Glucagon receptor (GCGR) signaling is essential for glucose and lipid homeostasis, making it a potential therapeutic target for metabolic disorders. Zebrafish possess two GCGR co-orthologs, GCGRa and GCGRb, however, their distinct function remain unclear. In this study we employed CRISPR/Cas9 gene editing to generate GCGRa⁻/⁻, GCGRb⁻/⁻, and double-knockout (GCGR⁻/⁻) zebrafish to dissect isoform-specific functions. RNA-Seq analysis was performed to characterize transcriptomic alterations, while an overfeeding protocol was used to assess metabolic tolerance, and ligand-response assays in cell lines evaluated isoform activation dynamics. Transcriptomic analysis revealed that both isoforms regulate overlapping but distinct metabolic pathways. Functional enrichment analysis linked GCGRa to lipid and energy metabolism, cholesterol biosynthesis and glucose homeostasis, through key signaling cascades such as glucagon, PPARγ and PI3K-AKT. In contrast, GCGRb loss altered fatty acid β-oxidation, GPCR signaling, and oxidative phosphorylation networks, implicating roles in metabolism and cellular stress. The GCGR⁻/⁻ primarily impacted core metabolic networks including lipid, gluconeogenesis and energy metabolism, indicating complementary and overlapping functions of both receptors in maintaining hepatic metabolic homeostasis. Ligand-response assays revealed that GCGRb, but not GCGRa, is activated by both glucagon (GCGa) and glucagon like-peptide-1 (GLP1a), supporting the post-duplication receptor diversification theory. Notably, all knockouts exhibited impaired growth under high-nutrient conditions, confirming GCGR’s role in diet-responsive development. This study provides the first systematic functional comparison of zebrafish GCGR isoforms, establishing zebrafish as a valuable model for investigating glucagon-based metabolic regulation and therapeutic interventions.

Prediabetes represents a critical window of immune–metabolic dysregulation during which insulin resistance, low-grade inflammation, and barrier dysfunction emerge before overt diabetes. Human β-defensins (HBDs), classically described as antimicrobial peptides, are increasingly recognized as modulators of epithelial integrity, inflammatory signaling, and host–microbiota interactions process central to early metabolic deterioration. Evidence from genetic, experimental, and clinical studies indicates that alterations in HBD expression accompany insulin resistance, β-cell stress, and gut barrier impairment, with tissue-specific patterns observed across the dysglycemic spectrum. While most human data derive from established diabetes, animal models and limited human observations suggest that defensin dysregulation may arise earlier and contribute to the transition from prediabetes to diabetes. Importantly, HBDs are detectable in saliva, serum, and tissues, supporting their feasibility as accessible biomarkers of early immune–metabolic stress. However, heterogeneity in assay platforms, sample matrices, and study design currently limit clinical translation. This review synthesizes current evidence linking β-defensins to early metabolic dysfunction, distinguishes associative human findings from mechanistic experimental data, and highlights critical gaps in prediabetes-focused research. We propose that β-defensins represent promising early immune–metabolic indicators whose validation in longitudinal prediabetes cohorts may improve risk stratification and enable earlier intervention.

Matrix metalloproteinase 9 (MMP-9) plays a key role in the pathogenesis of inflammatory diseases and is upregulated by TNF-α. ITE (2-[1'H-indole-3'-carbonyl]-thiazole-4-carboxylic acid methyl ester) functions as an endogenous ligand for the aryl hydrocarbon receptor and is involved in inflammation. It is still uncertain whether ITE could affect TNF-α-induced MMP-9 expression in monocytic cells. In this study, we explored the effect of ITE on TNF-α-induced MMP-9 expression and the underlying mechanisms involved. Our results show that pretreatment of THP-1 monocytic cells with ITE significantly blocked TNF-α-induced MMP-9 expression at both the mRNA and protein secretion levels. Similar results were seen in primary human monocytes. The inhibition of MMP-9 by ITE occurs independently of TNFR1/2 modulation and apoptotic processes. RNA transcription data revealed that ITE suppresses the genes associated with inflammatory pathways. Mechanistically, histone modification profiling identified H3K9 acetylation as an epigenetic regulatory mark of TNF-α-induced MMP-9 expression. ChIP-qPCR data revealed that ITE pretreatment decreased TNF-α-triggered transcriptionally permissive acetylation marks at H3K9 in the MMP-9 promoter. Pharmacological inhibition of histone acetylation mimics the action of ITE in suppressing TNF-α-induced MMP-9 gene expression. Conversely, the acetylation induced by trichostatin A effectively reverses the inhibitory action of ITE. Moreover, increased TNF-α-induced binding of NF-κB or AP-1 at the MMP-9 promoter region was inhibited by ITE, resulting in suppression of MMP-9 gene expression. In conclusion, our study demonstrates that ITE reduces the TNF-α-induced MMP-9 expression via the H3K9 acetylation/NF-κB/AP-1 axis, highlighting a potential mechanism for mitigating MMP-9-related inflammatory disorders.

