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Traumatic brain injury (TBI) is a significant public health concern characterized by a complex cascade of cellular events. TBI induces adenosine monophosphate-activated protein kinase (AMPK) dysfunction impairs energy balance activates inflammatory cytokines and leads to neuronal damage. AMPK is a key regulator of cellular energy homeostasis during inflammatory responses. Recent research has revealed its key role in modulating the inflammatory process in TBI. Following TBI the activation of AMPK can influence various important pathways and mechanisms including metabolic pathways and inflammatory signaling. Our study investigated the effects of post-TBI loss of AMPK function on functional outcomes inflammasome activation, and inflammatory cytokine production. Male C57BL/6 adult wild-type (WT) and AMPK knockout (AMPK-KO) mice were subjected to a controlled cortical impact (CCI) model of TBI or sham surgery. The mice were tested for behavioral impairment at 24 h post-TBI thereafter, mice were anesthetized, and their brains were quickly removed for histological and biochemical evaluation. In vitro we investigated inflammasome activation in mixed glial cells stimulated with lipopolysaccharides+ Interferon-gamma (LI) (0.1 µg/20 ng/ml LPS/IFNg) for 6 h to induce an inflammatory response. Estimating the nucleotide-binding domain, leucine-rich-containing family pyrin domain containing western blotting ELISA and qRT-PCR performed 3 (NLRP3) inflammasome activation and cytokine production. Our findings suggest that TBI leads to reduced AMPK phosphorylation in WT mice and that the loss of AMPK correlates with worsened behavioral deficits at 24 h post-TBI in AMPK-KO mice as compared to WT mice. Moreover compared with the WT mice AMPK-KO mice exhibit exacerbated NLRP3 inflammasome activation and increased expression of proinflammatory mediators such as IL-1b IL-6 TNF-a iNOS and Cox 2. These results align with the in vitro studies using brain glial cells under inflammatory conditions, demonstrating greater activation of inflammasome components in AMPK-KO mice than in WT mice. Our results highlighted the critical role of AMPK in TBI outcomes. We found that the absence of AMPK worsens behavioral deficits and heightens inflammasome-mediated inflammation thereby exacerbating brain injury after TBI. Restoring AMPK activity after TBI could be a promising therapeutic approach for alleviating TBI-related damage.
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Upon antigenic stimulation, CD4 + T-cells undergo clonal expansion, elevating their bioenergetic demands and utilization of nutrients like glucose and glutamine. The nuclear factor erythroid 2-related factor 2 (Nrf2) is a well-known regulator of oxidative stress, but its involvement in modulating the metabolism of CD4 + T-cells remains unexplored. Here, we elucidate the role of Nrf2 beyond the traditional antioxidation, in modulating activation-driven expansion of CD4 + T-cells by influencing their nutrient metabolism. T-cell-specific activation of Nrf2 enhances early activation and IL-2 secretion, upregulates TCR-signaling, and increases activation-driven proliferation of CD4 + T-cells. Mechanistically, high Nrf2 inhibits glucose metabolism through glycolysis but promotes glutamine metabolism via glutaminolysis to support increased T-cell proliferation. Further, Nrf2 expression is temporally regulated in activated CD4 + T-cells with elevated expression during the early activation, but decreased expression thereafter. Overall, our findings uncover a novel role of Nrf2 as a metabolic modulator of CD4 + T-cells, thus providing a framework for improving Nrf2-targeting therapies and T-cell immunotherapies.
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Zika virus (ZIKV) infection during pregnancy causes severe neurological and ocular abnormalities in infants, yet no vaccine or antivirals are available. Our transcriptomic analysis of ZIKV-infected retinal pigment epithelial (RPE) cells revealed alterations in the cholesterol pathway. Thus, we investigated the functional roles of ATP binding cassette transporter G1 (ABCG1) and sterol response element binding protein 2 (SREPB-2), two key players in cholesterol metabolism, during ocular ZIKV infection. Our in vitro data showed that increased ABCG1 activity via liver X receptors (LXRs), reduced ZIKV replication, while ABCG1 knockdown increased replication with elevated intracellular cholesterol. Conversely, inhibiting SREBP-2 or its knockdown reduced ZIKV replication by lowering cholesterol levels. In vivo, LXR agonist or SREBP-2 inhibitor treatment mitigated ZIKV-induced chorioretinal lesions in mice, concomitant with decreased expression of inflammatory mediators and increased activation of antiviral response genes. In summary, our study identifies ABCG1's antiviral role and SREBP-2's proviral effects in ocular ZIKV infection, offering cholesterol metabolism as a potential target to develop antiviral therapies.
