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1.
Nat Genet ; 51(11): 1588-1595, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31676868

RESUMO

The early stages of type 1 diabetes (T1D) are characterized by local autoimmune inflammation and progressive loss of insulin-producing pancreatic ß cells. Here we show that exposure to proinflammatory cytokines reveals a marked plasticity of the ß-cell regulatory landscape. We expand the repertoire of human islet regulatory elements by mapping stimulus-responsive enhancers linked to changes in the ß-cell transcriptome, proteome and three-dimensional chromatin structure. Our data indicate that the ß-cell response to cytokines is mediated by the induction of new regulatory regions as well as the activation of primed regulatory elements prebound by islet-specific transcription factors. We find that T1D-associated loci are enriched with newly mapped cis-regulatory regions and identify T1D-associated variants disrupting cytokine-responsive enhancer activity in human ß cells. Our study illustrates how ß cells respond to a proinflammatory environment and implicate a role for stimulus response islet enhancers in T1D.


Assuntos
Cromatina/genética , Citocinas/farmacologia , Diabetes Mellitus Tipo 1/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Redes Reguladoras de Genes , Células Secretoras de Insulina/metabolismo , Transcriptoma , Cromatina/química , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/patologia , Elementos Facilitadores Genéticos , Estudo de Associação Genômica Ampla , Humanos , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/patologia , Fatores de Transcrição
2.
Nat Genet ; 51(10): 1486-1493, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31548716

RESUMO

Immune-disease-associated variants are enriched in active chromatin regions of T cells and macrophages. However, whether these variants function in specific cell states is unknown. Here we stimulated T cells and macrophages in the presence of 13 cytokines and profiled active and open chromatin regions. T cell activation induced major chromatin remodeling, while the presence of cytokines fine-tuned the magnitude of changes. We developed a statistical method that accounts for subtle changes in the chromatin landscape to identify SNP enrichment across cell states. Our results point towards the role of immune-disease-associated variants in early rather than late activation of memory CD4+ T cells, with modest differences across cytokines. Furthermore, variants associated with inflammatory bowel disease are enriched in type 1 T helper (TH1) cells, whereas variants associated with Alzheimer's disease are enriched in different macrophage cell states. Our results represent an in-depth analysis of immune-disease-associated variants across a comprehensive panel of activation states of T cells and macrophages.


Assuntos
Cromatina/metabolismo , Citocinas/farmacologia , Estudo de Associação Genômica Ampla , Doenças do Sistema Imunitário/imunologia , Macrófagos/imunologia , Células Th1/imunologia , Cromatina/genética , Humanos , Doenças do Sistema Imunitário/tratamento farmacológico , Doenças do Sistema Imunitário/genética , Ativação Linfocitária , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Células Th1/efeitos dos fármacos , Células Th1/metabolismo
3.
DNA Cell Biol ; 38(10): 1025-1029, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31532239

RESUMO

Neutrophil trafficking into damaged or infected tissues is essential for the initiation of inflammation, clearance of pathogens and damaged cells, and ultimately tissue repair. Neutrophil recruitment is highly dependent on the stepwise induction of adhesion molecules and promigratory chemokines and cytokines. A number of studies in animal models have shown the efficacy of cannabinoid receptor 2 (CB2) agonists in limiting inflammation in a range of preclinical models of inflammation, including colitis, atherosclerosis, multiple sclerosis, and ischemia-reperfusion injury. Recent work in preclinical models of inflammation raises two questions: by what mechanisms do CB2 agonists provide anti-inflammatory effects during acute inflammation and what challenges exist in the translation of CB2 modulating therapeutics into the clinic.


Assuntos
Aterosclerose/genética , Colite/genética , Esclerose Múltipla/genética , Neutrófilos/metabolismo , Receptor CB2 de Canabinoide/genética , Traumatismo por Reperfusão/genética , Animais , Aterosclerose/tratamento farmacológico , Aterosclerose/metabolismo , Aterosclerose/patologia , Agonistas de Receptores de Canabinoides/uso terapêutico , Antagonistas de Receptores de Canabinoides/uso terapêutico , Colite/tratamento farmacológico , Colite/metabolismo , Colite/patologia , Citocinas/metabolismo , Citocinas/farmacologia , Modelos Animais de Doenças , Regulação da Expressão Gênica , Humanos , Inflamação , Ligantes , Camundongos , Camundongos Knockout , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Infiltração de Neutrófilos , Neutrófilos/efeitos dos fármacos , Neutrófilos/patologia , Receptor CB2 de Canabinoide/agonistas , Receptor CB2 de Canabinoide/antagonistas & inibidores , Receptor CB2 de Canabinoide/deficiência , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia
4.
Int J Mol Sci ; 20(18)2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31510070

