RESUMEN
Despite all the scientific progress in recent decades to unravel the immune processes and the way the parasite bypasses the immune system, Chagas disease is still a major public health problem, affecting an estimated 3.5 million people. Among the components that may participate in the response against the parasite, testosterone has been gaining more and more visibility. Studies indicate that the parasite itself seems to carry out steroidogenesis, in which, in co-culture with androgen precursors, T. cruzi has been shown to produce TS, but the purpose of the TS synthesized by the parasite and how this can influence its invasion glycoproteins is still unclear unknown. The aim of this study was to evaluate the influence of testosterone in Trypanosoma cruzi infection on the immune response of bone marrow-derived macrophages. Bone marrow from male rats was extracted and cultured with RMPI medium containing 30% L929 cell supernatant for macrophage differentiation. The cells were incubated for 10 days and, after this period, they were seeded in 96 wells in the amount of 1 x 105 cells per well. TS was added at different concentrations of 20 µM, 10 µM, 5 µM and 1 µM and then infected with the Y strain of T. cruzi, at a rate of 10 parasites per cell, with the culture remaining for six, 12 and 24 h. The supernatant was collected and the production of nitric oxide (NO), tumor necrosis factor (TNF) and the number of cell parasites was assessed by staining with 4'-6'-diamino-2-phenylindole (DAPI) and ranked by high Content Screening (HSC). The parasite was then cultured with the addition of TS, at the mentioned concentrations, leaving it for six and 12 h and then performing the RT-PCR of the mucins. DAPI staining revealed a significant increase in the number of parasites in cells containing TS. The exception was observed when 1 µM of hormone/well was used. A reduction in TNF production was found with 20 and 10 µM of TS for 6 h stimulation, although increased levels were observed with 5 and 1 µM, similar to the infected control. However, there was an increase in TNF production and not after 12 h. The relative expression of parasite glycoprotein 82 was increased with the presence of TS in the medium, regardless of time. Our data suggest that TS may contribute to cellular immunosuppression, increasing parasite infection in the cell, as well as inflammatory mediators that lead to cell and tissue damage in infected individuals, as well as the possible use of TS to allow their invasion into the cell hosts.
Asunto(s)
Macrófagos , Óxido Nítrico , Testosterona , Trypanosoma cruzi , Animales , Masculino , Macrófagos/parasitología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratas , Testosterona/biosíntesis , Testosterona/farmacología , Óxido Nítrico/biosíntesis , Óxido Nítrico/metabolismo , Glicoproteínas/metabolismo , Glicoproteínas/farmacología , Ratas Wistar , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesis , Mediadores de Inflamación/metabolismo , Proteínas Protozoarias/metabolismo , Células Cultivadas , Células de la Médula Ósea/parasitología , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/inmunologíaRESUMEN
Trypanosoma cruzi (T. cruzi) causes Chagas, which is a neglected tropical disease (NTD). WHO estimates that 6 to 7 million people are infected worldwide. Current treatment is done with benznidazole (BZN), which is very toxic and effective only in the acute phase of the disease. In this work, we designed, synthesized, and characterized thirteen new phenoxyhydrazine-thiazole compounds and applied molecular docking and in vitro methods to investigate cell cytotoxicity, trypanocide activity, nitric oxide (NO) production, cell death, and immunomodulation. We observed a higher predicted affinity of the compounds for the squalene synthase and 14-alpha demethylase enzymes of T. cruzi. Moreover, the compounds displayed a higher predicted affinity for human TLR2 and TLR4, were mildly toxic in vitro for most mammalian cell types tested, and LIZ531 (IC50 2.8 µM) was highly toxic for epimastigotes, LIZ311 (IC50 8.6 µM) for trypomastigotes, and LIZ331 (IC50 1.9 µM) for amastigotes. We observed that LIZ311 (IC50 2.5 µM), LIZ431 (IC50 4.1 µM) and LIZ531 (IC50 5 µM) induced 200 µg/mL of NO and JM14 induced NO production in three different concentrations tested. The compound LIZ331 induced the production of TNF and IL-6. LIZ311 induced the secretion of TNF, IFNγ, IL-2, IL-4, IL-10, and IL-17, cell death by apoptosis, decreased acidic compartment formation, and induced changes in the mitochondrial membrane potential. Taken together, LIZ311 is a promising anti-T. cruzi compound is not toxic to mammalian cells and has increased antiparasitic activity and immunomodulatory properties.