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Introduction: Multiple sclerosis (MS) is the most common inflammatory neurodegenerative disease of the central nervous system (CNS) in young adults and results in progressive neurological defects. The relapsing-remitting phenotype (RRMS) is the most common disease course in MS and may progress to the progressive form (PPMS). Objectives: There is a gap in knowledge regarding whether the relapsing form can be distinguished from the progressive course or healthy subjects (HS) based on an altered serum metabolite profile. In this study, we performed global untargeted metabolomics with the 2D GCxGC-MS platform to identify altered metabolites between RRMS, PPMS, and HS. Methods: We profiled 235 metabolites in the serum of patients with RRMS (n=41), PPMS (n=31), and HS (n=91). A comparison of RRMS and HS patients revealed 22 significantly altered metabolites at p<0.05 (false discovery rate [FDR]=0.3). The PPMS and HS comparisons revealed 28 altered metabolites at p<0.05 (FDR=0.2). Results: Pathway analysis using MetaboAnalyst revealed enrichment of four metabolic pathways in both RRMS and PPMS (hypergeometric test p<0.05): 1) galactose metabolism; 2) amino sugar and nucleotide sugar metabolism; 3) phenylalanine, tyrosine, and tryptophan biosynthesis; and 4) aminoacyl-tRNA biosynthesis. The Qiagen IPA enrichment test identified the sulfatase 2 (SULF2) (p=0.0033) and integrin subunit beta 1 binding protein 1 (ITGB1BP1) (p=0.0067) genes as upstream regulators of altered metabolites in the RRMS vs. HS groups. However, in the PPMS vs. HS comparison, valine was enriched in the neurodegeneration of brain cells (p=0.05), and heptadecanoic acid, alpha-ketoisocaproic acid, and glycerol participated in inflammation in the CNS (p=0.03). Conclusion: Overall, our study suggested that RRMS and PPMS may contribute metabolic fingerprints in the form of unique altered metabolites for discriminating MS disease from HS, with the potential for constructing a metabolite panel for progressive autoimmune diseases such as MS.
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Bacterial endophthalmitis is a blinding infectious disease typically acquired during ocular surgery. We previously reported significant alterations in retinal metabolism during Staphylococcus (S) aureus endophthalmitis. However, the changes in retinal lipid composition during endophthalmitis are unknown. Here, using a mouse model of S. aureus endophthalmitis and an untargeted lipidomic approach, we comprehensively analyzed temporal alterations in total lipids and oxylipin in retina. Our data showed a time-dependent increase in the levels of lipid classes, sphingolipids, glycerolipids, sterols, and non-esterified fatty acids, whereas levels of phospholipids decreased. Among lipid subclasses, phosphatidylcholine decreased over time. The oxylipin analysis revealed increased prostaglandin-E2, hydroxyeicosatetraenoic acids, docosahexaenoic acid, eicosapentaenoic acid, and α-linolenic acid. In-vitro studies using mouse bone marrow-derived macrophages showed increased lipid droplets and lipid-peroxide formation in response to S. aureus infection. Collectively, these findings suggest that S. aureus-infection alters the retinal lipid profile, which may contribute to the pathogenesis of bacterial endophthalmitis.
