RESUMEN
Itaconic acid is an emerging platform chemical with extensive applications. Itaconic acid is currently produced by Aspergillus terreus through biological fermentation. However, A. terreus is a fungal pathogen that needs additional morphology controls, making itaconic acid production on industrial scale problematic. Here, we reprogrammed the Generally Recognized As Safe (GRAS) yeast Yarrowia lipolytica for competitive itaconic acid production. After preventing carbon sink into lipid accumulation, we evaluated itaconic acid production both inside and outside the mitochondria while fine-tuning its biosynthetic pathway. We then mimicked the regulation of nitrogen limitation in nitrogen-replete conditions by down-regulating NAD+-dependent isocitrate dehydrogenase through weak promoters, RNA interference, or CRISPR interference. Ultimately, we optimized fermentation parameters for fed-batch cultivations and produced itaconic acid titers of 130.1 grams per liter in 1-liter bioreactors and 94.8 grams per liter in a 50-liter bioreactor on semipilot scale. Our findings provide effective approaches to harness the GRAS microorganism Y. lipolytica for competitive industrial-scale production of itaconic acid.
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Reactores Biológicos , Fermentación , Succinatos , Yarrowia , Yarrowia/metabolismo , Yarrowia/genética , Succinatos/metabolismo , Ingeniería Metabólica/métodos , Nitrógeno/metabolismo , Vías Biosintéticas , Isocitrato Deshidrogenasa/metabolismo , Isocitrato Deshidrogenasa/genéticaRESUMEN
Microbial alkane degradation pathways provide biological routes for converting these hydrocarbons into higher-value products. We recently reported the functional expression of a methyl-alkylsuccinate synthase (Mas) system in Escherichia coli, allowing for the heterologous anaerobic activation of short-chain alkanes. However, the enzymatic activation of methane via natural or engineered alkylsuccinate synthases has yet to be reported. To address this, we employed high-throughput screening to engineer the itaconate (IA)-responsive regulatory protein ItcR (WT-ItcR) from Yersinia pseudotuberculosis to instead respond to methylsuccinate (MS, the product of methane addition to fumarate), resulting in genetically encoded biosensors for MS. Here, we describe ItcR variants that, when regulating fluorescent protein expression in E. coli, show increased sensitivity, improved overall response, and enhanced specificity toward exogenously added MS relative to the wild-type repressor. Structural modeling and analysis of the ItcR ligand binding pocket provide insights into the altered molecular recognition. In addition to serving as biosensors for screening alkylsuccinate synthases capable of methane activation, MS-responsive ItcR variants also establish a framework for the directed evolution of other molecular reporters, targeting longer-chain alkylsuccinate products or other succinate derivatives.
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Técnicas Biosensibles , Metano , Metano/metabolismo , Escherichia coli/metabolismo , Anaerobiosis , Yersinia pseudotuberculosis/metabolismo , Evolución Molecular Dirigida , Proteínas Bacterianas/metabolismo , Succinatos/metabolismoRESUMEN
The Krebs cycle enzyme aconitate decarboxylase 1 (ACOD1) mediates itaconate synthesis in monocytes and macrophages. Previously, we reported that administration of 4-octyl itaconate to lupus-prone mice abrogated immune dysregulation and clinical features. In this study, we explore the role of the endogenous ACOD1/itaconate pathway in the development of TLR7-induced lupus (imiquimod [IMQ] model). We found that, in vitro, ACOD1 was induced in mouse bone marrow-derived macrophages and human monocyte-derived macrophages following TLR7 stimulation. This induction was partially dependent on type I IFN receptor signaling and on specific intracellular pathways. In the IMQ-induced mouse model of lupus, ACOD1 knockout (Acod1-/-) displayed disruptions of the splenic architecture, increased serum levels of anti-dsDNA and proinflammatory cytokines, and enhanced kidney immune complex deposition and proteinuria, when compared with the IMQ-treated wild-type mice. Consistent with these results, Acod1-/- bone marrow-derived macrophages treated in vitro with IMQ showed higher proinflammatory features. Furthermore, itaconate serum levels in systemic lupus erythematosus patients were decreased compared with healthy individuals, in association with disease activity and specific perturbed cardiometabolic parameters. These findings suggest that the ACOD1/itaconate pathway plays important immunomodulatory and vasculoprotective roles in systemic lupus erythematosus, supporting the potential therapeutic role of itaconate analogs in autoimmune diseases.
