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1.
Physiol Rep ; 10(11): e15342, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35674115

RESUMO

The hallmark of pulmonary hypertension (PH) is vascular remodeling. We have previously shown that human pulmonary microvascular endothelial cells (hPMVEC) respond to hypoxia with epidermal growth factor (EGF) mediated activation of the receptor tyrosine kinase, EGF receptor (EGFR), resulting in arginase-2 (Arg2)-dependent proliferation. We hypothesized that the release of EGF by hPMVEC could result in the proliferation of human pulmonary arterial smooth muscle cells (hPASMC) via activation of EGFR on the hPASMC leading to Arg2 up-regulation. To test this hypothesis, we used conditioned media (CM) from hPMVEC grown either in normoxia (NCM) or hypoxia (HCM). Human PASMC were incubated in normoxia with either HCM or NCM, and HCM caused significant induction of Arg2 and viable cell numbers. When HCM was generated with either an EGF-neutralizing antibody or an EGFR blocking antibody the resulting HCM did not induce Arg2 or increase viable cell numbers in hPASMC. Adding an EGFR blocking antibody to HCM, prevented the HCM-induced increase in Arg2 and viable cell numbers. HCM induced robust phosphorylation of hPASMC EGFR. When hPASMC were transfected with siRNA against EGFR the HCM-induced increase in viable cell numbers was prevented. When hPASMC were treated with the arginase antagonist nor-NOHA, the HCM-induced increase in viable cell numbers was prevented. These data suggest that hypoxic hPMVEC releases EGF, which activates hPASMC EGFR leading to Arg2 protein expression and an increase in viable cell numbers. We speculate that EGF neutralizing antibodies or EGFR blocking antibodies represent potential therapeutics to prevent and/or attenuate vascular remodeling in PH associated with hypoxia.


Assuntos
Fator de Crescimento Epidérmico , Hipertensão Pulmonar , Arginase/metabolismo , Hipóxia Celular/fisiologia , Proliferação de Células , Células Endoteliais/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Humanos , Hipertensão Pulmonar/metabolismo , Hipóxia/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Artéria Pulmonar/metabolismo , Remodelação Vascular
2.
Nanomedicine ; 43: 102565, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35595014

RESUMO

It was hypothesized that the catalyst nanoceria can increase inflammation/oxidative stress from the basal and reduce it from the elevated state. Macrophages clear nanoceria. To test the hypothesis, M0 (non-polarized), M1- (classically activated, pro-inflammatory), and M2-like (alternatively activated, regulatory phenotype) RAW 264.7 macrophages were nanoceria exposed. Inflammatory responses were quantified by IL-1ß level, arginase activity, and RT-qPCR and metabolic changes and oxidative stress by the mito and glycolysis stress tests (MST and GST). Morphology was determined by light microscopy, macrophage phenotype marker expression, and a novel three-dimensional immunohistochemical method. Nanoceria blocked IL-1ß and arginase effects, increased M0 cell OCR and GST toward the M2 phenotype and altered multiple M1- and M2-like cell endpoints toward the M0 level. M1-like cells had greater volume and less circularity/roundness. M2-like cells had greater volume than M0 macrophages. The results are overall consistent with the hypothesis.


Assuntos
Arginase , Nanoestruturas , Arginase/metabolismo , Cério , Humanos , Inflamação , Estresse Oxidativo
3.
J Am Soc Nephrol ; 33(6): 1077-1086, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35577558

RESUMO

BACKGROUND: After kidney injury, macrophages transition from initial proinflammatory activation to a proreparative phenotype characterized by expression of arginase-1 (Arg1), mannose receptor 1 (Mrc1), and macrophage scavenger receptor 1 (Msr1). The mechanism by which these alternatively activated macrophages promote repair is unknown. METHODS: We characterized the macrophage and renal responses after ischemia-reperfusion injury with contralateral nephrectomy in LysM-Cre;Arg1fl/fl mice and littermate controls and used in vitro coculture of macrophages and tubular cells to determine how macrophage-expressed arginase-1 promotes kidney repair. RESULTS: After ischemia-reperfusion injury with contralateral nephrectomy, Arg1-expressing macrophages were almost exclusively located in the outer stripe of the medulla adjacent to injured S3 tubule segments containing luminal debris or casts. Macrophage Arg1 expression was reduced by more than 90% in injured LysM-Cre;Arg1fl/fl mice, resulting in decreased mouse survival, decreased renal tubular cell proliferation and decreased renal repair compared with littermate controls. In vitro studies demonstrate that tubular cells exposed apically to dead cell debris secrete high levels of GM-CSF and induce reparative macrophage activation, with those macrophages in turn secreting Arg1-dependent factor(s) that directly stimulate tubular cell proliferation. CONCLUSIONS: GM-CSF-induced, proreparative macrophages express arginase-1, which is required for the S3 tubular cell proliferative response that promotes renal repair after ischemia-reperfusion injury.


