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1.
Nat Biomed Eng ; 3(10): 817-829, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31358881

RESUMO

Homeostatic antigen presentation by hepatic antigen-presenting cells, which results in tolerogenic T-cell education, could be exploited to induce antigen-specific immunological tolerance. Here we show that antigens modified with polymeric forms of either N-acetylgalactosamine or N-acetylglucosamine target hepatic antigen-presenting cells, increase their antigen presentation and induce antigen-specific tolerance, as indicated by CD4+ and CD8+ T-cell deletion and anergy. These synthetically glycosylated antigens also expanded functional regulatory T cells, which are necessary for the durable suppression of antigen-specific immune responses. In an adoptive-transfer mouse model of type-1 diabetes, treatment with the glycosylated autoantigens prevented T-cell-mediated diabetes, expanded antigen-specific regulatory T cells and resulted in lasting tolerance to a subsequent challenge with activated diabetogenic T cells. Glycosylated autoantigens targeted to hepatic antigen-presenting cells might enable therapies that promote immune tolerance in patients with autoimmune diseases.


Assuntos
Acetilgalactosamina/imunologia , Acetilgalactosamina/farmacologia , Acetilglucosamina/imunologia , Acetilglucosamina/farmacologia , Apresentação do Antígeno/efeitos dos fármacos , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/imunologia , Tolerância Imunológica/efeitos dos fármacos , Transferência Adotiva , Animais , Apresentação do Antígeno/imunologia , Autoantígenos/farmacologia , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Modelos Animais de Doenças , Feminino , Fígado/efeitos dos fármacos , Fígado/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Baço , Linfócitos T/efeitos dos fármacos
2.
J Pharmacol Sci ; 139(3): 193-200, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30827890

RESUMO

Ischemia/reperfusion (I/R)-induced oxidative stress is a serious clinical problem in the reperfusion therapy for ischemic diseases. Tumstatin is an endogenous bioactive peptide cleaved from type IV collagen α3 chain. We previously reported that T3 peptide, an active subfragment of tumstatin, exerts cytoprotective effects on H2O2-induced apoptosis through the inhibition of intracellular reactive oxygen species (ROS) production in H9c2 cardiomyoblasts. In this study, we investigated whether T3 peptide has cardioprotective effects against I/R injury by using in vitro and ex vivo experimental models. H9c2 cardiomyoblasts were stimulated with oxygen and glucose deprivation (OGD) for 12 h followed by reoxygenation for 1-8 h (OGD/R; in vitro model). The cells were treated with T3 peptide (30-1000 ng/ml) during OGD. Ten minutes after the pre-perfusion of T3 peptide (300 ng/ml), Langendorff perfused rat hearts were exposed to ischemia for 30 min followed by reperfusion for 1 h (ex vivo model). T3 peptide inhibited OGD/R-induced apoptosis through the inhibition of mitochondrial ROS production and dysfunction in H9c2 cardiomyoblasts. T3 peptide also prevented I/R-induced cardiac dysfunction, arrhythmia and myocardial infarction in the perfused rat heart. In conclusion, we for the first time demonstrated that T3 peptide exerts cardioprotective effects against I/R injury.


Assuntos
Apoptose/efeitos dos fármacos , Autoantígenos/administração & dosagem , Cardiotônicos/administração & dosagem , Colágeno Tipo IV/administração & dosagem , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Animais , Arritmias Cardíacas/etiologia , Arritmias Cardíacas/prevenção & controle , Autoantígenos/química , Autoantígenos/farmacologia , Cardiotônicos/farmacologia , Linhagem Celular , Colágeno Tipo IV/química , Colágeno Tipo IV/farmacologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Masculino , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/prevenção & controle , Traumatismo por Reperfusão Miocárdica/complicações , Peptídeos/administração & dosagem , Peptídeos/farmacologia , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
3.
Haematologica ; 104(4): 669-677, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30442724

RESUMO

Myelosuppression is a major and frequently dose-limiting side effect of anticancer therapy and is responsible for most treatment-related morbidity and mortality. In addition, repeated cycles of DNA damage and cell death of hematopoietic stem and progenitor cells, followed by compensatory proliferation and selection pressure, lead to genomic instability and pave the way for therapy-related myelodysplastic syndromes and secondary acute myeloid leukemia. Protection of hematopoietic stem and progenitor cells from chemo- and radiotherapy in patients with solid tumors would reduce both immediate complications and long-term sequelae. Epidermal growth factor (EGF) and prostaglandin E2 (PGE2) were reported to prevent chemo- or radiotherapy-induced myelosuppression in mice. We tested both molecules for potentially protective effects on human CD34+ cells in vitro and established a xenograft mouse model to analyze stress resistance and regeneration of human hematopoiesis in vivo EGF was neither able to protect human stem and progenitor cells in vitro nor to promote hematopoietic regeneration following sublethal irradiation in vivo PGE2 significantly reduced in vitro apoptotic susceptibility of human CD34+ cells to taxol and etoposide. This could, however, be ascribed to reduced proliferation rather than to a change in apoptosis signaling and BCL-2 protein regulation. Accordingly, 16,16-dimethyl-PGE2 (dmPGE2) did not accelerate regeneration of the human hematopoietic system in vivo Repeated treatment of sublethally irradiated xenograft mice with known antiapoptotic substances, such as human FLT3L and thrombopoietin (TPO), which suppress transcription of the proapoptotic BCL-2 proteins BIM and BMF, also only marginally promoted human hematopoietic regeneration in vivo.