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Carboxiliasas , Lupus Eritematoso Sistémico , Macrófagos , Ratones Noqueados , Succinatos , Animales , Lupus Eritematoso Sistémico/inmunología , Ratones , Humanos , Femenino , Macrófagos/inmunología , Succinatos/farmacología , Enfermedades Cardiovasculares/inmunología , Biomarcadores , Ratones Endogámicos C57BL , Transducción de Señal/inmunología , Adulto , Masculino , Modelos Animales de Enfermedad , Persona de Mediana Edad , Citocinas/metabolismo , Receptor Toll-Like 7/metabolismo , HidroliasasRESUMEN
Itaconate is a promising platform chemical with broad applicability, including the synthesis of poly(methyl methacrylate). Most studies on microbial itaconate production entail the use of crop-based feedstock, which imposes constraints due to its limited supply. Brown macroalgae have recently gained attention as next-generation biomass owing to their high biomass productivity and carbohydrate content and amenability to mass production. Therefore, the use of macroalgae for itaconate production warrants exploration. In this study, the direct production of itaconate from brown macroalgae was demonstrated using engineered Vibrio sp. dhg, which has emerged as an efficient platform host for brown macroalgal biorefineries. Specifically, to enhance production, cis-aconitate decarboxylase (Cad) from Aspergillus terreus was heterologously expressed and isocitrate dehydrogenase (icd) was deleted. Notably, the resulting strain, VIC, achieved itaconate titers of 2.5 and 1.5 g/L from a mixture of alginate and mannitol (10 g/L of each) and 40 g/L of raw Saccharina japonica (S. japonica), respectively. Overall, this study highlights the utility of brown macroalgae as feedstock, as well as that of Vibrio sp. dhg as a platform strain for improving itaconate bioproduction.
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Ingeniería Metabólica , Phaeophyceae , Algas Marinas , Succinatos , Vibrio , Vibrio/metabolismo , Vibrio/genética , Vibrio/crecimiento & desarrollo , Algas Marinas/metabolismo , Algas Marinas/química , Phaeophyceae/metabolismo , Phaeophyceae/química , Succinatos/metabolismo , Aspergillus/metabolismo , Aspergillus/genética , Aspergillus/enzimología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , BiomasaAsunto(s)
Artritis Reumatoide , Fibroblastos , Succinatos , Sinoviocitos , Humanos , Artritis Reumatoide/patología , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Fibroblastos/metabolismo , Sinoviocitos/efectos de los fármacos , Sinoviocitos/metabolismo , Sinoviocitos/patología , Succinatos/farmacologíaRESUMEN
Polymeric derivatives of itaconic acid are gaining interest as biobased alternatives to petroleum-based monomers due to their versatility, renewable nature, commercial availability, and cost-effectiveness. Itaconate ester monomer's challenges incorporating in (meth)acrylic waterborne polymers are the low propagation rate, unfavorable reactivity ratios, and the depropagation process. To overcome these challenges, the seeded semibatch emulsion polymerization of 100% biobased dibutyl itaconate, methyl methacrylate, and butyl acrylate was investigated at different temperatures. Consequently, 30 wt % DBI was successfully incorporated within waterborne (meth)acrylates in short reaction times (4 h), obtaining high DBI incorporation (>90%). The results demonstrate that DBI incorporation influences the instantaneous monomer conversion, polymer's microstructure, and mechanical properties. By incorporating a biobased itaconate cross-linker, kinetics and mechanical characteristics of the polymers were improved. This combined approach can be implemented without altering industrial processes, resolving the commercialization dilemma for itaconate monomers to synthesize high-performance biobased polymers for adhesive and coating industries.