RESUMO

A key role of the mitochondrial Translocator Protein 18 KDa (TSPO) in neuroinflammation has been recently proposed. However, little is known about TSPO-activated pathways underlying the modulation of reactive microglia. In the present work, the TSPO activation was explored in an in vitro human primary microglia model (immortalized C20 cells) under inflammatory stimulus. Two different approaches were used with the aim to (i) pharmacologically amplify or (ii) silence, by the lentiviral short hairpin RNA, the TSPO physiological function. In the TSPO pharmacological stimulation model, the synthetic steroidogenic selective ligand XBD-173 attenuated the activation of microglia. Indeed, it reduces and increases the release of pro-inflammatory and anti-inflammatory cytokines, respectively. Such ligand-induced effects were abolished when C20 cells were treated with the steroidogenesis inhibitor aminoglutethimide. This suggests a role for neurosteroids in modulating the interleukin production. The highly steroidogenic ligand XBD-173 attenuated the neuroinflammatory response more effectively than the poorly steroidogenic ones, which suggests that the observed modulation on the cytokine release may be influenced by the levels of produced neurosteroids. In the TSPO silencing model, the reduction of TSPO caused a more inflamed phenotype with respect to scrambled cells. Similarly, during the inflammatory response, the TSPO silencing increased and reduced the release of pro-inflammatory and anti-inflammatory cytokines, respectively. In conclusion, the obtained results are in favor of a homeostatic role for TSPO in the context of dynamic balance between anti-inflammatory and pro-inflammatory mediators in the human microglia-mediated inflammatory response. Interestingly, our preliminary results propose that the TSPO expression could be stimulated by NF-κB during activation of the inflammatory response.


Assuntos
Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Microglia/efeitos dos fármacos , Purinas/farmacologia , Interferência de RNA , Receptores de GABA/metabolismo , Aminoglutetimida/farmacologia , Anti-Inflamatórios/farmacologia , Inibidores da Aromatase/farmacologia , Sequência de Bases , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Citocinas/farmacologia , Expressão Gênica/efeitos dos fármacos , Humanos , Mediadores da Inflamação/farmacologia , Microglia/metabolismo , NF-kappa B/metabolismo , Fenótipo , Receptores de GABA/genética
5.
Mol Med Rep ; 20(5): 4358-4366, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31545423

RESUMO

Natural killer (NK) cells are a group of large granular lymphocytes that play an important regulatory role in innate immunity and adaptive immunity. Immune­related pancytopenia (IRP) is a type of pancytopenia resulting from bone marrow hematopoietic cells that were destroyed or suppressed by auto­antibodies. The specific mechanism of IRP is not clear. In the present study, it was identified that the percentage of NK cells in peripheral blood lymphocytes was decreased in patients with IRP. Subsequently, high purity NK cells were extracted from 6 untreated patients with IRP using the immunomagnetic beads sorting, magnetic­activated cell­sorting method, which were then cultured then in RPMI­1640 medium containing 20% FBS. NK cell expansion agents, with or without recombinant interleukin (IL)­15, were used to amplify high­purity NK cells on the basic of recombinant IL­2. Expression of the activated receptors NKG2­D type II integral membrane protein (NKG2D) and natural killer cell p46­related protein (NKp46), and the inhibitory receptors CD158a and NKG2­A/NKG2­B type II integral membrane protein (NKG2A), in CD56+ NK cells were detected by flow cytometry before and after cell culture. It was observed that treatment with an NK cell expansion agent combined with the stimulation of recombinant IL­2 and recombinant IL­15 could increase the number whilst maintaining the purity of NK cells. There were no significant changes in the expression of NKG2D, NKp46, NKG2A and CD158a in patients with IRP before and after NK cell culture. This new amplification method lays a foundation for clinical NK cell immunotherapy and anti­tumor applications.


Assuntos
Interleucina-15/farmacologia , Interleucina-2/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Proteínas Recombinantes/farmacologia , Adolescente , Adulto , Idoso , Biomarcadores , Técnicas de Cultura de Células , Criança , Citocinas/farmacologia , Feminino , Citometria de Fluxo , Humanos , Imunoterapia Adotiva , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Masculino , Pessoa de Meia-Idade , Pancitopenia/diagnóstico , Pancitopenia/etiologia , Pancitopenia/terapia , Adulto Jovem
6.
J Orthop Res ; 37(9): 1979-1987, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31062877

RESUMO

The synovium plays a key role in the development of osteoarthritis, as evidenced by pathological changes to the tissue observed in both early and late stages of the disease. One such change is the attachment of cartilage wear particles to the synovial intima. While this phenomenon has been well observed clinically, little is known of the biological effects that such particles have on resident cells in the synovium. The present work investigates the hypothesis that cartilage wear particles elicit a pro-inflammatory response in diseased and healthy human fibroblast-like synoviocytes, like that induced by key cytokines in osteoarthritis. Fibroblast-like synoviocytes from 15 osteoarthritic human donors and a subset of three non-osteoarthritic donors were exposed to cartilage wear particles, interleukin-1α or tumor necrosis factor-α for 6 days and analyzed for proliferation, matrix production, and release of pro-inflammatory mediators and degradative enzymes. Wear particles significantly increased proliferation and release of nitric oxide, interleukin-6 and -8, and matrix metalloproteinase-9, -10, and -13 in osteoarthritic synoviocytes, mirroring the effects of both cytokines, with similar trends in non-osteoarthritic cells. These results suggest that cartilage wear particles are a relevant physical factor in the osteoarthritic environment, perpetuating the pro-inflammatory and pro-degradative cascade by modulating synoviocyte behavior at early and late stages of the disease. Future work points to therapeutic strategies for slowing disease progression that target cell-particle interactions. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1979-1987, 2019.


