Search Results (42)

Background/Objectives: Metabolic dysfunction-associated steatotic liver disease (MASLD) involves the interplay of hepatic lipid accumulation and immune-mediated inflammatory signaling, yet human-relevant in vitro systems that capture both processes simultaneously in a scalable format remain limited. The objective of this study was to develop and characterize a matrix-free 3D hepatocyte–macrophage co-culture model enabling simultaneous assessment of lipid accumulation and NF-κB-mediated inflammatory activation under glucolipotoxic stress. Methods: A 3D liver co-culture model was established by combining HepG2 hepatocyte-like cells with phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophage-like cells stably expressing a NF-κB–Luc2 reporter. Spheroids were generated using a hanging-drop method in standard 96-well plates and matured for 8–10 days. Mature spheroids were subjected to acute 24 h glucolipotoxic challenge combining high glucose and palmitic acid and assessed for neutral lipid accumulation, NF-κB reporter activation (luciferase), and macrophage marker expression (qPCR). Results: Time-course characterization demonstrated progressive hepatocyte marker remodeling (albumin, alpha-fetoprotein, CYP3A4) and dynamic macrophage phenotype shifts (CD14, CD206, MARCO, TREM2). Acute glucolipotoxic challenge induced dose-dependent increases in neutral lipid accumulation and NF-κB reporter activation, accompanied by coordinated macrophage-associated transcriptional changes consistent with lipid-handling and tissue-remodeling programs. Post-challenge metabolic activity was retained under the selected stress conditions. As a proof-of-concept demonstration, three botanical extracts showed distinct attenuation profiles across the lipid and inflammatory endpoints. Conclusions: This 3D hepatocyte–macrophage co-culture model provides orthogonal readouts of steatosis and NF-κB-mediated inflammatory activation under glucolipotoxic stress, offering a reproducible, fit-for-purpose screening tool for investigating early glucolipotoxic hepatic responses and evaluating candidate compounds in a defined in vitro setting. Full article

Background: Insulin resistance (IR) has emerged as a key player in the pathogenesis of cognitive impairment in Parkinson’s disease (PD). This study aims to systematically compare glucolipotoxicity-based (TyG, AIP) versus adiposity-driven (TyG-BMI, METS-IR) IR indices for their associations with PD dementia and to develop a clinically applicable nomogram using an interpretable machine learning framework. Methods: This cross-sectional study analyzed 251 PD patients: 42 with normal cognition, 160 with mild cognitive impairment (PD-MCI) and 49 with dementia (PDD). Logistic and linear regression examined associations between IR indices and cognitive impairment across different domains. Six machine learning models were compared for dementia classification, with the optimal model interpreted using SHapley Additive exPlanations (SHAP) to construct a nomogram. Results: Each standard deviation increase in TyG and AIP was linked to 79% (OR 1.79, 95%CI 1.04–3.07) and 75% (OR 1.75, 95%CI 1.05–2.91) higher risk of PDD, respectively, but not PD-MCI. In contrast, TyG-BMI and METS-IR showed no significant associations with either condition. TyG showed linear negative correlations with memory and orientation, and inverted U-shaped associations with visuospatial function and attention. AIP exhibited linear negative correlation with memory. The logistic regression model achieved the highest performance (AUC of 0.759) among six machine learning models. Crucially, SHAP analysis visually quantified TyG as a top modifiable predictor, facilitating the construction of an interpretable clinical nomogram. Conclusions: Glucolipotoxicity-based indices (TyG, AIP), unlike BMI-dependent markers (TyG-BMI, METS-IR), are robustly linked to PD dementia through domain-specific linear or nonlinear patterns. This suggests metabolic dysregulation predicts risk independent of weight loss. Furthermore, integrating SHAP-based interpretability transforms complex algorithms into a transparent, actionable tool for early risk stratification. Full article