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Polimerizacion , Succinatos , Succinatos/química , Acrilatos/química , Polímeros/química , Agua/químicaRESUMEN
Ustilago maydis and Ustilago cynodontis are natural producers of a broad range of valuable molecules including itaconate, malate, glycolipids, and triacylglycerols. Both Ustilago species are insensitive toward medium impurities, and have previously been engineered for efficient itaconate production and stabilized yeast-like growth. Due to these features, these strains were already successfully used for the production of itaconate from different alternative feedstocks such as molasses, thick juice, and crude glycerol. Here, we analyzed the amylolytic capabilities of Ustilago species for metabolization of starch, a highly abundant and low-cost polymeric carbohydrate widely utilized as a substrate in several biotechnological processes. Ustilago cynodontis was found to utilize gelatinized potato starch for both growth and itaconate production, confirming the presence of extracellular amylolytic enzymes in Ustilago species. Starch was rapidly degraded by U. cynodontis, even though no α-amylase was detected. Further experiments indicate that starch hydrolysis is caused by the synergistic action of glucoamylase and α-glucosidase enzymes. The enzymes showed a maximum activity of around 0.5 U ml-1 at the fifth day after inoculation, and also released glucose from additional substrates, highlighting potential broader applications. In contrast to U. cynodontis, U. maydis showed no growth on starch accompanied with no detectable amylolytic activity.
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Almidón , Succinatos , Ustilago , Ustilago/metabolismo , Ustilago/genética , Ustilago/enzimología , Ustilago/crecimiento & desarrollo , Almidón/metabolismo , Succinatos/metabolismo , Glucano 1,4-alfa-Glucosidasa/metabolismo , HidrólisisRESUMEN
Itaconic acid and its derivative 4-octyl itaconate (OI) represent a novel anti-inflammatory medication that has demonstrated efficacy in multiple inflammation models because of its minimal side effects. Recently, natural polymers conjugated with small molecule drugs, known as polymer-drug conjugates (PDCs), have emerged as a promising approach to sustained drug release. In this work, we reported an approach to prepare a PDC containing an OI and make it into an injectable hydrogel. Chitosan (CS) was selected for PDC synthesis because of its abundant free amino groups that can be conjugated with molecules containing carboxyl groups by carbodiimide chemistry. We used an ethanol/water cosolvent system to synthesize a CS-OI conjugate via EDC/NHS catalysis. The CS-OI conjugate had improved water solubility and unique anti-inflammatory activity and did not show compromised antibacterial activity compared with unmodified CS. Beta-glycerophosphate (ß-GP) cross-linked CS-OI hydrogel exhibited good injectability with sustainable OI release and effectively modulated inflammatory response in a rat model. Therefore, this study provides valuable insights into the design of PDC hydrogels with inflammatory modulatory properties.
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Antiinflamatorios , Quitosano , Hidrogeles , Inflamación , Succinatos , Quitosano/química , Animales , Succinatos/química , Succinatos/farmacología , Hidrogeles/química , Hidrogeles/farmacología , Inflamación/tratamiento farmacológico , Antiinflamatorios/farmacología , Antiinflamatorios/química , Antiinflamatorios/administración & dosificación , Ratas , Masculino , Ratas Sprague-Dawley , Ratones , Antibacterianos/farmacología , Antibacterianos/química , Antibacterianos/administración & dosificaciónRESUMEN
Mesotrione, as a widely used herbicide, is present in the environment in detectable amounts, causing serious damage. Here, we aimed to investigate the effect of mesotrione on Caco-2 cells and the possibility of its toxicity mitigation by cichoric acid. Therefore, we analyzed the cytotoxicity of both these compounds and the selected oxidative stress parameters, apoptosis and interaction of both the tested compounds with the cell membrane and their accumulation within the cells. In cytotoxicity studies, the stimulating activity of mesotrione was observed, and simultaneously, the inhibitory effect of cichoric acid was noticed. This effect was related to the results of oxidative stress analysis and apoptosis measurements. The activity level of key enzymes (glutathione peroxidase, catalase and superoxide dismutase) in Caco-2 cells exposed to cichoric acid was higher as compared to that of the control. The treatment with mesotrione did not induce apoptosis in the Caco-2 cells. The penetration of the studied compounds into the Caco-2 cells was measured by using an HPLC methodology, and the results indicate mesotrione's high penetration capacity. The distribution of charge on the surface of the cell membranes changed under the influence of both compounds. Considering the mutual interactions of beneficial and potentially toxic food ingredients, it should be noted that, despite the observed favorable trend, cichoric acid is not able to overcome the toxic and cancer-stimulating effects of this pesticide.
