Search

The medicinal uses of Calotropis procera are diverse, yet some of them are based on effects that still lack scientific support. Control of diabetes is one of them. Recently, latex proteins from C. procera latex (LP) have been shown to promote in vivo glycemic control by the inhibition of hepatic glucose production via AMP-activated protein kinase (AMPK). Glycemic control has been attributed to an isolated fraction of LP (CpPII), which is composed of cysteine peptidases (95%) and osmotin (5%) isoforms. Those proteins are extensively characterized in terms of chemistry, biochemistry and structural aspects. Furthermore, we evaluated some aspects of the mitochondrial function and cellular mechanisms involved in CpPII activity. The effect of CpPII on glycemic control was evaluated in fasting mice by glycemic curve and glucose and pyruvate tolerance tests. HepG2 cells was treated with CpPII, and cell viability, oxygen consumption, PPAR activity, production of lactate and reactive oxygen species, mitochondrial density and protein and gene expression were analyzed. CpPII reduced fasting glycemia, improved glucose tolerance and inhibited hepatic glucose production in control animals. Additionally, CpPII increased the consumption of ATP-linked oxygen and mitochondrial uncoupling, reduced lactate concentration, increased protein expression of mitochondrial complexes I, III and V, and activity of peroxisome-proliferator-responsive elements (PPRE), reduced the presence of reactive oxygen species (ROS) and increased mitochondrial density in HepG2 cells by activation of AMPK/PPAR. Our findings strongly support the medicinal use of the plant and suggest that CpPII is a potential therapy for prevention and/or treatment of type-2 diabetes. A common epitope sequence shared among the proteases and osmotin is possibly the responsible for the beneficial effects of CpPII.

Graphical abstract, Highlights • “Multiblock metabolomics” elucidates the global metabolic network in a whole body. • “Multiblock metabolomics” combines LC/MS-based metabolomics with multiblock PCA. • “Multiblock metabolomics” highlights and elicits organ-specific metabolism. • TGs with less unsaturated fatty acids were highly accumulated in the diabetic liver., Principal component analysis (PCA) is a useful tool for omics analysis to identify underlying factors and visualize relationships between biomarkers. However, this approach is limited in addressing life complexity and further improvement is required. This study aimed to develop a new approach that combines mass spectrometry-based metabolomics with multiblock PCA to elucidate the whole-body global metabolic network, thereby generating comparable metabolite maps to clarify the metabolic relationships among several organs. To evaluate the newly developed method, Zucker diabetic fatty (ZDF) rats (n = 6) were used as type 2 diabetic models and Sprague Dawley (SD) rats (n = 6) as controls. Metabolites in the heart, kidney, and liver were analyzed by capillary electrophoresis and liquid chromatography mass spectrometry, respectively, and the detected metabolites were analyzed by multiblock PCA. More than 300 metabolites were detected in the heart, kidney, and liver. When the metabolites obtained from the three organs were analyzed with multiblock PCA, the score and loading maps obtained were highly synchronized and their metabolism patterns were visually comparable. A significant finding in this study was the different expression patterns in lipid metabolism among the three organs; notably triacylglycerols with polyunsaturated fatty acids or less unsaturated fatty acids showed specific accumulation patterns depending on the organs.

Background & aims Macrophages are key regulators of inflammation and cancer promotion in the liver, and their recruitment and activation is linked to chemokine receptor signaling. However, the exact roles of the chemokine receptors CCR2 and CCR5 for macrophage functions in the liver is obscure. Methods To study CCR2 and CCR5 in inflammatory liver injury, we used mice with a hepatocyte-specific knock-out of the nuclear factor κB (NF-κB) essential modulator (NEMO), termed NEMOLPC-KO mice, and generated NEMOLPC-KOCcr2-/- and NEMOLPC-KOCcr5-/- mice. NEMOLPC-KO mice develop hepatitis and fibrosis after two and liver tumors after six months. Results We found that both CCR2 and CCR5 deficiency led to reduced fibrosis, while CCR5 deficiency increased steatosis and tumor burden in NEMOLPC-KO mice. CCR2 was required for recruitment of hepatic macrophages, whereas CCR5 promoted stellate cell activation. The reduction of monocytes and macrophages by either anti-Gr1 antibody or clodronate-loaded liposomes (CLL), but not of CD8+ T cells or NK cells, significantly aggravated liver injury in NEMOLPC-KO mice and was further increased in NEMOLPC-KOCcr5-/- mice. CLL-induced liver injury was dampened by the adoptive transfer of ex vivo generated macrophages, whereas the adoptive transfer of control CD115+ immature monocytes or B cells did not reduce liver injury. Conclusions Although CCR2 and CCR5 principally promote liver fibrosis, they exert differential functions on hepatic macrophages during liver disease progression in NEMOLPC-KO mice. While CCR2 controls the recruitment of monocytes to injured livers, CCR5-dependent functions of liver macrophages limit hepatic injury, thereby reducing steatosis and hepatocarcinogenesis., Graphical abstract

Background & Aims p21-activated kinase-1 (PAK1) belongs to a family of serine-threonine kinases and contributes to cellular pathways such as nuclear factor-κB (NF-κB), mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), and Wingless-related integration site(Wnt)/β-catenin, all of which are involved in intestinal homeostasis. Overexpression of PAK1 is linked to inflammatory bowel disease as well as colitis-associated cancer (CAC), and similarly was observed in interleukin (IL)10 knockout (KO) mice, a model of colitis and CAC. Here, we tested the effects of PAK1 deletion on intestinal inflammation and carcinogenesis in IL10 KO mice. Methods IL10/PAK1 double-knockout (DKO) mice were generated and development of colitis and CAC was analyzed. Large intestines were measured and prepared for histology or RNA isolation. Swiss rolls were stained with H&E and periodic acid-Schiff. Co-immunoprecipitation and immunofluorescence were performed using intestinal organoids, SW480, and normal human colon epithelial cells 1CT. Results When compared with IL10 KO mice, DKOs showed longer colons and prolonged crypts, despite having higher inflammation and numbers of dysplasia. Crypt hyperproliferation was associated with Notch1 activation and diminished crypt differentiation, indicated by a reduction of goblet cells. Gene expression analysis indicated up-regulation of the Notch1 target hairy and enhancer of split-1 and the stem cell receptor leucin-rich repeat-containing G-protein–coupled receptor 5 in DKO mice. Interestingly, the stem cell marker olfactomedin-4 was present in colonic tissue. Increased β-catenin messenger RNA and cytoplasmic accumulation indicated aberrant Wnt signaling. Co-localization and direct interaction of Notch1 and PAK1 was found in colon epithelial cells. Notch1 activation abrogated this effect whereas silencing of PAK1 led to Notch1 activation. Conclusions PAK1 contributes to the regulation of crypt homeostasis under inflammatory conditions by controlling Notch1. This identifies a novel PAK1–Notch1 axis in intestinal pathophysiology of inflammatory bowel disease and CAC., Graphical abstract