PURPOSE:MicroRNA (miRNA) profiling of visceral adipose tissue in Type-2 diabetes mellitus (T2DM) remains limited. We compared the expression of obesity-associated miRNAs in visceral fat from individuals with T2DM versus metabolically healthy obesity (MHO) and examined Raf kinase inhibitory protein (RKIP) as a candidate miR-543 target. METHODS:Visceral fat biopsies were obtained from adults with T2DM (n = 8) and MHO (n = 11). Thirteen miRNAs previously linked to obesity were quantified, and RKIP expression was evaluated at the mRNA level and by immunohistochemistry, including phosphorylated RKIP (pRKIP). RESULTS:Of the 13 miRNAs analyzed, miR-23a-3p and miR-543 were upregulated in T2DM (p = 0.032 and p = 0.009, respectively), whereas miR-320a-3p was downregulated (p = 0.009). RKIP mRNA levels did not differ between groups; however, in MHO adipocytes, RKIP mRNA correlated positively with miR-543 expression (r = 0.655, p = 0.034). Total RKIP protein was comparable between groups, while pRKIP was significantly higher in T2DM adipocytes (p = 0.012). CONCLUSIONS:Posttranscriptional regulation in visceral adipocytes differs between T2DM and MHO. Increased miR-543 and elevated pRKIP in T2DM suggest a rapid shift in regulatory signaling consistent with enhanced lipolysis and a proinflammatory milieu. Further studies are warranted to delineate the pRKIP-associated pathways in adipose tissue remodeling in T2DM.

Introduction and Objective: Metabolic-associated steatotic liver disease (MASLD) is an obesity-associated condition marked by hepatic steatosis and chronic inflammatory signalling driven by proinflammatory cytokines, including IL-6 and TNF-α. Although TNF-α is a major mediator of liver inflammation and injury, the mechanisms by which IL-6 regulates TNF-α expression, signaling, and activation remain unclear. Methods: Male C57BL/6J (wild-type; WT) and IL-6 knockout (IL-6KO) mice were fed either a high-fat diet (HFD) or standard chow for 16 weeks. Recombinant mouse IL-6 was administered to IL-6 KO mice to assess rescue of TACE/ADAM17 expression and TNF-α signaling. Metabolic and immunological profiling was done. Analyses were performed using RNA-seq, qRT-PCR, flow-cytometry, IHC, Western blotting, ELISA, and Oil Red O staining. Results: Under high fat diet conditions, IL-6KO mice showed significantly lower systemic and hepatic TNF-α levels, reduced hepatic steatosis, and improved insulin sensitivity relative to WT mice. Hepatic TNF-α signaling was significantly impaired in HFD-fed IL-6KO mice, as evidenced by reduced TNFR1/2 expression and attenuated activation of ERK1/2, MEK1/2, JNK, c-Jun, p38, and NF-κB. Ex-vivo TNF-α stimulation failed to fully activate downstream signaling in liver tissue from IL-6KO mice, confirming impaired TNF-α responsiveness relative to WT mice. Correspondingly, inflammatory outputs including macrophage infiltration and lobular inflammation were attenuated. Hepatic TACE/ADAM17 expression and activity were reduced in IL-6KO mice, while TIMP3 was increased. IL-6 administration restored hepatic TACE/ADAM17 function and reactivated downstream inflammatory signaling. Conclusion: These data indicate that IL-6 plays a critical role in shaping TNF-α pathogenic activity in liver, suggesting that targeting IL-6 signalling may attenuate TNF-α-mediated liver injury in MASLD and steatohepatitis. Disclosure N. Benobaid: None. S. Kochumon: None. F. Bahman: None. S. Shenouda: None. N. Almansour: None. F. Alrashed: None. A. Al Madhoun: None. F. Al-Mulla: None. E. Rosen: Consultant; Ended; Novartis AG. Research Support; Current; Roche Pharmaceuticals, Alnylam Pharmaceuticals, Inc. R. Ahmad: None. Funding Kuwait Foundation for the Advancement of Sciences (KFAS) (RA AM-2023-021)