Assuntos
Cartilagem/fisiologia , Citocinas/farmacologia , Inflamação/etiologia , Sinoviócitos/imunologia , Idoso , Idoso de 80 Anos ou mais , Feminino , Fibroblastos/imunologia , Humanos , Interleucina-1/farmacologia , Masculino , Pessoa de Meia-Idade , Osteoartrite/etiologia , Fator de Necrose Tumoral alfa/farmacologia
7.
Radiat Res ; 192(1): 92-97, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31063041

RESUMO

In response to concerns over possible radiological or nuclear incidents, the Radiation and Nuclear Countermeasures Program within the National Institute of Allergy and Infectious Diseases (NIAID) was tasked by the U.S. Department of Health and Human Services to support development of medical countermeasures (MCM) to treat the acute and delayed injuries that can result from radiation exposure. To date, the only three drugs approved by the U.S. Food and Drug Administration for treatment of acute radiation syndrome are growth factors targeting granulocyte (Neupogen® or Neulasta®) or granulocyte and macrophage (Leukine®) hematopoietic cell lineages. Although these are currently stockpiled for deployment in response to a mass casualty scenario, these growth factors will likely be administered in a scarce-resources environment and availability may be limited. Therefore, there is growing interest in understanding the role that these growth factors play in mitigating radiation damage, to optimize their use and maximize the number of people who can be treated. For these reasons, the NIAID and the Radiation Injury Treatment Network organized a workshop to explore the use of growth factors and other cytokines as MCMs in the treatment of radiation-induced injuries. Subject matter experts from government, industry and academia gathered at this workshop to discuss the concept of operations, triage and treatment, administration to diverse civilian populations, growth factors under development for radiation indications, and how the practice of medicine can inform other potential approaches.


Assuntos
Citocinas/farmacologia , Emergências , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Saúde Pública , Lesões por Radiação/tratamento farmacológico , Citocinas/uso terapêutico , Descoberta de Drogas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/uso terapêutico
8.
Int J Med Microbiol ; 309(5): 274-282, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31113736

RESUMO

Amyloid curli fibrils produced by Escherichia coli are well-known virulence factor influencing E. coli adhesion and biofilm formation. However, the impact of curli on intestinal epithelial barrier stimulated with proinflammatory cytokines is unknown. In the study, we examined the effect of curli produced by nonpathogenic E. coli K-12 and wild-type E. coli EC32 strains, and purified CsgA proteins on differentiated Caco-2 cell monolayers stimulated with a mixture of IL-1ß, TNF-α, and INFγ cytokines as a model of 'inflamed intestinal epithelial barrier' in vitro. The results of the study indicated that curliated E. coli adhered better to polarized Caco-2 cells than their curli-deficient mutants and the adherence was further augmented by stimulation of epithelial cells with proinflammatory cytokines. Interestingly, curli reduced internalization but enhanced intracellular survival of the wild-type E. coli strain EC32 within intestinal epithelial cells. Curli-expressing E. coli, as well as purified CsgA proteins, attenuated IL-8 secretion by unstimulated Caco-2 cells, although the effect was barely observed on cytokine-stimulated cells. The findings of the study revealed that curli fibrils are an important virulence factor enabling curliated E. coli to effectively colonize intestinal epithelium especially in individuals with inflammatory intestinal disorders.


Assuntos
Aderência Bacteriana , Citocinas/farmacologia , Células Epiteliais/microbiologia , Proteínas de Escherichia coli/genética , Escherichia coli/patogenicidade , Intestinos/citologia , Células CACO-2 , Técnicas de Cultura de Células , Células Epiteliais/efeitos dos fármacos , Escherichia coli/genética , Humanos , Interferon gama/farmacologia , Interleucina-1beta/farmacologia , Intestinos/imunologia , Fator de Necrose Tumoral alfa/farmacologia , Fatores de Virulência/genética
9.
Mol Biol (Mosk) ; 53(2): 200-217, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31099771