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Endoftalmite , Staphylococcus aureus , Humanos , Staphylococcus aureus/fisiologia , Lipidômica , Oxilipinas , Endoftalmite/microbiologia , Endoftalmite/patologia , Retina/patologiaRESUMO
Unresolved and uncontrolled inflammation is considered a hallmark of pathogenesis in chronic inflammatory diseases like multiple sclerosis (MS), suggesting a defective resolution process. Inflammatory resolution is an active process partially mediated by endogenous metabolites of dietary polyunsaturated fatty acids (PUFA), collectively termed specialized pro-resolving lipid mediators (SPMs). Altered levels of resolution mediators have been reported in several inflammatory diseases and may partly explain impaired inflammatory resolution. Performing LC-MS/MS-based targeted lipid mediator profiling, we observed distinct changes in fatty acid metabolites in serum from 30 relapsing-remitting MS (RRMS) patients relative to 30 matched healthy subjects (HS). Robust linear regression revealed 12 altered lipid mediators after adjusting for confounders (p <0.05). Of these, 15d-PGJ2, PGE3, and LTB5 were increased in MS while PGF2a, 8,9-DiHETrE, 5,6-DiHETrE, 20-HETE, 15-HETE, 12-HETE, 12-HEPE, 14-HDoHE, and DHEA were decreased in MS compared to HS. In addition, 12,13-DiHOME and 12,13-DiHODE were positively correlated with expanded disability status scale values (EDSS). Using Partial Least Squares, we identified several lipid mediators with high VIP scores (VIP > 1: 32% - 52%) of which POEA, PGE3, DHEA, LTB5, and 12-HETE were top predictors for distinguishing between RRMS and HS (AUC =0.75) based on the XGBoost Classifier algorithm. Collectively, these findings suggest an imbalance between inflammation and resolution. Altogether, lipid mediators appear to have potential as diagnostic and prognostic biomarkers for RRMS.
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The metabolic needs of the premature/premyelinating oligodendrocytes (pre-OLs) and mature oligodendrocytes (OLs) are distinct. The metabolic control of oligodendrocyte maturation from the pre-OLs to the OLs is not fully understood. Here, we show that the terminal maturation and higher mitochondrial respiration in the OLs is an integrated process controlled through pyruvate dehydrogenase complex (Pdh). Combined bioenergetics and metabolic studies show that OLs show elevated mitochondrial respiration than the pre-OLs. Our signaling studies show that the increased mitochondrial respiration activity in the OLs is mediated by the activation of Pdh due to inhibition of the pyruvate dehydrogenase kinase-1 (Pdhk1) that phosphorylates and inhibits Pdh activity. Accordingly, when Pdhk1 is directly expressed in the pre-OLs, they fail to mature into the OLs. While Pdh converts pyruvate into the acetyl-CoA by its oxidative decarboxylation, our study shows that Pdh-dependent acetyl-CoA generation from pyruvate contributes to the acetylation of the bHLH family transcription factor, oligodendrocyte transcription factor 1 (Olig1) which is known to be involved in the OL maturation. Pdh inhibition via direct expression of Pdhk1 in the pre-OLs blocks the Olig1-acetylation and OL maturation. Using the cuprizone model of demyelination, we show that Pdh is deactivated during the demyelination phase, which is however reversed in the remyelination phase upon cuprizone withdrawal. In addition, Pdh activity status correlates with the Olig1-acetylation status in the cuprizone model. Hence, the Pdh metabolic node activation allows a robust mitochondrial respiration and activation of a molecular program necessary for the terminal maturation of oligodendrocytes. Our findings open a new dialogue in the developmental biology that links cellular development and metabolism. These findings have far-reaching implications in the development of therapies for a variety of demyelinating disorders including multiple sclerosis.