Assuntos
Fator Estimulador de Colônias de Granulócitos e Macrófagos , Traumatismo por Reperfusão , Animais , Arginase/genética , Arginase/metabolismo , Macrófagos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Regeneração , Traumatismo por Reperfusão/metabolismo
4.
Mol Biol Rep ; 49(5): 4155-4160, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35505270

RESUMO

BACKGROUND: Arginase enzyme is essential for the catalysis of the last step of the urea cycle, resulting in the conversion of L-arginine to L-ornithine and urea. Arginase deficiency could lead to hyperarginemia, an autosomal recessive disorder of the urea cycle that could result in developmental manifestations after the first year of life, followed by gradually progressive atonic cerebral palsy, spastic quadriplegia, and mental decline. ARG1 mutations have been reported in hyperarginemia patients of Western countries because they exhibited reduced arginase activity. Hence, it is important to assess ARG1 mutations in cerebral palsy cases with hyperarginemia in different populations. METHODS AND RESULTS: This study involved two unrelated pediatric patients from two non-consanguineous East Indian families, exhibiting a range of manifestations, including hypotonia of all limbs, mental retardation, and multiple episodes of seizure. The onset of the disease ranged from 1 to 3 years of age. Hyperammonemia (> 250 micromoles) and serum hyperarginemia (> 350 micromoles) were observed in both the patients. Whole-genome sequencing, followed by Sanger sequencing of both the patients confirmed the presence of a homozygous 3' splice site variation in intron 3 of the ARG1 gene (chr6: g.131902357A>T) that affects the invariant AG acceptor splice site of exon 4 (c.330-2A>T; ENST00000356962.2). CONCLUSION: The study reported the identification of a novel ARG1 mutation in two different unrelated pediatric cases from Odisha, India associated with hyperarginemia. The pathogenicity of the mutation was robustly supported by the clinical phenotype, complete co-segregation with the disease, and biochemical observations.


Assuntos
Arginase , Paralisia Cerebral , Arginase/genética , Arginase/metabolismo , Paralisia Cerebral/enzimologia , Paralisia Cerebral/genética , Criança , Humanos , Íntrons , Mutação , Ureia/metabolismo
5.
Pharmacol Res ; 179: 106229, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35470065

RESUMO

Acute liver injury (ALI) is characterized by massive hepatocyte necrosis and subsequent recruitment of myeloid cells to liver. Mesenchymal stem cells (MSCs) have therapeutic potential for ALI through their immunoregulation on macrophages, but the mechanism is not completely clear due to the heterogeneity and controversy of liver macrophages. Here, we detected the survival rate, biochemical indexes, histopathology, and inflammatory chemokine levels to assess the efficacy of MSC treatment on CCl4-induced ALI of C57BL/6 mice. Furthermore, flow cytometry and single-cell RNA sequencing (scRNA-Seq) were used to precisely distinguish macrophage populations and reveal the immunoregulation of MSCs. MSC treatment could effectively alleviate ALI and mitigate the recruitment of mononuclear phagocytes. Flow cytometry and scRNA-Seq analyses collectively indicated that there were monocytes with high Ly6C expression and heterogeneous monocyte-derived macrophages (MoMF) with low Ly6C expression in liver. Ly6Chi pro-inflammatory monocytes and Ly6Clo MoMF with powerful phagocytosis dominated during the acute injury period. MSC treatment promoted the transition from Ly6Chi to Ly6Clo population, inhibit the proinflammatory function of monocytes and promote the lysosomal function of MoMF. Furthermore, MSCs attenuated the recruitment of neutrophils by reducing the expression of CXCL2 of MoMF. MoMF with high expression of arginase 1 appeared during the recovery period, and MSCs could increase their expression of arginase 1, which may promote liver repair. To sum up, we demonstrated the characteristics of distinct MoMF during different periods of ALI and revealed their functional changes after MSC treatment, providing immunotherapeutic targets for MSC treatment of ALI.


Assuntos
Células-Tronco Mesenquimais , Análise de Célula Única , Animais , Arginase/metabolismo , Arginase/farmacologia , Homeostase , Fígado , Macrófagos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL
6.
Nature ; 603(7901): 515-521, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35264792

RESUMO

Nitrogen availability is a growth-limiting factor in many habitats1, and the global nitrogen cycle involves prokaryotes and eukaryotes competing for this precious resource. Only some bacteria and archaea can fix elementary nitrogen; all other organisms depend on the assimilation of mineral or organic nitrogen. The nitrogen-rich compound guanidine occurs widely in nature2-4, but its utilization is impeded by pronounced resonance stabilization5, and enzymes catalysing hydrolysis of free guanidine have not been identified. Here we describe the arginase family protein GdmH (Sll1077) from Synechocystis sp. PCC 6803 as a Ni2+-dependent guanidine hydrolase. GdmH is highly specific for free guanidine. Its activity depends on two accessory proteins that load Ni2+ instead of the typical Mn2+ ions into the active site. Crystal structures of GdmH show coordination of the dinuclear metal cluster in a geometry typical for arginase family enzymes and allow modelling of the bound substrate. A unique amino-terminal extension and a tryptophan residue narrow the substrate-binding pocket and identify homologous proteins in further cyanobacteria, several other bacterial taxa and heterokont algae as probable guanidine hydrolases. This broad distribution suggests notable ecological relevance of guanidine hydrolysis in aquatic habitats.