Prediabetes is accompanied by early β-cell stress and oxidative imbalance before overt hyperglycemia. Circulating extracellular vesicle (EV) microRNAs (miRNAs) may capture early metabolic disturbances, but their mechanistic relevance remains unclear. Plasma EV miRNA profiles were analyzed across normoglycemia, prediabetes, and newly diagnosed type 2 diabetes, with validation in an independent cohort (n = 150). Functional studies were performed in pancreatic β-cells exposed to glucolipotoxic stress to examine miRNA regulation, IFN-α signaling, mitochondrial redox status, and insulin secretion. Six EV miRNAs, including miR-186-5p, were consistently reduced in prediabetes and correlated with glycemic and insulin resistance indices. In β-cells, glucolipotoxic stress selectively suppressed miR-186-5p, leading to derepression of IFNA2, activation of IFN-α–JAK/STAT signaling, increased mitochondrial ROS, impaired ATP/ADP dynamics, and reduced glucose-stimulated insulin secretion. Restoration of miR-186-5p or pharmacologic JAK inhibition mitigated these defects, and luciferase assays confirmed IFNA2 as a direct target of miR-186-5p. EV-associated miR-186-5p represents an early marker of metabolic stress in prediabetes and provides mechanistic insight into IFN-α–driven oxidative and secretory dysfunction in β-cells. Full article

Insulin-producing pancreatic β cells play a crucial role in the regulation of glucose homeostasis, and their failure is a key event for diabetes development. Prolonged exposure to palmitate in the presence of elevated glucose levels, termed gluco-lipotoxicity, is known to induce β cell apoptosis. Autophagy has been proposed to be regulated by gluco-lipotoxicity in order to favor β cell survival. However, the role of palmitate metabolism in gluco-lipotoxcity-induced autophagy is presently unknown. We therefore treated INS-1 cells for 6 and 24 h with palmitate in the presence of low and high glucose concentrations and then monitored autophagy. Gluco-lipotoxicity induces accumulation of LC3-II levels in INS-1 at 6 h which returns to basal levels at 24 h. Using the RFP-GFP-LC3 probe, gluco-lipotoxicity increased both autophagosomes and autolysosmes structures, reflecting early stimulation of an autophagy flux. Triacsin C, a potent inhibitor of the long fatty acid acetyl-coA synthase, completely prevents LC3-II formation and recruitment to autophagosomes, suggesting that autophagic response requires palmitate metabolism. In contrast, etomoxir and bromo-palmitate, inhibitors of fatty acid mitochondrial β-oxidation, are unable to prevent gluco-lipotoxicity-induced LC3-II accumulation and recruitment to autophagosomes. Moreover, bromo-palmitate and etomoxir potentiate palmitate autophagic response. Even if gluco-lipotoxicity raised ceramide levels in INS-1 cells, ceramide synthase 4 overexpression does not potentiate LC3-II accumulation. Gluco-lipotoxicity also still stimulates an autophagic flux in the presence of an ER stress repressor. Finally, selective inhibition of sphingosine kinase 1 (SphK1) activity precludes gluco-lipotoxicity to induce LC3-II accumulation. Moreover, SphK1 overexpression potentiates autophagic flux induced by gluco-lipotxicity. Altogether, our results indicate that early activation of autophagy by gluco-lipotoxicity is mediated by SphK1, which plays a protective role in β cells. Full article

Progressive decline in pancreatic beta-cell function is central to the pathogenesis of type 2 diabetes (T2D). Here, we explore the relationship between the beta cell and its nutritional environment, asking how an excess of energy substrate leads to altered energy production and subsequent insulin secretion. Alterations in intracellular metabolic homeostasis are key markers of islets with T2D, but changes in cellular metabolite exchanges with their environment remain unknown. We answered this question using nuclear magnetic resonance-based quantitative metabolomics and evaluated the consumption or secretion of 31 extracellular metabolites from healthy and T2D human islets. Islets were also cultured under high levels of glucose and/or palmitate to induce gluco-, lipo-, and glucolipotoxicity. Biochemical analyses revealed drastic alterations in the pyruvate and citrate pathways, which appear to be associated with mitochondrial oxoglutarate dehydrogenase (OGDH) downregulation. We repeated these manipulations on the rat insulinoma-derived beta-pancreatic cell line (INS-1E). Our results highlight an OGDH downregulation with a clear effect on the pyruvate and citrate pathways. However, citrate is directed to lipogenesis in the INS-1E cells instead of being secreted as in human islets. Our results demonstrate the ability of metabolomic approaches performed on culture media to easily discriminate T2D from healthy and functional islets. Full article