COVID-19 pandemic caused by SARS-CoV-2 has already claimed millions of lives worldwide due to the absence of a suitable anti-viral therapy. The CoV envelope (E) protein, which has not received much attention so far, is a 75 amino acid long integral membrane protein involved in assembly and release of the virus inside the host. Here we have used artificial intelligence (AI) and pattern recognition techniques for initial screening of FDA approved pharmaceuticals and nutraceuticals to target this E protein. Subsequently, molecular docking simulations have been performed between the ligands and target protein to screen a set of 9 ligand molecules. Finally, we have provided detailed insight into their mechanisms of action related to the varied symptoms of infected patients., Highlights • This work focuses on integral membrane Envelope (E) Protein as the target which is the most conserved among all. • We implemented artificial intelligence (AI) and pattern recognition techniques for initial screening • Next level screening incorporates their (a) existing functions (b) FDA approval status and (c) severity of the side effects. • Blind molecular docking simulation has been done and 9 ligand molecules have been screened. • A detailed insight into the mode of action of these 9 drugs related to the varied symptoms of infected patients is provided.

In 30% of epileptic individuals, intractable epilepsy represents a problem for the management of seizures and severely affects the patient's quality of life due to pharmacoresistance with commonly used antiseizure drugs (ASDs). Surgery is not the best option for all resistant patients due to its post-surgical consequences. Therefore, several alternative or complementary therapies have scientifically proven significant therapeutic potential for the management of seizures in intractable epilepsy patients with seizure-free occurrences. Various non-pharmacological interventions include metabolic therapy, brain stimulation therapy, and complementary therapy. Metabolic therapy works out by altering the energy metabolites and include the ketogenic diets (KD) (that is restricted in carbohydrates and mimics the metabolic state of the body as produced during fasting and exerts its antiepileptic effect) and anaplerotic diet (which revives the level of TCA cycle intermediates and this is responsible for its effect). Neuromodulation therapy includes vagus nerve stimulation (VNS), responsive neurostimulation therapy (RNS) and transcranial magnetic stimulation therapy (TMS). Complementary therapies such as biofeedback and music therapy have demonstrated promising results in pharmacoresistant epilepsies. The current emphasis of the review article is to explore the different integrated mechanisms of various treatments for adequate seizure control, and their limitations, and supportive pieces of evidence that show the efficacy and tolerability of these non-pharmacological options.

Background & Aims Nonalcoholic steatohepatitis (NASH) is a multisystem condition, implicating liver and adipose tissue. Although the general involvement of the innate and adaptive immune system has been established, we aimed to define the exact role of the functionally diverse T-cell subsets in NASH pathogenesis through diet reversal and immunologic modulation. Methods Multiple experimental set-ups were used in 8-week-old C57BL/6J mice, including prolonged high-fat high-fructose diet (HFHFD) feeding, diet reversal from HFHFD to control diet, and administration of anti-CD8a and anti–interleukin 17A antibodies. Plasma alanine aminotransferase, glucose, and lipid levels were determined. Liver and adipose tissue were assessed histologically. Cytotoxic T (Tc), regulatory T, T helper (Th) 1, and Th17 cells were characterized in liver and visceral adipose tissue (VAT) via flow cytometry and RNA analysis. Results HFHFD feeding induced the metabolic syndrome and NASH, which coincided with an increase in hepatic Th17, VAT Tc, and VAT Th17 cells, and a decrease in VAT regulatory T cells. Although diet reversal induced a phenotypical metabolic and hepatic normalization, the observed T-cell disruptions persisted. Treatment with anti-CD8a antibodies decreased Tc cell numbers in all investigated tissues and induced a biochemical and histologic attenuation of the HFHFD-induced NASH. Conversely, anti–interleukin 17A antibodies decreased hepatic inflammation without affecting other features of NASH or the metabolic syndrome. Conclusions HFHFD feeding induces important immune disruptions in multiple hepatic and VAT T-cell subsets, refractory to diet reversal. In particular, VAT Tc cells are critically involved in NASH pathogenesis, linking adipose tissue inflammation to liver disease., Graphical abstract

Alexander disease (AxD) is a rare and fatal neurodegenerative disorder caused by mutations in the gene encoding glial fibrillary acidic protein (GFAP). In this report, a mouse model of AxD (GFAPTg;Gfap+/R236H) was analyzed that contains a heterozygous R236H point mutation in murine Gfap as well as a transgene with a GFAP promoter to overexpress human GFAP. Using label-free quantitative proteomic comparisons of brain tissue from GFAPTg;Gfap+/R236H versus wild-type mice confirmed upregulation of the glutathione metabolism pathway and indicated proteins were elevated in the peroxisome proliferator-activated receptor (PPAR) signaling pathway, which had not been reported previously in AxD. Relative protein-level differences were confirmed by a targeted proteomics assay, including proteins related to astrocytes and oligodendrocytes. Of particular interest was the decreased level of the oligodendrocyte protein, 2-hydroxyacylsphingosine 1-beta-galactosyltransferase (Ugt8), since Ugt8-deficient mice exhibit a phenotype similar to GFAPTg;Gfap+/R236H mice (e.g., tremors, ataxia, hind-limb paralysis). In addition, decreased levels of myelin-associated proteins were found in the GFAPTg;Gfap+/R236H mice, consistent with the role of Ugt8 in myelin synthesis. Fabp7 upregulation in GFAPTg;Gfap+/R236H mice was also selected for further investigation due to its uncharacterized association to AxD, critical function in astrocyte proliferation, and functional ability to inhibit the anti-inflammatory PPAR signaling pathway in models of amyotrophic lateral sclerosis (ALS). Within Gfap+ astrocytes, Fabp7 was markedly increased in the hippocampus, a brain region subjected to extensive pathology and chronic reactive gliosis in GFAPTg;Gfap+/R236H mice. Last, to determine whether the findings in GFAPTg;Gfap+/R236H mice are present in the human condition, AxD patient and control samples were analyzed by Western blot, which indicated that Type I AxD patients have a significant fourfold upregulation of FABP7. However, immunohistochemistry analysis showed that UGT8 accumulates in AxD patient subpial brain regions where abundant amounts of Rosenthal fibers are located, which was not observed in the GFAPTg;Gfap+/R236H mice., Graphical abstract, Highlights • Models of AxD provide insight into functions of astrocytes related to neuropathology. • PPAR pathway found upregulated in AxD mice may be neuroprotective. Fatty acid binding protein (Fabp7) was upregulated in AxD mice and human CNS samples. • Depleted levels in the classical constituents of myelin were observed in AxD mice. • UGT8 may be related to myelin deficits in AxD., In Brief The article contains the first whole brain proteomic survey from a mouse model of Alexander disease (AxD). Several novel findings include activation of the PPAR signaling pathway, which has been reported to be protective in models of amyotrophic lateral sclerosis (ALS). Another finding related to the gliosis phenotype in AxD mice was the upregulation of fatty acid binding protein 7 (FABP7), which induces an NF-κB inflammatory response and counteracts the anti-inflammatory effects of PPAR signaling.