Introduction and Objective: Metabolic dysfunction-associated steatotic liver disease (MASLD) results from dysregulated hepatic lipid handling. Here, we uncover an inflammation independent metabolic role for Toll-like receptor 2 (TLR2) that protects against hepatic steatosis and establish proof of concept for its therapeutic potential. Methods: C57BL/6J (WT) and TLR2 knockout (KO) male mice were fed a high-fat diet (HFD) to induce MASLD for 16 weeks. TLR2KO and WT mice were then treated with Pam3CSK4. Hepatic steatosis, plasma lipid profiles, and glucose tolerance (GT) were assessed. In vivo and in vitro samples were examined by RNA-seq, qRT-PCR, FACS, histology, ChIP-qPCR, W-blot, and siRNA transfection. Human liver biopsies from patients with MASLD were used for translational validation. Results: TLR2 deficiency led to obesity, insulin resistance, and marked hepatic steatosis that occurred independently of hepatic inflammation. Mechanistically, loss of TLR2 increased hepatocyte lipid accumulation via a cell-autonomous pathway involving upregulation of PPARγ-CD36. Genetic deletion of PPARγ or CD36 mitigated lipid accumulation induced by TLR2 loss. Activation of TLR2 in mice with the selective agonist Pam3CSK4 suppressed hepatic PPARγ-CD36 signaling, reduced lipid uptake and storage, and prevented steatosis. In obese WT mice, Pam3CSK4 treatment reversed established hepatic steatosis, improved GT, reduced TG, increased plasma HDL-C, and enhanced lipoprotein export, accompanied by induction of hepatic lipid oxidation and export genes and proteins (Ppara, Cpt1a, Acox1, ApoB, and Mttp). These metabolic benefits were absent in TLR2KO mice, confirming strict TLR2 dependency. Human MASLD livers exhibited low TLR2, increased fat, and elevated PPARγ-CD36. Conclusion: TLR2 exerts a protective role in hepatic lipid homeostasis by suppressing the PPARγ-CD36 lipid uptake axis while promoting lipid oxidation and export. Activation of TLR2 by Pam3CSK4 reverses hepatic steatosis in preclinical murine models, highlighting its therapeutic promise for MASLD. Disclosure S. Kochumon: None. F. Alrashed: None. F. Alzaid: None. F. Bahman: None. N. Benobaid: None. T.K. Jacob: None. R. Nizam: None. N. Akhter: None. A. Al Madhoun: None. F. Al-Mulla: None. E. Rosen: Consultant; Ended; Novartis AG. Research Support; Current; Roche Pharmaceuticals, Alnylam Pharmaceuticals, Inc. R. Ahmad: None. Funding Kuwait Foundation for the Advancement of Sciences (KFAS). Grant# RA AM 2023-021

BACKGROUND:The aryl hydrocarbon receptor (AhR) is linked to inflammation, but its plasma agonist activity and association with metabolic and inflammatory markers in obesity remain unclear. This cross-sectional study aimed to determine the level of plasma AhR agonistic activity and its association with systemic inflammation and metabolic dysregulation in obesity. METHODS:Plasma samples were collected from 80 non-diabetic (39-obese, 23-overweight, and 18-normal/healthy weight) individuals. AhR agonist activity was assessed using a cell-based luciferase reporter assay. Plasma AhR was quantified by ELISA. Inflammatory markers were assessed using a multiplex Luminex platform. RESULTS:Our findings indicate that plasma AhR agonist activity is elevated in obese (92.77 ± 4.002 fold activation) compared with normal/healthy weight (51.39 ± 2.335) and overweight participants (67.54 ± 5.24 fold activation). Moreover, the AhR protein was also elevated in obese (94.88 ± 7.62 pg/ml) compared to normal/healthy weight (65.88 ± 6.78 pg/ml) and overweight participants (67.54 ± 5.24 pg/ml), which was positively correlated with AhR activity (r = 0.441, p < 0.0001). AhR activity was positively correlated with inflammatory markers including IL-1β, IL-6, TNF-α, TNF-β, and MCP-1, as well as metabolic markers such as BMI, total cholesterol, TG, insulin, FBG, and HbA1c. In contrast, it was negatively associated with HDL cholesterol. Notably, HOMA-IR was positively correlated with AhR activity. In the regression model, TNF-α, MCP-1, BMI, and HDL cholesterol emerged as significant predictors of AhR activity. CONCLUSIONS:Our findings demonstrate that elevated plasma AhR agonist activity is associated with obesity, systemic inflammation, and metabolic dysregulation. These results highlight AhR activity as a biomarker of interest and support further studies to clarify its mechanistic role and potential clinical relevance in metabolic disorders.