Assuntos
Autoantígenos/farmacologia , Dinoprostona/farmacologia , Fator de Crescimento Epidérmico/farmacologia , Hematopoese/efeitos dos fármacos , Iodeto Peroxidase/farmacologia , Proteínas de Ligação ao Ferro/farmacologia , Proteínas de Membrana/farmacologia , Animais , Avaliação de Medicamentos , Humanos , Camundongos , Camundongos Knockout
4.
Pharmazie ; 73(12): 715-720, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30522555

RESUMO

Tumstatin7 (CNYYSNS) is an antitumor peptide derived from the NC1 domain of Type IV collagen that has been associated with tumor angiogenesis. In this work, we generated a peptide composed of tumstatin7 fused to TAT, a cell-internalizing peptide consisting of 11 amino acids. Tumstatin7-TAT was internalized by cells and triggered cell death. The new peptide was more potent in inducing B16F10 melanoma cell apoptosis in vitro than the shorter tumstatin7. Whereas tumstatin7-TAT significantly reduced tumor cell viability, tumstatin7 showed only weak effects even at the highest treatment concentration applied. Both tumstatin7-TAT and tumstatin7 inhibited cell migration in an in vitro wound healing model, and the former was more effective than the latter in inhibiting tumor growth in vivo. Combining the cell-internalizing property of TAT with the tumor-specific property of tumstatin7 may provide a useful adjunct to tumor therapy.


Assuntos
Autoantígenos/farmacologia , Colágeno Tipo IV/farmacologia , Produtos do Gene tat/metabolismo , Melanoma Experimental/tratamento farmacológico , Peptídeos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Autoantígenos/administração & dosagem , Autoantígenos/química , Movimento Celular/efeitos dos fármacos , Colágeno Tipo IV/administração & dosagem , Colágeno Tipo IV/química , Feminino , Humanos , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/administração & dosagem , Peptídeos/química , Cicatrização/efeitos dos fármacos
5.
Biologicals ; 56: 45-53, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30327235

RESUMO

The present work elucidates the production of recombinant human asparaginase (rhASP) under optimized fermentation and downstream processes in Escherichia coli. The maximum biomass yield of 6.7 g/L was achieved with fed-batch fermentation. The highest rhASP inclusion bodies recovery yield (91%) was achieved with the optimized lysis conditions. The 8.0 M urea at pH 8.5 has shown efficient solubilization (94%) of rhASP inclusion bodies. The refolding efficiency of rhASP increased at pH 8.5 (84%) and temperature 25°C (86%). The diluted rhASP solution was concentrated and partially purified (92%) using cross flow filtration. A single step ion exchange chromatography is successfully achieved the maximum purity of ≥ 97%. The molecular mass of purified rhASP is confirmed as 34.1 kDa by mass spectrometry. The secondary structure of rhASP is characterized by FT-IR spectroscopy based on the structural elements. Finally, cell proliferative assay of purified rhASP is signifies the similar biological activity over the standard.


Assuntos
Asparaginase/biossíntese , Autoantígenos/biossíntese , Proteínas Recombinantes/biossíntese , Asparaginase/química , Asparaginase/isolamento & purificação , Asparaginase/farmacologia , Autoantígenos/química , Autoantígenos/isolamento & purificação , Autoantígenos/farmacologia , Técnicas de Cultura Celular por Lotes , Proliferação de Células/efeitos dos fármacos , Cromatografia por Troca Iônica , Escherichia coli , Fermentação , Humanos , Corpos de Inclusão/enzimologia , Redobramento de Proteína , Estrutura Secundária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologia
6.
J Theor Biol ; 455: 212-221, 2018 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-30036530

RESUMO

Cooperation between VEGFR2 and integrin αVß3 is critical for neovascularization in wound healing, cardiovascular ischemic diseases, ocular diseases, and tumor angiogenesis. In the present study, we developed a rule-based computational model to investigate the potential mechanism by which the Src-induced integrin association with VEGFR2 enhances VEGFR2 activation. Simulations demonstrated that the main function of integrin is to reduce the degradation of VEGFR2 and hence stabilize the activation signal. In addition, receptor synthesis rate and recruitment from internal compartment were found to be sensitive determinants of the activation state of VEGFR2. The model was then applied to simulate the effect of integrin-binding peptides such as tumstatin and cilengitide on VEGFR2 signaling. Further, computational modeling proposed potential molecular mechanisms for the angiogenesis-modulating activity of other integrin-binding peptides. The model highlights the complexity of the crosstalk between αVß3 integrin and VEGFR2 and the necessity of utilizing models to elucidate potential mechanisms in angiogenesis-modulating peptide therapy.


