusion, our findings highlight sex-specific molecular signatures that reflect known differences in glucose and lipid metabolism between female and male skeletal muscle. After just 8 weeks of endurance training, these sex differences were attenuated, suggesting a convergence towards a shared beneficial adaptation at the molecular level.

The pig is a valuable animal model in diabetes research; however, standardized protocols are essential for evaluating in vivo metabolism. Here, we present a protocol for in vivo assessment of glucose control and insulin secretion and sensitivity in the pig. We describe steps for catheter implantation, testing of intravenous glucose tolerance, performance of hyperinsulinemic-euglycemic clamps (HECs) and hyperglycemic clamps (HGCs), and blood processing. We then detail procedures for analysis of plasma glucose, insulin, glucagon, and C-peptide concentrations as well as data analysis. For complete details on the use and execution of this protocol, please refer to Renner et al.1 and Renner et al.2.

Ensuring the quality and reproducibility of biological resources is essential for advancing biomedical research and upholding animal welfare standards. The European Mouse Mutant Archive (EMMA), part of the INFRAFRONTIER research infrastructure, plays a key role in this effort by cryopreserving scientifically validated mutant mouse and rat strains and making them accessible to the global scientific community. To further enhance its processes and promote transparency, INFRAFRONTIER/EMMA has developed a set of ten Quality Principles specifically tailored to the unique requirements of cryopreserved rodent mutant strains. These principles guide EMMA's workflows by providing a structured yet flexible quality framework across its distributed nodes. They encompass both general standards-such as adherence to the 3Rs (Replace, Reduce, Refine), staff competence, and continuous improvement-and more specific areas including scientific evaluation, data curation, and intellectual property rights. Each principle is presented with contextual background, defined requirements, practical recommendations, and key references. This initiative aims to strengthen the reliability, ethical integrity, and reproducibility of preclinical research resources.

ABCB5 is a member of the ATP-binding cassette transporter superfamily that is expressed as a full transporter (ABCB5FL) and half transporter (ABCB5β). The ABCB5FL transporter mediates low-level multidrug resistance in cancer and is normally expressed in the prostate and testis, while ABCB5β has been found to be a marker of melanoma and limbal stem cells and is expressed in pigmented cells. To explore ABCB5’s role in normal physiology, we generated Abcb5-deficient C57BL/6J mice by the deletion of Abcb5 exon 2, knocking out both forms of ABCB5, which were completely phenotyped. The mice were fertile and demonstrated altered bioenergetics and fat metabolism, along with alterations in their blood composition, including anisocytosis and decreased white blood cells and platelet counts. This study uncovers further avenues of investigation into the role of Abcb5 in intermediary metabolism, particularly in relation to atherogenesis.

PURPOSE: Analyze phenotypic data from knockout mice with late-adult retinal pathologic phenotypes to identify genes associated with development of adult-onset retinal diseases. METHODS: The International Mouse Phenotyping Consortium (IMPC) database was queried for genes associated with abnormal retinal phenotypes in the late-adult knockout mouse pipeline (49-80 weeks postnatal age). We identified human orthologs and performed protein-protein analysis and biological pathways analysis with known inherited retinal disease (IRD) and age-related macular degeneration (AMD) genes using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING), PLatform for Analysis of single cell Eye in a Disk (PLAE), Protein Analysis Through Evolutionary Relationships (PANTHER), and Kyoto Encyclopedia of Genes and Genomes (KEGG). RESULTS: Screening of 587 late-adult mouse genes yielded 12 with abnormal retinal phenotypes, which corresponded to 20 human orthologs. Three of the 12 mouse genes and two of the 20 human orthologs were previously implicated in retinal pathology or physiology in a literature review. Although all of the genes demonstrated retinal pathology when deleted from the mouse genome, most do not have established roles in human retinal disease. Furthermore, human protein-protein analysis and biological pathway analysis yielded only a few relationships between the candidate gene list and that of known IRD and AMD genes, suggesting they may represent novel retinal functions. CONCLUSIONS: We identified 12 mouse genes with significant late-adult abnormal retinal pathology, eight of which have not been previously implicated in either mouse or human retinal physiology or pathology. These serve as novel retinal disease gene candidates for late-onset retinal disease.