CARD9, a scaffolding protein, has been implicated in the pathogenesis of metabolic diseases, including obesity and diabetes. We recently reported novel roles for CARD9 in islet β-cell dysregulation under duress of gluco (HG)- and glucolipotoxic (GLT) stress. CARD9 expression was also increased in β-cells following exposure to HG and GLT stress. The current study is aimed at understanding the putative roles of histone deacetylation in HG- and GLT-induced expression of CARD9. Using two structurally distinct inhibitors of histone deacetylases (HDACs), namely trichostatin (TSA) and suberoylanilide hydroxamic acid (SAHA), we provide the first evidence to suggest that the increased expression of CARD9 seen under duress of HG and GLT stress is under the regulatory control of histone deacetylation. Interestingly, the expression of protein kinase Cδ (PKCδ), a known upstream regulator of CARD9 activation, is also increased under conditions of metabolic stress. However, it is resistant to TSA and SAHA, suggesting that it is not regulated via histone deacetylation. Based on these data, we propose that targeting the appropriate HDACs, which mediate the expression (and function) of CARD9, might be the next step to further enhance our current understanding of the roles of CARD9 in islet dysfunction under metabolic stress and diabetes. Full article

Obesity is strongly associated with insulin sensitivity in type 2 diabetes (T2D), mainly because free fatty acids (FFAs) are released from excess fat tissue. Long-term exposure to high levels of FFAs and glucose leads to glucolipotoxicity, causing damage to pancreatic β-cells, thus accelerating the progression of T2D. Therefore, the prevention of β-cell dysfunction and apoptosis is essential to prevent the development of T2D. Unfortunately, there are currently no specific clinical strategies for protecting β-cells, highlighting the need for effective therapies or preventive approaches to improve the survival of β-cells in T2D. Interestingly, recent studies have shown that the monoclonal antibody denosumab (DMB), used in osteoporosis, displays a positive effect on blood glucose regulation in patients with T2D. DMB acts as an osteoprotegerin (OPG) by inhibiting the receptor activator of the NF-κB ligand (RANKL), preventing the maturation and function of osteoclasts. However, the exact mechanism by which the RANK/RANKL signal affects glucose homeostasis has not been fully explained. The present study used human 1.4 × 10⁷ β-cells to simulate the T2D metabolic condition of high glucose and free fatty acids (FFAs), and it investigated the ability of DMB to protect β-cells from glucolipotoxicity. Our results show that DMB effectively attenuated the cell dysfunction and apoptosis caused by high glucose and FFAs in β-cells. This may be caused by blocking the RANK/RANKL pathway that reduced mammalian sterile 20-like kinase 1 (MST1) activation and indirectly increased pancreatic and duodenal homeobox 1 (PDX-1) expression. Furthermore, the increase in inflammatory cytokines and ROS caused by the RANK/RANKL signal also played an important role in glucolipotoxicity-induced cytotoxicity, and DMB can also protect β-cells by reducing the mechanisms mentioned above. These findings provide detailed molecular mechanisms for the future development of DMB as a potential protective agent of β-cells. Full article

A loss of functional beta cell mass is a final etiological event in the development of frank type 2 diabetes (T2D). To preserve or expand beta cells and therefore treat/prevent T2D, growth factors have been considered therapeutically but have largely failed to achieve robust clinical success. The molecular mechanisms preventing the activation of mitogenic signaling pathways from maintaining functional beta cell mass during the development of T2D remain unknown. We speculated that endogenous negative effectors of mitogenic signaling cascades impede beta cell survival/expansion. Thus, we tested the hypothesis that a stress-inducible epidermal growth factor receptor (EGFR) inhibitor, mitogen-inducible gene 6 (Mig6), regulates beta cell fate in a T2D milieu. To this end, we determined that: (1) glucolipotoxicity (GLT) induces Mig6, thereby blunting EGFR signaling cascades, and (2) Mig6 mediates molecular events regulating beta cell survival/death. We discovered that GLT impairs EGFR activation, and Mig6 is elevated in human islets from T2D donors as well as GLT-treated rodent islets and 832/13 INS-1 beta cells. Mig6 is essential for GLT-induced EGFR desensitization, as Mig6 suppression rescued the GLT-impaired EGFR and ERK1/2 activation. Further, Mig6 mediated EGFR but not insulin-like growth factor-1 receptor nor hepatocyte growth factor receptor activity in beta cells. Finally, we identified that elevated Mig6 augmented beta cell apoptosis, as Mig6 suppression reduced apoptosis during GLT. In conclusion, we established that T2D and GLT induce Mig6 in beta cells; the elevated Mig6 desensitizes EGFR signaling and induces beta cell death, suggesting Mig6 could be a novel therapeutic target for T2D. Full article