Alterations of the lipid profile of the stratum corneum have an important role in the pathogenesis of atopic dermatitis (AD) because they contribute to epidermal barrier impairment. However, they have not previously been envisioned as a cellular response to altered metabolic requirements in AD epidermis. In this study, we report that the lipid composition in the epidermis of flaky tail, that is, ft/ft mice mimics that of human lesional AD (ADL) epidermis, both showing a shift toward shorter lipid species. The amounts of C24 and C26 free fatty acids and C24 and C26 ceramides—oxidized exclusively in peroxisomes—were reduced in the epidermis of ft/ft mice despite increased lipid synthesis, similar to that seen in human ADL edpidermis. Increased ACOX1 protein and activity in granular keratinocytes of ft/ft epidermis, altered lipid profile in human epidermal equivalents overexpressing ACOX1, and increased ACOX1 immunostaining in skin biopsies from patients with ADL suggest that peroxisomal β-oxidation significantly contributes to lipid signature in ADL epidermis. Moreover, we show that increased anaerobic glycolysis in ft/ft mouse epidermis is essential for keratinocyte proliferation and adenosine triphosphate synthesis but does not contribute to local inflammation. Thus, this work evidenced a metabolic shift toward enhanced peroxisomal β-oxidation and anaerobic glycolysis in ADL epidermis.

International audience

Metabolic flexibility is the capacity of cells to alter fuel metabolism in response to changes in metabolic demand or nutrient availability. It is critical for maintaining cellular bioenergetics and is involved in the pathogenesis of cardiovascular disease and metabolic disorders. However, the regulation and function of metabolic flexibility in lymphatic endothelial cells (LECs) remain unclear. We have previously shown that glycolysis is the predominant metabolic pathway to generate ATP in LECs and that fibroblast growth factor receptor (FGFR) signaling controls lymphatic vessel formation by promoting glycolysis. Here, we found that chemical inhibition of FGFR activity or knockdown of FGFR1 induces substantial upregulation of fatty acid β-oxidation (FAO) while reducing glycolysis and cellular ATP generation in LECs. Interestingly, such compensatory elevation was not observed in glucose oxidation and glutamine oxidation. Mechanistic studies show that FGFR blockade promotes the expression of carnitine palmitoyltransferase 1A (CPT1A), a rate-limiting enzyme of FAO; this is achieved by dampened extracellular signal–regulated protein kinase activation, which in turn upregulates the expression of the peroxisome proliferator–activated receptor alpha. Metabolic analysis further demonstrates that CPT1A depletion decreases total cellular ATP levels in FGFR1-deficient rather than wildtype LECs. This result suggests that FAO, which makes a negligible contribution to cellular energy under normal conditions, can partially compensate for energy deficiency caused by FGFR inhibition. Consequently, CPT1A silencing potentiates the effect of FGFR1 knockdown on impeding LEC proliferation and migration. Collectively, our study identified a key role of metabolic flexibility in modulating the effect of FGFR signaling on LEC growth.

Highlights • Simvastatin-romidepsin combination kills bladder cancer cells synergistically. • The combination induces histone acetylation by activating AMPK. • AMPK activation and histone acetylation are associated with ER stress induction. • Positive feedback cycle between ER stress induction and PPARγ expression., The HMG-CoA reductase inhibitor simvastatin activates AMP-activated protein kinase (AMPK) and thereby induces histone acetylation. We postulated that combining simvastatin with the histone deacetylase (HDAC) inhibitor romidepsin would kill bladder cancer cells by inducing histone acetylation cooperatively. The combination of romidepsin and simvastatin induced robust apoptosis and killed bladder cancer cells synergistically. In murine subcutaneous tumor models using MBT-2 cells, a 15-day treatment with 0.5 mg/kg romidepsin and 15 mg/kg simvastatin was well tolerated and inhibited tumor growth significantly. Mechanistically, the combination induced histone acetylation by activating AMPK. The combination also decreased the expression of HDACs, thus further promoting histone acetylation. This AMPK activation was essential for the combination's action because compound C, an AMPK inhibitor, suppressed the combination-induced histone acetylation and the combination's ability to induce apoptosis. We also found that the combination increased the expression of peroxisome proliferator-activated receptor (PPAR) γ, leading to reactive oxygen species production. Furthermore, the combination induced endoplasmic reticulum (ER) stress and this ER stress was shown to be associated with increased AMPK expression and histone acetylation, thus playing an important role in the combination's action. Our study also suggests there is a positive feedback cycle between ER stress induction and PPARγ expression.