Assuntos
Autoantígenos , Colágeno Tipo IV , Células Endoteliais/metabolismo , Integrina alfaVbeta3/metabolismo , Modelos Biológicos , Neovascularização Patológica , Transdução de Sinais/efeitos dos fármacos , Venenos de Serpentes , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Autoantígenos/farmacologia , Colágeno Tipo IV/farmacocinética , Colágeno Tipo IV/farmacologia , Humanos , Proteínas de Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Venenos de Serpentes/farmacocinética , Venenos de Serpentes/farmacologia
7.
Brain Struct Funct ; 223(7): 3463-3471, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29936552

RESUMO

The antiphospholipid syndrome (APS) is an autoimmune disease characterized by the presence of antiphospholipid antibodies, which may trigger vascular thrombosis with consecutive infarcts. However, cognitive dysfunctions representing one of the most commonest neuropsychiatric symptoms are frequently present despite the absence of any ischemic brain lesions. Data on the structural and functional basis of the neuropsychiatric symptoms are sparse. To examine the effect of APS on hippocampal neurogenesis and on white matter, we induced experimental APS (eAPS) in adult female Balb/C mice by immunization with ß2-glycoprotein 1. To investigate cell proliferation in the dentate gyrus granular cell layer (DG GCL), eAPS and control mice (n = 5, each) were injected with 5-bromo-2'-deoxyuridine (BrdU) once a day for 10 subsequent days. Sixteen weeks after immunization, eAPS resulted in a significant reduction of BrdU-positive cells in the DG GCL compared to control animals. However, double staining with doublecortin and NeuN revealed a largely preserved neurogenesis. Ultrastructural analysis of corpus callosum (CC) axons in eAPS (n = 6) and control mice (n = 7) revealed no significant changes in CC axon diameter or g-ratio. In conclusion, decreased cellular proliferation in the hippocampus of eAPS mice indicates a limited regenerative potential and may represent one neuropathological substrate of cognitive changes in APS while evidence for alterations of white matter integrity is lacking.


Assuntos
Síndrome Antifosfolipídica/induzido quimicamente , Síndrome Antifosfolipídica/patologia , Proliferação de Células , Giro Denteado/patologia , Animais , Anticorpos Antifosfolipídeos/metabolismo , Autoantígenos/farmacologia , Escala de Avaliação Comportamental , Bromodesoxiuridina/administração & dosagem , Bromodesoxiuridina/metabolismo , Diferenciação Celular/fisiologia , Corpo Caloso/ultraestrutura , Modelos Animais de Doenças , Feminino , Fluorescência , Camundongos , Camundongos Endogâmicos BALB C , Neurogênese , Radiossensibilizantes/administração & dosagem , Radiossensibilizantes/metabolismo , beta 2-Glicoproteína I/farmacologia
8.
J Nutr Biochem ; 58: 71-79, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29885599

RESUMO

Selective inhibition of T cells has been implied to prevent and/or treat autoimmune and inflammatory diseases. Some food compounds that have such immune-modulating functions may serve as nutritional approach to this purpose. In this study, we chose naringenin, a citrus fruits-derived compound with antiinflammatory property, to test this possibility. In this in vitro study, we stimulated mouse T cells with anti-CD3/CD28 (polyclonal TCR activation) or autoantigen MOG35-55 in the presence of naringenin. We found that naringenin dose-dependently suppressed anti-CD3/CD28 and MOG35-55-induced T cell proliferation, production of T cell cytokines IFN-γ, IL-17, IL-6 and TNF-α. We further showed that inhibited T cell proliferation was associated with T cell cycle arrest at G0/G1 phase, which was in turn related to delayed degradation of the cyclin-dependent kinase inhibitor p27kip1 and the down-regulation of retinoblastoma protein phosphorylation in activated T cells. Finally, it was revealed that all these T cell-suppressive effects might be related to naringenin's interference with IL-2/IL-2R-mediated signaling pathway and STAT5 phosphorylation in activated T cells. Our results confirmed T cell-suppressive activity of naringenin previously reported by us and others, but for the first time, it was shown that the working mechanism may involve its ability to modulate cell cycle progression, cell cycle-related proteins and IL-2/IL-2R signaling pathway. Together, these results further support proposed potential of naringenin being a preventive/therapeutic agent in T-cell-mediated autoimmune inflammatory disorders.