RESUMO

Heat shock proteins (HSPs) are important factors of protein homeostasis and possess chaperone properties, providing for a folding and intracellular transport of proteins and facilitating the recovery or utilization of proteins partly denatured on exposure to various stress factors. Proteins of the Hsp70 family are the most universal molecular chaperones and interact with the greatest number of protein substrates. Several proteins of the Hsp70 family are released into the extracellular space, where they play an important role in intercellular communications and act as alarmins, or "danger signals," to modulate the immune response. The secreted Hsp70 can additionally act as an effective neuroprotector, increasing the survival of neurons in various proteinopathies, as has been demonstrated in Alzheimer's and Parkinson's disease models. In this regard, recombinant Hsp70 and inducers of endogenous Hsp70 synthesis may be considered as candidate therapeutics with immune-modulating and neuroprotective properties.Heat shock proteins (HSPs) are important factors of protein homeostasis and possess chaperone properties, providing for a folding and intracellular transport of proteins and facilitating the recovery or utilization of proteins partly denatured on exposure to various stress factors. Proteins of the Hsp70 family are the most universal molecular chaperones and interact with the greatest number of protein substrates. Several proteins of the Hsp70 family are released into the extracellular space, where they play an important role in intercellular communications and act as alarmins, or "danger signals," to modulate the immune response. The secreted Hsp70 can additionally act as an effective neuroprotector, increasing the survival of neurons in various proteinopathies, as has been demonstrated in Alzheimer's and Parkinson's disease models. In this regard, recombinant Hsp70 and inducers of endogenous Hsp70 synthesis may be considered as candidate therapeutics with immune-modulating and neuroprotective properties.


Assuntos
Citocinas/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Homeostase , Proteostase , Citocinas/imunologia , Citocinas/farmacologia , Citocinas/uso terapêutico , Proteínas de Choque Térmico HSP70/imunologia , Proteínas de Choque Térmico HSP70/farmacologia , Proteínas de Choque Térmico HSP70/uso terapêutico , Homeostase/efeitos dos fármacos , Humanos , Proteostase/efeitos dos fármacos
10.
Radiat Res ; 192(1): 99-120, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31081742

RESUMO

Due to the threat of a radiological or nuclear incident that could impact citizens, the U.S. Department of Health and Human Services tasked the National Institute of Allergy and Infectious Diseases (NIAID) with identifying and funding early- to mid-stage medical countermeasure (MCM) development to treat radiation-induced injuries. Given that the body's natural response to radiation exposure includes production of growth factors and cytokines, and that the only drugs approved by the U.S. Food and Drug Administration to treat acute radiation syndrome are growth factors targeting either the granulocyte (Neupogen® or Neulasta®) or granulocyte and macrophage (Leukine®) hematopoietic cell lineages, there is interest in understanding the role that these factors play in responding to and/or ameliorating radiation damage. Furthermore, in an environment where resources are scarce, such as what might be expected during a radiation public health emergency, availability of growth factor or other treatments may be limited. For these reasons, the NIAID partnered with the Radiation Injury Treatment Network (RITN), whose membership includes medical centers with expertise in the management of bone marrow failure, to explore the use of growth factors and other cytokines as MCMs to mitigate/treat radiation injuries. A workshop was convened that included government, industry and academic subject matter experts, with presentations covering the anticipated concept of operations during a mass casualty incident including triage and treatment, growth factors under development for a radiation indication, and how the practice of medicine can inform other potential approaches, as well as considerations for administration of these products to diverse civilian populations. This report reviews the information presented, and provides an overview of the discussions from a guided breakout session.


Assuntos
Citocinas/farmacologia , Emergências , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Saúde Pública , Lesões por Radiação/tratamento farmacológico , Animais , Citocinas/uso terapêutico , Descoberta de Drogas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/uso terapêutico
11.
Cells ; 8(4)2019 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-31022954

RESUMO

We previously established a method for a directed differentiation of human pluripotent stem cells into classical brown adipocytes (BA) by forming aggregates via massive floating culture in the presence of a specific cytokine cocktail. However, use of recombinant cytokines requires significant cost. Moreover, an enforced differentiation by exogenously added cytokines may amend skewed differentiation propensity of patient's pluripotent stem cells, providing unsatisfactory disease models. Therefore, an exogenous cytokine-free method, where cytokines required for differentiation are provided in an auto/paracrine manner mimicking natural developmental process, is beneficial. Here we show that, if human pluripotent stem cells are cultured as size-controlled spheroids (100-120 µm radius, 2000-2500 cells/spheroid) in a mutually segregated manner with half-change of the medium every other day, they differentiate into classical BA via an authentic MYF5-positive myoblast route in the absence of exogenous cytokines. Differentiated BA exerted thermogenic activity in transplanted mice in response to beta-adrenergic receptor agonist stimuli. The cytokine-free differentiation method has further advantages in exploring BATokines, BA-derived physiologically active substances. Indeed, we have found that BA produces an unknown small (<1000 Da), highly hydrophilic molecule that augments insulin secretion from pancreatic beta cells. Our upgraded technique will contribute to an advancement of stem cell study for diverse purposes.