Avoiding the loss of functional beta cell mass is critical for preventing or treating diabetes. Currently, the molecular mechanisms underlying beta cell death are partially understood, and there is a need to identify new targets for developing novel therapeutics to treat diabetes. Previously, our group established that Mig6, an inhibitor of EGF signaling, mediates beta cell death under diabetogenic conditions. The objective here was to clarify the mechanisms linking diabetogenic stimuli to beta cell death by investigating Mig6-interacting proteins. Using co-immunoprecipitation and mass spectrometry, we evaluated the binding partners of Mig6 under both normal glucose (NG) and glucolipotoxic (GLT) conditions in beta cells. We identified that Mig6 interacted dynamically with NumbL, whereas Mig6 associated with NumbL under NG, and this interaction was disrupted under GLT conditions. Further, we demonstrated that the siRNA-mediated suppression of NumbL expression in beta cells prevented apoptosis under GLT conditions by blocking the activation of NF-κB signaling. Using co-immunoprecipitation experiments, we observed that NumbL’s interactions with TRAF6, a key component of NFκB signaling, were increased under GLT conditions. The interactions among Mig6, NumbL, and TRAF6 were dynamic and context-dependent. We proposed a model wherein these interactions activated pro-apoptotic NF-κB signaling while blocking pro-survival EGF signaling under diabetogenic conditions, leading to beta cell apoptosis. These findings indicated that NumbL should be further investigated as a candidate anti-diabetic therapeutic target. Full article

Diabetic nephropathy (DN) exacerbates renal tissue damage and is a major cause of end-stage renal disease. Reactive oxygen species play a vital role in hyperglycemia-induced renal injury. This study examined whether the oral hypoglycemic drug acarbose (Ab) could attenuate the progression of DN in type 2 diabetes mellitus mice. In this study, 50 mg/kg body weight of Ab was administered to high-fat diet (HFD)-fed db/db mice. Their body weight was recorded every week, and the serum glucose concentration was monitored every 2 weeks. Following their euthanasia, the kidneys of mice were analyzed through hematoxylin and eosin, periodic acid Schiff, Masson’s trichrome, and immunohistochemistry (IHC) staining. The results revealed that Ab stabilized the plasma glucose and indirectly improved the insulin sensitivity and renal functional biomarkers in diabetic mice. In addition, diabetes-induced glomerular hypertrophy, the saccharide accumulation, and formation of collagen fiber were reduced in diabetic mice receiving Ab. Although the dosages of Ab cannot decrease the blood sugar in db/db mice, our results indicate that Ab alleviates glucolipotoxicity-induced DN by inhibiting kidney fibrosis-related proteins through the Ras/ERK pathway. Full article

Mulberry leaf (Morus alba L.) is used as a traditional medicine and potential health food to treat various metabolic diseases, such as hypertension, diabetes, and hyperlipidemia. However, we sought the mechanisms by which functional components of mulberry leaves mediate diabetic steatohepatitis. We applied an in vitro model of HepG2 cells induced by glucolipotoxicity and evaluated the effects of MLE and its major components nCGA, Crp, and CGA. The results showed that MLE and nCGA reduced liver fat accumulation by inhibiting SREBP-1/FASN, SREBP-2/HMG-CoAR, and activating PPARα/CPT-1. Additionally, MLE and nCGA decreased inflammatory responses associated with NF-κB, TNF-α, and IL-6 to alleviate steatohepatitis. Furthermore, we showed that MLE and nCGA exerted anti-glucolipotoxicity effects by downregulating miR-34a, thus activating SIRT1/AMPK signaling, and subsequently suppressing hepatic lipid accumulation. Full article