Leigh syndrome is a highly heterogeneous condition caused by pathological mutations in either nuclear or mitochondrial DNA regions encoding molecules involved in mitochondrial oxidative phosphorylation, in which many organs including the brain can be affected. Among these organs, a high incidence of poor bone health has been recognized in primary mitochondrial diseases including Leigh syndrome. However, the direct association between mitochondrial dysfunction and poor bone health has not been fully elucidated. Mitochondrial biosynthesis is a potential therapeutic target for this syndrome, as it can ameliorate the impairment of oxidative phosphorylation without altering these gene mutations. A recent study has shown the impaired osteogenesis in the dental pulp stem cells derived from the deciduous teeth of a child with Leigh syndrome, harboring the heteroplasmic mutation G13513A in the mitochondrial DNA region encoding the ND5 subunit of the respiratory chain complex I. The present study aimed to investigate whether mitochondrial biogenesis could be a therapeutic target for improving osteogenesis, using the same stem cells in a patient-specific cellular model. For this purpose, bezafibrate was used because it has been reported to induce mitochondrial biogenesis as well as to improve bone metabolism and osteoporosis. Bezafibrate clearly improved the differentiation of patient-derived stem cells into osteoblasts and the mineralization of differentiated osteoblasts. The mRNA expression of peroxisome proliferator-activated receptor-gamma coactivator-1α, ATP production, and mitochondrial Ca2+ levels were all significantly increased by bezafibrate in the patient-derived cells. In addition, the increased amount and morphological shift from the fragmentary to network shape associated with DRP1 downregulation were also observed in the bezafibrate-treated patient-derived cells. These results suggest that mitochondrial biogenesis may be a potential therapeutic target for improving osteogenesis in patients with Leigh syndrome, and bezafibrate may be one of the candidate treatment agents., Highlights • Dental pulp stem cells from a child with Leigh syndrome have impaired osteogenesis. • Bezafibrate-PGC-1α pathway improves osteogenesis via mitochondrial biogenesis. • Bezafibrate also induces DRP1 downregulation and mitochondrial network formation. • Dental pulp stem cells may help to establish treatment strategies for Leigh syndrome.

The prevalence and diagnosis of nonalcoholic fatty liver disease (NAFLD) is on the rise worldwide and currently has no FDA-approved pharmacotherapy. The increase in disease burden of NAFLD and a more severe form of this progressive liver disease, nonalcoholic steatohepatitis (NASH), largely mirrors the increase in obesity and type 2 diabetes (T2D) and reflects the hepatic manifestation of an altered metabolic state. Indeed, metabolic syndrome, defined as a constellation of obesity, insulin resistance, hyperglycemia, dyslipidemia and hypertension, is the major risk factor predisposing the NAFLD and NASH. There are multiple potential pharmacologic strategies to rebalance aspects of disordered metabolism in NAFLD. These include therapies aimed at reducing hepatic steatosis by directly modulating lipid metabolism within the liver, inhibiting fructose metabolism, altering delivery of free fatty acids from the adipose to the liver by targeting insulin resistance and/or adipose metabolism, modulating glycemia, and altering pleiotropic metabolic pathways simultaneously. Emerging data from human genetics also supports a role for metabolic drivers in NAFLD and risk for progression to NASH. In this review, we highlight the prominent metabolic drivers of NAFLD pathogenesis and discuss the major metabolic targets of NASH pharmacotherapy.

Type 2 diabetes (T2D) has become a global epidemic. Although several drugs are available to manage T2D, problems associated with person-to-person variability in drug efficacy and potential side-effects remain unresolved. Owing to the emerging role of the gut microbiome in obesity and T2D, the interaction between gut microbes and anti-diabetic drugs and its influence on drugs' functions remains of immediate research interest. On one hand, drugs can manipulate gut microbiome composition and metabolic capacity. Conversely, the metabolic activities of the microbiome and its metabolites can also influence drug metabolism and effects. Hence, understanding this bi-directional drug-microbiome interaction and how it influences the clinical outcomes of antidiabetic drugs can pave the way to develop next-generation strategies to ameliorate diabetes. This review presents evidences demonstrating the putative interactions between anti-diabetic drugs and the gut microbiome, and discusses the potential of microbiome modulators to manipulate drug-microbiome interactions and the drug metabolism., Graphical abstract Unlabelled Image, Highlights • Gut microbes interact with medications used to treat T2D, that at least partially mediates potential benefits of these drugs. • Anti-diabetic drugs impact gut microbiome and its metabolic activity and vice-versa. • Understanding the dynamics of drug-microbiome cross-talk would offer better therapeutic outcomes for diabetes.

PMID: 34438017 WOS:000702705800007 The SARS-CoV-2 virus has caused a worldwide COVID-19 pandemic. In less than a year and a half, more than 200 million people have been infected and more than four million have died. Despite some improvement in the treatment strategies, no definitive treatment protocol has been developed. The pathogenesis of the disease has not been clearly elucidated yet. A clear understanding of its pathogenesis will help develop effective vaccines and drugs. The immunopathogenesis of COVID-19 is characteristic with acute respiratory distress syndrome and multiorgan involvement with impaired Type I interferon response and hyperinflammation. The destructive systemic effects of COVID-19 cannot be explained simply by the viral tropism through the ACE2 and TMPRSS2 receptors. In addition, the recently identified mutations cannot fully explain the defect in all cases of Type I interferon synthesis. We hypothesize that retinol depletion and resulting impaired retinoid signaling play a central role in the COVID-19 pathogenesis that is characteristic for dysregulated immune system, defect in Type I interferon synthesis, severe inflammatory process, and destructive systemic multiorgan involvement. Viral RNA recognition mechanism through RIG-I receptors can quickly consume a large amount of the body's retinoid reserve, which causes the retinol levels to fall below the normal serum levels. This causes retinoid insufficiency and impaired retinoid signaling, which leads to interruption in Type I interferon synthesis and an excessive inflammation. Therefore, reconstitution of the retinoid signaling may prove to be a valid strategy for management of COVID-19 as well for some other chronic, degenerative, inflammatory, and autoimmune diseases.

Liver steatosis is emerging as a major cause of chronic liver disease worldwide, mainly due to the increasing rate of obesity, type 2 diabetes, and metabolic syndrome. Because of the increased incidence of liver steatosis, many organs are currently declined for transplantation despite high demand and waiting list mortality. Defatting strategies have recently emerged as a means of rapidly reducing liver steatosis to expand the pool of available organs. This review summarises advances in defatting strategies in experimental and human models of liver steatosis over the last 20 years. CA extern