Assuntos
Adipócitos Marrons/citologia , Diferenciação Celular/fisiologia , Células-Tronco Pluripotentes/citologia , Adipócitos Marrons/metabolismo , Adipócitos Marrons/fisiologia , Animais , Técnicas de Cultura de Células/métodos , Células Cultivadas , Citocinas/farmacologia , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células Secretoras de Insulina/citologia , Camundongos , Células-Tronco Pluripotentes/metabolismo , Esferoides Celulares/metabolismo , Termogênese
12.
Can J Physiol Pharmacol ; 97(8): 699-707, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31026403

RESUMO

Obesity is associated with skeletal muscle insulin resistance and the development of metabolic syndrome. Undifferentiated skeletal muscle cells are sensitive to oxidative stress. Berberine hydrochloride (BBR) improves insulin resistance and exhibits anti-inflammatory properties. However, the underlying mechanism and the cell signaling pathways involved remain largely elusive. We therefore investigated the anti-inflammatory effects of BBR and the signaling pathways using skeletal C2C12 myoblast cells. Undifferentiated C2C12 myoblast cells were treated with interleukin-1ß alone or in combination with tumor necrosis factor-α in the presence or absence of BBR. We found that BBR reduced the cytokine-induced expression of inducible nitric oxide synthase and stress-related kinases including p-38 mitogen-activated protein kinase, nuclear factor kappa B (NF-κB), and stress-activated protein kinases/Jun amino-terminal kinases (SAPK/JNK) in C2C12 myoblast cells. Furthermore, BBR reversed cytokine-mediated suppression of AMP-activated protein kinase (AMPKα), sirtuin-1 (SIRT-1), and PPAR-γ coactivator-1α (PGC-1α). In addition, cytokine-induced reduction of mitochondrial marker proteins and function were rescued after BBR treatment. Catalase, an antioxidant enzyme, was elevated after BBR treatment. Our results demonstrate that BBR ameliorates cytokine-induced inflammation. The anti-inflammatory effect of BBR in skeletal progenitor cells is mediated through pathways including activation of the AMPKα-SIRT-1-PGC-1α, inhibition of the mitogen-activated protein kinase 4 (MKK4)-SAPK/JNK-C-JUN, as well as protection of mitochondrial bioenergetics. BBR may be a potential medication for metabolic syndrome.


Assuntos
Anti-Inflamatórios/farmacologia , Berberina/farmacologia , Citocinas/farmacologia , Mioblastos/efeitos dos fármacos , Mioblastos/patologia , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Citoproteção/efeitos dos fármacos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Camundongos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mioblastos/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
13.
J Immunol Res ; 2019: 1529189, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30882002

RESUMO

Paracoccidioidomycosis (PCM), a systemic mycosis endemic in Latin America, occurs after inhalation of mycelial components of Paracoccidioides spp. When the fungus reaches the lungs and interacts with the alveolar macrophages and other cells, phagocytic cells such as neutrophils and monocytes are immediately recruited to the injured site. The interaction between surface molecules of pathogens and homologous receptors, present on the surface membrane of phagocytes, modulates the innate immune cell activation. Studies have shown the importance of fungal recognition by the Dectin-1 receptor, which can induce a series of cellular protective responses against fungi. The objective of the present study was to evaluate Dectin-1 receptor expression and the effector mechanisms of human monocytes and neutrophils activated or not with different cytokines, such as IFN-γ, TNF-α, and GM-CSF, followed by the challenge with Paracoccidioides brasiliensis (P. brasiliensis or Pb265). Therefore, analysis of Dectin-1 receptor expression was done by flow cytometry whereas the effector mechanisms were evaluated by fungal recovery by colony-forming unit (CFU) counting and hydrogen peroxide (H2O2) production. Our results showed that, after treatment with IFN-γ, TNF-α, and GM-CSF and challenge with Pb265, cells, especially monocytes, demonstrated an increase in Dectin-1 expression. Both types of cells treated with the cytokines exhibited a decreased fungal recovery and, conversely, an increased production of H2O2. However, when cultures were treated with an anti-Dectin-1 monoclonal antibody, to block the P. brasiliensis binding, a decrease in H2O2 production and an increase in fungal recovery were detected. This effect was observed in all cultures treated with the specific monoclonal antibody. These results show the involvement of the Dectin-1 receptor in fungal recognition and its consequent participation in the induction of the killing mechanisms against P. brasiliensis.