Asunto(s)
Enfermedad de Chagas , Simulación del Acoplamiento Molecular , Óxido Nítrico , Tiazoles , Tripanocidas , Trypanosoma cruzi , Trypanosoma cruzi/efectos de los fármacos , Tiazoles/farmacología , Tiazoles/química , Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/inmunología , Humanos , Animales , Ratones , Óxido Nítrico/metabolismo , Óxido Nítrico/biosíntesis , Tripanocidas/farmacología , Tripanocidas/química , Concentración 50 Inhibidora , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Hidrazinas/farmacología , Hidrazinas/química , Citocinas/metabolismo , Ratones Endogámicos BALB CRESUMEN
Yersinia enterocolitica (Ye) inserts outer proteins (Yops) into cytoplasm to infect host cells. However, in spite of considerable progress, the mechanisms implicated in this process, including the association of Yops with host proteins, remain unclear. Here, we evaluated the functional role of Galectin-1 (Gal1), an endogenous ß-galactoside-binding protein, in modulating Yop interactions with host cells. Our results showed that Gal1 binds to Yops in a carbohydrate-dependent manner. Interestingly, Gal1 binding to Yops protects these virulence factors from trypsin digestion. Given that early control of Ye infection involves activation of macrophages, we evaluated the role of Gal1 and YopP in the modulation of macrophage function. Although Gal1 and YopP did not influence production of superoxide anion and/or TNF by Ye-infected macrophages, they coordinately inhibited nitric oxide (NO) production. Notably, recombinant Gal1 (rGal1) did not rescue NO increase observed in Lgals1-/- macrophages infected with the YopP mutant Ye ∆yopP. Whereas NO induced apoptosis in macrophages, no significant differences in cell death were detected between Gal1-deficient macrophages infected with Ye ∆yopP, and WT macrophages infected with Ye wt. Strikingly, increased NO production was found in WT macrophages treated with MAPK inhibitors and infected with Ye wt. Finally, rGal1 administration did not reverse the protective effect in Peyer Patches (PPs) of Lgals1-/- mice infected with Ye ∆yopP. Our study reveals a cooperative role of YopP and endogenous Gal1 during Ye infection.
Asunto(s)
Proteínas de la Membrana Bacteriana Externa/metabolismo , Galectina 1/metabolismo , Inmunidad , Óxido Nítrico/biosíntesis , Yersinia enterocolitica/inmunología , Secuencia de Aminoácidos , Animales , Proteínas de la Membrana Bacteriana Externa/química , Ratones Endogámicos C57BL , Ratones Noqueados , Péptido Hidrolasas/metabolismo , Unión Proteica , Proteolisis , Proteómica , Factores de Virulencia/metabolismo , Yersinia enterocolitica/patogenicidadRESUMEN
Polyphenols play a therapeutic role in vascular diseases, acting in inherent illness-associate conditions such as inflammation, diabetes, dyslipidemia, hypertension, and oxidative stress, as demonstrated by clinical trials and epidemiological surveys. The main polyphenol cardioprotective mechanisms rely on increased nitric oxide, decreased asymmetric dimethylarginine levels, upregulation of genes encoding antioxidant enzymes via the Nrf2-ARE pathway and anti-inflammatory action through the redox-sensitive transcription factor NF-κB and PPAR-γ receptor. However, poor polyphenol bioavailability and extensive metabolization restrict their applicability. Polyphenols carried by nanoparticles circumvent these limitations providing controlled release and better solubility, chemical protection, and target achievement. Nano-encapsulate polyphenols loaded in food grade polymers and lipids appear to be safe, gaining resistance in the enteric route for intestinal absorption, in which the mucoadhesiveness ensures their increased uptake, achieving high systemic levels in non-metabolized forms. Nano-capsules confer a gradual release to these compounds, as well as longer half-lives and cell and whole organism permanence, reinforcing their effectiveness, as demonstrated in pre-clinical trials, enabling their application as an adjuvant therapy against cardiovascular diseases. Polyphenol entrapment in nanoparticles should be encouraged in nutraceutical manufacturing for the fortification of foods and beverages. This study discusses pre-clinical trials evaluating how nano-encapsulate polyphenols following oral administration can aid in cardiovascular performance.
Asunto(s)
Antioxidantes/farmacología , Cardiotónicos/farmacología , Composición de Medicamentos/métodos , Hipertensión/tratamiento farmacológico , Isquemia Miocárdica/tratamiento farmacológico , Polifenoles/farmacología , Elementos de Respuesta Antioxidante , Antioxidantes/química , Antioxidantes/farmacocinética , Arginina/análogos & derivados , Arginina/antagonistas & inhibidores , Arginina/metabolismo , Cardiotónicos/química , Cardiotónicos/farmacocinética , Diabetes Mellitus/tratamiento farmacológico , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Diabetes Mellitus/fisiopatología , Portadores de Fármacos , Dislipidemias/tratamiento farmacológico , Dislipidemias/genética , Dislipidemias/metabolismo , Dislipidemias/fisiopatología , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hipertensión/genética , Hipertensión/metabolismo , Hipertensión/fisiopatología , Isquemia Miocárdica/genética , Isquemia Miocárdica/metabolismo , Isquemia Miocárdica/fisiopatología , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Nanocápsulas/administración & dosificación , Nanocápsulas/química , Óxido Nítrico/antagonistas & inhibidores , Óxido Nítrico/biosíntesis , Estrés Oxidativo/efectos de los fármacos , Polifenoles/química , Polifenoles/farmacocinética , Transducción de SeñalRESUMEN
Leishmania is an obligate intracellular parasite that primarily inhabits macrophages. The destruction of the parasite in the host cell is a fundamental mechanism for infection control. In addition, inhibition of the leishmanicidal activity of macrophages seems to be related to the ability of some species to inhibit the production of nitric oxide (NO) by depleting arginine. Some species of Leishmania have the ability to produce NO from a constitutive nitric oxide synthase-like enzyme (cNOS-like). However, the localization of cNOS-like in Leishmania has not been described before. As such, this study was designed to locate cNOS-like enzyme and NO production in promastigotes of Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis. NO production was initially quantified by flow cytometry, which indicated a significant difference in NO production between L. (L.) amazonensis (GMFC = 92.17 +/- 4.6) and L. (V.) braziliensis (GMFC = 18.89 +/- 2.29) (P < 0.05). Analysis of cNOS expression by immunoblotting showed more expression in L. (L.) amazonensis versus L. (V.) braziliensis. Subsequently, cNOS-like immunolabeling was observed in promastigotes in regions near vesicles, the flagellar pocket and mitochondria, and small clusters of particles appeared to be fusing with vesicles suggestive of glycosomes, peroxisome-like-organelles that compartmentalize the glycolytic pathway in trypanosomatid parasites. In addition, confocal microscopy analysis demonstrated colocalization of cNOS-like and GAPDH, a specific marker for glycosomes. Thus, L. (L.) amazonensis produces greater amounts of NO than L. (V.) braziliensis, and both species present the cNOS-like enzyme inside glycosomes.