Assuntos
Hidrolases , Synechocystis , Arginase/metabolismo , Proteínas de Bactérias/metabolismo , Guanidina/metabolismo , Hidrolases/metabolismo , Nitrogênio/metabolismo
7.
Biomed Pharmacother ; 149: 112840, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35316752

RESUMO

Arginase is a key hydrolase in the urea cycle that hydrolyses L-arginine to urea and L-ornithine. Increasing number of studies in recent years demonstrate that two mammalian arginase isoforms, arginase 1 (ARG1) and arginase 2 (ARG2), were aberrantly upregulated in various types of cancers, and played crucial roles in the regulation of tumor growth and metastasis through various mechanisms such as regulating L-arginine metabolism, influencing tumor immune microenvironment, etc. Thus, arginase receives increasing focus as an attractive target for cancer therapy. In this review, we provide a comprehensive overview of the physiological and biological roles of arginase in a variety of cancers, and shed light on the underlying mechanisms of arginase mediating cancer cells growth and development, as well as summarize the recent clinical research advances of targeting arginase for cancer therapy.


Assuntos
Arginase , Neoplasias , Animais , Arginase/metabolismo , Arginina/metabolismo , Mamíferos , Neoplasias/tratamento farmacológico , Microambiente Tumoral , Ureia
8.
PLoS One ; 17(2): e0263993, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35167596

RESUMO

BACKGROUND: Leishmania (L) parasite, the causative agent of zoonotic cutaneous leishmaniasis (ZCL), effectively stimulates the mammalian cells to mount strong humoral responses by enhancing T-helper-2 (Th2)-associated cytokines for its survival. The best strategy to decrease the intensity of infection in the host is induction of cellular immunity. METHODS: We evaluated the effects of the empty bacterial pcDNA3 plasmid on mice infected with L. major and quantified the immune mediators including IFN-γ, IL-4, IL-10, IgG2a, IgG1, arginase activity and nitric oxide (NO) in the mice. Moreover, the footpad lesion size and parasite load were assessed. RESULTS: We observed that pcDNA3 could modulate the immune responses in favor of host cells and decrease the disease severity. Th2- associated mediators, including arginase, IL-4, and IL-10 are downregulated, while cellular responses are upregulated in line with an increase in the levels of nitric oxide (NO) and interfero-gamma (IFN-γ). Interestingly, pcDNA3 induced specific Th1-associated antibodies, IgG2a isotype; however, it suppressed the production of humoral IgG1. The stimulation of the immune response by the empty pcDNA3 is able to shift the immune function to predominant cellular responses caused by Th1, and it had a positive effect on the treatment of zoonotic cutaneous leishmaniasis (ZCL). CONCLUSIONS: Altogether, we introduced the pcDNA3 as a potential interfering factor in the modulation of the immune system against ZCL. Since this vector has been widely used as a control group in different studies, we suggest that the potential function of the empty vector should be deeply assessed, as it exerts anti-parasitic effects on mice infected with L. major.


Assuntos
Leishmania major/imunologia , Leishmaniose Cutânea/prevenção & controle , Plasmídeos/imunologia , Células Th2/imunologia , Animais , Arginase/metabolismo , Feminino , Imunoglobulina G/metabolismo , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-4/metabolismo , Leishmania major/patogenicidade , Leishmaniose Cutânea/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Óxido Nítrico/metabolismo , Plasmídeos/genética
9.
Toxicol In Vitro ; 80: 105326, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35134483

RESUMO

Mogroside V is the main bioactive component of Siraitia grosvenorii (Swingle), and has a potential anti-inflammatory function. However, the effect of mogroside V on fine particulate matter (PM2.5)-induced inflammation has not been reported. In the present study, the biological effect of mogroside V on inflammation was investigated in PM2.5- treated porcine alveolar macrophages (3D4/21). The results showed that mogroside V significantly inhibited PM2.5-induced nitric oxide (NO) production and rescued the arginase activity inhibited by PM2.5. In the presence of mogroside V, the upregulation of IL-18, TNF-α and COX-2 by PM2.5 in 3D4/21 cells was inhibited. Mogroside V attenuated PM2.5-induced phosphorylation of NF-κB p65 and the expression of NLRP3. Mogroside V reduced intracellular ROS levels induced by PM2.5. In the transcriptomic analysis, inflammation-related genes in 3D4/21 cells were not significantly affected after treatment with mogroside V. These results indicated that mogroside V can alleviate the inflammatory response of porcine alveolar macrophages induced by PM2.5 from pig house and that mogroside V may play the role through the antioxidant function of eliminating ROS. Mogroside V has a clear anti-inflammatory function in the presence of inflammation.