Assuntos
Citocinas/farmacologia , Lectinas Tipo C/metabolismo , Paracoccidioidomicose/imunologia , Fagócitos/imunologia , Contagem de Colônia Microbiana , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Humanos , Peróxido de Hidrogênio/análise , Interferon gama/farmacologia , Lectinas Tipo C/genética , Masculino , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Paracoccidioides , Fagócitos/efeitos dos fármacos
14.
Life Sci ; 223: 47-53, 2019 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-30858120

RESUMO

Psoriasis is an inflammatory skin disease with preference for the skin and joints that occurs due to hyper-proliferation and abnormal apoptosis of keratinocytes. DUSP1 expression in dermal mesenchymal stem cells (MSCs) is obviously lower in psoriasis patients than that in healthy individuals. The present study aimed to explore the roles of DUSP1 in the proliferation and apoptosis of HaCaT cells treated with a cocktail of M5. We showed that DUSP1 was markedly reduced in psoriasis patients and M5-treated HaCaT cells compared with the control subjects. MTT and BrdU assays revealed that overexpression of DUSP1 significantly suppressed the proliferation of HaCaT cells. Furthermore, DUSP1 decreased M5-induced the upregulation of cyclin D1 and Rb. In addition, we demonstrated that forced overexpression of DUSP1 caused an augment of cell apoptosis rate, c-caspase 3 protein level and the Bax/Bcl-2 ratio. Finally, we determined that enhancing DUSP1 expression resulted in the reduction of p-ERK, p-Elk-1 and Egr-1 protein levels using western blot, and the Chromatin immunoprecipitation (ChIP) assay displayed that p-Elk-1 binds to the promoter of Egr-1 in HaCaT cells. The roles of DUSP1 in cell proliferation and apoptosis were abolished by overexpression of Egr-1. In summary, gain function of DUSP1 regulates proliferation and apoptosis of HaCaT cells through the ERK/Elk-1/Egr-1 signaling pathway.


Assuntos
Apoptose/fisiologia , Proliferação de Células/fisiologia , Fosfatase 1 de Especificidade Dupla/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Queratinócitos/patologia , Sistema de Sinalização das MAP Quinases/fisiologia , Psoríase/patologia , Proteínas Elk-1 do Domínio ets/metabolismo , Apoptose/efeitos dos fármacos , Estudos de Casos e Controles , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Meios de Cultura , Citocinas/farmacologia , Fosfatase 1 de Especificidade Dupla/genética , Proteína 1 de Resposta de Crescimento Precoce/genética , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Psoríase/metabolismo , Pele/metabolismo , Pele/patologia
15.
Nanomedicine ; 18: 66-77, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30831276

RESUMO

Taking advantages of drug delivery system, immunostimulatory and chemotherapeutic agents with different physiochemical properties can be co-delivered to realize synergistic antitumor effect. Here the immunostimulatory cytokine interleukin-2 (IL-2) was firstly adsorbed in doxorubicin (DOX) loaded nanovesicles (NV-DOXIL-2) with high encapsulation efficiency by a facile solvent free method. After intravenous injection to melanoma bearing mice, NV-DOXIL-2 can accumulate in tumor and remarkably suppress tumor growth with negligible systemic toxicity. To extend the comprehensive application of this strategy, interferon-γ (IFN-γ) was further introduced to the combinatorial system to develop cytokine cocktails adsorbed NVs. This kind of NVs can significantly inhibit the primary tumor growth and lung metastasis of triple-negative breast cancer. With exploration of underlying mechanism, the cytokine cocktails adsorbed NVs can facilitate maturation of dendritic cells, promote the infiltration and activation of CD8+ T lymphocytes and natural killer cells, and increase the recruitment of CD45+ immune cells and Ly6G+ neutrophils.


Assuntos
Citocinas/uso terapêutico , Doxorrubicina/uso terapêutico , Imunoterapia , Nanopartículas/química , Neoplasias/imunologia , Neoplasias/terapia , Animais , Linhagem Celular Tumoral , Citocinas/farmacologia , Doxorrubicina/farmacologia , Composição de Medicamentos , Interferon gama/farmacologia , Interferon gama/uso terapêutico , Interleucina-2/farmacologia , Interleucina-2/uso terapêutico , Neoplasias Pulmonares/secundário , Camundongos , Nanopartículas/ultraestrutura , Neoplasias/tratamento farmacológico
16.
Toxicol In Vitro ; 58: 51-59, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30876886

RESUMO

Many drugs can induce liver injury, characterized by hepatocellular, cholestatic or mixed hepatocellular-cholestatic lesions. While an inflammatory stress is known to aggravate hepatocellular injury caused by some drugs much less evidence exists for cholestatic features. In this study, the influence of pro-inflammatory cytokines (IL-6, IL-1ß and TNF-α), either individually or combined, on cytotoxic and cholestatic properties of antibiotics was evaluated using differentiated HepaRG cells. Six antibiotics of various chemical structures and known to cause cholestasis and/or hepatocellular injury in clinic were investigated. Caspase-3 activity was increased with all these tested hepatotoxic drugs and except with erythromycin, was further augmented in presence of cytokines mainly when these were co-added as a mixture. TNF-α and IL-1ß aggravated cytotoxicity of TVX more than IL-6. Bile canaliculi (BC) dilatation induced by cholestatic drugs was increased by co-treatment with IL-6 and IL-1ß but not with TNF-α. Reduced accumulation of carboxy-dichlorofluorescein, a substrate of the multi-drug resistance-associated protein 2, in antibiotic-induced dilatated BC, was further extended in presence of individual or mixed cytokines. In conclusion, our data demonstrate that pro-inflammatory cytokines either individually or in mixture, can modulate cholestatic and/or cytotoxic responses to antibiotics and that the extent of these effects is dependent on the cytokine and the cholestatic antibiotic.