Asunto(s)
Leishmania braziliensis/enzimología , Leishmania mexicana/enzimología , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico/biosíntesis , Proteínas Protozoarias/metabolismo , Leishmaniasis Cutánea/metabolismo , Leishmaniasis Mucocutánea/metabolismo , Especificidad de la EspecieRESUMEN
BACKGROUND: Toxoplasma gondii causes toxoplasmosis and is controlled by activated macrophages. However, infection of macrophages by tachyzoites induces TGF-ß signaling (TGF-s) inhibiting nitric oxide (NO) production. NO inhibition may be a general escape mechanism of distinct T. gondii strains. OBJECTIVES: To evaluate in activated macrophages the capacity of T. gondii strains of different virulence and genetics (RH, type I; ME-49, type II; VEG, type III; P-Br, recombinant) to evade the NO microbicidal defense system and determine LC3 loading to the parasitophorous vacuole. METHODS: Activated peritoneal macrophages were infected with the different T. gondii strains, NO-production was evaluated by the Griess reagent, and inducible nitric oxide synthase expression, TGF-s, and LC3 localisation assayed by immunofluorescence. FINDINGS: Only RH persisted in macrophages, while VEG was more resistant than P-Br and ME-49. All strains induced TGF-s, degradation of inducible nitric oxide synthase, and NO-production inhibition from 2 to 24 h of infection, but only RH sustained these alterations for 48 h. By 24 h of infection, TGF-s lowered in macrophages infected by ME-49, and P-Br, and NO-production recovered, while VEG sustained TGF-s and NO-production inhibition longer. LC3 loading to parasitophorous vacuole was strain-dependent: higher for ME-49, P-Br and VEG, lower for RH. All strains inhibited NO-production, but only RH sustained this effect probably because it persisted in macrophages due to additional evasive mechanisms as lower LC3 loading to parasitophorous vacuole. MAIN CONCLUSIONS: These results support that T. gondii can escape the NO microbicidal defense system at the initial phase of the infection, but only the virulent strain sustain this evasion mechanism.
Asunto(s)
Macrófagos Peritoneales/parasitología , Macrófagos/parasitología , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico/biosíntesis , Toxoplasma/fisiología , Animales , Macrófagos/metabolismo , Ratones , Toxoplasmosis Animal/parasitologíaRESUMEN
L-Arginine (L-ARG) supplementation has been suggested as a therapeutic option in several diseases, including Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke-like syndrome (MELAS), arguably the most common mitochondrial disease. It is suggested that L-ARG, a nitric oxide (NO) precursor, can restore NO levels in blood vessels, improving cerebral blood flow. However, NO also participates in mitochondrial processes, such as mitochondrial biogenesis, the regulation of the respiratory chain, and oxidative stress. This study investigated the effects of L-ARG on mitochondrial function, nitric oxide synthesis, and nitro-oxidative stress in cell lines harboring the MELAS mitochondrial DNA (mtDNA) mutation (m.3243A>G). We evaluated mitochondrial enzyme activity, mitochondrial mass, NO concentration, and nitro-oxidative stress. Our results showed that m.3243A>G cells had increased NO levels and protein nitration at basal conditions. Treatment with L-ARG did not affect the mitochondrial function and mass but reduced the intracellular NO concentration and nitrated proteins in m.3243A>G cells. The same treatment led to opposite effects in control cells. In conclusion, we showed that the main effect of L-ARG was on protein nitration. Lowering protein nitration is probably involved in the mechanism related to L-ARG supplementation benefits in MELAS patients.