Assuntos
Anti-Inflamatórios/farmacologia , Macrófagos Alveolares/efeitos dos fármacos , Material Particulado/toxicidade , Triterpenos/farmacologia , Animais , Arginase/metabolismo , Linhagem Celular , Ciclo-Oxigenase 2/metabolismo , Citocinas/genética , Inflamassomos/genética , Macrófagos Alveolares/metabolismo , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Óxido Nítrico/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Suínos , Receptor 4 Toll-Like/metabolismo , Transcriptoma/efeitos dos fármacos
10.
PLoS Pathog ; 18(2): e1010302, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35120185

RESUMO

Helminth neuroinfections represent serious medical conditions, but the diversity of the host-parasite interplay within the nervous tissue often remains poorly understood, partially due to the lack of laboratory models. Here, we investigated the neuroinvasion of the mouse spinal cord by Trichobilharzia regenti (Schistosomatidae). Active migration of T. regenti schistosomula through the mouse spinal cord induced motor deficits in hindlimbs but did not affect the general locomotion or working memory. Histological examination of the infected spinal cord revealed eosinophilic meningomyelitis with eosinophil-rich infiltrates entrapping the schistosomula. Flow cytometry and transcriptomic analysis of the spinal cord confirmed massive activation of the host immune response. Of note, we recorded striking upregulation of the major histocompatibility complex II pathway and M2-associated markers, such as arginase or chitinase-like 3. Arginase also dominated the proteins found in the microdissected tissue from the close vicinity of the migrating schistosomula, which unselectively fed on the host nervous tissue. Next, we evaluated the pathological sequelae of T. regenti neuroinvasion. While no demyelination or blood-brain barrier alterations were noticed, our transcriptomic data revealed a remarkable disruption of neurophysiological functions not yet recorded in helminth neuroinfections. We also detected DNA fragmentation at the host-schistosomulum interface, but schistosomula antigens did not affect the viability of neurons and glial cells in vitro. Collectively, altered locomotion, significant disruption of neurophysiological functions, and strong M2 polarization were the most prominent features of T. regenti neuroinvasion, making it a promising candidate for further neuroinfection research. Indeed, understanding the diversity of pathogen-related neuroinflammatory processes is a prerequisite for developing better protective measures, treatment strategies, and diagnostic tools.


Assuntos
Arginase/metabolismo , Eosinófilos/metabolismo , Schistosomatidae/imunologia , Medula Espinal/parasitologia , Infecções por Trematódeos/imunologia , Infecções por Trematódeos/metabolismo , Animais , Biomarcadores/metabolismo , Quimiocinas/metabolismo , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Interações Hospedeiro-Parasita , Imunidade , Complexo Principal de Histocompatibilidade , Camundongos , Camundongos Endogâmicos C57BL , Neuroglia/parasitologia , Neurônios/parasitologia , Infecções por Trematódeos/patologia
11.
Cells ; 11(3)2022 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-35159317

RESUMO

Embryonic stem cell-expressed Ras (ERas) is an atypical constitutively active member of the Ras family and controls distinct signaling pathways, which are critical, for instance, for the maintenance of quiescent hepatic stellate cells (HSCs). Unlike classical Ras paralogs, ERas has a unique N-terminal extension (Nex) with as yet unknown function. In this study, we employed affinity pull-down and quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses and identified 76 novel binding proteins for human and rat ERas Nex peptides, localized in different subcellular compartments and involved in various cellular processes. One of the identified Nex-binding proteins is the nonmitochondrial, cytosolic arginase 1 (ARG1), a key enzyme of the urea cycle and involved in the de novo synthesis of polyamines, such as spermidine and spermine. Here, we show, for the first time, a high-affinity interaction between ERas Nex and purified ARG1 as well as their subcellular colocalization. The inhibition of ARG1 activity strikingly accelerates the activation of HSCs ex vivo, suggesting a central role of ARG1 activity in the maintenance of HSC quiescence.


Assuntos
Arginase , Células Estreladas do Fígado , Proteína Oncogênica p21(ras) , Animais , Arginase/metabolismo , Cromatografia Líquida , Células-Tronco Embrionárias/metabolismo , Células Estreladas do Fígado/metabolismo , Humanos , Proteína Oncogênica p21(ras)/metabolismo , Ratos , Espectrometria de Massas em Tandem
12.
Neurochem Res ; 47(5): 1255-1268, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35098420