Normothermic ex-vivo kidney perfusion (NEVKP) results in significantly improved graft function in porcine auto-transplant models of donation after circulatory death injury compared with static cold storage (SCS); however, the molecular mechanisms underlying these beneficial effects remain unclear. We performed an unbiased proteomics analysis of 28 kidney biopsies obtained at three time points from pig kidneys subjected to 30 min of warm ischemia, followed by 8 h of NEVKP or SCS, and auto-transplantation. 70/6593 proteins quantified were differentially expressed between NEVKP and SCS groups (false discovery rate < 0.05). Proteins increased in NEVKP mediated key metabolic processes including fatty acid ß-oxidation, the tricarboxylic acid cycle, and oxidative phosphorylation. Comparison of our findings with external datasets of ischemia-reperfusion and other models of kidney injury confirmed that 47 of our proteins represent a common signature of kidney injury reversed or attenuated by NEVKP. We validated key metabolic proteins (electron transfer flavoprotein subunit beta and carnitine O-palmitoyltransferase 2, mitochondrial) by immunoblotting. Transcription factor databases identified members of the peroxisome proliferator-activated receptors (PPAR) family of transcription factors as the upstream regulators of our dataset, and we confirmed increased expression of PPARA, PPARD, and RXRA in NEVKP with reverse transcription polymerase chain reaction. The proteome-level changes observed in NEVKP mediate critical metabolic pathways. These effects may be coordinated by PPAR-family transcription factors and may represent novel therapeutic targets in ischemia-reperfusion injury., Graphical abstract, Highlights • Quantitative proteomics of NEVKP and cold storage pig kidneys at three time points. • Proteins increased in NEVKP are associated with key steps of mitochondrial metabolism. • NEVKP attenuates proteins increased in the kidney response to injury in prior studies. • In silico and mRNA analyses suggest PPAR-family members as likely regulators in NEVKP., In Brief The molecular changes associated with normothermic ex-vivo kidney perfusion (NEVKP) compared with static cold storage were studied using discovery proteomics in a porcine model. NEVKP resulted in increased expression of mitochondrial proteins (ETFB, CPT2) responsible for critical metabolic steps of ATP-synthesis. PPARGC1A, PPARA/D, and RXRA were computationally predicted as upstream regulators of proteins increased in NEVKP and showed increased mRNA expression in NEVKP-treated kidneys. PPAR-family members and their target proteins may represent new therapeutic targets to ameliorate ischemia-reperfusion injury.

Background & Aims The pathogenesis of Wilson disease (WD) involves hepatic and brain copper accumulation resulting from pathogenic variants affecting the ATP7B gene and downstream epigenetic and metabolic mechanisms. Prior methylome investigations in human WD liver and blood and in the Jackson Laboratory (Bar Harbor, ME) C3He-Atp7btx-j/J (tx-j) WD mouse model revealed an epigenetic signature of WD, including changes in histone deacetylase (HDAC) 5. We tested the hypothesis that histone acetylation is altered with respect to copper overload and aberrant DNA methylation in WD. Methods We investigated class IIa HDAC4 and HDAC5 and H3K9/H3K27 histone acetylation in tx-j mouse livers compared with C3HeB/FeJ (C3H) control in response to 3 treatments: 60% kcal fat diet, D-penicillamine (copper chelator), and choline (methyl group donor). Experiments with copper-loaded hepatoma G2 cells were conducted to validate in vivo studies. Results In 9-week tx-j mice, HDAC5 levels increased significantly after 8 days of a 60% kcal fat diet compared with chow. In 24-week tx-j mice, HDAC4/5 levels were reduced 5- to 10-fold compared with C3H, likely through mechanisms involving HDAC phosphorylation. HDAC4/5 levels were affected by disease progression and accompanied by increased acetylation. D-penicillamine and choline partially restored HDAC4/5 and H3K9ac/H3K27ac to C3H levels. Integrated RNA and chromatin immunoprecipitation sequencing analyses revealed genes regulating energy metabolism and cellular stress/development, which, in turn, were regulated by histone acetylation in tx-j mice compared with C3H mice, with Pparα and Pparγ among the most relevant targets. Conclusions These results suggest dietary modulation of class IIa HDAC4/5, and subsequent H3K9/H3K27 acetylation/deacetylation can regulate gene expression in key metabolic pathways in the pathogenesis of WD., Graphical abstract

Graphical abstract, Macrophages represent the first line of anti-pathogen defense - they encounter invading pathogens to perform the phagocytic activity, to deliver the plethora of pro- and anti-inflammatory cytokines, and to shape the tissue microenvironment. Throughout pneumonia course, alveolar macrophages and infiltrated blood monocytes produce increasing cytokine amounts, which activates the antiviral/antibacterial immunity but can also provoke the risk of the so-called cytokine “storm” and normal tissue damage. Subsequently, the question of how the cytokine spectrum is shaped and balanced in the pneumonia context remains a hot topic in medical immunology, particularly in the COVID19 pandemic era. The diversity in cytokine profiles, involved in pneumonia pathogenesis, is determined by the variations in cytokine-receptor interactions, which may lead to severe cytokine storm and functional decline of particular tissues and organs, for example, cardiovascular and respiratory systems. Cytokines and their receptors form unique profiles in individual patients, depending on the (a) microenvironmental context (comorbidities and associated treatment), (b) lung monocyte heterogeneity, and (c) genetic variations. These multidisciplinary strategies can be proactively considered beforehand and during the pneumonia course and potentially allow the new age of personalized immunotherapy.

Visual Abstract, Highlights • Neutrophil adhesion to the vascular endothelium in the femoral artery of low-density lipoprotein receptor–null mice fed with HFD was significantly enhanced compared with wild-type mice fed with HFD or low-density lipoprotein receptor–null mice fed with NC. • HFD induced citrullinated histone H3 of neutrophils in LDL receptor–null mice through the up-regulation of CXCL1 in blood. • Hypercitrullination of histone H3 in neutrophils by CXCL1 enhanced neutrophil adhesion in vitro and in vivo. • Pemafibrate decreased neutrophil adhesion and citrullination of histone H3 through the reduction of CXCL1., Summary Neutrophil adhesion on the atheroprone femoral artery of high-fat diet–fed low-density lipoprotein receptor–null mice was enhanced more than in wild-type mice. The inhibition of histone H3 citrullination of neutrophils reversed the enhancement of neutrophil adhesion, suggesting that hypercitrullination contributes to enhanced neutrophil adhesion. Furthermore, pemafibrate reduced the citrullination of histone H3 in these mice. Therefore, the hypercitrullination of histone H3 in neutrophils contributes to atherosclerotic vascular inflammation.