Introduction and Objective: Obesity-driven metabolic inflammation promotes insulin resistance and fatty liver disease, with TNF-α as a key mediator. Although histone acetylation is linked to TNF-α activation, direct causal evidence is lacking. This study presents a proof-of-concept investigation to determine whether histone acetylation directly drives persistent TNF-α transcription in obesity. Methods: To establish a causal link between histone acetylation and sustained TNF-α transcription in obesity, male C57BL/6J-mice were fed a high-fat diet for 16 weeks to induce diet-induced obesity (DIO) and subsequently treated for 4 weeks with either the histone acetyltransferase inhibitor anacardic acid or a vehicle control. Metabolic and immunological profiling was done. Chromatin immunoprecipitation (ChIP-qPCR), RNA sequencing, qRT-PCR, IHC, and ELISA techniques were employed. Results: DIO markedly increased TNF-α expression and elevated global and promoter-specific H3K9/H3K18 acetylation in liver and adipose tissue. Enrichment of these histone marks at the TNF-α promoter correlated with body-weight gain, insulin resistance, and hepatic steatosis. DIO mice treated with anacardic acid showed a significant reduction in H3K9/H3K18 acetylation at the TNF-α promoter and decreased NF-κB promoter binding, resulting in lower TNF-α transcription. These epigenetic improvements were accompanied by reduced inflammatory markers, including CCL2 expression, macrophage (F4/80+) infiltration in both liver and visceral fat tissue. Anacardic acid intervention improves metabolic impairment. Conclusion: DIO establishes a transcriptionally permissive chromatin environment at the TNF-α locus through H3K9/H3K18 acetylation, enabling sustained NF-κB-dependent TNF-α expression and metabolic deterioration. Pharmacological inhibition of H3K9/H3K18 acetylation reverses transcriptional changes, providing direct causal evidence for the role of chromatin remodeling in obesity-driven inflammation, insulin resistance and hepatic steatosis in obesity. Disclosure F. Bahman: None. S. Kochumon: None. N. Benobaid: None. T.K. Jacob: None. A. Al Madhoun: None. F. Al-Mulla: None. R. Ahmad: None. Funding Kuwait Foundation for the Advancement of Sciences(KFAS)

Myosteatosis, defined as pathological lipid accumulation within and between skeletal muscle fibers, is increasingly recognized as a determinant of impaired muscle quality, metabolic inflexibility, and adverse clinical outcomes. Although well described in ageing, obesity, and cancer, its relevance to type 1 diabetes (T1D) remains underexplored. T1D is characterized by lifelong insulin deficiency, persistent autoimmune activation, and glycemic variability, conditions that profoundly disrupt cellular energy metabolism and substrate utilization in skeletal muscle, even in the absence of obesity or overt sarcopenia. This review integrates evidence from human imaging, metabolic phenotyping, immunological profiling, and multi-omics analyses to define myosteatosis as an immunometabolic phenotype in T1D. Central to this framework is dysregulation of the AMP-activated protein kinase (AMPK)-peroxisome proliferator-activated receptor (PPAR)-mitochondrial axis, which normally coordinates fatty-acid oxidation, mitochondrial biogenesis, and energy efficiency in skeletal muscle. In T1D, chronic immune activation and metabolic stress suppress AMPK and PPARδ signaling, impair PGC-1α–dependent mitochondrial function, and reduce oxidative capacity, promoting intramyocellular lipid accumulation despite preserved muscle mass. These defects are reinforced by persistent inflammatory signaling (IL-6, TNF-α, IL-1β; NF-κB, JNK, and NLRP3 pathways), accumulation of lipotoxic intermediates (ceramides and diacylglycerols), dysregulated myokine secretion (increased myostatin with reduced IL-15 and irisin), and infiltration of pro-inflammatory macrophages and CD8+ T cells. Mitochondrial stress, reflected by impaired phosphocreatine recovery, altered acylcarnitine profiles, increased oxidative damage, and reduced NAD+–SIRT1/3 activity, further consolidates immunometabolic dysfunction and lipid deposition. Collectively, this review positions myosteatosis as a clinically relevant and potentially modifiable consequence of immune-driven failure of cellular energy utilization in T1D. Because direct mechanistic data from T1D skeletal muscle remain scarce, the framework presented here is deliberately hypothesis-generating: it is assembled substantially by inference from type 2 diabetes (T2D), obesity and ageing models, and we map the resulting evidence gaps explicitly in order to define a research agenda rather than to assert a validated T1D-specific mechanism. Targeting the AMPK-PPAR-mitochondrial axis and its inflammatory and lipotoxic modifiers may enable earlier detection and mechanism-based interventions to preserve muscle metabolic resilience and functional capacity in autoimmune diabetes.