Assuntos
Flavanonas/farmacologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Animais , Autoantígenos/farmacologia , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Citocinas/metabolismo , Relação Dose-Resposta a Droga , Encefalomielite Autoimune Experimental/patologia , Feminino , Flavanonas/administração & dosagem , Interleucina-2/metabolismo , Interleucina-2/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Receptores de Interleucina-2/metabolismo , Proteína do Retinoblastoma/metabolismo , Fator de Transcrição STAT5/metabolismo , Linfócitos T/imunologia
9.
Calcif Tissue Int ; 102(5): 522-532, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29204673

RESUMO

Many autoimmune diseases are associated with deranged bone metabolism. The resulting localized or systemic bone loss can compromise the quality of life of patients by causing local bone deformities or fragility fractures. There is emerging evidence that antibodies have a direct impact on key players of bone homeostasis, in particular osteoclasts. Clinical and pre-clinical studies provide insight into the function of autoantibodies related to Rheumatoid Arthritis (rheumatoid factor, anti-citrullinated protein antibodies, and anti-carbamylated protein antibodies) and their inflammation-independent interaction with bone cells. Furthermore, we summarize the current knowledge about neutralizing antibodies to the antiresorptive protein osteoprotegerin, which have been described in patients with Coeliac Disease, Rheumatoid Arthritis, and Spondyloarthritis.


Assuntos
Autoanticorpos/imunologia , Autoantígenos/farmacologia , Doenças Ósseas Metabólicas/tratamento farmacológico , Reabsorção Óssea/tratamento farmacológico , Osso e Ossos/metabolismo , Animais , Anticorpos Anti-Proteína Citrulinada/uso terapêutico , Doenças Ósseas Metabólicas/imunologia , Reabsorção Óssea/imunologia , Osso e Ossos/imunologia , Humanos
10.
J Cell Mol Med ; 21(12): 3288-3297, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28608951

RESUMO

The extracellular matrix (ECM) creates the microenvironment of the tissue; an altered ECM in the asthmatic airway may be central in airway inflammation and remodelling. Tumstatin is a collagen IV-derived matrikine reduced in the asthmatic airway wall that reverses airway inflammation and remodelling in small and large animal models of asthma. This study hypothesized that the mechanisms underlying the broad asthma-resolving effects of tumstatin were due to autocrine remodelling of the ECM. Neutrophils and endothelial cells were seeded on decellularized ECM of non-asthmatic (NA) or asthmatic (A) airway smooth muscle (ASM) cells previously exposed to tumstatin in the presence or absence of a broad matrix metalloproteinase inhibitor, Marimastat. Gene expression in NA and A ASM induced by tumstatin was assessed using RT-PCR arrays. The presence of tumstatin during ECM deposition affected neutrophil and endothelial cell properties on both NA and A ASM-derived matrices and this was only partly due to MMP activity. Gene expression patterns in response to tumstatin in NA and A ASM cells were different. Tumstatin may foster an anti-inflammatory and anti-angiogenic microenvironment by modifying ASM-derived ECM. Further work is required to examine whether restoring tumstatin levels in the asthmatic airway represents a potential novel therapeutic approach.


Assuntos
Inibidores da Angiogênese/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Autoantígenos/farmacologia , Colágeno Tipo IV/farmacologia , Matriz Extracelular/efeitos dos fármacos , Metaloproteinases da Matriz/genética , Miócitos de Músculo Liso/efeitos dos fármacos , Remodelação das Vias Aéreas , Asma/genética , Asma/metabolismo , Asma/patologia , Brônquios/efeitos dos fármacos , Brônquios/metabolismo , Brônquios/patologia , Quimiotaxia/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Matriz Extracelular/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Ácidos Hidroxâmicos/farmacologia , Interleucina-8/farmacologia , Metaloproteinases da Matriz/metabolismo , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Neutrófilos/patologia
11.
Mol Cell Neurosci ; 80: 89-99, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28254618