Diabetic neuropathy (DN) is a type of sensory nerve damage that can occur in patients with diabetes. Although the understanding of pathophysiology is incomplete, DN is often associated with structural and functional alterations of the affected neurons. Among all possible causes of nerve damage, Schwann cells (SCs) are thought to play a key role in repairing peripheral nerve injury, suggesting that functional deficits occurring in SCs may potentially exhibit their pathogenic roles in DN. Therefore, elucidating the mechanisms that underlie this pathology can be used to develop novel therapeutic targets. In this regard, glucagon-like peptide-1 receptor agonists (GLP-1 RAs) have recently attracted great attention in ameliorating SCs’ dysfunction. However, the detailed mechanisms remain uncertain. In the present study, we investigated how GLP-1 RA Liraglutide protects against RSC96 SCs dysfunction through a diabetic condition mimicked by high glucose and high free fatty acid (FFA). Our results showed that high glucose and high FFAs reduced the viability of RSC96 SCs by up to 51%, whereas Liraglutide reduced oxidative stress by upregulating antioxidant enzymes, and thus protected cells from apoptosis. Liraglutide also inhibited NFκB-mediated inflammation, inducing SCs to switch from pro-inflammatory cytokine production to anti-inflammatory cytokine production. Moreover, Liraglutide upregulated the production of neurotrophic factors and myelination-related proteins, and these protective effects appear to be synergistically linked to insulin signaling. Taken together, our findings demonstrate that Liraglutide ameliorates diabetes-related SC dysfunction through the above-mentioned mechanisms, and suggest that modulating GLP-1 signaling in SCs may be a promising strategy against DN. Full article

Ceramides belong to the sphingolipid family and represent the central hub of the sphingolipid network. In obesity, oversupply of saturated fatty acids including palmitate raises ceramide levels which can be detrimental to cells. Elevated ceramides can cause insulin resistance, endoplasmic reticulum stress, and mitochondrial dysfunction. Studies over the last few decades have highlighted the role played by ceramides in pancreatic islet β-cell apoptosis, especially under glucolipotoxic and inflammatory conditions. This review focuses on ceramides and adiponectin receptor signaling, summarizing recent advancements in our understanding of their roles in islet β-cells and the discovery of zinc-dependent lipid hydrolase (ceramidase) activity of adiponectin receptors. The therapeutic potential of targeting these events to prevent islet β-cell loss for treating diabetes is discussed. Full article

Mulberry leaf (Morus alba L.) has been used as a health food and in traditional medicine to treat several metabolic diseases, including diabetes, hypertension, and hyperlipidemia. However, the mechanism by which mulberry leaf and its functional components mediate atherosclerosis remains unclear. This study aimed to determine the effect of mulberry leaf extract (MLE) and its major component, neochlorogenic acid (nCGA), on the proliferation and migration of rat aortic vascular smooth muscle cells (VSMCs, A7r5 cell line) under diabetic cultured conditions (oleic acid and high glucose, OH). Our findings showed that MLE and nCGA significantly inhibited cell proliferation and migration in A7r5 cells as determined by a scratch wound assay and a Transwell assay. Furthermore, we observed MLE and nCGA inhibited cell proliferation and migration, such as reducing the phosphoinositide 3-kinases (PI3K)/protein kinase B (Akt), focal adhesion kinase (FAK), and small GTPase proteins using Western blot analysis. In conclusion, we confirmed the anti-atherosclerotic effects of MLE and nCGA in reducing vascular smooth muscle cell (VSMC) migration and proliferation under diabetic cultured conditions via inhibition of FAK/small GTPase proteins, PI3K/Akt, and Ras-related signaling. Full article

Since the discovery of insulin a century ago, insulin injection has been a primary treatment for both type 1 (T1D) and type 2 diabetes (T2D). T2D is a complicated disea se that is triggered by the dysfunction of insulin-producing β cells and insulin resistance in peripheral tissues. Insulin injection partially compensates for the role of endogenous insulin which promotes glucose uptake, lipid synthesis and organ growth. However, lacking the continuous, rapid, and accurate glucose regulation by endogenous functional β cells, the current insulin injection therapy is unable to treat the root causes of the disease. Thus, new technologies such as human pluripotent stem cell (hPSC)-derived islets are needed for both identifying the key molecular and genetic causes of T2D and for achieving a long-term treatment. This perspective review will provide insight into the efficacy of hPSC-derived human islets for treating and understanding T2D. We discuss the evidence that β cells should be the primary target for T2D treatment, the use of stem cells for the modeling of T2D and the potential use of hPSC-derived islet transplantation for treating T2D. Full article