The high prevalence of insulin resistance and metabolic syndrome in Kuwait increases the risk of type 2 diabetes (T2D) and cardiovascular diseases development. Although the genetic contribution to insulin resistance and metabolic syndrome is established, the role of a disintegrin and metalloprotease 12 (ADAM12) in insulin resistance and T2D remains unclear. Our GWAS has identified four novel ADAM12 variants, namely, ADAM12-K170R rs112264074 [NP_003465.3:p.Lys170Arg], ADAM12-R176W rs140497576 [NP_003465.3:p.Arg176Trp], ADAM12-M662V rs115100580 [NP_003465.3:p.Met662Val], and ADAM12-I908V rs41303603 [NP_003465.3:p.Ile908Val]. These variants were associated with different metabolic traits, including FBG, HbA1c, low- (LDL) and high-density lipoprotein (HDL), total cholesterol (TC), and diastolic and systolic blood pressure. Using an independent replication cohort, the association of ADAM12-I908V with FBG was confirmed, and an association with HbA1c, LDL, and TC was found. In addition, ADAM12-K170R showed associations with waist-hip ratio in diabetic patients, and ADAM12-M662V associated with HDL, TC, and HbA1c in healthy controls, while ADAM12-R176W variant was not detected in the replication cohort. Moreover, we examined the impact of ADAM12 variants on its proteolytic activity and results show that 176W and 662V variants had higher activity in cell lysate supernatants, but only 662V and 908V variants had higher activity in intact cells, suggesting that enzyme activity dysregulation may contribute to the development of metabolic syndrome and T2D.

Interleukin-6 (IL-6) is a central mediator of chronic low-grade inflammation associated with metabolic disease. Because obesity is characterized by elevated circulating insulin and metabolic endotoxemia, we investigated whether insulin modulates lipopolysaccharide (LPS) induced IL-6 expression in adipocytes and examined the underlying epigenetic mechanisms. Insulin priming markedly enhanced LPS-induced Il6 mRNA expression (25.33 ± 0.833-fold) and protein levels (181.8 ± 2.754 pg/ml) in 3T3-L1 mouse adipocytes. Similar synergistic effects were observed in primary mouse (Il6 mRNA; 1.364 ± 0.287-fold and protein; 298.6 ± 13.79-pg/ml) and human adipocytes (Il6 mRNA; 12.99 ± 0.912-fold and protein; 1441 ± 68.69-pg/ml). In vivo, mice treated with insulin followed by LPS exposure exhibited significantly higher Il6 expression in peripheral blood mononuclear cells and adipose tissue compared to either treatment alone. Pharmacological inhibition of PI3K signaling suppressed this effect and AKT phosphorylation. Mechanistically, epigenetic profiling revealed that insulin increased histone H3 lysine 9 acetylation (H3K9ac), an active chromatin marker, in a PI3K-dependent manner. Chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR) analysis demonstrated an enhanced H3K9 acetylation at the NF-κB and CREB loci at the distal region and CREB/NF-IL6 locus at the proximal region of the Il6 promoter following combined insulin and LPS stimulation; this effect was significantly attenuated upon blockade of insulin signaling. This synergistic induction was dependent on H3K9 acetylation, indicating that metabolic and inflammatory signals converge at the Il6 promoter to promote chromatin remodeling and transcriptional co-activator recruitment. Collectively, these findings demonstrate that insulin synergizes with LPS to amplify IL-6 mediated inflammation in adipocytes through epigenetic remodeling of the Il6 locus, linking hyperinsulinemia to chronic inflammation in obesity and insulin resistance.