Asunto(s)
Arginina/farmacología , ADN Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Enfermedades Mitocondriales/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Línea Celular , Humanos , Enfermedades Mitocondriales/genética , Mutación , Óxido Nítrico/biosíntesisRESUMEN
Several reports have suggested that photobiomodulation, owing to its analgesic, anti-inflammatory, and healing effects, may be an effective therapeutic option for local effects of snakebites when the availability and accessibility of conventional serum therapy are inefficient and far from medical care centers. Although there have been studies that demonstrate the application of photobiomodulation in the treatment of local adverse events due to snakebites from snakes of the genus Bothrops, its role in the activation of leukocytes, particularly macrophages, has not been evaluated. Here, we assessed the effect of light-emitting diode (LED) treatment on macrophage activation induced by B. jararacussu venom (BjV). LED treatment caused an increase in the viability of macrophages incubated with BjV. This treatment reduced reactive oxygen species (ROS) and nitric oxide (NO) production by macrophages after incubation with BjV. However, LED treatment did not interfere with IL-1ß and IL-10 production by macrophages after incubation with BjV. In conclusion, this study showed that LED treatment has the potential to be used in combination with conventional serum therapy to prevent or minimize the progression of local to severe symptoms after Bothrops envenomation.
Asunto(s)
Bothrops , Venenos de Crotálidos/toxicidad , Terapia por Luz de Baja Intensidad/instrumentación , Macrófagos/efectos de la radiación , Semiconductores , Mordeduras de Serpientes/inmunología , Mordeduras de Serpientes/radioterapia , Animales , Supervivencia Celular/efectos de los fármacos , Interleucina-10/metabolismo , Interleucina-1beta/metabolismo , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Espacio Intracelular/efectos de la radiación , Macrófagos/inmunología , Masculino , Ratones , Óxido Nítrico/biosíntesis , Especies Reactivas de Oxígeno/metabolismo , Mordeduras de Serpientes/metabolismo , Mordeduras de Serpientes/patología , Superóxidos/metabolismoRESUMEN
BACKGROUND Toxoplasma gondii causes toxoplasmosis and is controlled by activated macrophages. However, infection of macrophages by tachyzoites induces TGF-β signaling (TGF-s) inhibiting nitric oxide (NO) production. NO inhibition may be a general escape mechanism of distinct T. gondii strains. OBJECTIVES To evaluate in activated macrophages the capacity of T. gondii strains of different virulence and genetics (RH, type I; ME-49, type II; VEG, type III; P-Br, recombinant) to evade the NO microbicidal defense system and determine LC3 loading to the parasitophorous vacuole. METHODS Activated peritoneal macrophages were infected with the different T. gondii strains, NO-production was evaluated by the Griess reagent, and inducible nitric oxide synthase expression, TGF-s, and LC3 localisation assayed by immunofluorescence. FINDINGS Only RH persisted in macrophages, while VEG was more resistant than P-Br and ME-49. All strains induced TGF-s, degradation of inducible nitric oxide synthase, and NO-production inhibition from 2 to 24 h of infection, but only RH sustained these alterations for 48 h. By 24 h of infection, TGF-s lowered in macrophages infected by ME-49, and P-Br, and NO-production recovered, while VEG sustained TGF-s and NO-production inhibition longer. LC3 loading to parasitophorous vacuole was strain-dependent: higher for ME-49, P-Br and VEG, lower for RH. All strains inhibited NO-production, but only RH sustained this effect probably because it persisted in macrophages due to additional evasive mechanisms as lower LC3 loading to parasitophorous vacuole. MAIN CONCLUSIONS These results support that T. gondii can escape the NO microbicidal defense system at the initial phase of the infection, but only the virulent strain sustain this evasion mechanism.
Asunto(s)
Animales , Ratones , Toxoplasma/fisiología , Macrófagos Peritoneales/parasitología , Óxido Nítrico Sintasa/metabolismo , Macrófagos/parasitología , Óxido Nítrico/biosíntesis , Toxoplasmosis Animal/parasitología , Macrófagos/metabolismoRESUMEN
Histoplasma capsulatum is the agent of histoplasmosis, one of the most frequent mycoses in the world. The infection initiates with fungal spore inhalation, transformation into yeasts in the lungs and establishment of a granulomatous disease, which is characterized by a Th1 response. The production of Th1 signature cytokines, such as IFN-γ, is crucial for yeast clearance from the lungs, and to prevent dissemination. Recently, it was demonstrated that IL-17, a Th17 signature cytokine, is also important for fungal control, particularly in the absence of Th1 response. IL-22 is another cytokine with multiple functions on host response and disease progression. However, little is known about the role of IL-22 during histoplasmosis. In this study, we demonstrated that absence of IL-22 affected the clearance of yeasts from the lungs and increased the spreading to the spleen. In addition, IL-22 deficient mice (Il22-/-) succumbed to infection, which correlated with reductions in the numbers of CD4+ IFN-γ+ T cells, reduced IFN-γ levels, and diminished nitric oxide synthase type 2 (NOS2) expression in the lungs. Importantly, treatment with rIFN-γ mitigated the susceptibility of Il22-/- mice to H. capsulatum infection. These data indicate that IL-22 is crucial for IFN-γ/NO production and resistance to experimental histoplasmosis.