RESUMO

Parkin is an E3 ubiquitin ligase whose mutations cause autosomal recessive juvenile Parkinson's disease (PD). Unlike the human phenotype, parkin knockout (KO) mice show no apparent dopamine neuron degeneration, although they demonstrate reduced expression and activity of striatal mitochondrial proteins believed to be necessary for neuronal survival. Instead, parkin-KO mice show reduced striatal evoked dopamine release, abnormal synaptic plasticity, and non-motor symptoms, all of which appear to mimic the preclinical features of Parkinson's disease. Extensive studies have screened candidate synaptic proteins responsible for reduced evoked dopamine release, and synaptotagmin XI (Syt XI), an isoform of Syt family regulating membrane trafficking, has been identified as a substrate of parkin in humans. However, its expression level is unaltered in the striatum of parkin-KO mice. Thus, the target(s) of parkin and the molecular mechanisms underlying the impaired dopamine release in parkin-KO mice remain unknown. In this study, we focused on Syt IV because of its highly homology to Syt XI, and because they share an evolutionarily conserved lack of Ca2+-binding capacity; thus, Syt IV plays an inhibitory role in Ca2+-dependent neurotransmitter release in PC12 cells and neurons in various brain regions. We found that a proteasome inhibitor increased Syt IV protein, but not Syt XI protein, in neuron-like, differentiated PC12 cells, and that parkin interacted with and polyubiquitinated Syt IV, thereby accelerating its protein turnover. Parkin overexpression selectively degraded Syt IV protein, but not Syt I protein (indispensable for Ca2+-dependent exocytosis), thus enhancing depolarization-dependent exocytosis. Furthermore, in parkin-KO mice, the level of striatal Syt IV protein was increased. Our data indicate a crucial role for parkin in the proteasomal degradation of Syt IV, and provide a potential mechanism of parkin-regulated, evoked neurotransmitter release.


Assuntos
Neurônios/metabolismo , Proteólise , Sinaptotagminas/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/fisiologia , Animais , Autoantígenos/farmacologia , Células COS , Chlorocebus aethiops , Corpo Estriado/citologia , Exocitose/genética , Proteínas de Membrana/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Crescimento Neural/farmacologia , Oligopeptídeos/farmacologia , Células PC12/efeitos dos fármacos , Células PC12/ultraestrutura , Inibidores de Proteassoma/farmacologia , Transporte Proteico , Proteólise/efeitos dos fármacos , Ratos , Sinaptotagminas/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação/efeitos dos fármacos , Proteína 2 Associada à Membrana da Vesícula/metabolismo
12.
Biomaterials ; 118: 51-62, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27940382

RESUMO

Autoimmune diseases occur when the immune system incorrectly recognizes self-molecules as foreign; in the case of multiple sclerosis (MS), myelin is attacked. Intriguingly, new studies reveal toll-like receptors (TLRs), pathways usually involved in generating immune responses against pathogens, play a significant role in driving autoimmune disease in both humans and animal models. We reasoned polyplexes formed from myelin self-antigen and regulatory TLR antagonists might limit TLR signaling during differentiation of myelin-specific T cells, inducing tolerance by biasing T cells away from inflammatory phenotypes. Complexes were formed by modifying myelin peptide with cationic amino acids to create peptides able to condense the anionic nucleic-acid based TLR antagonist. These immunological polyplexes eliminate synthetic polymers commonly used to condense polyplexes and do not rely on gene expression; however, the complexes mimic key features of traditional polyplexes such as tunable loading and co-delivery. Using these materials and classic polyplex analysis techniques, we demonstrate condensation of both immune signals, protection from enzymatic degradation, and tunable physicochemical properties. We show polyplexes reduce TLR signaling, and in primary dendritic cell and T cell co-culture, reduce myelin-driven inflammation. During mouse models of MS, these tolerogenic polyplexes improve the progression, severity, and incidence of disease.


Assuntos
Autoantígenos/imunologia , Autoantígenos/uso terapêutico , Autoimunidade/imunologia , Esclerose Múltipla/imunologia , Esclerose Múltipla/terapia , Ácidos Nucleicos Peptídicos/uso terapêutico , Receptores Toll-Like/imunologia , Animais , Autoantígenos/farmacologia , Autoimunidade/efeitos dos fármacos , Células Cultivadas , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Ácidos Nucleicos , Ácidos Nucleicos Peptídicos/imunologia , Ácidos Nucleicos Peptídicos/farmacologia , Transdução de Sinais , Receptores Toll-Like/antagonistas & inibidores , Resultado do Tratamento
13.
Sci Rep ; 6: 26309, 2016 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-27199164

RESUMO

Tumstatin, a protein fragment of the alpha-3 chain of Collagen IV, is known to be significantly reduced in the airways of asthmatics. Further, there is evidence that suggests a link between the relatively low level of tumstatin and the induction of angiogenesis and inflammation in allergic airway disease. Here, we show that the intra-segmental administration of tumstatin can impede the development of vascular remodelling and allergic inflammatory responses that are induced in a segmental challenge model of experimental asthma in sheep. In particular, the administration of tumstatin to lung segments chronically exposed to house dust mite (HDM) resulted in a significant reduction of airway small blood vessels in the diameter range 10(+)-20 µm compared to controls. In tumstatin treated lung segments after HDM challenge, the number of eosinophils was significantly reduced in parenchymal and airway wall tissues, as well as in the bronchoalveolar lavage fluid. The expression of VEGF in airway smooth muscle was also significantly reduced in tumstatin-treated segments compared to control saline-treated segments. Allergic lung function responses were not attenuated by tumstatin administration in this model. The data are consistent with the concept that tumstatin can act to suppress vascular remodelling and inflammation in allergic airway disease.