Assuntos
Antibacterianos/efeitos adversos , Canalículos Biliares/efeitos dos fármacos , Colestase/induzido quimicamente , Citocinas/farmacologia , Canalículos Biliares/fisiologia , Proteína C-Reativa/metabolismo , Caspase 3/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Colestase/metabolismo , Fluoresceínas/metabolismo , Humanos
17.
Bull Exp Biol Med ; 166(4): 444-447, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30788736

RESUMO

The effects of a natural complex of cytokines IL-1, IL-2, IL-6, TNF, MIF, and GTFß on myocardial blood flow were studied under control conditions and during acute experimental aortal stenosis. Systemic administration of the cytokine complex under control conditions led to moderate impairment of the blood flow in the myocardium associated with plethora and perivascular edema. The number of functioning vessels in the myocardium significantly increased under these conditions, which reflected enhancement of the coronary blood flow. The comparison of the myocardial blood flow under conditions of acute heart overload alone and in combination with systemic administration of the cytokine complex revealed similar changes. In both cases, moderate plethora in all compartments of the vascular network, moderate perivascular edema, and moderate blood stasis in the myocardial capillaries were seen. The only difference the increase in the density of functioning capillaries that was significantly more pronounced after cytokine administration. These data indicate that the increase in the blood cytokine level induced dilatation of myocardial vessels and intensification of blood flow in normal and under conditions of acute hemodynamic heart overload. Against the background of pronounced vasodilatation, the dyscirculatory changes in the myocardium were moderate. It was assumed that the increase in the duration or frequency of hypercytokinemia episodes can induce more severe blood flow disturbances in the myocardium.


Assuntos
Circulação Coronária/efeitos dos fármacos , Citocinas/farmacologia , Coração/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Miocárdio/metabolismo , Animais , Estenose da Valva Aórtica/metabolismo , Estenose da Valva Aórtica/fisiopatologia , Feminino , Cobaias , Interleucina-1/farmacologia , Interleucina-2/farmacologia , Interleucina-6/farmacologia , Masculino , Consumo de Oxigênio/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia
18.
Mol Biol Cell ; 30(5): 566-578, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30625033

RESUMO

Junctional adhesion molecule-A (JAM-A), an epithelial tight junction protein, plays an important role in regulating intestinal permeability through association with a scaffold signaling complex containing ZO-2, Afadin, and the small GTPase Rap2. Under inflammatory conditions, we report that the cytoplasmic tail of JAM-A is tyrosine phosphorylated (p-Y280) in association with loss of barrier function. While barely detectable Y280 phosphorylation was observed in confluent monolayers of human intestinal epithelial cells under basal conditions, exposure to cytokines TNFα, IFNγ, IL-22, or IL-17A, resulted in compromised barrier function in parallel with increased p-Y280. Phosphorylation was Src kinase dependent, and we identified Yes-1 and PTPN13 as a major kinase and phosphatase for p-JAM-A Y280, respectively. Moreover, cytokines IL-22 or IL-17A induced increased activity of Yes-1. Furthermore, the Src kinase inhibitor PP2 rescued cytokine-induced epithelial barrier defects and inhibited phosphorylation of JAM-A Y280 in vitro. Phosphorylation of JAM-A Y280 and increased permeability correlated with reduced JAM-A association with active Rap2. Finally, we observed increased phosphorylation of Y280 in colonic epithelium of individuals with ulcerative colitis and in mice with experimentally induced colitis. These findings support a novel mechanism by which tyrosine phosphorylation of JAM-A Y280 regulates epithelial barrier function during inflammation.


Assuntos
Células Epiteliais/metabolismo , Inflamação/patologia , Intestinos/patologia , Molécula A de Adesão Juncional/metabolismo , Fosfotirosina/metabolismo , Sequência de Aminoácidos , Animais , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/metabolismo , Colite Ulcerativa/patologia , Citocinas/farmacologia , Sulfato de Dextrana , Células HEK293 , Humanos , Intestinos/química , Camundongos Endogâmicos C57BL , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Proteína Tirosina Fosfatase não Receptora Tipo 13/metabolismo , Proteínas Proto-Oncogênicas c-yes/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo , Proteínas rap de Ligação ao GTP/metabolismo
19.
Domest Anim Endocrinol ; 67: 42-53, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30690257