Nuclear receptors (NRs) activate transcription of target genes in response to binding of ligands to their ligand-binding domains (LBDs). Typically, in vitro assays use either gene expression or the recruitment of coactivators to the isolated LBD of the NR of interest to measure NR activation. However, this approach ignores that NRs function as homo- as well as heterodimers and that the LBD harbors the main dimerization interface. Cofactor recruitment is thereby interconnected with oligomerization status as well as ligand occupation of the partnering LBD through allosteric cross talk. Here we present a modular set of homogeneous time-resolved FRET–based assays through which we investigated the activation of PPARγ in response to ligands and the formation of heterodimers with its obligatory partner RXRα. We introduced mutations into the RXRα LBD that prevent coactivator binding but do not interfere with LBD dimerization or ligand binding. This enabled us to specifically detect PPARγ coactivator recruitment to PPARγ:RXRα heterodimers. We found that the RXRα agonist SR11237 destabilized the RXRα homodimer but promoted formation of the PPARγ:RXRα heterodimer, while being inactive on PPARγ itself. Of interest, incorporation of PPARγ into the heterodimer resulted in a substantial gain in affinity for coactivator CBP-1, even in the absence of ligands. Consequently, SR11237 indirectly promoted coactivator binding to PPARγ by shifting the oligomerization preference of RXRα toward PPARγ:RXRα heterodimer formation. These results emphasize that investigation of ligand-dependent NR activation should take NR dimerization into account. We envision these assays as the necessary assay tool kit for investigating NRs that partner with RXRα.

Introduction Peroxisome proliferator–activated receptor (PPAR) subfamily play an important role in chondrogenesis. Previous study has reported that mixture of GW0742 (PPAR-δ agonist), hyaluronic acid (HA) and mesenchymal stem cells (MSCs) enhance chondrogenesis. The purpose of this study is to compare with efficacies of commercially available HA and demonstrate correlation of PPAR-γ and PPAR-δ. Methods In this experimental study, MSCs were cultured with chondrogenic media and clinical HA gels (Euflexxa®, Synvisc®, Orthovisc® and Supartz®) using micormass culture method. Expression of type Ⅰ, Ⅱ collagen and matrix metalloprotease-13 (MMP-13) was measured by immunoblotting. MSCs were cultured with chondrogenic media and/or HA and/or GW0742 and/or rosiglitazone (PPAR-γ agonist) and/or human osteoarthritis synovial fluid. Immunoblotting was used to measure expression of type Ⅱ collagen and PPAR-γ. To identify the effective dose for chondrogenesis and adipogenesis, either 0.1, 1, 5 or 10 μM of rosiglitazone was added to MSCs in chondrogenic media or adipogenic media. Results Clinical HA gels inhibited expression of type Ⅰ collagen and enhanced the expression of MMP-13. Type Ⅱ collagen expression was significantly elevated in all treatment groups except Supartz®. GW0742 decreased the expression of PPAR-γ with/without inflammation condition. Rosiglitazone enhanced adipogenesis in a dose-dependent manner and enhanced the expression of type Ⅱ collagen under inflammation condition. Otherwise, the expression of type Ⅱ collagen and formation of chondrocyte spheroids showed a dose-dependent manner with a peak at 1 μM of rosiglitazone. Conclusions PPAR-γ has a considerable anti-inflammatory effect and a strong pro-adipogenic effect, which inhibits the chondrogenic effect. PPAR-γ is related with PPAR-δ and shows a chondrogenic effect at lower concentrations. And clinical HA gels shows various efficacy of chondrogenesis. This study suggested that PPAR-γ and PPAR-δ are key regulatory factors of chondrogenesis., Graphical abstract Image 1

Non-alcoholic fatty liver disease (NAFLD) represents the most common chronic liver disease in industrialized countries. NAFLD progresses through the inflammatory phase of non-alcoholic steatohepatitis (NASH) to fibrosis and cirrhosis, with some cases developing liver failure or hepatocellular carcinoma (HCC). Liver biopsy remains the gold standard approach to a definitive diagnosis of NAFLD and the distinction between simple steatosis and NASH. The pathogenesis of NASH is still not clear. Several theories have been proposed ranging from the "Two Hit Theory" to the "Multiple Hit Theory". However, the general consensus is that the gut microbiota, oxidative stress, and mitochondrial damage play key roles in the pathogenesis of NASH. The interaction between the gut epithelia and some commensal bacteria induces the rapid generation of reactive oxygen species (ROS). The main goal of any therapy addressing NASH is to reverse or prevent progression to liver fibrosis/cirrhosis. This problem represents the first "Achilles' heel" of the new molecules being evaluated in most ongoing clinical trials. The second is the inability of these molecules to reach the mitochondria, the primary sites of energy production and ROS generation. Recently, a variety of non-pharmacological and pharmacological treatment approaches for NASH have been evaluated including vitamin E, the thiazolidinediones, and novel molecules related to NASH pathogenesis (including obeticholic acid and elafibranor). Recently, a new isoform of human manganese superoxide dismutase (MnSOD) was isolated and obtained in a synthetic recombinant form designated rMnSOD. This protein has been shown to be a powerful antioxidant capable of mediating ROS dismutation, penetrating biological barriers via its uncleaved leader peptide, and reducing portal hypertension and fibrosis in rats affected by liver cirrhosis. Based on these distinctive characteristics, it can be hypothesized that this novel recombinant protein (rMnSOD) potentially represents a new and highly efficient adjuvant therapy to counteract the progression from NASH to HCC.

Visual Abstract, Highlights • This study shows that apoA-I may be an alternative strategy to improve the biocompatibility of stents. • Systemic infusions of apoA-I reduce in-stent neointimal hyperplasia in a murine model of stenting. • The cellular phenotype of the neointima post-stenting is altered by apoA-I infusions such that the smooth muscle cell phenotype is preserved, and there are fewer macrophages. • There was an increase in endothelial cell content of the arteries post-stenting in the mice infused with apoA-I, indicating an enhancement of endothelialization. • Systemic infusions of apoA-I inhibit platelet activation., Summary Even the most advanced drug-eluting stents evoke unresolved issues, including chronic inflammation, late thrombosis, and neoatherosclerosis. This highlights the need for novel strategies that improve stent biocompatibility. Our studies show that apolipoprotein A-I (apoA-I) reduces in-stent restenosis and platelet activation, and enhances endothelialization. These findings have therapeutic implications for improving stent biocompatibility.