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Apoptosis , Ácidos Cafeicos , Ciclohexanonas , Estrés Oxidativo , Humanos , Células CACO-2 , Apoptosis/efectos de los fármacos , Ciclohexanonas/farmacología , Estrés Oxidativo/efectos de los fármacos , Ácidos Cafeicos/farmacología , Succinatos/farmacología , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Adenocarcinoma/tratamiento farmacológico , Adenocarcinoma/patología , Adenocarcinoma/metabolismo , Herbicidas/toxicidad , Superóxido Dismutasa/metabolismo , Supervivencia Celular/efectos de los fármacos , Catalasa/metabolismo , Glutatión Peroxidasa/metabolismo , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismoRESUMEN
Nirmatrelvir, a pivotal component of the oral antiviral Paxlovid for COVID-19, targets the SARS-CoV-2 main protease (Mpro) as a covalent inhibitor. Here, we employed combined computational methods to explore how the prevalent Omicron variant mutation P132H, alone and in combination with A173V (P132H-A173V), affects nirmatrelvir's efficacy. Our findings suggest that P132H enhances the noncovalent binding affinity of Mpro for nirmatrelvir, whereas P132H-A173V diminishes it. Although both mutants catalyze the rate-limiting step more efficiently than the wild-type (WT) Mpro, P132H slows the overall rate of covalent bond formation, whereas P132H-A173V accelerates it. Comprehensive analysis of noncovalent and covalent contributions to the overall binding free energy of the covalent complex suggests that P132H likely enhances Mpro sensitivity to nirmatrelvir, while P132H-A173V may confer resistance. Per-residue decompositions of the binding and activation free energies pinpoint key residues that significantly affect the binding affinity and reaction rates, revealing how the mutations modulate these effects. The mutation-induced conformational perturbations alter drug-protein local contact intensities and the electrostatic preorganization of the protein, affecting noncovalent binding affinity and the stability of key reaction states, respectively. Our findings inform the mechanisms of nirmatrelvir resistance and sensitivity, facilitating improved drug design and the detection of resistant strains.
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Antivirales , Proteasas 3C de Coronavirus , Mutación , SARS-CoV-2 , SARS-CoV-2/enzimología , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/genética , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Proteasas 3C de Coronavirus/metabolismo , Proteasas 3C de Coronavirus/química , Proteasas 3C de Coronavirus/genética , Antivirales/farmacología , Antivirales/química , Humanos , Tratamiento Farmacológico de COVID-19 , Simulación de Dinámica Molecular , Inhibidores de Proteasas/farmacología , Inhibidores de Proteasas/química , Inhibidores de Proteasas/metabolismo , Leucina/química , Termodinámica , Sulfonamidas/farmacología , Sulfonamidas/química , Sulfonamidas/metabolismo , Unión Proteica , Succinatos/química , Succinatos/farmacología , Succinatos/metabolismo , Lactamas , Nitrilos , ProlinaRESUMEN
The role of oxidative stress and ferroptosis in osteoarthritis (OA) pathogenesis is increasingly recognized. Notably, 4-octyl Itaconate (OI) has been documented to counteract oxidative stress and inflammatory responses, highlighting its therapeutic potential in OA. This study explored the effects of OI on GPX4 methylation, oxidative stress, and ferroptosis in chondrocytes affected by OA. Our results demonstrated that OI mitigated IL-1ß-induced chondrocyte degeneration in a dose-dependent manner. It also suppressed reactive oxygen species (ROS) production and sustained GPX4 expression, thereby attenuating the degenerative impact of IL-1ß and Erastin on chondrocytes by curtailing ferroptosis. Moreover, we observed that blocking GPX4 methylation could alleviate IL-1ß-induced degeneration, oxidative stress, and ferroptosis in chondrocytes. The regulatory mechanism of OI on GPX4 expression in chondrocytes involved the inhibition of GPX4 methylation. In a mouse model of OA, OI's protective effects against OA were comparable to those of Ferrostatin-1. Thus, OI reduced chondrocyte degeneration, oxidative stress, and ferroptosis by inhibiting GPX4 methylation, offering a novel mechanistic insight into its therapeutic application in OA.