Dietary fats are consumed as mixtures, yet it remains unclear whether fatty acid composition, independent of fat content, dictates human macrophage polarization. We compared two defined mixtures containing identical fatty acids (palmitic, oleic, and linoleic acids) in different ratios: a palmitate-enriched mixture (4:3:3) and an unsaturated fat-dominant mixture (2:4:4). In primary human monocyte-derived macrophages, palmitate enrichment increased CD14+CD11b+HLA-DR+ pro-inflammatory polarization, whereas the unsaturated fat-dominant mixture increased CD14+CD11b+CD163+ anti-inflammatory polarization. Mechanistic studies in THP-1-derived macrophages recapitulated these phenotype shifts and identified a reciprocal nuclear-receptor program: palmitate enrichment induced peroxisome proliferator-activated receptor gamma (PPARγ), together with ER-stress mediators EIF2AK3 and DDIT3, while the unsaturated fat-dominant mixture preferentially induced PPARα and IRF4. Pharmacologic modulation demonstrated functional dependence on PPARγ: GW9662 attenuated palmitate-driven M1-like polarization, whereas rosiglitazone disrupted the protective program under unsaturated fat-dominant conditions. These findings show that fatty acid composition, at equivalent total lipid concentration, is a dominant determinant of human macrophage inflammatory fate and highlight PPARγ as a context-dependent lipid sensor.

Overweight and obesity have emerged as global health crises and are increasingly recognized as drivers of central nervous system (CNS) dysfunction. Beyond excess energy storage, white adipose tissue (WAT) functions as an active endocrine and immune organ that, during obesity, undergoes inflammatory remodeling and releases cytokines, lipid mediators, adipokines, and extracellular vesicles that influence brain physiology. These peripheral signals disrupt key brain interfaces, including the blood-brain barrier (BBB), perivascular and glymphatic clearance pathways, promoting endothelial dysfunction, altered astrocyte-pericyte support, impaired amyloid-β clearance, and region-specific glial activation. Obesity-associated neuroinflammation is characterized by microglial priming and astrocyte reactivity across the hypothalamus, hippocampus, and other circuits governing metabolism, cognition, and reward, with growing evidence for sex-dependent vulnerability. We further highlight adipokines as key mediators of adipose-brain communication. In obesity, leptin resistance impairs central energy regulation, reduced adiponectin contributes to neuroinflammation and synaptic dysfunction, and elevated resistin enhances TLR4-dependent inflammatory signaling and BBB permeability, collectively linking metabolic stress to neurodegenerative processes. Finally, we review therapeutic strategies targeting the adipose-brain axis, including exercise and dietary interventions that improve neuroplasticity and barrier integrity, and pharmacological approaches such as orlistat and incretin-based therapies. Emerging multi-incretin agonists, including tirzepatide and retatrutide, raise important questions regarding direct CNS actions beyond metabolic benefits, underscoring the need to integrate barrier biology and neuroimmune mechanisms in future studies.

Introduction and Objective: Persistent IL-6 overexpression in obesity drives hepatic inflammation and steatosis; thus, targeting IL-6 is critical for controlling obesity-associated liver inflammation. The tryptophan metabolite ITE (2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester) exhibits immunomodulatory properties, yet its role in regulating hepatic IL-6 in obesity remains unclear. This study investigates the effects of ITE on IL-6 expression, associated liver inflammation, and the underlying molecular mechanisms. Methods: Male C57BL/6 mice were fed chow or a high-fat diet for 16 weeks, followed by 4 weeks of vehicle or ITE treatment. Hepatic IL-6 and inflammatory markers were assessed by immunohistochemistry, Western blotting, and quantitative RT-PCR. Histone modification profiling was performed. Chromatin immunoprecipitation coupled with qPCR (ChIP-qPCR) was used to evaluate histone methylation marks and transcription factors binding at the Il6 promoter. Results: ITE treatment significantly reduced hepatic IL-6 expression and inflammatory responses, including macrophage accumulation (F4/80), compared with vehicle-treated obese mice. Histone profiling of liver tissue from ITE-treated mice revealed increased enrichment of transcriptionally repressive epigenetic marks, specifically H3K79 and H3K4 trimethylation. ChIP-qPCR confirmed enhanced deposition of H3K79me3 and H3K4me3 at the Il6 promoter. Moreover, obesity-induced recruitment of NF-κB and AP-1 transcription factors to the Il6 promoter was markedly reduced following ITE treatment, resulting in a less transcriptionally permissive chromatin state and suppression of Il6 gene expression. Conclusion: In conclusion, our study demonstrates that ITE reduces the IL-6 expression in liver via H3K79/H3K4 Trimethylation resulting reduced binding of NF-κB/AP-1 to Il-6 promoter, highlighting a potential mechanism for mitigating IL6 -related inflammatory disorders. Disclosure F. Bahman: None. N. Akhter: None. G. Alhamar: None. F. Al-Mulla: None. R. Ahmad: None. Funding KFAS (RA-AM-2023-023)