RESUMO

Peroxisome proliferator-activated receptors (PPARs) are members of a nuclear receptor family of ligand-dependent transcription factors. Three isoforms of PPAR named PPARα, PPARß/δ, and PPARγ have been described, each encoded by a separate gene: PPARA, PPARD, and PPARG, respectively. In the present study, we examined the profiles of PPAR and retinoid X receptor (RXR; PPAR heterodimer partner) mRNA expression and PPAR DNA binding activity in porcine trophoblast tissue collected on days 15, 20, 25, and 30 of pregnancy and in day-20 embryos. Placenta trophoblast cells isolated on day 25 of pregnancy were used to determine effects of (1) cytokines on PPAR and RXR mRNA expression and (2) PPAR agonists on prostaglandin (PG) E2 synthesis and the expression of genes involved in steroidogenesis, fatty acid binding, and PG transport, as well as on cell proliferation. The mRNA expression of PPARA and RXRB was greater in trophoblast tissue collected on days 25 and 30 of pregnancy compared with day 15 (P < 0.05), while DNA binding activity of PPARα decreased between day 15 and 25 (P < 0.05). Increased concentrations of PPARD and RXRA transcripts were observed in trophoblasts collected on day 20 compared to trophoblasts from days 15 and 30 (P < 0.05). Moreover, concentrations of DNA-bound PPARß/δ and PPARγ proteins increased in day-30 trophoblasts compared to day 15 (P < 0.01) and day 20 (P < 0.05), respectively. On day 20 of gestation, the mRNA expression of PPARD, PPARG, and RXRA and protein levels of PPARα and PPARγ isoforms were greater in trophoblast than embryonic tissue (P < 0.01). Interleukin 1ß and/or interferon γ, but not IL6 and leukemia inhibitory factor, upregulated PPAR and RXR mRNA expression in placenta trophoblast cells in vitro (P < 0.05). Rosiglitazone (a PPARγ agonist) stimulated prostaglandin E synthase mRNA expression in trophoblast cells and PGE2 accumulation in incubation medium (P < 0.05). Moreover, activation of PPAR isoforms differentially affected the expression of genes involved in steroidogenesis, fatty acid binding, and PG transport in studied cells. Finally, PPARα and PPARγ agonists stimulated trophoblast cell proliferation (P < 0.05), and this effect was abolished by the addition of a respective PPAR antagonist (P < 0.05). Overall, these results point to a role of PPAR isoforms in porcine placenta development and function.


Assuntos
Receptores Ativados por Proliferador de Peroxissomo/genética , Receptores Ativados por Proliferador de Peroxissomo/fisiologia , Sus scrofa/embriologia , Trofoblastos/química , Trofoblastos/fisiologia , Animais , Proliferação de Células/efeitos dos fármacos , Citocinas/farmacologia , DNA/metabolismo , Dinoprostona/biossíntese , Implantação do Embrião/fisiologia , Embrião de Mamíferos/química , Embrião de Mamíferos/fisiologia , Feminino , Expressão Gênica/efeitos dos fármacos , Placenta , Placentação/fisiologia , Gravidez , RNA Mensageiro/análise , Receptores X Retinoide/genética , Sus scrofa/fisiologia , Trofoblastos/citologia
20.
Anatol J Cardiol ; 21(2): 91-97, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30694801

RESUMO

OBJECTIVE: Omentin is a recently identified novel adipocytokine mainly expressed in the epicardial adipose tissue. Although it has favorable effects on cardiovascular disease, the impact of omentin on the hearts is still an understudied issue. The aim of the present study was to investigate the possible effects of omentin on isolated rat heart. METHODS: Using the Langendorff method, 28 adult male Sprague-Dawley rat hearts were isolated and perfused with modified Krebs-Henseleit solution (mK-Hs). Concentrations of 100, 200, and 400 ng/mL omentin were given to the hearts for 30 min. The control group (n=7) was perfused with mK-Hs alone. Gene expressions in the left ventricle tissue were determined by real-time polymerase chain reaction. Left ventricular cyclic adenosine monophosphate and cyclic guanosine monophosphate (cGMP) concentrations were determined by using enzyme-linked immunosorbent assay. RESULTS: All concentrations of omentin significantly decreased left ventricular developed pressure and maximal rate of pressure development that are the indexes of cardiac contractility. At the same time, omentin decreased both phosphoinositide 3-kinase γ (PI3Kγ) and sarcolemmal L-type Ca2+ channel (CaV1.2) mRNA levels. Moreover, this peptide at concentrations of 200 and 400 ng/mL increased endothelial nitric oxide synthase (eNOS) mRNA. Furthermore, concentrations of 200 and 400 ng/mL omentin increased the amount of cGMP. CONCLUSION: We conclude that acute omentin treatment decreases cardiac contractility. Elevated eNOS mRNA and cGMP levels with reduced CaV1.2 mRNA are likely to lead to negative inotropy.


Assuntos
Citocinas/genética , Citocinas/farmacologia , Ventrículos do Coração , Lectinas/genética , Lectinas/farmacologia , Contração Miocárdica/efeitos dos fármacos , Animais , Regulação da Expressão Gênica , Masculino , Reação em Cadeia da Polimerase , Ratos , Ratos Sprague-Dawley
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