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Condrocitos , Ferroptosis , Interleucina-1beta , Ratones Endogámicos C57BL , Osteoartritis , Estrés Oxidativo , Fosfolípido Hidroperóxido Glutatión Peroxidasa , Succinatos , Animales , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Ferroptosis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Succinatos/farmacología , Succinatos/uso terapéutico , Interleucina-1beta/metabolismo , Osteoartritis/tratamiento farmacológico , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Fosfolípido Hidroperóxido Glutatión Peroxidasa/genética , Ratones , Masculino , Humanos , Metilación/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Células Cultivadas , Modelos Animales de EnfermedadRESUMEN
Cotinine, the primary metabolite of nicotine in the human body, is an emerging pollutant in aquatic environments. It causes environmental problems and is harmful to the health of humans and other mammals; however, the mechanisms of its biodegradation have been elucidated incompletely. In this study, a novel Gram-negative strain that could degrade and utilize cotinine as a sole carbon source was isolated from municipal wastewater samples, and its cotinine degradation characteristics and kinetics were determined. Pseudomonas sp. JH-2 was able to degrade 100 mg/L (0.56 mM) of cotinine with high efficiency within 5 days at 30 â, pH 7.0, and 1% NaCl. Two intermediates, 6-hydroxycotinine and 6-hydroxy-3-succinoylpyridine (HSP), were identified by high-performance liquid chromatography and liquid chromatograph mass spectrometer. The draft whole genome sequence of strain JH-2 was obtained and analyzed to determine genomic structure and function. No homologs of proteins predicted in Nocardioides sp. JQ2195 and reported in nicotine degradation Pyrrolidine pathway were found in strain JH-2, suggesting new enzymes that responsible for cotinine catabolism. These findings provide meaningful insights into the biodegradation of cotinine by Gram-negative bacteria.
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Biodegradación Ambiental , Cotinina , Pseudomonas , Aguas Residuales , Pseudomonas/metabolismo , Pseudomonas/genética , Pseudomonas/aislamiento & purificación , Pseudomonas/clasificación , Cotinina/metabolismo , Cotinina/análogos & derivados , Aguas Residuales/microbiología , Nicotina/metabolismo , Nicotina/análogos & derivados , Piridinas/metabolismo , Genoma Bacteriano , Filogenia , SuccinatosRESUMEN
OBJECTIVE: To describe and compare prothrombin time (PT), activated partial thromboplastin time (aPTT), thromboelastography (TEG), HCT, and platelet count measurements in a hemorrhage/over-resuscitation model. DESIGN: Randomized crossover study. SETTING: University teaching hospital. ANIMALS: Six cats. INTERVENTIONS: Anesthetized cats underwent 3 treatments at 2-month intervals. The treatments were as follows: NHR-no controlled hemorrhage and sham resuscitation; LRS-controlled hemorrhage and lactated Ringer's solution (LRS) for resuscitation; and Voluven-controlled hemorrhage and 6% tetrastarch 130/0.4 for resuscitation. The LRS and Voluven were administered at 60 and 20 mL/kg/h, respectively, for 120 minutes. Blood samples were drawn for PT, aPTT, TEG, HCT, and platelet count measurements at a healthy check (T - 7d), after controlled hemorrhage (T0), at 60 and 120 minutes of resuscitation (T60 and T120), and at 24 hours after completion of resuscitation (T24h). Data were analyzed using a general linear mixed model approach (significance was P < 0.05). MEASUREMENTS AND MAIN RESULTS: Total median blood loss (controlled hemorrhage and blood sampling from T0 to T120) at T120 was 11.4, 31.0, and 30.8 mL/kg for NHR, LRS, and Voluven, respectively. PT and aPTT during LRS and Voluven were prolonged at T60 and T120 compared to NHR (P < 0.001). On TEG, the reaction time, kinetic time, and alpha-angle were within reference intervals for cats at all time points in all treatments, while maximum amplitude was less than the reference interval (40 mm) at T0, T60, and T120 during Voluven and at T60 and T120 during LRS compared to NHR (both P < 0.001). The HCT and platelet count were significantly lower at T60 and T120 during LRS and Voluven compared to NHR (P < 0.001). CONCLUSIONS: Hypocoagulopathy was observed during hemorrhage and liberal fluid resuscitation. Prolongation of PT and aPPT and decreased clot strength may have been caused by hemodilution and platelet loss.