Asunto(s)
Histoplasmosis/inmunología , Interferón gamma/inmunología , Interleucinas/inmunología , Animales , Femenino , Histoplasmosis/patología , Interferón gamma/biosíntesis , Interleucinas/deficiencia , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico/biosíntesis , Óxido Nítrico/inmunología , Interleucina-22RESUMEN
The NMDA receptor is crucial to several functions in CNS physiology and some of its effects are mediated by promoting nitric oxide production from L-arginine and activation of signaling pathways and the transcription factor CREB. Our previous work demonstrated in retinal cells that increasing intracellular free L-arginine levels directly correlates to nitric oxide (NO) generation and can be promoted by protein synthesis inhibition and increase of free L-arginine concentration. Eukaryotic elongation factor 2 kinase (eEF2K), a calcium/calmodulin-dependent kinase, is also known to be activated by NMDA receptors leading to protein synthesis inhibition. Here we explored how does eEF2K participate in NMDA-induced NO signaling. We found that when this enzyme is inhibited, NMDA loses its ability to promote NO synthesis. On the other hand, when NO synthesis is increased by protein synthesis inhibition with cycloheximide or addition of exogenous L-arginine, eEF2K has no participation, showcasing a specific link between this enzyme and NMDA-induced NO signaling. We have previously shown that inhibition of the canonical NO signaling pathway (guanylyl cyclase/cGMP/cGK) blocks CREB activation by glutamate in retinal cells. Interestingly, pharmacological inhibition of eEF2K fully prevents CREB activation by NMDA, once again demonstrating the importance of eEF2K in NMDA receptor signaling. In summary, we demonstrated here a new role for eEF2K, directly controlling NMDA-dependent nitrergic signaling and modulating L-arginine availability in neurons, which can potentially be a new target for the study of physiological and pathological processes involving NMDA receptors in the central nervous system.
Asunto(s)
Sistema Nervioso Central/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Quinasa del Factor 2 de Elongación/metabolismo , N-Metilaspartato/farmacología , Óxido Nítrico/biosíntesis , Animales , Arginina/farmacología , Pollos , Cicloheximida/farmacología , Quinasa del Factor 2 de Elongación/antagonistas & inhibidores , Indazoles/farmacología , Masculino , Fosforilación/efectos de los fármacos , Piridinas/farmacología , Pirimidinas/farmacología , RatasRESUMEN
BACKGROUND: The mechanism of resistance to SbIII in Leishmania is complex, multifactorial and involves not only biochemical mechanisms, but also other elements, such as the immune system of the host. OBJECTIVES: In this study, putative changes in the immunological profile of human monocytes infected with wild-type (WT) and antimony (SbIII)-resistant Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum lines were evaluated. METHODS: Susceptibility assays WT and SbIII-resistant L. braziliensis and L. infantum were performed using lines THP-1 human monocytic lineage. Phagocytic capacity, cytokine profile, intracellular nitric oxide (NO) production and surface carbohydrate residues profile were performed in peripheral blood monocytes by flow cytometry. FINDINGS: The phagocytic capacity and intracellular NO production by classical (CD14++CD16-) and proinflammatory (CD14++CD16+) monocytes were higher in the presence of L. infantum lines compared to L. braziliensis lines. The results also highlight proinflammatory monocytes as the cellular subpopulation of major relevance in a phagocytosis event and NO expression. It is important to note that L. infantum induced a proinflammatory cytokine profile characterised by higher levels of TNF-α in culture supernatant than L. braziliensis. Conversely, both Leishmania lines induce high levels of IL-6 in culture supernatant. Analysis of the expression profile of surface carbohydrates showed that L. braziliensis presents 4.3-fold higher expression of galactose(ß1,4)N-acetylglucosamine than L. infantum line. Interestingly, the expression level of α-N-acetylgalactosamine residues was 2-fold lower in the SbIII-resistant L. braziliensis line than its counterpart WT line, indicating differences in surface glycoconjugates between these lines. MAIN CONCLUSIONS: Our results showed that L. braziliensis and L. infantum induce different innate immune responses and a highly inflammatory profile, which is characteristic of infection by L. infantum, the species associated with visceral disease.
