Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer subtype with limited targeted therapies, leading to higher mortality compared with other breast cancer subtypes. In our previous study we identified N-acyl sphingosine amidohydrolase 1 (ASAH1) to be overexpressed in TNBC cells and demonstrated its role in promoting TNBC growth and progression. ASAH1 is a metabolic enzyme that converts ceramide into sphingosine and free fatty acids, thereby creating a favorable environment for tumor growth. To gain a more comprehensive understanding of the metabolic alterations associated with ASAH1 inhibition, we performed a large-scale metabolomic analysis of TNBC cells expressing ASAH1 shRNAs. Our analysis revealed a significant increase in carnosine levels following ASAH1 inhibition. Subsequent studies demonstrated that exogenous treatment of TNBC cells with carnosine inhibited their growth. Moreover, combining carnosine with ASAH1 inhibitors (carmofur or ceranib-2) resulted in potent synergistic inhibition of TNBC growth. Mechanistically, the combined treatment led to a greater reduction in mitochondrial membrane potential, increased mitochondrial superoxide production and further increased apoptosis. Collectively, these results identify a new metabolism-based combination therapeutic strategy for the effective treatment of TNBC.

To investigate the link between skin features and diabetes, we performed a multi-modal study combining a diabetic mouse model, a human pilot cohort, and keratinocyte response assays. Our goal was to identify molecular signatures in skin, cells, and sweat that correlate with glycemic control and could serve as early biomarkers of hyperglycemia. This integrative approach advances understanding of diabetes-associated dermatological changes and supports the development of targeted, non-invasive diagnostic tools for early detection.

Following glucose tolerance tests, proteomic and metabolomic profiling was performed. We identified dihydrolipoyl-succinyltransferase (DLST) as a promising biomarker whose protein and transcript levels consistently correlated with glycemic status across models. In HaCaT keratinocytes, DLST was found to drive metabolic reprogramming toward glutamine utilization as part of an antioxidant protective response. Chromatin immunoprecipitation assay revealed that Sp1 binds to the DLST promoter region, while functional studies, including Sp1 overexpression and knowdown, further confirmed that Sp1 regulates DLST at transcriptional level. Furthermore, DLST knockdown reproduced metabolic shifts in glutamine pathways, findings supported by proteomic analyses and stable isotopic flux analysis.

Reduced DLST expression in healthy skin corresponded with glutamine metabolism changes observed in keratinocytes upon oral glucose ingestion. While healthy skin and sweat displayed opposing metabolic trends, pre-type 2 diabetes samples showed aligned trends, suggesting sweat metabolomics could serve as an early systemic metabolic indicator.

These findings identify DLST as a glucose-responsive biomarker reflecting skin and sweat responses to systemic glucose, providing a foundation for the development of targeted, non-invasive diagnostic tools for early detection of diabetes.

Objective

Cholecystokinin (CCK) has been implicated in coordinating lipid digestion and satiety. However, the molecular mechanisms underlying nutrient-dependent CCK-release from human I-cells are not well understood, at least in part due to the cells’ scattered distribution and a lack of reliable CCK-assays. Here we used human intestinal organoids to characterize lipid-sensing mechanisms underlying CCK release.

Methods

Human duodenal organoids were genetically engineered using CRISPR-Cas9 to insert either the fluorescent protein Venus or the cAMP reporter Epac-S-H187 at the CCK locus. Transcriptomic profiling of CCK-positive and negative cells was performed following fluorescent-activated cell sorting. Intracellular calcium and cAMP as well as CCK-GRAB-sensor secretory responses were assessed during live-cell imaging, and CCK secretion was quantified by LC-MS/MS. To evaluate lipid-sensing pathways, FFAR1, FFAR4, and GPR119 knockout organoids were generated.

Results

Transcriptomic analysis identified expression of lipid-sensing GPCRs in I-cells, including FFAR1, FFAR4, FFAR2, GPR119, GPBAR1, OR51E1 and OR51E2. Fatty acids and FFAR1 agonists increased intracellular calcium, whereas agonists of GPBAR1 and GPR119, and short-chain fatty acids elevated cAMP. CCK release was triggered by FFAR1, GPBAR and GPR119 agonists, and fatty acids with chain-length > C₈. FFAR1 knockout organoids exhibited impaired Ca²⁺ and CCK secretory responses to fatty acids, whereas responses of FFAR4 KO organoids were like wild-type.

Conclusions

In human I-cells, FFAR1 plays a crucial role in lipid-induced CCK release, whereas FFAR4 appears to be redundant in this context. These findings improve understanding of human CCK physiology and identify potential therapeutic targets for metabolic regulation and appetite control.