Assuntos
Asma/fisiopatologia , Autoantígenos/farmacologia , Colágeno Tipo IV/farmacologia , Pulmão/patologia , Remodelação Vascular/efeitos dos fármacos , Resistência das Vias Respiratórias/efeitos dos fármacos , Alérgenos/administração & dosagem , Animais , Asma/imunologia , Autoantígenos/administração & dosagem , Líquido da Lavagem Broncoalveolar/citologia , Doença Crônica , Colágeno Tipo IV/administração & dosagem , Dermatophagoides pteronyssinus/imunologia , Feminino , Inflamação/patologia , Pulmão/irrigação sanguínea , Pulmão/imunologia , Músculo Liso/metabolismo , Carneiro Doméstico , Fator A de Crescimento do Endotélio Vascular/metabolismo
14.
Oncol Rep ; 35(6): 3403-8, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27109498

RESUMO

The present study was aimed to investigate the effect of tumstatin on inhibition of proliferation and induction of apoptosis in Saos-2 human osteosarcoma cells and to understand the mechanism involved. Inhibition of cell proliferation was analyzed by MTT assay and induction of apoptosis through nuclear fragmentation assay. Viability of Saos-2 cells was reduced to 19% on treatment with 25 µM concentration of tumstatin after 48 h. Presence of characteristic apoptotic nuclei, rounded cell shape and shrunken size were caused by tumstatin treatment at 25 µM concentration. The level of mRNA corresponding to PTEN, FasR and FasL was increased significantly in tumstatin treated Saos-2 cells compared to untreated control. Investigation of the mechanism revealed NF-κB activation by phosphorylation on serine 536. The activated NF-κB was translocated into the nucleus from the cytoplasm on treatment with tumstatin. Degradation of the IκBα by tumstatin was found to be much slower compared to that induced by treatment with TNF-α. Thus, tumstatin inhibits proliferation and induces apoptosis in Saos-2 cells through activation of NF-κB and its translocation to the nucleus. Therefore, tumstatin can play an important role in the treatment of osteosarcoma.


Assuntos
Apoptose/efeitos dos fármacos , Autoantígenos/farmacologia , Neoplasias Ósseas/tratamento farmacológico , Colágeno Tipo IV/farmacologia , Osteossarcoma/tratamento farmacológico , Fator de Transcrição RelA/metabolismo , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Osteossarcoma/patologia , PTEN Fosfo-Hidrolase/genética , Fosforilação
15.
Reprod Biol ; 15(3): 163-71, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26370459

RESUMO

Exposure to di-(2-ethylhexyl) phthalate (DEHP) induces spermatogenic disturbance (SD) through oxidative stress, and affects the immune system by acting as an adjuvant. Recently, we reported that in mice, a low dose of DEHP, which did not affect spermatogenesis, was able to alter the testicular immune microenvironment. Experimental autoimmune orchitis (EAO) can be induced by repeated immunization with testicular antigens, and its pathology is characterized by production of autoantibodies and SD. In the present study, we investigated the effect of a low-dose DEHP on the susceptibility of mice to EAO. The exposure to DEHP-containing feed (0.01%) caused a modest functional damage to the blood-testis barrier (BTB) with an increase in testicular number of interferon gamma (IFN-γ)-positive cells and resulted in the production of autoantibodies targeting haploid cells, but did not affect spermatogenesis. While only single immunization with testicular antigens caused very mild EAO, the concurrent DEHP exposure induced severe EAO with significant increases in number of interferon gamma-positive cells and macrophages, as well as lymphocytic infiltration and serum autoantibody titer accompanied by severe SD. To summarize, the exposure of mice to the low-dose DEHP does not induce significant SD, but it may cause an increase in IFN-γ positive cells and modest functional damage to the BTB in the testis. These changes lead to an autoimmune response against haploid cell autoantigens, resulting in increased susceptibility to EAO.