Epidemiological evidence shows that obesity increases the risk of developing metabolic diseases. Nevertheless, the mechanisms behind this connection remain underappreciated. The substantial impact of these disorders on global health has led to extensive research efforts aimed at identifying the pathophysiological links between them. Chronic low-grade inflammation, induced by altered secretion of adipokines and other bioactive molecules, from adipose tissue, is believed to causally link obesity to various metabolic disorders. Multiple studies have indicated that TLR4 regulates inflammation, adipogenesis, thermogenesis, and glucose metabolism through its interaction with endotoxins, particularly in the context of obesity. The increased expression of TLR4 observed in obesity is believed to contribute to the development of type 2 diabetes (T2D), as it disrupts key physiological processes that regulate metabolic inflammation. This review aims to summarize recent research on the pathobiological roles of TLR4-mediated inflammation in obesity and its contribution to the development of metabolic disorders. Overall, current evidence supports a central role for TLR4 as a mediator of obesity-associated metabolic inflammation, highlighting TLR4 and its downstream pathways as promising targets for preventing or treating obesity related metabolic diseases.

Low-fat diets are widely promoted as health-protective; however, the consequences of removing sucrose within a low-fat dietary framework remain unclear. Here, we investigated the effects of a sucrose-free low-fat diet (SF-LFD) compared with a sucrose-containing low-fat control diet (C-LFD) in mice (n=6/group) over 16 weeks. Despite unchanged body and liver weights, SF-LFD feeding resulted in impaired glucose tolerance, reduced insulin sensitivity, and broad alterations in circulating metabolic hormones, including elevated C-peptide, incretins, ghrelin, and resistin, as well as reduced fasting insulin. 16S rRNA sequencing revealed that SF-LFD markedly disrupted gut microbial diversity and composition, with depletion of short-chain fatty acid–producing commensals, including Lactobacillus murinus and members of the Lachnospiraceae family, and enrichment of taxa associated with inflammatory or stress-adapted states, including Helicobacter ganmani, Odoribacter splanchnicus, and Alistipes species. This dysbiosis was accompanied by pronounced colonic inflammation characterized by crypt architectural disruption, loss of goblet cells, submucosal expansion, increased CD3+ T-cell and F4/80+ macrophage infiltration, and robust upregulation of inflammatory mediators, including Il1b, Il6, Ccl2, Rorγt, and Tbx21. SF-LFD feeding induced hepatic microvesicular steatosis, lobular inflammation, recruitment of F4/80+ and CD11c+ immune cells and increased hepatic expression of IL1b and IL6. Together, these findings suggest that sucrose elimination from a low-fat diet disrupts gut microbiota, impairs metabolic homeostasis, and promotes gut and liver inflammation, revealing an unrecognized dietary trigger of metabolic dysfunction.