INTRODUCTION: Maturity-onset Diabetes of the Young (MODY) is a rare form of diabetes and arises from mutations in key regulatory genes of the pancreatic beta cell, leading to their functional impairment and early-onset diabetes. Research into PDX1-MODY, a form of MODY caused by mutations in the PDX1 gene, enhances understanding of gene-specific mechanisms underlying glucose dysregulation and provides insights into possible approaches to restore normal metabolic function. However, no currently published mouse model accurately depicts the genetic cause of PDX1-MODY in human patients. METHODS: Using CRISPR-Cas9 technology, we generated the first mouse model carrying one of the most prevalent pathological PDX1 point mutation found in human patients, P33T, and conducted an 18-week in vivo phenotyping experiment assessing homozygous PDX1P33T and wild-type littermates on both chow and high fat diet (HFD). Additionally, transcriptomic and proteomic analyses were performed on isolated pancreatic islets. Islet architecture was investigated via fluorescent microscopy. RESULT: Contrary to expectations, our comprehensive phenotypic analysis of the mouse model carrying the homozygous PDX1P33T mutation revealed no significant differences in metabolic parameters compared to wild-type controls, and no pathological outcomes were observed as seen in human patients. Notably, male PDX1P33T mice exhibited an increase in islet size and number on chow diet, with omics analyses suggesting reprogramming toward stress resilience, but failed to adapt respectively on HFD. DISCUSSION: Our work indicates substantial differences between mouse and human PDX1 function in the pancreas. Further refinement of animal models is necessary to better elucidate the pathophysiology of PDX1-MODY.

BACKGROUND AND AIMS: Fasting hypoglycemia has clinical implications for children with growth hormone (GH)-insensitivity syndrome. This study investigates the pathophysiology of juvenile hypoglycemia in a large animal model for GH receptor (GHR) deficiency (the GHR-KO pig) and elucidates mechanisms underlying the transition to normoglycemia in adulthood. METHODS: Insulin sensitivity was assessed in juvenile and adult GHR-KO pigs and wild-type (WT) controls via hyperinsulinemic-euglycemic clamp (HEC) tests. Glucose turnover was measured using D-[6,6-2H2] glucose and 2H2O. Clinical chemical and targeted metabolomics parameters in blood serum were correlated with qPCR and western blot analyses of liver and adipose tissue. RESULTS: GHR-KO pigs showed increased insulin sensitivity (p=0.0019), especially at young age (M-value +34% vs. WT), insignificantly reduced insulin levels, and reduced endogenous glucose production (p=0.0007), leading to fasting hypoglycemia with depleted liver glycogen, elevated β-hydroxybutyrate, but no increase in NEFA levels. Low hormone-sensitive lipase phosphorylation in adipose tissue suggested impaired lipolysis in young GHR-KO pigs. Metabolomics indicated enhanced fatty acid beta-oxidation and use of glucogenic amino acids, likely serving as compensatory pathways to maintain energy homeostasis. In adulthood, insulin sensitivity remained elevated but less pronounced (M-value +20%), while insulin levels were significantly reduced, enabling normoglycemia and improved NEFA availability. Increased fat mass, not sex hormones, appeared key to this metabolic transition, as early castration had no effect. CONCLUSION: Juvenile hypoglycemia in GH insensitivity results from excessive insulin sensitivity, reduced glucose production, and impaired lipolysis. Normoglycemia in adulthood emerges through increased adiposity and moderated insulin sensitivity, independently of sex hormones. These findings elucidate the age-dependent metabolic adaptations in GH insensitivity.

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