RESUMO

Alzheimer's disease (AD) is an insidious neurodegenerative disorder representing a serious continuously escalating medico-social problem. The AD-associated progressive dementia is followed by gradual formation of amyloid plaques and neurofibrillary tangles in the brain. Though, converging evidence indicates apparent metabolic dysfunctions as key AD characteristic. In particular, late-onset AD possesses a clear metabolic signature. Considerable brain insulin signaling impairment and a decline in glucose metabolism are common AD attributes. Thus, positron emission tomography (PET) with glucose tracers is a reliable non-invasive tool for early AD diagnosis and treatment efficacy monitoring. Various approaches and agents have been trialed to modulate insulin signaling. Accumulating data point to arginase inhibition as a promising direction to treat AD via diverse molecular mechanisms involving, inter alia, the insulin pathway. Here, we use a transgenic AD mouse model, demonstrating age-dependent brain insulin signaling abnormalities, reduced brain insulin receptor levels, and substantial energy metabolism alterations, to evaluate the effects of arginase inhibition with Norvaline on glucose metabolism. We utilize fluorodeoxyglucose whole-body micro-PET to reveal a significant treatment-associated increase in glucose uptake by the brain tissue in-vivo. Additionally, we apply advanced molecular biology and bioinformatics methods to explore the mechanisms underlying the effects of Norvaline on glucose metabolism. We demonstrate that treatment-associated improvement in glucose utilization is followed by significantly elevated levels of insulin receptor and glucose transporter-3 expression in the mice hippocampi. Additionally, Norvaline diminishes the rate of Tau protein phosphorylation. Our results suggest that Norvaline interferes with AD pathogenesis. These findings open new avenues for clinical evaluation and innovative drug development.


Assuntos
Doença de Alzheimer , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Arginase/metabolismo , Arginase/farmacologia , Arginase/uso terapêutico , Encéfalo/metabolismo , Glucose/metabolismo , Camundongos , Camundongos Transgênicos , Valina/análogos & derivados , Proteínas tau/metabolismo
13.
Arch Pharm Res ; 45(1): 11-28, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-35060088

RESUMO

Atopic dermatitis (AD) and mood disorder comorbidities are typical, but the exact mechanism underlying their interplay has not been clarified. In this study, we aimed to identify the possible mechanisms of anxiety/depressive-like behaviors observed in AD, focusing on microglia. AD was induced by Dermatophagoides farinae body extract (Dfb) in NC/Nga mice and anxiety/depressive-like behaviors were analyzed by behavioral assessments such as open field test (OFT), tail suspension test (TST), sucrose preference test (SPT), and social interaction. As clinical symptoms of AD induced, anxiety/depressive-like behaviors were increased in the OFT and TST and serum glucocorticoid was elevated. AD mice showed an increased mRNA expression of interleukin-4 (IL-4) in lymph nodes but decreased arginase 1 (Arg1) mRNA expression without a change of IL-4 in the hippocampus. In addition, AD mice showed microglia with a shortened branch of de-ramified form and astrocytes with longer processes and decreased branching in the hippocampus, especially in the dentate gyrus (DG). The immunofluorescence study of the DG confirmed that Arg1 reduction was associated with microglia, but not astrocytes. Furthermore, glucocorticoid receptor reduction, increased 5-HT1AR, reduced phosphorylated cAMP response element-binding protein (pCREB), and brain-derived neurotrophic factor (BDNF) expression were identified in the hippocampus of AD mice. Notably, an immunofluorescence study confirmed that pCREB was decreased in the DG of AD mice. Collectively, our data suggest that the reduced Arg1 positive microglia might contribute to anxiety/depressive-like behaviors via pCREB/BDNF reduction in AD.


Assuntos
Ansiedade/complicações , Arginase/metabolismo , Depressão/complicações , Dermatite Atópica/complicações , Microglia/enzimologia , Animais , Antígenos de Dermatophagoides/efeitos adversos , Ansiedade/patologia , Arginase/fisiologia , Western Blotting , Depressão/patologia , Dermatite Atópica/patologia , Dermatite Atópica/psicologia , Modelos Animais de Doenças , Imunofluorescência , Elevação dos Membros Posteriores , Masculino , Camundongos , Microglia/patologia , Teste de Campo Aberto , Reação em Cadeia da Polimerase Via Transcriptase Reversa
14.
Cell Rep ; 38(2): 110215, 2022 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-35021079

RESUMO

Macrophages are known to mediate anti-helminth responses, but it remains uncertain which subsets are involved or how macrophages actually kill helminths. Here, we show rapid monocyte recruitment to the lung after infection with the nematode parasite Nippostrongylus brasiliensis. In this inflamed tissue microenvironment, these monocytes differentiate into an alveolar macrophage (AM)-like phenotype, expressing both SiglecF and CD11c, surround invading parasitic larvae, and preferentially kill parasites in vitro. Monocyte-derived AMs (Mo-AMs) express type 2-associated markers and show a distinct remodeling of the chromatin landscape relative to tissue-derived AMs (TD-AMs). In particular, they express high amounts of arginase-1 (Arg1), which we demonstrate mediates helminth killing through L-arginine depletion. These studies indicate that recruited monocytes are selectively programmed in the pulmonary environment to express AM markers and an anti-helminth phenotype.