Abstract Introduction Chronic low-grade inflammation is a hallmark of obesity and a key driver of metabolic dysfunction and cardiovascular disease. However, the immunological mechanisms linking inflammation to early metabolic and vascular risk in prediabetes remain unclear. Activated CD8+HLA-DR+ T cells represent a marker of sustained immune activation, yet their contribution to β-cell stress and cardiometabolic dysfunction has not been defined. Methods A cross-sectional study of 198 adults was conducted, stratified as lean healthy (n = 88), healthy obese (n = 69), and prediabetic obese (n = 41). Circulating CD8+HLA-DR+ T cells were quantified by flow cytometry and correlated with metabolic (glucose, insulin, C-peptide, HOMA-B%) and cardiovascular markers (lipids, blood pressure, MMP-9). Plasma cytokines were measured using a 38-plex Luminex assay. Findings were validated using public whole-blood transcriptomic data (GSE145412). Results Prediabetic obese individuals exhibited significantly higher frequencies of CD8+HLA-DR+ T cells compared with healthy obese and lean groups (p < 0.001). These activated T cells correlated positively with C-peptide, IL-6, TNF-α, IFN-γ, and IL-17A, and inversely with HOMA-B%, linking immune activation to β-cell stress and inflammation. CD8+HLA-DR+ frequency also associated with diastolic pressure, triglycerides, and MMP-9; multivariable analysis identified MMP-9 as an independent correlate (p = 0.04). Transcriptomic validation confirmed parallel immune-metabolic signatures in obese diabetic subjects. Conclusion Activated CD8+HLA-DR+ T cells emerge as key immune biomarkers linking inflammation, β-cell dysfunction, and cardiovascular risk in prediabetic obesity. Their association with MMP-9 highlights a potential immunometabolic axis driving vascular remodeling. These findings identify CD8+HLA-DR+ T-cell activation as a promising early indicator and therapeutic target for cardiometabolic risk prevention. Funding Source Kuwait Foundation for the Advancement of Science (KFAS)-RA MoH-2022-002 Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)

Furin, a proprotein convertase, regulates glucose homeostasis by processing the insulin receptor (IR) precursor. Although the association of furin genetic variants with cardiac and neuronal diseases is well-established, studies investigating the association with type 2 diabetes (T2D) are scarce. This study aimed to examine the association of furin variants with T2D in an Arab cohort. In addition, it sought to elucidate the functional impact of these diabetes-associated variants on furin stability and kinetic activity. Of the 15 rare missense variants in furin identified in global genomic studies, only one, rs148110342_C > T_(R81C), was found in our study cohort, with a minor allele frequency of 2.4%. Allele-based association testing, adjusted for age, sex, and body mass index, revealed significant associations between the rs148110342 and being T2D and borderline associations with fasting plasma glucose and HbA1c levels. Enzyme kinetic studies showed that the R81C variant has higher Km values, indicating lower enzymatic activity compared with wild-type furin. In silico structural modeling of the interactions between the R81C variant prodomain and the furin catalytic subunit revealed an increase in hydrogen bonding, which might explain the observed reduction in enzymatic activity. Furthermore, cell culture studies suggested that the R81C variant impairs furin's autocatalytic processing and its ability to cleave the precursor insulin receptor. A significant reduction in phosphorylation of ERK1/2 and AKT occurred in HEPG2 cells transfected with R81C variants, suggesting a downregulation of the IR signaling pathway. These findings suggest that the furin R81C variant can potentially impact insulin signaling and thereby contribute to T2D pathogenesis.NEW & NOTEWORTHY This study contributes novel insights into the role of furin variants in T2D risk. The rare rs148110342_C > T_(R81C) variant of furin exhibits a minor allele frequency of 2.4% in Arabs. We observed significant associations between the variant and being diabetic. The variant furin revealed lower enzyme kinetic activity, impairment of furin's autocatalytic processing, and a significant reduction in ERK1/2 and AKT phosphorylation. These findings suggest that the variant downregulates the IR signaling pathway.

Gut microbiota research has highlighted its pivotal role in human health and disease. Its composition is shaped by diet, genetics, age, and environmental factors. When the balance of these microbes is disrupted (dysbiosis), it can contribute to health problems like metabolic, inflammatory, and mental disorders. The microbiota supports digestion, fermentation, and vitamin production, which are essential for overall health. The gut microbiota has emerged as a critical modulator of immune function, with increasing evidence highlighting its role in establishing and maintaining immune tolerance. Despite significant advances in understanding the interactions between the gut microbiome and immune system, gaps remain in the literature regarding the specific mechanisms through which microbiota influences immune tolerance. This review aims to address these knowledge gaps by synthesizing current research on the microbiota impact on immune tolerance, emphasizing key factors such as microbial diversity, metabolic byproducts, and the microbiota interaction with immune cells, specifically focusing on the role of microbial tryptophan metabolites in PD-1/PD-L1 tolerance. We also highlight critical areas for future research, including the identification of microbial species or strains that can modulate immune tolerance, the influence of diet and environmental factors on microbiota composition, and the development of microbiota-based therapies. By bridging these gaps, this review seeks to provide a comprehensive understanding of the mechanistic role of microbiota immune tolerance and its potential as a novel therapeutic target for autoimmune and inflammatory diseases.