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Esclerose Múltipla , Remielinização , Humanos , Cuprizona , Reprogramação Metabólica , Acetilcoenzima A , Oligodendroglia/fisiologia , Oxirredutases , Piruvatos , Fatores de TranscriçãoRESUMO
Multiple sclerosis (MS) is the most common inflammatory neurodegenerative disease in young adults, resulting in neurological defects and disability. The endogenous mechanisms to resolve inflammation are intact but become defective in patients, resulting in lack of resolution mediators and unresolved chronic inflammation. Docosahexaenoic acid (DHA) metabolism being impaired in MS, we hypothesize that supplementing its downstream metabolite maresin 1 (MaR1) will alleviate inflammation and demyelination in preclinical mouse model of MS; experimental allergic encephalomyelitis (EAE). Restoration of MaR1 by its exogenous administration in EAE mice propagated inflammatory resolution and had a protective effect on neurological deficits, prevented disease progression, and reduced disease severity by reducing immune cell infiltration (CD4+IL17+ and CD4+IFN-γ+) into the CNS. It significantly reduced the proinflammatory cytokine IL17 and promoted an anti-inflammatory response via IL10 and IL4. Neutralization of IL10 abolished the protective effect of MaR1 in EAE confirming IL10 is mediating MaR1 effect in EAE. Furthermore, it improved the pathophysiology and exerted neuroprotective effects by mitigating disease signs in EAE as evidenced by lower levels of NFL in the plasma of treated group compared to control and higher MBP expression in the brain from the MaR1 treated mice, decreased inflammatory infiltrates, and less demyelination and vacuolization in the spinal cord tissue sections of treated mice. SCENITH data confirmed that MaR1 maintains myelin by regulating oligodendrocyte metabolism. Also, it induces metabolic reprogramming in infiltrating CD4 cells and macrophages, which modulate their phenotype. Metabolic changes induced macrophages by MaR1 restores the impaired efferocytosis in EAE, promoting clearance of damaged myelin and dead cells; thereby lowering the disability with disease course. Overall, MaR1 supplementation has anti-inflammatory and neuroprotective effects in preclinical animal models and induces metabolic reprogramming in disease associated cell-types, promotes efferocytosis, implying that it could be a new therapeutic molecule in MS and other autoimmune diseases. Highlights: Inflammation is dysregulated in EAE due to impaired synthesis of DHA derived proresolving lipid mediator MaR1.Administration of the resolution agonist MaR1 propagates resolution processes and improves neurological outcome in RR model of EAE.MaR1 ameliorates clinical signs of EAE by attenuating pro-inflammatory cytokine IL17 mediated response and promoting anti-inflammatory response through IL10.MaR1 supplementation improves the pathophysiology in EAE and shows neuroprotection as indicated by the lower levels of NFL in the plasma and higher expression of MBP in the brain of treated mice.MaR1 induces metabolic reprogramming in disease-associated cell types.MaR1 promotes efferocytosis in EAE through metabolic reprogramming of macrophages. Significance: Inflammatory process is a protective response to several challenges like injury or infection. However, it must resolve over time to maintain tissue homeostasis. Impaired or delayed resolution leads to damaging effects, including chronic inflammation, tissue damage, and disease progression as occurs in multiple sclerosis (MS). We report that inflammation is dysregulated in preclinical animal model of MS, experimental autoimmune encephalomyelitis (EAE), partially due to impaired synthesis of proresolving lipid mediators. We show that the administration of the resolution agonist known as maresin 1 (MaR1) in EAE actively propagates resolution processes and improves neurological outcome. We conclude that MaR1 is a potential interventional candidate to attenuate dysregulated inflammation and to restore neurological deficits in EAE.
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Metabolism and energy processes governing oligodendrocyte function during neuroinflammatory disease are of great interest. However, how varied cellular environments affect oligodendrocyte activity during neuroinflammation is unknown. We demonstrate that activated microglial energy metabolism controls oligodendrocyte mitochondrial respiration and activity. Lipopolysaccharide/interferon gamma promote glycolysis and decrease mitochondrial respiration and myelin protein synthesis in rat brain glial cells. Enriched microglia showed an early burst in glycolysis. In microglia-conditioned medium, oligodendrocytes did not respire and expressed less myelin. SCENITH revealed metabolic derangement in microglia and O4-positive oligodendrocytes in endotoxemia and experimental autoimmune encephalitogenic models. The early burst of glycolysis in microglia was mediated by PDPK1 and protein kinase B/AKT signaling. We found that microglia-produced NO and itaconate, a tricarboxylic acid bifurcated metabolite, reduced mitochondrial respiration in oligodendrocytes. During inflammation, we discovered a signaling pathway in microglia that could be used as a therapeutic target to restore mitochondrial function in oligodendrocytes and induce remyelination.
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In various cancer models, dietary interventions have been shown to inhibit tumor growth, improve anticancer drug efficacy, and enhance immunity, but no such evidence exists for epithelial ovarian cancer (EOC), the most lethal gynecologic cancer. The anticancer immune responses induced by 16-h intermittent fasting (IF) were studied in mice with EOC. IF consistently reduced metabolic growth factors and cytokines that stimulate tumor growth, creating a tumor-hostile environment. Immune profiling showed that IF dramatically alters anti-cancer immunity by increasing CD4+ and CD8+ cells, Th1 and cytotoxic responses, and metabolic fitness. ß-hydroxy butyrate (BHB), a bioactive metabolite produced by IF, partially imitates its anticancer effects by inducing CD8+ effector function. In a direct comparison, IF outperformed exogenous BHB treatment in survival and anti-tumor immune response, probably due to increased ketogenesis. Thus, IF and one of its metabolic mediators BHB suppress EOC growth and sustain a potent anti-tumor T cell response.