Assuntos
Pulmão/imunologia , Macrófagos Alveolares/imunologia , Infecções por Strongylida/imunologia , Animais , Arginase/metabolismo , Diferenciação Celular , Citocinas , Feminino , Pulmão/parasitologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Nippostrongylus , Infecções por Strongylida/parasitologia
15.
Clin Nutr ; 41(1): 21-32, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34864452

RESUMO

BACKGROUND & AIMS: Folic acid supplementation is widely accepted during pregnancy, as it exerts a protective effect on neural tube defects. However, the long-term underlying effects of folic acid supplementation during pregnancy (FASDP) on offspring remain unclear. METHODS: Thirty pregnant female rats were randomly divided into normal control group, folic acid appropriate supplementation group (2.5 × FA group) and folic acid oversupplementation group (5 × FA group) and fed with corresponding folic acid concentration AIN93G diet. UPLC-Q-TOF-MS, UPLC-TQ-MS and GC-MS were performed to detect the serum metabolites profiles in adult male offspring and explore the effects of FASDP. Moreover, molecular biology technologies were used to clarify the underlying mechanism. RESULTS: We demonstrate that 2.5-folds folic acid leads to dyslipidemic-diabetic slightly in male offspring, while 5-folds folic acid aggravates the disorder and prominent hepatic lipid accumulations. Using untargeted and targeted metabolomics, total 63 differential metabolites and 12 significantly differential KEGG pathways are identified. Of note, arginine biosynthesis, arginine and proline metabolism are the two most significant pathways. Mechanistic investigations reveal that the increased levels of arginase-1 (Arg1) causes the lipid metabolism disorder by regulating nitric oxide synthase-3 (NOS3)-adenosine monophosphate activated protein kinase-α (AMPKα) pathway, resulting in lipid accumulation in hepatocytes. CONCLUSIONS: Our data suggest that maternal folic acid oversupplementation during pregnancy contributes to lipid metabolism disorder in male offspring by regulating Arg1-NOS3-AMPKα pathway.


Assuntos
Arginase/metabolismo , Suplementos Nutricionais/efeitos adversos , Ácido Fólico/efeitos adversos , Transtornos do Metabolismo dos Lipídeos/induzido quimicamente , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Dieta/métodos , Feminino , Ácido Fólico/administração & dosagem , Cromatografia Gasosa-Espectrometria de Massas , Hepatócitos/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Masculino , Metabolômica , Óxido Nítrico Sintase Tipo III/metabolismo , Gravidez , Ratos , Transdução de Sinais/efeitos dos fármacos
16.
Cardiovasc Res ; 118(1): 254-266, 2022 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33483748

RESUMO

AIMS: Rheumatoid arthritis (RA) is a chronic inflammatory disease affecting joints and blood vessels. Despite low levels of low-density lipoprotein cholesterol (LDL-C), RA patients exhibit endothelial dysfunction and are at increased risk of death from cardiovascular complications, but the molecular mechanism of action is unknown. We aimed in the present study to identify the molecular mechanism of endothelial dysfunction in a mouse model of RA and in patients with RA. METHODS AND RESULTS: Endothelium-dependent relaxations to acetylcholine were reduced in aortae of two tumour necrosis factor alpha (TNFα) transgenic mouse lines with either mild (Tg3647) or severe (Tg197) forms of RA in a time- and severity-dependent fashion as assessed by organ chamber myograph. In Tg197, TNFα plasma levels were associated with severe endothelial dysfunction. LOX-1 receptor was markedly up-regulated leading to increased vascular oxLDL uptake and NFκB-mediated enhanced Arg2 expression via direct binding to its promoter resulting in reduced NO bioavailability and vascular cGMP levels as shown by ELISA and chromatin immunoprecipitation. Anti-TNFα treatment with infliximab normalized endothelial function together with LOX-1 and Arg2 serum levels in mice. In RA patients, soluble LOX-1 serum levels were also markedly increased and closely related to serum levels of C-reactive protein. Similarly, ARG2 serum levels were increased. Similarly, anti-TNFα treatment restored LOX-1 and ARG2 serum levels in RA patients. CONCLUSIONS: Increased TNFα levels not only contribute to RA, but also to endothelial dysfunction by increasing vascular oxLDL content and activation of the LOX-1/NFκB/Arg2 pathway leading to reduced NO bioavailability and decreased cGMP levels. Anti-TNFα treatment improved both articular symptoms and endothelial function by reducing LOX-1, vascular oxLDL, and Arg2 levels.