Asunto(s)
Antimonio/farmacología , Antiprotozoarios/farmacología , Leishmania braziliensis/inmunología , Leishmania infantum/inmunología , Monocitos/parasitología , Óxido Nítrico/biosíntesis , Fagocitosis/inmunología , Adulto , Resistencia a Medicamentos , Femenino , Citometría de Flujo , Humanos , Inmunidad Innata , Leishmania braziliensis/efectos de los fármacos , Leishmania infantum/efectos de los fármacos , Masculino , Monocitos/inmunología , Adulto JovenRESUMEN
In gestational diabetes mellitus (GDM) pregnancies, a compromised fetal liver may impact offspring's metabolic health. Here, we aimed to address prooxidant, proinflammatory and profibrotic markers in the livers from GDM rats and their fetuses, and to analyze the expression of miR-122 (a relevant microRNA in liver pathophysiology) in fetal and maternal plasma of GDM rats, as well as in the fetal livers of neonatal streptozotocin-induced (nSTZ) diabetic rats, the rats that generate GDM through intrauterine programming. GDM and nSTZ rats were evaluated on day 21 of pregnancy. We found increased nitric oxide production and lipoperoxidation in the livers from GDM rats and their fetuses compared to controls. Livers from GDM fetuses also showed increased levels of connective tissue growth factor and matrix metalloproteinase-2. The expression of miRNA-122 was downregulated in the plasma from GDM rats and their male fetuses, as well as in the livers from male fetuses of nSTZ diabetic rats. miR-122 levels were regulated both in vitro through PPARγ activation and in vivo through a maternal diet enriched in PPAR ligands. Our findings revealed a prooxidant/proinflammatory environment in the livers from GDM rats and their fetuses and a dysregulation of miR-122, likely relevant in the programming of offspring's diseases.
Asunto(s)
Diabetes Mellitus Experimental/genética , Diabetes Gestacional/genética , Feto/embriología , Regulación de la Expresión Génica , Inflamación/genética , Hígado/embriología , MicroARNs/sangre , Útero/patología , Animales , Biomarcadores/metabolismo , MicroARN Circulante/sangre , MicroARN Circulante/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/metabolismo , Diabetes Gestacional/sangre , Diabetes Gestacional/metabolismo , Modelos Animales de Enfermedad , Femenino , Inflamación/sangre , Peroxidación de Lípido , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , MicroARNs/genética , Óxido Nítrico/biosíntesis , Aceite de Oliva , Oxidantes/metabolismo , Embarazo , Ratas Wistar , EstreptozocinaRESUMEN
Nitric oxide (NO) is related to a wide range of physiological processes such as vasodilation, macrophages cytotoxicity and wound healing. The human skin contains NO precursors (NOx). Those are mainly composed of nitrite (NO2-), nitrate (NO3-), and S-nitrosothiols (RSNOs) which forms a large NO store. These NOx stores in human skin can mobilize NO to blood stream upon ultraviolet (UV) light exposure. The main purpose of this study was to evaluate the most effective UV light wavelength to generate NO and compare it to each NO precursor in aqueous solution. In addition, the UV light might change the RSNO content on human skin. First, we irradiated pure aqueous solutions of NO2- and NO3- and mixtures of NO2- and glutathione and NO3- and S-nitrosoglutathione (GSNO) to identify the NO release profile from those species alone. In sequence, we evaluated the NO generation profile on human skin slices. Human skin was acquired from redundant plastic surgical samples and the NO and RSNO measurements were performed using a selective NO electrochemical sensor. The data showed that UV light could trigger the NO generation in skin with a peak at 280-285 nm (UVB range). We also observed a significant RSNO formation in irradiated human skin, with a peak at 320 nm (UV region) and at 700 nm (visible region). Pre-treatment of the human skin slice using NO2- and thiol (RSHs) scavengers confirmed the important role of these molecules in RSNO formation. These findings have important implications for clinical trials with potential for new therapies.
Asunto(s)
Óxido Nítrico/biosíntesis , S-Nitrosotioles/metabolismo , Piel/metabolismo , Piel/efectos de la radiación , Rayos Ultravioleta , Humanos , Procesos FotoquímicosRESUMEN
Being the base of several non-communicable diseases, including cancer, inflammation is a complex process generated by tissue damage or change in the body homeostatic state. Currently, the therapeutic treatment for chronic inflammation related diseases is based on the use of selective cyclooxygenase II enzyme, COX-2, inhibitors or Coxibs, which have recently regained attention giving their preventive role in colon cancer. Thus, the discovery of new molecules that selectively inhibit COX-2 and other inflammatory mediators is a current challenge in the medicinal chemistry field. 1-Phenylbenzimidazoles have shown potential COX inhibitory activity, because they can reproduce the interaction profile of known COX inhibitors. Therefore, in the present investigation a series of 1,2-diphenylbenzimidazoles (DPBI) with different aromatic substitutions in the para position were synthesized and their interaction with COX-2 and nitric oxide synthase, iNOS, was determined in silico, in vitro and in vivo. Compound 2-(4-bromophenyl)-1-(4-nitrophenyl)-1H-benzo[d]imidazole showed the best inhibition towards COX-2, while compounds N-(4-(2-(4-bromophenyl)-1H-benzo[d]imidazol-1-yl)phenyl)acetamide and N-(4-(2-(4-chlorophenyl)-1H-benzo[d]imidazol-1-yl)phenyl)acetamide diminished the production of NO in vitro. Additionally, they had a significant anti-inflammatory activity in vivo when given orally.