Assuntos
Doenças Autoimunes/induzido quimicamente , Autoimunidade/efeitos dos fármacos , Dietilexilftalato/farmacologia , Orquite/induzido quimicamente , Testículo/efeitos dos fármacos , Animais , Autoantígenos/farmacologia , Masculino , Camundongos , Testículo/imunologia
16.
Sci Rep ; 5: 14150, 2015 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-26412210

RESUMO

Anti-cytokine therapeutic antibodies have been demonstrated to be effective in the treatment of several auto-immune disorders. However, The problems in antibody manufacture and the immunogenicity caused by multiple doses of antibodies inspire people to use auto-cytokine as immunogen to induce anti-cytokine antibodies. Nevertheless, the tolerance for inducing immune response against self-antigen has hindered the wide application of the strategy. To overcome the tolerance, here we proposed a strategy using the inter-species cytokine as immunogen for active immunization (TISCAI) to induce anti-cytokine antibody. As a proof of concept, an inter-species cytokine RANKL was successfully used as immunogen to induce anti-RANKL immune response. Furthermore, to prevent undesirable side-effects, the human RANKL was mutated based on the crystal structure of the complex of human RANKL and its rodent counterpart receptor RANK. We found, the antibodies produced blocked the osteoclast development in vitro and osteoporosis in OVX rat models. The results demonstrated this strategy adopted is very useful for general anti-cytokine immunotherapy for different diseases settings.


Assuntos
Imunoterapia , Osteoporose/genética , Osteoporose/imunologia , Ligante RANK/genética , Ligante RANK/imunologia , Vacinas , Animais , Anticorpos/imunologia , Autoantígenos/imunologia , Autoantígenos/farmacologia , Reabsorção Óssea/genética , Reabsorção Óssea/metabolismo , Diferenciação Celular/efeitos dos fármacos , Modelos Animais de Doenças , Humanos , Imunização , Camundongos , Modelos Moleculares , Osteoclastos/citologia , Osteoclastos/imunologia , Osteoclastos/metabolismo , Osteoporose/diagnóstico , Osteoporose/terapia , Ovariectomia , Ligação Proteica , Conformação Proteica , Ligante RANK/química , Ligante RANK/metabolismo , Ratos , Receptor Ativador de Fator Nuclear kappa-B/química , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Microtomografia por Raio-X
17.
J Thromb Haemost ; 13(8): 1479-93, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26039631

RESUMO

BACKGROUND: Aspirin and P2Y12 antagonists are antiplatelet compounds that are used clinically in patients with thrombosis. However, some patients are 'resistant' to antiplatelet therapy, which increases their risk of developing acute coronary syndromes. These patients often present with an underlying condition that is associated with altered levels of circulating platelet primers and platelet hyperactivity. Platelet primers cannot stimulate platelet activation, but, in combination with physiologic stimuli, significantly enhance platelet function. OBJECTIVES: To explore the role of platelet primers in resistance to antiplatelet therapy, and to evaluate whether phosphoinositide 3-kinase (PI3K) contributes to this process. METHODS AND RESULTS: We used platelet aggregation, thromboxane A2 production and ex vivo thrombus formation as functional readouts of platelet activity. Platelets were treated with the potent P2Y12 inhibitor AR-C66096, aspirin, or a combination of both, in the presence or absence of the platelet primers insulin-like growth factor-1 (IGF-1) and thrombopoietin (TPO), or the Gz-coupled receptor ligand epinephrine. We found that platelet primers largely overcame the inhibitory effects of antiplatelet compounds on platelet functional responses. IGF-1-mediated and TPO-mediated, but not epinephrine-mediated, enhancements in the presence of antiplatelet drugs were blocked by the PI3K inhibitors wortmannin and LY294002. CONCLUSIONS: These results demonstrate that platelet primers can contribute to antiplatelet resistance. Furthermore, our data demonstrate that there are PI3K-dependent and PI3K-independent mechanisms driving primer-mediated resistance to antiplatelet therapy.


Assuntos
Trifosfato de Adenosina/análogos & derivados , Aspirina/farmacologia , Plaquetas/efeitos dos fármacos , Resistência a Medicamentos , Ativação Plaquetária/efeitos dos fármacos , Inibidores da Agregação de Plaquetas/farmacologia , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Trifosfato de Adenosina/farmacologia , Autoantígenos/farmacologia , Biomarcadores/metabolismo , Coagulação Sanguínea/efeitos dos fármacos , Plaquetas/metabolismo , Antígenos CD36/metabolismo , Epinefrina/farmacologia , Humanos , Fator de Crescimento Insulin-Like I/farmacologia , Iodeto Peroxidase/farmacologia , Proteínas de Ligação ao Ferro/farmacologia , Fosfatidilinositol 3-Quinase/metabolismo , Agregação Plaquetária/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Receptor PAR-1/metabolismo , Tromboxano A2/metabolismo
18.
Clin Exp Immunol ; 180(1): 58-69, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25412700