Assuntos
Aorta Torácica/efeitos dos fármacos , Arginase/metabolismo , Artrite Reumatoide/tratamento farmacológico , Células Endoteliais/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Receptores Depuradores Classe E/metabolismo , Inibidores do Fator de Necrose Tumoral/uso terapêutico , Fator de Necrose Tumoral alfa/metabolismo , Vasodilatação/efeitos dos fármacos , Adulto , Animais , Animais Geneticamente Modificados , Aorta Torácica/enzimologia , Aorta Torácica/imunologia , Aorta Torácica/fisiopatologia , Arginase/genética , Artrite Reumatoide/enzimologia , Artrite Reumatoide/imunologia , Artrite Reumatoide/fisiopatologia , Estudos de Casos e Controles , Modelos Animais de Doenças , Células Endoteliais/enzimologia , Células Endoteliais/imunologia , Endotélio Vascular/enzimologia , Endotélio Vascular/imunologia , Endotélio Vascular/fisiopatologia , Feminino , Humanos , Lipoproteínas LDL/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Receptores Depuradores Classe E/genética , Transdução de Sinais , Fator de Necrose Tumoral alfa/genética
17.
Chem Biol Interact ; 351: 109690, 2022 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-34637778

RESUMO

The currently available treatment options for leishmaniasis are associated with high costs, severe side effects, and high toxicity. In previous studies, thiohydantoins demonstrated some pharmacological activities and were shown to be potential hit compounds with antileishmanial properties. The present study further explored the antileishmanial effect of acetyl-thiohydantoins against Leishmania amazonensis and determined the main processes involved in parasite death. We observed that compared to thiohydantoin nuclei, acetyl-thiohydantoin treatment inhibited the proliferation of promastigotes. This treatment caused alterations in cell cycle progression and parasite size and caused morphological and ultrastructural changes. We then investigated the mechanisms involved in the death of the protozoan; there was an increase in ROS production, phosphatidylserine exposure, and plasma membrane permeabilization and a loss of mitochondrial membrane potential, resulting in an accumulation of lipid bodies and the formation of autophagic vacuoles on these parasites and confirming an apoptosis-like process. In intracellular amastigotes, selected acetyl-thiohydantoins reduced the percentage of infected macrophages and the number of amastigotes/macrophages by increasing ROS production and reducing TNF-α levels. Moreover, thiohydantoins did not induce cytotoxicity in murine macrophages (J774A.1), human monocytes (THP-1), or sheep erythrocytes. In silico and in vitro analyses showed that acetyl-thiohydantoins exerted in vitro antileishmanial effects on L. amazonensis promastigotes in apoptosis-like and amastigote forms by inducing ROS production and reducing TNF-α levels, indicating that they are good candidates for drug discovery studies in leishmaniasis treatment. Additionally, we carried out molecular docking analyses of acetyl-thiohydantoins on two important targets of Leishmania amazonensis: arginase and TNF-alpha converting enzyme. The results suggested that the acetyl groups in the N1-position of the thiohydantoin ring and the ring itself could be pharmacophoric groups due to their affinity for binding amino acid residues at the active site of both enzymes via hydrogen bond interactions. These results demonstrate that thiohydantoins are promising hit compounds that could be used as antileishmanial agents.


Assuntos
Tioidantoínas/farmacologia , Tripanossomicidas/farmacologia , Proteína ADAM17/metabolismo , Animais , Arginase/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Leishmania/efeitos dos fármacos , Leishmania/enzimologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Simulação de Acoplamento Molecular , Proteínas de Protozoários/metabolismo , Ovinos , Tioidantoínas/síntese química , Tioidantoínas/metabolismo , Tioidantoínas/toxicidade , Tripanossomicidas/síntese química , Tripanossomicidas/metabolismo , Tripanossomicidas/toxicidade , Fator de Necrose Tumoral alfa/metabolismo
18.
Clin Lung Cancer ; 23(2): e140-e147, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34393062

RESUMO

BACKGROUND: Key regulators of antitumor immunity such as arginase-1 and the adenosine pathway may have an important role in modulating the effect of immunotherapy. Here, we investigated the expression profile of these immune-related biomarkers in thymic epithelial tumors (TETs) and small cell lung cancer (SCLC), 2 solid tumors where immune checkpoint inhibitors have activity. MATERIALS AND METHODS: Immunohistochemical staining was performed using tissue microarrays of 123 TET (110 thymoma and 13 thymic carcinoma) and 125 SCLC cases. The expression profile of the following immune-related biomarkers was assessed: arginase-1, CD39, CD73, A2AR, PD-L2, and CD15. The expression profile was also correlated with clinical data. RESULTS: No sample was positive for arginase-1. In the adenosine pathway, the prevalence of positive staining for CD39, CD73, and A2AR was 4.9%, 2.5%, and 69.2%, in TETs and 0%, 1.7%, and 50.8%, in SCLC. The multivariate analysis showed that CD39 expression was significantly associated with worse disease related survival (hazard ratio [HR], 10.36; 95% confidence interval [CI]: 2.01-53.47; P= .005) and a shorter time-to progression (HR, 11.35; 95% CI, 2.11-61.23; P = .005) in TETs. Other biomarkers were not associated with disease related survival or time to progression in TETs. No biomarker was associated with survival in SCLC. CONCLUSION: Arginase-1 was not detectable in TETs and SCLC. Expression of markers in the adenosine pathway were present in both TETs and SCLC. CD39 expression in tumor cells may identify subsets of patients with TETs with an unfavorable prognosis.