Objective The peroxisome proliferator-activated receptor-γ coactivator-1α1 (PGC-1α1) regulates genes involved in energy metabolism. Increasing adipose tissue energy expenditure through PGC-1α1 activation is potentially beneficial for systemic metabolism. Pharmacological PGC-1α1 activators could be valuable tools in the fight against obesity and metabolic disease. Finding such compounds has been challenging partly because PGC-1α1 is a transcriptional coactivator with no known ligand-binding properties. While, PGC-1α1 activation is regulated by several mechanisms, protein stabilization is a crucial limiting step due to its short half-life under unstimulated conditions. Methods We designed a cell-based high-throughput screening system to identify PGC-1α1 protein stabilizers. Positive hits were tested for their ability to induce endogenous PGC-1α1 protein accumulation and activate target gene expression in brown adipocytes. Select compounds were analyzed for their effects on global gene expression and cellular respiration in adipocytes. Results Among 7,040 compounds screened, we highlight four small molecules with high activity as measured by: PGC-1α1 protein accumulation, target gene expression, and uncoupled mitochondrial respiration in brown adipocytes. Conclusions We identify compounds that induce PGC-1α1 protein accumulation and show that this increases uncoupled respiration in brown adipocytes. This screening platform establishes the foundation for a new class of therapeutics with potential use in obesity and associated disorders., Graphical abstract, Highlights • A high-throughput platform to identify PGC-1α1 activators. • PGC-1α1 protein stabilizers work as activators in brown adipocytes. • Small molecule PGC-1α1 activators induce Ucp1 expression and cellular respiration.

[Objective] The very low-density lipoprotein receptor (VLDLR) plays an important role in the development of hepatic steatosis. In this study, we investigated the role of Peroxisome Proliferator-Activated Receptor (PPAR)b/d and fibroblast growth factor 21 (FGF21) in hepatic VLDLR regulation. [Methods] Studies were conducted in wild-type and Pparb/d-null mice, primary mouse hepatocytes, human Huh-7 hepatocytes, and liver biopsies from control subjects and patients with moderate and severe hepatic steatosis. [Results] Increased VLDLR levels were observed in liver of Pparb/d-null mice and in Pparb/d-knocked down mouse primary hepatocytes through mechanisms involving the heme-regulated eukaryotic translation initiation factor 2a (eIF2a) kinase (HRI), activating transcription factor (ATF) 4 and the oxidative stress-induced nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathways. Moreover, by using a neutralizing antibody against FGF21, Fgf21-null mice and by treating mice with recombinant FGF21, we show that FGF21 may protect against hepatic steatosis by attenuating endoplasmic reticulum (ER) stress-induced VLDLR upregulation. Finally, in liver biopsies from patients with moderate and severe hepatic steatosis, we observed an increase in VLDLR levels that was accompanied by a reduction in PPARb/d mRNA abundance and DNA-binding activity compared with control subjects. [Conclusions]: Overall, these findings provide new mechanisms by which PPARb/d and FGF21 regulate VLDLR levels and influence hepatic steatosis development., This study was partly supported by funds from the Spanish Ministry of the Economy and Competitiveness (SAF2015-64146-R to MVC, SAF2014-55725 to FV, and SAF2015-65267-R to AMV) and European Union ERDF funds. CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM) and CIBER Fisiopatologia de la Obesidad y Nutrición (CIBERobn) are Carlos III Health Institute projects. WW is supported by Start-Up Grants from the Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore, and by the Région Midi-Pyrénées, France, and TQL is supported by a CONACyT (National Council for Science and Technology in Mexico) Ph.D. scholarship.

Peroxisome proliferator-activated receptor (PPAR) α is widely expressed in the vasculature and has pleiotropic and lipid-lowering independent effects, but its role in the growth and function of vascular smooth muscle cells (VSMCs) during vascular pathophysiology is still unclear. Herein, VSMC-specific PPARα-deficient mice (PparaΔSMC) were generated by Cre-LoxP site-specific recombinase technology and VSMCs were isolated from mice aorta. PPARα deficiency attenuated VSMC apoptosis induced by angiotensin (Ang) II and hydrogen peroxide, and increased the migration of Ang II-challenged cells., Highlights • PPARα deficiency has both excitatory and inhibitory effects on VSMC function. • PPARα deficiency inhibits VSMC apoptosis but induces Ang II-induced migration. • The dual role of PPARα may lie behind the mixed clinical effects of fibrates.

Pregnane X receptor (PXR) plays an important role in xenobiotic metabolism. While ligand binding induces PXR-dependent gene transcription, PXR shows constitutive transcriptional activity in the absence of ligands when expressed in cultured cells. This constitutive activity sometimes hampers investigation of PXR activation by compounds of interest. In this study, we investigated the molecular mechanism of PXR activation. In the reported crystal structures of unliganded PXR, helix 12 (H12), including a coactivator binding motif, was stabilized, while it is destabilized in the unliganded structures of other nuclear receptors, suggesting a role for H12 stabilization in the basal activity of PXR. Since Phe420, located in the loop between H11 and H12, is thought to interact with Leu411 and Ile414 to stabilize H12, we substituted alanine at Phe420 (PXR-F420A) and separately inserted three alanine residues directly after Phe420 (PXR-3A) and investigated their influence on PXR-mediated transcription. Reporter gene assays demonstrated that the mutants showed drastically reduced basal activity and enhanced responses to various ligands, which was further enhanced by coexpression of the coactivator peroxisome proliferator-activated receptor gamma coactivator 1α. Mutations of both Leu411 and Ile414 to alanine also suppressed basal activity. Mammalian two-hybrid assays showed that PXR-F420A and PXR-3A bound to corepressors and coactivators in the absence and presence of ligands, respectively. We conclude that the intramolecular interactions of Phe420 with Leu411 and Ile414 stabilize H12 to recruit coactivators even in the absence of ligands, contributing to the basal transcriptional activity of PXR. We propose that the generated mutants might be useful for PXR ligand screening.

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by synovial inflammation and progressive joint destruction and is a primary cause of disability worldwide. Despite the existence of numerous anti-rheumatic drugs, a significant number of patients with RA do not respond or are intolerant to current treatments. Mesenchymal stem/stromal cell (MSCs) therapy represents a promising therapeutic tool to treat RA, mainly attributable to the immunomodulatory effects of these cells. This review comprises a comprehensive analysis of the scientific literature related to preclinical studies of MSC-based therapy in RA to analyse key aspects of current protocols as well as novel approaches which aim to improve the efficacy of MSC-based therapy.