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Estudios Cruzados , Tiempo de Protrombina , Resucitación , Lactato de Ringer , Tromboelastografía , Animales , Gatos , Tromboelastografía/veterinaria , Tromboelastografía/métodos , Lactato de Ringer/administración & dosificación , Lactato de Ringer/farmacología , Recuento de Plaquetas/veterinaria , Tiempo de Protrombina/veterinaria , Hematócrito/veterinaria , Tiempo de Tromboplastina Parcial/veterinaria , Resucitación/veterinaria , Resucitación/métodos , Hemorragia/veterinaria , Hemorragia/sangre , Enfermedades de los Gatos/sangre , Derivados de Hidroxietil Almidón/farmacología , Derivados de Hidroxietil Almidón/administración & dosificación , Masculino , Femenino , Gelatina/administración & dosificación , Gelatina/farmacología , SuccinatosRESUMEN
Dicaffeoyltartaric acid (diCT) and 3,5-dicaffeoylquinic acid (3,5-diCQ) are described for their aphicidal properties on several aphid species. Intending to valorize diCT and 3,5-diCQ as biocontrol products and because of the high adaptive capacities of aphids to xenobiotics, we sought to determine the existence of adaptation first in Myzus persicae (Sulzer) (Hemiptera: Aphididae) and then other aphids. Resistance of aphids to these biopesticides could be promoted by (i) the existence of resistance to synthetic insecticides that may confer cross-resistance and (ii) the presence of these compounds in wild plants likely which may have led to pre-existing adaptation in aphids. We assessed the resistance levels to diCT and 3,5-diCQ in 7 lab strains (including some resistant to synthetic aphicides) and 7 wild populations of M. persicae using biotests. The activities of detoxification enzymes contributing to insecticide resistance were also measured. Additionally, we followed the same method to characterize susceptibility to these caffeic derivatives in wild populations of Nasonovia ribisnigri (Mosley) (Hemiptera: Aphididae), Brevicoryne brassicae(Linnaeus) (Hemiptera: Aphididae) and, Aphis craccivora(Koch) (Hemiptera: Aphididae). Our results show variability in susceptibility to diCT between populations of M. persicae, but resistance ratios (RR) were low (RRâ =â 3.59). We found no cross-resistance between synthetic insecticides and diCT. Carboxylesterase and glutathione-S-transferase did not seem to be involved in its detoxification. A clone of A. craccivora collected from peanut, a species rich in diCT, was not susceptible to either diCT or 3,5-diCQ, suggesting a common molecular target for these 2 molecules and the existence of a high-effect resistance mechanism. These active botanical substances remain good candidates for M. persicae biocontrol in agriculture.
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Áfidos , Ácidos Cafeicos , Resistencia a los Insecticidas , Insecticidas , Áfidos/efectos de los fármacos , Animales , Insecticidas/farmacología , Ácidos Cafeicos/farmacología , Ácido Quínico/análogos & derivados , Ácido Quínico/farmacología , SuccinatosRESUMEN
Diabetes and derived complications, especially diabetic nephropathy and neuropathy annually cause great morbimortality worldwide. 5-hydroxytryptamine (5-HT) acts as a modulator of renal sympathetic input and vascular tone. In this line, 5-HT2 receptor blockade has been linked with reduced incidence and progression of diabetic microvascular alterations. In this work, we aimed to determine, in diabetic rats, whether 5-HT2 blockade ameliorates renal function and to characterize the serotonergic modulatory action on renal sympathetic neurotransmission. Diabetes was induced in male Wistar rats by alloxan administration (150â¯mg/kg, s.c.), and sarpogrelate (30â¯mg/kg·day, p.o.; 5-HT2 antagonist) was administered for 14 days (DM-S). Normoglycemic and diabetic (DM) animals were maintained as aged-matched controls. At 28th day, DM-S animals were anesthetized and prepared for the in situ autoperfusion of the kidney. Renal vasoconstrictor responses were induced electrically or by i.a. noradrenaline (NA) administration. The role of 5-HT and selective 5-HT agonist/antagonist were studied on these renal vasopressor responses. Sarpogrelate treatment decreased renal sympathetic-induced vasopressor responses, reduced renal hypertrophy and kidney damage markers increased in DM. Intraarterial 5-HT inhibited the sympathetic-induced renal vasoconstrictions, effect reproduced by 5-CT, AS-19, L-694,247 and LY 344864 (5-HT1/5/7, 5-HT7, 5-HT1D and 5-HT1F receptor agonists, respectively). Blocking 5-HT1D/1F/7 receptors completely abolished the 5-CT sympatho-inhibition. NA vasoconstrictions were not altered by any of the 5-HT agonists tested. Thus, in experimental diabetes, chronic sarpogrelate treatment reduces renal damage markers, kidney hypertrophy and renal sympathetic hyperactivity and modifies serotonergic modulation of renal sympathetic neurotransmission, causing a sympatho-inhibition by prejunctional 5-HT1D/1F and 5-HT7 activation.