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Dietary isoflavones, a type of phytoestrogens, have gained importance owing to their health-promoting benefits. However, the beneficial effects of isoflavones are mediated by smaller metabolites produced with the help of gut bacteria that are known to metabolize these phytoestrogenic compounds into Daidzein and Genistein and biologically active molecules such as S-Equol. Identifying and measuring these phytoestrogens and their metabolites is an important step towards understanding the significance of diet and gut microbiota in human health and diseases. We have overcome the reported difficulties in quantitation of these isoflavones and developed a simplified, sensitive, non-enzymatic, and sulfatases-free extraction methodology. We have subsequently used this method to quantify these metabolites in the urine of mice using UPLC-MS/MS. The extraction and quantitation method was validated for precision, linearity, accuracy, recoveries, limit of detection (LOD), and limit of quantification (LOQ). Linear calibration curves for Daidzein, Genistein, and S-Equol were set up by performing linear regression analysis and checked using the correlation coefficient (r2 > 0.995). LOQs for Daidzein, Genistein, and S-Equol were 2, 4, and 2 ng/mL, respectively. This UPLC-MS/MS swift method is suitable for quantifying isoflavones and the microbial-derived metabolite S-Equol in mice urine and is particularly useful for large numbers of samples.
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Genisteína , Isoflavonas , Humanos , Camundongos , Animais , Genisteína/análise , Fitoestrógenos/urina , Equol , Cromatografia Líquida , Espectrometria de Massas em Tandem , Isoflavonas/análise , DietaRESUMO
Bacterial endophthalmitis is a severe complication of eye surgeries that can lead to vision loss. Current treatment involves intravitreal antibiotic injections that control bacterial growth but not inflammation. To identify newer therapeutic targets to promote inflammation resolution in endophthalmitis, we recently employed an untargeted metabolomics approach. This led to the discovery that the levels of S-nitroso-L-glutathione (GSNO) were significantly reduced in an experimental murine Staphylococcus aureus (SA) endophthalmitis model. In this study, we tested the hypothesis whether GSNO supplementation via different routes (oral, intravitreal) provides protection during bacterial endophthalmitis. Our results show that prophylactic administration of GSNO via intravitreal injections ameliorated SA endophthalmitis. Therapeutically, oral administration of GSNO was found to be most effective in reducing intraocular inflammation and bacterial burden. Moreover, oral GSNO treatment synergized with intravitreal antibiotic injections in reducing the severity of endophthalmitis. Furthermore, in vitro experiments using cultured human retinal Muller glia and retinal pigment epithelial (RPE) cells showed that GSNO treatment reduced SA-induced inflammatory mediators and cell death. Notably, both in-vivo and ex-vivo data showed that GSNO strengthened the outer blood-retinal barrier during endophthalmitis. Collectively, our study demonstrates GSNO as a potential therapeutic agent for the treatment of intraocular infections due to its dual anti-inflammatory and cytoprotective properties.
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Endoftalmite , Infecções Oculares Bacterianas , Infecções Estafilocócicas , Camundongos , Humanos , Animais , Modelos Animais de Doenças , Endoftalmite/tratamento farmacológico , Endoftalmite/microbiologia , Infecções Estafilocócicas/tratamento farmacológico , Inflamação/tratamento farmacológico , Staphylococcus aureus , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Glutationa , Infecções Oculares Bacterianas/tratamento farmacológico , Infecções Oculares Bacterianas/microbiologia , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Administração OralRESUMO
Epithelial Ovarian Cancer (EOC) is the most lethal gynecologic cancer with limited genetic alterations identified that can be therapeutically targeted. In tumor bearing mice, short-term fasting, fasting mimicking diet and calorie restriction enhance the activity of antineoplastic treatment by modulating systemic metabolism and boosting anti-tumor immunity. We tested the outcome of sixteen-hour intermittent fasting (IF) on mouse EOC progression with focus on fasting driven antitumor immune responses. IF resulted in consistent decrease of tumor promoting metabolic growth factors and cytokines, recapitulating changes that creates a tumor antagonizing environment. Immune profiling revealed that IF profoundly reshapes anti-cancer immunity by inducing increase in CD4+ and CD8+ cells, paralleled by enhanced antitumor Th1 and cytotoxic responses, by enhancing their metabolic fitness. Metabolic studies revealed that IF generated bioactive metabolite BHB which can be a potential substitute for simulating the antitumor benefits of IF. However, in a direct comparison, IF surpassed exogenous BHB therapy in improving survival and activating anti-tumor immune response. Thus, our data provides strong evidence for IF and its metabolic mediator BHB for ameliorating EOC progression and as a viable approach in maintaining and sustaining an effective anti-tumor T cell response.