Mitochondria are critical for regulation of the activation, differentiation, and survival of macrophages and other immune cells. In response to various extracellular signals, such as microbial or viral infection, changes to mitochondrial metabolism and physiology could underlie the corresponding state of macrophage activation. These changes include alterations of oxidative metabolism, mitochondrial membrane potential, and tricarboxylic acid (TCA) cycling, as well as the release of mitochondrial reactive oxygen species (mtROS) and mitochondrial DNA (mtDNA) and transformation of the mitochondrial ultrastructure. Here, we provide an updated review of how changes in mitochondrial metabolism and various metabolites such as fumarate, succinate, and itaconate coordinate to guide macrophage activation to distinct cellular states, thus clarifying the vital link between mitochondria metabolism and immunity. We also discuss how in disease settings, mitochondrial dysfunction and oxidative stress contribute to dysregulation of the inflammatory response. Therefore, mitochondria are a vital source of dynamic signals that regulate macrophage biology to fine-tune immune responses.

Cells adapt to an exposure to xenobiotics by upregulating the biosynthesis of proteins involved in xenobiotic metabolism. This is achieved largely via activation of cellular xenosensors that modulate gene expression. Biotransformation of xenobiotics frequently comes with the generation of reactive oxygen species (ROS). ROS, in turn, are known modulators of signal transduction processes. FOXO (forkhead box, class O) transcription factors are among the proteins deeply involved in the cellular response to stress, including oxidative stress elicited by the formation of ROS. On the one hand, FOXO activity is modulated by ROS, while on the other, FOXO target genes include many that encode antioxidant proteins – thereby establishing a regulatory circuit. Here, the role of ROS and of FOXOs in the regulation of xenosensor transcriptional activities will be discussed. Constitutive androstane receptor (CAR), pregnane X receptor (PXR), peroxisome proliferator-activated receptors (PPARs), arylhydrocarbon receptor (AhR) and nuclear factor erythroid 2-related factor 2 (Nrf2) all interact with FOXOs and/or ROS. The two latter not only fine-tune the activities of xenosensors but also mediate interactions between them. As a consequence, the emerging picture of an interplay between xenosensors, ROS and FOXO transcription factors suggests a modulatory role of ROS and FOXOs in the cellular adaptive response to xenobiotics., Graphical abstract fx1, Highlights • Exposure of cells to xenobiotics may trigger formation of reactive oxygen species. • Xenosensors respond to xenobiotics by upregulation of xenobiotic metabolism. • FOXO transcription factors modulate the activities of several xenosensors. • ROS affect FOXO activity, and FOXO target genes include antioxidant proteins. • FOXOs bridge xenobiotic-induced ROS generation and xenosensor regulation.

Objective Alternative activation (M2) of adipose tissue resident macrophage (ATM) inhibits obesity-induced metabolic inflammation. The underlying mechanisms remain unclear. Recent studies have shown that dysregulated lipid homeostasis caused by increased lipolysis in white adipose tissue (WAT) in the obese state is a trigger of inflammatory responses. We investigated the role of M2 macrophages in lipotoxicity-induced inflammation. Methods We used microarray experiments to profile macrophage gene expression regulated by two M2 inducers, interleukin-4 (Il-4), and peroxisome proliferator-activated receptor delta/gamma (Pparδ/Pparγ) agonists. Functional validation studies were performed in bone marrow-derived macrophages and mice deprived of the signal transducer and activator of transcription 6 gene (Stat6; downstream effector of Il-4) or Pparδ/Pparγ genes (downstream effectors of Stat6). Palmitic acid (PA) and β-adrenergic agonist were employed to induce macrophage lipid loading in vitro and in vivo, respectively. Results Profiling of genes regulated by Il-4 or Pparδ/Pparγ agonists reveals that alternative activation promotes the cell survival program, while inhibiting that of inflammation-related cell death. Deletion of Stat6 or Pparδ/Pparγ increases the susceptibility of macrophages to PA-induced cell death. NLR family pyrin domain containing 3 (Nlrp3) inflammasome activation by PA in the presence of lipopolysaccharide is also increased in Stat6−/− macrophages and to a lesser extent, in Pparδ/γ−/− macrophages. In concert, β-adrenergic agonist-induced lipolysis results in higher levels of cell death and inflammatory markers in ATMs derived from myeloid-specific Pparδ/γ−/− or Stat6−/− mice. Conclusions Our data suggest that ATM cell death is closely linked to metabolic inflammation. Within WAT where concentrations of free fatty acids fluctuate, M2 polarization regulated by the Stat6-Ppar axis enhances ATM's tolerance to lipid-mediated stress, thereby maintaining the homeostatic state., Highlights • Cell survival is transcriptionally regulated by macrophage alternative activation. • Fatty acid-triggered cell death is increased in Pparδ/γ−/− or Stat6−/− macrophages. • Il-4-Stat6 signaling suppresses lipotoxicity-induced inflammasome activation. • The Stat6-Pparδ/γ axis protects ATMs against lipolysis-induced cell death in vivo.

Objective Recently, we have shown that Bezafibrate (BEZ), the pan-PPAR (peroxisome proliferator-activated receptor) activator, ameliorated diabetes in insulin deficient streptozotocin treated diabetic mice. In order to study whether BEZ can also improve glucose metabolism in a mouse model for fatty liver and type 2 diabetes, the drug was applied to TallyHo mice. Methods TallyHo mice were divided into an early (ED) and late (LD) diabetes progression group and both groups were treated with 0.5% BEZ (BEZ group) or standard diet (SD group) for 8 weeks. We analyzed plasma parameters, pancreatic beta-cell morphology, and mass as well as glucose metabolism of the BEZ-treated and control mice. Furthermore, liver fat content and composition as well as hepatic gluconeogenesis and mitochondrial mass were determined. Results Plasma lipid and glucose levels were markedly reduced upon BEZ treatment, which was accompanied by elevated insulin sensitivity index as well as glucose tolerance, respectively. BEZ increased islet area in the pancreas. Furthermore, BEZ treatment improved energy expenditure and metabolic flexibility. In the liver, BEZ ameliorated steatosis, modified lipid composition and increased mitochondrial mass, which was accompanied by reduced hepatic gluconeogenesis. Conclusions Our data showed that BEZ ameliorates diabetes probably via reduced steatosis, enhanced hepatic mitochondrial mass, improved metabolic flexibility and elevated hepatic insulin sensitivity in TallyHo mice, suggesting that BEZ treatment could be beneficial for patients with NAFLD and impaired glucose metabolism., Highlights • Bezafibrate treatment reduced steatosis and ameliorated hepatic insulin resistance. • Bezafibrate treatment enhanced hepatic mitochondrial mass and metabolic flexibility. • Bezafibrate treatment increased beta-cell mass. • Bezafibrate treatment protected mice from developing diabetes. • Bezafibrate treatment normalized hyperglycemia in the manifest diabetic state.