RESUMO

T helper type 17 (Th17) cells play a pathogenic role in autoimmune disease, while interleukin (IL)-10-producing Th10 cells serve a protective role. The balance between the two subsets is regulated by the local cytokine milieu and by the relative expression of intact forkhead box protein 3 (FoxP3) compared to FoxP3Δ2, missing exon 2. Th17 and Th10 cell differentiation has usually been studied using polyclonal stimuli, and little is known about the ability of physiologically relevant self-antigens to induce Th17 or Th10 cell differentiation in autoimmune thyroid disease. We subjected mononuclear cells from healthy donors and patients with Hashimoto's thyroiditis (HT) or Graves' disease (GD) to polyclonal stimulation, or stimulation with human thyroglobulin (TG), human thyroid peroxidase (TPO), or Esherichia coli lipopolysaccharide (LPS). TPO and LPS induced increased differentiation of naive CD4(+) CD45RA(+) CD45R0(-) T cells from HT patients into Th17 cells. Th10 cell proportions were decreased in HT after polyclonal stimulation, but were comparable to those of healthy donors after antigen-specific stimulation. Taken together, our data show that an increased Th17 : Th10 ratio was found in HT patients after stimulation with thyroid-specific self-antigens. We also observed an elevated baseline production of IL-6 and transforming growth factor (TGF)-ß1 and of mRNA encoding FoxP3Δ2 rather than intact FoxP3. This may contribute to the skewing towards Th17 cell responses in HT.


Assuntos
Processamento Alternativo/imunologia , Diferenciação Celular/imunologia , Fatores de Transcrição Forkhead/imunologia , Doença de Graves/imunologia , Doença de Hashimoto/imunologia , Células Th17/imunologia , Adulto , Idoso , Processamento Alternativo/efeitos dos fármacos , Antígenos CD/imunologia , Autoantígenos/imunologia , Autoantígenos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Escherichia coli/química , Feminino , Doença de Graves/patologia , Doença de Hashimoto/patologia , Humanos , Interleucina-10/imunologia , Interleucina-6/imunologia , Iodeto Peroxidase/imunologia , Iodeto Peroxidase/farmacologia , Proteínas de Ligação ao Ferro/imunologia , Proteínas de Ligação ao Ferro/farmacologia , Lipopolissacarídeos/química , Lipopolissacarídeos/farmacologia , Masculino , Pessoa de Meia-Idade , Isoformas de Proteínas/imunologia , Células Th17/patologia , Tireoglobulina/imunologia , Tireoglobulina/farmacologia , Fator de Crescimento Transformador beta1/imunologia
19.
Clin Exp Med ; 15(1): 31-9, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24474501

RESUMO

Juvenile idiopathic arthritis (JIA) is a heterogeneous group of diseases characterized by autoimmune arthritis of unknown cause with onset before age of 16 years. Methotrexate provides clinical benefits in JIA. For children who do not respond to methotrexate, treatment with anti-tumor necrosis factor (TNF)-α is an option. However, some patients do not respond or are intolerant to anti-TNF therapy. Induction of peripheral tolerance has long been considered a promising approach to the treatment of chronic autoimmune diseases. We aimed to evaluate the potentialities of two altered peptide ligands (APLs) derived from human heat-shock protein 60, an autoantigen involved in the pathogenesis of autoimmune arthritis, in JIA patients. Interferon (IFN)-γ, TNF-α and interleukin (IL)-10 levels were determined in ex vivo assays using peripheral blood mononuclear cells (PBMC) from these patients. Wild-type peptide and one of these APLs increased IFN-γ and TNF-α levels. Unlike, the other APLs (called APL2) increased the IL-10 level without affecting IFN-γ and TNF-α levels. On the other hand, APL2 induces a marked activation of T cells since it transforms cell cycle phase's distribution of CD4+ T cells from these patients. In addition, we evaluated the therapeutic effect of APL2 in collagen-induced arthritis model. Therapy with APL2 reduced arthritis scores and histological lesions in mice. This effect was associated to a decrease in TNF-α and IL-17 levels. These results indicate a therapeutic potentiality of APL2 for JIA.


Assuntos
Artrite Experimental/tratamento farmacológico , Artrite Juvenil/imunologia , Autoantígenos/farmacologia , Chaperonina 60/química , Leucócitos Mononucleares/efeitos dos fármacos , Proteínas Mitocondriais/química , Peptídeos/farmacologia , Adolescente , Animais , Antirreumáticos/farmacologia , Artrite Experimental/genética , Artrite Experimental/imunologia , Artrite Experimental/patologia , Artrite Juvenil/genética , Artrite Juvenil/patologia , Autoantígenos/química , Chaperonina 60/genética , Chaperonina 60/imunologia , Criança , Pré-Escolar , Regulação da Expressão Gênica , Humanos , Interferon gama/biossíntese , Interferon gama/metabolismo , Interleucina-10/biossíntese , Interleucina-10/metabolismo , Interleucina-17/antagonistas & inibidores , Interleucina-17/biossíntese , Interleucina-17/metabolismo , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/patologia , Metotrexato/farmacologia , Camundongos , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/imunologia , Peptídeos/síntese química , Tolerância Periférica , Cultura Primária de Células , Transdução de Sinais , Sulfassalazina/farmacologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/metabolismo
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