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Experimental autoimmune encephalomyelitis (EAE) remains a widely used pre-clinical animal model to study multiple sclerosis (MS). Blood-based cytokines and CNS biomarkers are increasingly used as predictors of neurodegeneration, disease activity, and disability in MS. However, there exists variation in animal model characterization and disease course across animal strains/studies due to understudied confounding factors, limiting the utility of these biomarkers to predict disease activity in EAE. In this study, we investigated the profile of blood-based analytes including, cytokines (IL6, IL17, IL12p70, IL10, and TNFα) and neural markers (NFL and GFAP) in the plasma of relapsing-remitting (RR) (SJL) and chronic (B6) models of EAE during different phases (acute, chronic, and progressive) of disease course using ultrasensitive single molecule array technology (SIMoA, Quanterix), which can detect ultra-low levels of a wide range of analytes. NFL showed a substantial increase during post-disease onset at peak, chronic, and progressive phases in both RR SJL and chronic B6 models of EAE. The increase was markedly pronounced in the chronic B6 model. The leakage of GFAP from CNS into the periphery was also higher after disease onset in EAE, however, it was highest during the acute phase in B6. Out of all cytokines, only IL10 showed consistently lower levels in both models of EAE along the disease duration. We report that NFL, GFAP, and IL10 may be more useful predictors of disease activity and neurological outcome in EAE, which would make them potential candidates for use as surrogate markers for preclinical testing of therapeutic interventions in MS.
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Pathogenic Th17 cells are crucial to CNS autoimmune diseases like multiple sclerosis (MS), though their control by endogenous mechanisms is unknown. RNAseq analysis of brain glial cells identified immuno-responsive gene 1 (Irg1), a mitochondrial-related enzyme-coding gene, as one of the highly upregulated gene under inflammatory conditions which were further validated in the spinal cord of animals with experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Moreover, Irg1 mRNA and protein levels in myeloid, CD4, and B cells were higher in the EAE group, raising questions about its function in CNS autoimmunity. We observed that Irg1 knockout (KO) mice exhibited severe EAE disease and greater mononuclear cell infiltration, including triple-positive CD4 cells expressing IL17a, GM-CSF, and IFNγ. Lack of Irg1 in macrophages led to higher levels of Class II expression and polarized myelin primed CD4 cells into pathogenic Th17 cells through the NLRP3/IL1ß axis. Our findings show that Irg1 in macrophages plays an important role in the formation of pathogenic Th17 cells, emphasizing its potential as a therapy for autoimmune diseases, including MS.
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The etiopathogenesis of multiple sclerosis (MS) is strongly affected by environmental factors such as diet and the gut microbiota. An isoflavone-rich (ISO) diet was previously shown to reduce the severity of MS in the animal model experimental autoimmune encephalomyelitis (EAE). Translation of this concept to clinical trial where dietary isoflavones may be recommended for MS patients will require preliminary evidence that providing the isoflavone-rich diet to people with MS (PwMS) who lack phytoestrogen-metabolizing bacteria has beneficial effects. We have previously shown that the gut microbiota of PwMS resembles the gut microbiota of mice raised under a phytoestrogen-free (phyto-free) diet in that it lacks phytoestrogen-metabolizing bacteria. To investigate the effects of phytoestrogens on the microbiota inflammatory response and EAE disease severity we switched the diet of mice raised under a phyto-free (PF) diet to an isoflavone-rich diet. Microbiota analysis showed that the change in diet from one that is ISO to one that is PF reduces beneficial bacteria such as Bifidobacterium species. In addition we observed functional differences in lipopolysaccharide (LPS) biosynthesis pathways. Moreover LPS extracted from feces of mice fed an ISO diet induced increased production of anti-inflammatory cytokines from bone marrow-derived macrophages relative to fecal-LPS isolated from mice fed a PF diet. Eventually mice whose diet was switched from a PF diet to an ISO diet trended toward reduced EAE severity and mortality. Overall we show that an isoflavone-rich diet specifically modulates LPS biosynthesis of the gut microbiota imparts an anti-inflammatory response and decreases disease severity.