Type 2 Diabetes (T2D) is a complex disease that has increased in prevalence in the last years affecting over 530 million people around the world. T2D increases mortality and morbidity by different mechanisms. As a complex disease, it is caused by many genetic changes and, consequently, T2D genetics has been analyzed from different points of view and using different strategies. Most studies have focused on inherited Single Nucleotide Variants (SNVs) present in genomic DNA, while other types of genetic variants have received less attention.

The aim of the present review is to examine the limitations of the studies conducted to date, and the importance of non-SNVs and non-germinal variants in the genetics of this disease. In this way, structural variants, mobile elements and tandem repeats have a crucial role in T2D. In addition, variations in mitochondrial DNA and somatic variations have been shown to play an important role in metabolic alterations that may lead to the development of T2D.

Pancreatic beta cells have the unique function of synthesizing and secreting high amounts of the inhibitory neurotransmitter γ-aminobutyric acid (GABA). The mechanism of GABA secretion, whether vesicular or channel-mediated, is debated. Our study reveals surprising temporal complexity in the pattern of islet GABA secretion. We used insulin secretion modulators to demonstrate that GABA release is not directly correlated with insulin secretion. VGAT reporter mice also showed that beta cells do not express the requisite vesicular GABA transporter (VGAT) for vesicular GABA release. Instead, GABA is secreted from the cytosol in pulses that depend on expression of the LRRC8D subunit of the volume regulatory anion channel (VRAC). We further demonstrate the dynamic coordination of GABA release with calcium influx in beta cells and dependence on beta cell depolarization. These results suggest a model where GABA is released during the peaks of beta cell calcium oscillations to provide feedback which strengthens and reinforces the oscillation waveform.

Background

β-klotho (KLB), the obligate co-receptor for fibroblast growth factor (FGF) 19 and 21 signalling, has emerged as a therapeutic target for metabolic disease. Current understanding of KLB function is based largely on germline knockout mouse models, where developmental abnormalities obscure its function in adult physiology. Here, we used somatic hepatocyte-specific gene editing in adult mice to define hepatic KLB function.

Methods

Hepatocyte-specific KLB deficiency was induced by retro-orbital delivery of adeno-associated virus particles containing guide RNAs targeting Klb in adult male mice expressing Cas9 in the liver. Following transduction, mice were maintained on a 4-week chow diet or a 12-week high-fat diet, prior to comprehensive metabolic and biochemical phenotyping.

Results

Adult-onset hepatic KLB deficiency did not alter body weight or composition, indicating that reduced bodyweight in germline models reflects developmental effects. In line with previous studies, KLB deficiency resulted in derepressed bile acid synthesis, elevated faecal bile acid excretion, and a shift towards a more hydrophobic bile acid pool. Importantly, we also uncovered a role for hepatic KLB in regulating lipid homeostasis. Chow-fed KLB-deficient mice exhibited reduced circulating very-low-density lipoprotein levels (VLDL) with marked hepatic triglyceride accumulation, suggesting altered VLDL metabolism as a potential mechanism driving steatosis.

Conclusions

Using an adult-onset liver-specific knockout model, we disentangled KLB function from developmental confounders and established KLB as a critical regulator of adult lipid and lipoprotein homeostasis. Our study redefines the physiological role of KLB in hepatic metabolism and has important implications for the interpretation of FGF19/KLB-directed therapies for metabolic diseases.

The molecular circadian clock coordinates cellular and systemic metabolism across the day, yet how it integrates with energy-sensing pathways in human skeletal muscle remains incompletely understood. AMP-activated protein kinase (AMPK) functions as a cellular energy sensor and has been implicated in circadian regulation; however, its role in coupling clock components to metabolic programming in human muscle has not been established.

Here, we investigated the interaction between AMPK signaling and the circadian machinery in primary human myotubes, focusing on the downstream clock components NPAS2 and DBP. Silencing of the AMPKα subunit altered core clock gene expression, increasing CRY1 and CRY2 while reducing NPAS2 and DBP expression, with rhythmicity preserved. To determine the functional relevance of these changes, we examined metabolic phenotypes following siRNA-mediated silencing of NPAS2. NPAS2 knockdown reduced basal oxidative metabolism, decreased abundance of electron transport chain subunits, and increased β-oxidation flux, while maximal mitochondrial respiratory capacity remained intact. These alterations occurred independently of canonical AMPK–ACC signaling but were associated with reduced HIF1A expression. In contrast, DBP silencing reduced glucose oxidation but did not recapitulate the broader metabolic remodeling observed with NPAS2 knockdown.

Together, these findings identify NPAS2 as a downstream mediator linking AMPK-dependent circadian signaling to skeletal muscle metabolic regulation. Disruption of this axis alters substrate utilization and impairs mitochondrial function, revealing a mechanism by which circadian clock components contribute to metabolic regulation in human skeletal muscle.