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Antiinflamatorios no Esteroideos/farmacología , Bencimidazoles/farmacología , Inhibidores Enzimáticos/farmacología , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Óxido Nítrico/antagonistas & inhibidores , Animales , Antiinflamatorios no Esteroideos/síntesis química , Antiinflamatorios no Esteroideos/química , Bencimidazoles/síntesis química , Bencimidazoles/química , Bovinos , Ciclooxigenasa 1/metabolismo , Ciclooxigenasa 2/metabolismo , Relación Dosis-Respuesta a Droga , Edema/tratamiento farmacológico , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Humanos , Inflamación/tratamiento farmacológico , Masculino , Estructura Molecular , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo II/metabolismo , Ratas , Ratas Wistar , Relación Estructura-ActividadRESUMEN
Endothelial nitric oxide synthase (eNOS) malfunctioning has been proposed to contribute to the endothelial damage produced by cigarette. Besides eNOS, neuronal NOS (nNOS) is also expressed in most vascular tissues and plays an important role in the endothelium-dependent vascular relaxation. We hypothesize that nNOS may contribute to the endothelium dysfunction produced by cigarette in smokers. Vascular function was assessed in human resistance mesenteric arteries using a wire myograph, the level of protein expression by Western blot, eNOS and nNOS localization by immunofluorescence. Measurement of NO was assessed by fluorescence microscopy. Arteries of smokers showed impaired endothelium-dependent vascular relaxation in response to acetylcholine. Pharmacological nonselective blockade of NOS with l-NAME and selective nNOS blockade with inhibitor 1 reduced the relaxation of the mesenteric artery of both smokers and nonsmokers. Interestingly, the inhibitory effect of NOS inhibitors was greater in nonsmokers than in smokers. The expression of total nNOS and eNOS and the level of phosphorylation at eNOS-pSer1177 were reduced in arteries of smokers as compared with nonsmokers. No differences between groups were observed in the expression of total COX-1, COX-2, catalase and SOD-1. Immunofluorescence analysis showed the presence of nNOS in the vascular endothelium in both groups. Acetylcholine-induced NO production was impaired in arteries from smokers as compared to nonsmokers. Selective inhibition of nNOS caused a decreased in NO production, which was greater in nonsmokers than in smokers. Our data show that a decrease in nNOS expression contributes to the endothelial dysfunction caused by cigarette smoking in human.
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Fumar Cigarrillos/efectos adversos , Endotelio Vascular/metabolismo , Óxido Nítrico Sintasa de Tipo I/biosíntesis , Adulto , Anciano , Inhibidores Enzimáticos/farmacología , Femenino , Humanos , Masculino , Persona de Mediana Edad , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico/análisis , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo I/antagonistas & inhibidoresRESUMEN
Moderate levels of a proinflammatory macrophages phenotype are indispensable and play an important role in the skeletal muscle repair process since this response depends on their secreted products concentration to influence and modulate muscle inflammation as well as the differentiation of myoblasts. This study investigated the effects of photobiomodulation (PBM) on undifferentiated and differentiation-induced C2C12 myoblasts cultivated in different concentrations of M1 phenotype macrophage-conditioned media of J774 cells (MCM1) also submitted to PBM using the same irradiation parameters. Irradiation was performed once with low-level laser (780 nm, 70 mW, 1 J) and was evaluated cell viability, proliferation and differentiation, nitric oxide (NO) synthesis and IL-6 and TNF-α protein levels 24 and 48 h after C2C12 irradiation. PBM treatment in undifferentiated myoblasts exhibited lower IL-6 levels in the presence of nonirradiated MCM1 at both concentrations. Myoblasts in proliferation condition cultivated with irradiated MCM1 showed lower IL-6 and TNF-α levels after 48 h in the presence of both concentrations evaluated. PBM induced a decrease in the synthesis of NO on undifferentiated and differentiation-induced myoblasts. PBM was able to reduce the level of proinflammatory protein and markers, which are important to allow the differentiation of myoblasts during the muscle repair process.
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Luz , Macrófagos/efectos de la radiación , Mioblastos/efectos de la radiación , Animales , Línea Celular , Proliferación Celular/efectos de la radiación , Medios de Cultivo Condicionados , Activación de Macrófagos/efectos de la radiación , Macrófagos/citología , Macrófagos/metabolismo , Ratones , Óxido Nítrico/biosíntesisRESUMEN
Taurine is a semiessential amino acid found at high concentrations in mammalian plasma and cells, where it regulates cellular functions such as ion flux, controls cell volume and serves as a substrate for conjugated bile acids (BAs). Exogenous administration of both taurine and taurine-conjugated BAs have also been implicated in the modulation of cardiovascular functions. This brief review summarizes the role of taurine and taurine-conjugated BAs in vascular relaxation through the modulation of endothelium-derived nitric oxide (NO). The effects of taurine on vascular health are controversial. However, in the presence of cardiometabolic risk factors, it has been proposed that taurine can increase vascular NO levels by increasing eNOS expression, eNOS phosphorylation on Ser1177, NO bioavailability, the level of antioxidative defense, and the l-arginine/NOS inhibitor asymmetric dimethylarginine (ADMA) ratio. The taurine-conjugated BA-mediated activation of Farnesoid X receptor (FXR), G protein-coupled BA receptor (TGR5) and/or muscarinic 3 receptor (M3) was also reported to increase vascular NO production. FXR activation increases eNOS expression and may reduce ADMA formation, while TGR5 increases mobilization of Ca2+ and phosphorylation of eNOS and Akt in endothelial cells. Furthermore, taurine and taurine-conjugated BAs might regulate NO synthesis and activity by enhancing H2S generation. Several studies have demonstrated the beneficial effects of both taurine and taurine-conjugated BAs in reversing the endothelial dysfunction associated with diabetes, atherosclerosis, hypertension, obesity, malnutrition, and smoking. In addition, taurine-conjugated BAs have emerged as a potential treatment for portal hypertension. Despite these favorable findings, there is a need to further explore the mechanisms and signaling pathways underlying the endothelial effects of taurine and taurine-conjugated BAs. Here, we summarize the main findings regarding the effects of taurine and taurine-conjugated BAs on the endothelial dysfunction associated with altered NO metabolism in cardiovascular diseases.