Assuntos
Arginase/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Epiteliais e Glandulares/metabolismo , Carcinoma de Pequenas Células do Pulmão/metabolismo , Neoplasias do Timo/metabolismo , Biomarcadores Tumorais/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Epiteliais e Glandulares/patologia , Prognóstico , Carcinoma de Pequenas Células do Pulmão/patologia , Neoplasias do Timo/patologia , Microambiente Tumoral
19.
Exp Gerontol ; 159: 111676, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34968674

RESUMO

Considering the efficacy of rapamycin in increasing lifespan and healthspan, attenuating the aging-dependent immunological decline, we compared the evolution of Trypanosoma cruzi infection and acute myocarditis in young and elderly mice untreated and chronically treated with this drug. Five groups were investigated: young uninfected and infected, elderly uninfected and infected with Trypanosoma cruzi untreated and treated with rapamycin (4 mg/kg every 3 days) from the 8th to the 96th week of age. Seven days after the last treatment, elderly mice were inoculated with T. cruzi. Young animals were infected at 8-weeks-old. Untreated elderly mice exhibited increase parasitemia, parasite load and myocarditis, which were associated to down-regulation in IL-2, IL-6, IFN-γ, TNF, anti-T. cruzi immunoglobulin G (IgG) total, IgG1 and IgG2a plasma levels, inducible nitric oxide synthase (iNOS) gene expression and nitric oxide (NO) cardiac production, as well as upregulation in Arginase-1 gene expression and arginase activity compared to young animals. These parameters were improved in rapamycin-pretreated elderly mice, which exhibited a better parasitological control, reduced heart inflammation and microstructural damage. These responses were associated with a better balance between Th1 and Th2 effectors similar to that observed in young animals, including an improved activation of Th1 cytokines and the iNOS pathway that positively regulates NO biosynthesis, contradicting the predominant activation of the arginase pathway in untreated elderly animals. Thus, our findings suggest that chronic pretreatment with rapamycin can attenuate immunosenescence in mice, contributing to prolong parasite resistance and attenuate acute myocarditis in elderly host challenged by T. cruzi.


Assuntos
Doença de Chagas , Miocardite , Trypanosoma cruzi , Envelhecimento , Animais , Arginase/metabolismo , Doença de Chagas/tratamento farmacológico , Camundongos , Miocardite/tratamento farmacológico , Miocardite/parasitologia , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Sirolimo/farmacologia , Trypanosoma cruzi/metabolismo
20.
Gene ; 813: 146110, 2022 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-34902507

RESUMO

The use of tyrosine kinase inhibitors seems to restore the broadly compromised immune system described in chronic myeloid leukaemia (CML) patients at diagnosis leading to a re-activation of the effector-mediated immune surveillance. Here, we describe the expression dynamics of immune factors during the first year on imatinib therapy. Gene expression was evaluated in 132 peripheral blood samples from 79 CML patients, including 34 who were serially followed. An aliquot of the stored sample used to monitor BCR-ABL1 levels was retro-transcribed to cDNA and gene expression was quantified by real-time PCR. An elevated expression of ARG1 was observed at diagnosis, while TBET, CIITA, IL10 and TGFB1 were significantly decreased. Once on therapy, each gene displayed a particular behaviour. ARG1 normalized to control levels at 3 months only in optimal molecular responders and was identified as the major contributor to the difference among patients. TBET reached normal levels after 12 months in optimal responders and non-responders, regardless the Th1-response previously associated, and CIITA continued downregulated. IL10 and TGFB1 achieved normal levels early at 3 months in both groups, afterwards IL10 was sustained while TGFB1 was slightly increased after 1 year in responders. Our findings are in agreement with an immune re-activation after imatinib initiation; however, some immune mediators may require a longer exposition. The follow-up of novel and reliable biomarkers, such as ARG1, one of the principal mechanisms of myeloid-derived-suppressor cells to inhibit immune system, may be useful to deepen the characterization of early responder patients.


Assuntos
Arginase/genética , Mesilato de Imatinib/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Adulto , Antineoplásicos/farmacologia , Arginase/metabolismo , Biomarcadores Farmacológicos/análise , Biomarcadores Farmacológicos/sangue , Feminino , Proteínas de Fusão bcr-abl/genética , Expressão Gênica , Humanos , Fatores Imunológicos/uso terapêutico , Interleucina-10/sangue , Interleucina-10/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/sangue , Masculino , Pessoa de Meia-Idade , Proteínas Nucleares/sangue , Proteínas Nucleares/genética , Inibidores de Proteínas Quinases/farmacologia , Transativadores/sangue , Transativadores/genética , Transcriptoma/genética , Fator de Crescimento Transformador beta1/sangue , Fator de Crescimento Transformador beta1/genética
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