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Diabetes Mellitus Experimental , Riñón , Ratas Wistar , Succinatos , Sistema Nervioso Simpático , Animales , Succinatos/farmacología , Masculino , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/fisiopatología , Riñón/efectos de los fármacos , Riñón/inervación , Sistema Nervioso Simpático/efectos de los fármacos , Sistema Nervioso Simpático/fisiopatología , Ratas , Antagonistas del Receptor de Serotonina 5-HT2/farmacología , Serotonina/metabolismo , Nefropatías Diabéticas/tratamiento farmacológico , Nefropatías Diabéticas/fisiopatología , Vasoconstricción/efectos de los fármacosRESUMEN
The presence of heterogeneity in responses to oncolytic virotherapy poses a barrier to clinical effectiveness, as resistance to this treatment can occur through the inhibition of viral spread within the tumor, potentially leading to treatment failures. Here we show that 4-octyl itaconate (4-OI), a chemical derivative of the Krebs cycle-derived metabolite itaconate, enhances oncolytic virotherapy with VSVΔ51 in various models including human and murine resistant cancer cell lines, three-dimensional (3D) patient-derived colon tumoroids and organotypic brain tumor slices. Furthermore, 4-OI in combination with VSVΔ51 improves therapeutic outcomes in a resistant murine colon tumor model. Mechanistically, we find that 4-OI suppresses antiviral immunity in cancer cells through the modification of cysteine residues in MAVS and IKKß independently of the NRF2/KEAP1 axis. We propose that the combination of a metabolite-derived drug with an oncolytic virus agent can greatly improve anticancer therapeutic outcomes by direct interference with the type I IFN and NF-κB-mediated antiviral responses.
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Viroterapia Oncolítica , Virus Oncolíticos , Succinatos , Animales , Humanos , Viroterapia Oncolítica/métodos , Succinatos/farmacología , Ratones , Línea Celular Tumoral , Interferón Tipo I/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Neoplasias del Colon/terapia , Neoplasias del Colon/inmunología , Neoplasias del Colon/tratamiento farmacológico , Antivirales/farmacología , FN-kappa B/metabolismo , Quinasa I-kappa B/metabolismo , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Inflamación/tratamiento farmacológico , Femenino , Virus de la Estomatitis Vesicular Indiana/fisiología , Virus de la Estomatitis Vesicular Indiana/efectos de los fármacos , Transducción de Señal/efectos de los fármacosRESUMEN
Increasingly prevalent, nontuberculous mycobacteria (NTM) infections affect approximately 20% of people with cystic fibrosis (CF). Previous studies of CF sputum identified lower levels of the host metabolite itaconate in those infected with NTM. Itaconate can inhibit the growth of M. tuberculosis (MTB) in vitro via the inhibition of the glyoxylate cycle enzyme (ICL), but its impact on NTM is unclear. To test itaconic acid's (IA) effect on NTM growth, laboratory and CF clinical strains of Mycobacterium abscessus and Mycobacterium avium were cultured in 7H9 minimal media supplemented with 1-10 mM of IA and short-chain fatty acids (SCFA). M. avium and M. abscessus grew when supplemented with SCFAs, whereas the addition of IA (≥ 10 mM) completely inhibited NTM growth. NTM supplemented with acetate or propionate and 5 mM IA displayed slower growth than NTM cultured with SCFA and ≤ 1 mM of IA. However, IA's inhibition of NTM was pH dependent; as similar and higher quantities (100 mM) of pH adjusted IA (pH 7) did not inhibit growth in vitro, while in an acidic minimal media (pH 6.1), 1 to 5 mM of non-pH adjusted IA inhibited growth. None of the examined isolates displayed the ability to utilize IA as a carbon source, and IA added to M. abscessus isocitrate lyase (ICL) decreased enzymatic activity. Lastly, the addition of cell-permeable 4-octyl itaconate (4-OI) to THP-1 cells enhanced NTM clearance, demonstrating a potential role for IA/itaconate in host defense against NTM infections.