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Ácidos y Sales Biliares/metabolismo , Células Endoteliales/metabolismo , Óxido Nítrico/biosíntesis , Taurina/metabolismo , Animales , HumanosRESUMEN
Mercury is a metal widely dispersed in nature that when in contact with human organism, it damages the cardiovascular system. Long-term mercury exposure for 30 days induces endothelial dysfunction without blood pressure changes in normotensive adult rats. However, it is not known whether exposure to mercury can exacerbate endothelial dysfunction and hypertension development in predisposed animals. Thus, we aimed to investigate the effects of long-term mercury exposure on the blood pressure (BP) and in the isolated aortas of young normotensive and prehypertensive spontaneously hypertensive rats (SHRs). Four-week-old male Wistar rats and SHRs were treated daily with mercury chloride (HgCl2) (1st dose, 4.6 µg/kg; subsequent dose, 0.07 µg/kg/day, im, 30 days) or vehicle. BP was assessed weekly and the vascular reactivity to phenylephrine was evaluated in isolated aorta from rats exposed or not to mercury. Mercury exposure did not affect BP in young Wistar rats but accelerated the development of hypertension in young SHRs. Vascular reactivity to phenylephrine increased only in the aorta from mercury-exposed SHRs. While HgCl2 exposure in SHRs did not alter nitric oxide production, we observed increased superoxide anion production and decreased superoxide dismutase-1 protein expression, and enhanced cyclooxygenase-2 (COX-2) participation with increased prostaglandin (PGE2) production and decreased prostacyclin. In the Wistar group, mercury exposure did not alter superoxide anion production or the COX-2 pathway. Mercury exposure accelerated the natural course of hypertension in young SHRs and increased oxidative stress associated with reduced participation of antioxidant enzymes, an activated COX-2 pathway, thereby producing endothelial dysfunction, which is a risk factor in prehypertensive individuals.
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Ciclooxigenasa 2/metabolismo , Endotelio Vascular/efectos de los fármacos , Hipertensión/inducido químicamente , Cloruro de Mercurio/toxicidad , Animales , Presión Sanguínea/efectos de los fármacos , Endotelio Vascular/metabolismo , Hipertensión/metabolismo , Masculino , Cloruro de Mercurio/administración & dosificación , Óxido Nítrico/biosíntesis , Estrés Oxidativo/efectos de los fármacos , Fenilefrina/metabolismo , Ratas , Ratas Endogámicas SHR , Ratas WistarRESUMEN
BACKGROUND The mechanism of resistance to SbIII in Leishmania is complex, multifactorial and involves not only biochemical mechanisms, but also other elements, such as the immune system of the host. OBJECTIVES In this study, putative changes in the immunological profile of human monocytes infected with wild-type (WT) and antimony (SbIII)-resistant Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum lines were evaluated. METHODS Susceptibility assays WT and SbIII-resistant L. braziliensis and L. infantum were performed using lines THP-1 human monocytic lineage. Phagocytic capacity, cytokine profile, intracellular nitric oxide (NO) production and surface carbohydrate residues profile were performed in peripheral blood monocytes by flow cytometry. FINDINGS The phagocytic capacity and intracellular NO production by classical (CD14++CD16-) and proinflammatory (CD14++CD16+) monocytes were higher in the presence of L. infantum lines compared to L. braziliensis lines. The results also highlight proinflammatory monocytes as the cellular subpopulation of major relevance in a phagocytosis event and NO expression. It is important to note that L. infantum induced a proinflammatory cytokine profile characterised by higher levels of TNF-α in culture supernatant than L. braziliensis. Conversely, both Leishmania lines induce high levels of IL-6 in culture supernatant. Analysis of the expression profile of surface carbohydrates showed that L. braziliensis presents 4.3-fold higher expression of galactose(β1,4)N-acetylglucosamine than L. infantum line. Interestingly, the expression level of α-N-acetylgalactosamine residues was 2-fold lower in the SbIII-resistant L. braziliensis line than its counterpart WT line, indicating differences in surface glycoconjugates between these lines. MAIN CONCLUSIONS Our results showed that L. braziliensis and L. infantum induce different innate immune responses and a highly inflammatory profile, which is characteristic of infection by L. infantum, the species associated with visceral disease.