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Encefalomielite Autoimune Experimental , Microbioma Gastrointestinal , Isoflavonas , Animais , Citocinas/metabolismo , Dieta , Encefalomielite Autoimune Experimental/microbiologia , Inflamação , Isoflavonas/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Fitoestrógenos/farmacologiaRESUMO
To ameliorate diabetes mellitus-associated heart failure with preserved ejection fraction (HFpEF), we plan to lower diabetes-mediated oxidative stress-induced 4-hydroxy-2-nonenal (4HNE) accumulation by pharmacological agents that either decrease 4HNE generation or increase its detoxification.A cellular reactive carbonyl species (RCS), 4HNE, was significantly increased in diabetic hearts due to a diabetes-induced decrease in 4HNE detoxification by aldehyde dehydrogenase (ALDH) 2, a cardiac mitochondrial enzyme that metabolizes 4HNE. Therefore, hyperglycemia-induced 4HNE is critical for diabetes-mediated cardiotoxicity and we hypothesize that lowering 4HNE ameliorates diabetes-associated HFpEF. We fed a high-fat diet to ALDH2*2 mice, which have intrinsically low ALDH2 activity, to induce type-2 diabetes. After 4 months of diabetes, the mice exhibited features of HFpEF along with increased 4HNE adducts, and we treated them with vehicle, empagliflozin (EMP) (3 mg/kg/d) to reduce 4HNE and Alda-1 (10 mg/kg/d), and ALDH2 activator to enhance ALDH2 activity as well as a combination of EMP + Alda-1 (E + A), via subcutaneous osmotic pumps. After 2 months of treatments, cardiac function was assessed by conscious echocardiography before and after exercise stress. EMP + Alda-1 improved exercise tolerance, diastolic and systolic function, 4HNE detoxification and cardiac liver kinase B1 (LKB1)-AMP-activated protein kinase (AMPK) pathways in ALDH2*2 mice with diabetes-associated HFpEF. This combination was even more effective than EMP alone. Our data indicate that ALDH2 activation along with the treatment of hypoglycemic agents may be a salient strategy to alleviate diabetes-associated HFpEF.
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Diabetes Mellitus Tipo 2 , Insuficiência Cardíaca , Proteínas Quinases Ativadas por AMP/metabolismo , Aldeído Desidrogenase/metabolismo , Aldeído-Desidrogenase Mitocondrial/genética , Aldeído-Desidrogenase Mitocondrial/metabolismo , Animais , Compostos Benzidrílicos , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Glucosídeos , Insuficiência Cardíaca/tratamento farmacológico , Insuficiência Cardíaca/etiologia , Hipoglicemiantes/farmacologia , Hipoglicemiantes/uso terapêutico , Camundongos , Volume SistólicoRESUMO
4-hydroxy-2-nonenal (4HNE), an oxidative stress byproduct, is elevated in diabetes which decreases coronary angiogenesis, and this was rescued by the 4HNE detoxifying enzyme, aldehyde dehydrogenase 2 (ALDH2). Adiponectin (APN), an adipocytokine, has pro-angiogenic properties and its loss of function is critical in diabetes and its complications. Coronary endothelial cell (CEC) damage is the initiating step of diabetes-mediated heart failure with preserved ejection fraction (HFpEF) pathogenesis. Thus, we hypothesize that ALDH2 restores 4HNE-induced downregulation of APN signaling in CECs and subsequent coronary angiogenesis in diabetic HFpEF. Treatment with disulfiram, an ALDH2 inhibitor, exacerbated 4HNE-mediated decreases in APN-induced increased coronary angiogenesis and APN-signaling cascades, whereas pretreatment with alda1, an ALDH2 activator, rescued the effect of 4HNE. We employed control mice (db/m), spontaneous type-2 diabetic mice (db/db), ALDH2*2 knock-in mutant mice with intrinsic low ALDH2 activity (AL), and diabetic mice with intrinsic low ALDH2 activity (AF) mice that were created by crossing db/db and AL mice to test our hypothesis in vivo. AF mice exhibited heart failure with preserved ejection fraction (HFpEF)/severe diastolic dysfunction at 6 months with a preserved systolic function compared with db/db mice as well as 3 months of their age. Decreased APN-mediated coronary angiogenesis, along with increased circulatory APN levels and decreased cardiac APN signaling (index of APN resistance) were higher in AF mice relative to db/db mice. Alda1 treatment improved APN-mediated angiogenesis in AF and db/db mice. In summary, 4HNE-induces APN resistance and a subsequent decrease in coronary angiogenesis in diabetic mouse heart which was rescued by ALDH2.