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Succinatos , Succinatos/farmacología , Succinatos/metabolismo , Humanos , Concentración de Iones de Hidrógeno , Micobacterias no Tuberculosas/efectos de los fármacos , Micobacterias no Tuberculosas/crecimiento & desarrollo , Células THP-1 , Infecciones por Mycobacterium no Tuberculosas/microbiología , Infecciones por Mycobacterium no Tuberculosas/tratamiento farmacológico , Mycobacterium avium/efectos de los fármacos , Mycobacterium avium/crecimiento & desarrollo , Mycobacterium abscessus/efectos de los fármacos , Mycobacterium abscessus/crecimiento & desarrollo , Mycobacterium abscessus/metabolismoRESUMEN
BACKGROUND: Quercetin has received extensive attention for its therapeutic potential treating respiratory syncytial virus (RSV) infection diseases. Recent studies have highlighted quercetin's ability of suppressing alveolar macrophages (AMs)-derived lung inflammation. However, the anti-inflammatory mechanism of quercetin against RSV infection still remains elusive. PURPOSE: This study aims to elucidate the mechanism about quercetin anti-inflammatory effect on RSV infection. METHODS: BALB/c mice were intranasally infected with RSV and received quercetin (30, 60, 120 mg/kg/d) orally for 3 days. Additionally, an in vitro infection model utilizing mouse alveolar macrophages (MH-S cells) was employed to validate the proposed mechanism. RESULTS: Quercetin exhibited a downregulatory effect on glycolysis and tricarboxylic acid (TCA) cycle metabolism in RSV-infected AMs. However, it increased itaconic acid production, a metabolite derived from citrate through activating immune responsive gene 1 (IRG1), and further inhibiting succinate dehydrogenase (SDH) activity. While the suppression of SDH activity orchestrated a cascading downregulation of Hif-1α/NLRP3 signaling, ultimately causing AMs polarization from M1 to M2 phenotypes. CONCLUSION: Our study demonstrated quercetin stimulated IRG1-mediated itaconic acid anabolism and further inhibited SDH/Hif-1α/NLRP3 signaling pathway, which led to M1 to M2 polarization of AMs so as to ameliorate RSV-induced lung inflammation.
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Subunidad alfa del Factor 1 Inducible por Hipoxia , Macrófagos Alveolares , Ratones Endogámicos BALB C , Proteína con Dominio Pirina 3 de la Familia NLR , Quercetina , Infecciones por Virus Sincitial Respiratorio , Succinatos , Animales , Succinatos/farmacología , Macrófagos Alveolares/efectos de los fármacos , Infecciones por Virus Sincitial Respiratorio/tratamiento farmacológico , Quercetina/farmacología , Ratones , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Succinato Deshidrogenasa/metabolismo , Glucólisis/efectos de los fármacos , Femenino , Transducción de Señal/efectos de los fármacos , Ciclo del Ácido Cítrico/efectos de los fármacos , Virus Sincitiales Respiratorios/efectos de los fármacos , Antiinflamatorios/farmacología , HidroliasasRESUMEN
Cichoric acid (CA), a widely utilized polyphenolic compound in medicine, has garnered significant attention due to its potential health benefits. Sepsis-induced acute kidney disease (AKI) is related with an elevated risk of end-stage kidney disease (ESKD). However, it remains unclear whether CA provides protection against septic AKI. The aim of this study is to investigated the protective effect and possible mechanisms of CA against LPS-induced septic AKI. Sepsis-induced AKI was induced in mice through intraperitoneal injection of lipopolysaccharide (LPS), and RAW264.7 macrophages were incubated with LPS. LPS exposure significantly increased the levels of M1 macrophage biomarkers while reducing the levels of M2 macrophage indicators. This was accompanied by the release of inflammatory factors, superoxide anion production, mitochondrial dysfunction, activation of succinate dehydrogenase (SDH), and subsequent succinate formation. Conversely, pretreatment with CA mitigated these abnormalities. CA attenuated hypoxia-inducible factor-1α (HIF-1α)-induced glycolysis by lifting the NAD+/NADH ratio in macrophages. Additionally, CA disrupted the K (lysine) acetyltransferase 2A (KAT2A)/α-tubulin complex, thereby reducing α-tubulin acetylation and subsequently inactivating the NLRP3 inflammasome. Importantly, administration of CA ameliorated LPS-induced renal pathological damage, apoptosis, inflammation, oxidative stress, and disturbances in mitochondrial function in mice. Overall, CA restrained HIF-1α-mediated glycolysis via inactivation of SDH, leading to NLRP3 inflammasome inactivation and the amelioration of sepsis-induced AKI.