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Diabetes Mellitus Experimental , Insuficiência Cardíaca , Adiponectina , Aldeído-Desidrogenase Mitocondrial/genética , Animais , Diabetes Mellitus Experimental/patologia , Camundongos , Volume SistólicoRESUMO
Metabolic aberrations impact the pathogenesis of multiple sclerosis (MS) and possibly can provide clues for new treatment strategies. Using untargeted metabolomics, we measured serum metabolites from 35 patients with relapsing-remitting multiple sclerosis (RRMS) and 14 healthy age-matched controls. Of 632 known metabolites detected, 60 were significantly altered in RRMS. Bioinformatics analysis identified an altered metabotype in patients with RRMS, represented by four changed metabolic pathways of glycerophospholipid, citrate cycle, sphingolipid, and pyruvate metabolism. Interestingly, the common upstream metabolic pathway feeding these four pathways is the glycolysis pathway. Real-time bioenergetic analysis of the patient-derived peripheral blood mononuclear cells showed enhanced glycolysis, supporting the altered metabolic state of immune cells. Experimental autoimmune encephalomyelitis mice treated with the glycolytic inhibitor 2-deoxy-D-glucose ameliorated the disease progression and inhibited the disease pathology significantly by promoting the antiinflammatory phenotype of monocytes/macrophage in the central nervous system. Our study provided a proof of principle for how a blood-based metabolomic approach using patient samples could lead to the identification of a therapeutic target for developing potential therapy.
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Desenvolvimento de Medicamentos , Glicólise , Metabolômica , Esclerose Múltipla Recidivante-Remitente , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Antimetabólitos/farmacologia , Antimetabólitos/uso terapêutico , Desoxiglucose/farmacologia , Desoxiglucose/uso terapêutico , Desenvolvimento de Medicamentos/métodos , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Leucócitos Mononucleares/metabolismo , Camundongos , Esclerose Múltipla Recidivante-Remitente/sangue , Esclerose Múltipla Recidivante-Remitente/tratamento farmacológico , Esclerose Múltipla Recidivante-Remitente/metabolismoRESUMO
Multiple sclerosis (MS) is a debilitating autoimmune disorder. Currently, there is a lack of effective treatment for the progressive form of MS, partly due to insensitive readout for neurodegeneration. The recent development of sensitive assays for neurofilament light chain (NfL) has made it a potential new biomarker in predicting MS disease activity and progression, providing an additional readout in clinical trials. However, NfL is elevated in other neurodegenerative disorders besides MS, and, furthermore, it is also confounded by age, body mass index (BMI), and blood volume. Additionally, there is considerable overlap in the range of serum NfL (sNfL) levels compared to healthy controls. These confounders demonstrate the limitations of using solely NfL as a marker to monitor disease activity in MS patients. Other blood and cerebrospinal fluid (CSF) biomarkers of axonal damage, neuronal damage, glial dysfunction, demyelination, and inflammation have been studied as actionable biomarkers for MS and have provided insight into the pathology underlying the disease process of MS. However, these other biomarkers may be plagued with similar issues as NfL. Using biomarkers of a bioinformatic approach that includes cellular studies, micro-RNAs (miRNAs), extracellular vesicles (EVs), metabolomics, metabolites and the microbiome may prove to be useful in developing a more comprehensive panel that addresses the limitations of using a single biomarker. Therefore, more research with recent technological and statistical approaches is needed to identify novel and useful diagnostic and prognostic biomarker tools in MS.