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1.
BMC Res Notes ; 12(1): 648, 2019 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-31590685

RESUMO

OBJECTIVES: Mycobacterium indicus pranii (MIP) is an atypical mycobacterium species with potent antitumor efficacy. Macrophages and dendritic cells (DCs) are antigen-presenting cells, playing key roles in the activation of antitumor immunity. We have previously shown the potent activation of macrophages and DCs by MIP, which is mediated by MyD88-TLR2 signaling axis. In the present study, we further examined the role of MyD88 and TLR2 in MIP-mediated tumor regression. RESULTS: Wild-type and MyD88-/- mice were implanted with B16F10 tumor cells, treated with MIP or phosphate-buffered saline (PBS) and monitored for tumor growth. As expected, MIP therapy led to significant tumor regression in wild-type mice. However, antitumor efficacy of MIP was lost in MyD88-/- animals. Both PBS-treated (control) and MIP-treated MyD88-/- mice developed tumors with comparable volume. Since MyD88 relays TLR engagement signals, we analyzed the antitumor efficacy of MIP in TLR2-/- and TLR4-/- mice. It was observed that MIP therapy reduced tumor burden in wild-type and TLR4-/- mice but not in TLR2-/- mice. Tumor volume in MIP-treated TLR2-/- mice were comparable with those in PBS-treated wild-type animals. These results implicated the MyD88-TLR2 signaling axis in the antitumor efficacy of MIP.


Assuntos
Melanoma Experimental/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Micobactérias não Tuberculosas/imunologia , Receptor 2 Toll-Like/imunologia , Carga Tumoral/imunologia , Animais , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Macrófagos/imunologia , Macrófagos/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Micobactérias não Tuberculosas/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Carga Tumoral/genética
2.
Int J Mol Sci ; 20(18)2019 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-31546763

RESUMO

Systemic lupus erythematosus (SLE) is a chronic, multifactorial autoimmune disease that predominantly affects young females. Dysregulation of different immune cell populations leads to self-tolerance breakdown and subsequent multiple organ damage as the disease develops. Plasmacytoid dendritic cells (pDCs) are potent producers of type I interferon (IFN), while myeloid dendritic cells (mDCs) are more specialized in antigen presentations. We have previously reported that bone-marrow (BM)-derived pDCs from the murine lupus model New Zealand black/white F1 (BWF1) possess abnormalities. Therefore, this study continues to investigate what aberrant properties peripheral pDCs and mDCs possess in BWF1 and how they mediate SLE progression, by comparing their properties in pre-symptomatic and symptomatic mice. Results showed that CD11chiCD11b+ myeloid DCs expanded during the disease state with down-regulation of co-stimulatory molecules and major histocompatibility complex class II molecules (MHC II), but their capacity to stimulate T cells was not hampered. During the disease state, this subset of mDCs displayed heightened toll-like receptors 7 and 9 (TLR 7/9) responses with increased interleukin 10 (IL-10) and C-X-C motif chemokine ligand 13 (CXCL13) expressions. Moreover, the expressions of myeloid differentiation primary response 88 (Myd88) and nuclear factor kappa B subunit 1 (Nfkb1) were higher in CD11chiCD11b+ DCs at the disease stage, leading to higher nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 phosphorylation activity. In summary, we reported aberrant phenotypic properties with enhanced TLR7/9 responses of CD11chiCD11b+ DCs in SLE mediated by aberrant NF-κB signaling pathway. Our findings add additional and novel information to our current understanding of the role of DCs in lupus immunopathogenesis. Lastly, molecular candidates in the NF-κB pathway should be exploited for developing therapeutic targets for SLE.


Assuntos
Antígenos CD11/imunologia , Antígeno CD11b/imunologia , Quimiocina CXCL13/imunologia , Células Dendríticas/imunologia , Interleucina-10/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Glicoproteínas de Membrana/imunologia , Subunidade p50 de NF-kappa B/imunologia , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia , Animais , Antígenos CD11/genética , Antígeno CD11b/genética , Quimiocina CXCL13/genética , Células Dendríticas/patologia , Modelos Animais de Doenças , Feminino , Interleucina-10/genética , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/patologia , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Células Mieloides/imunologia , Células Mieloides/patologia , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Subunidade p50 de NF-kappa B/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 7 Toll-Like/genética , Receptor Toll-Like 9/genética
3.
Fish Shellfish Immunol ; 94: 220-229, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31494279

RESUMO

Myeloid differentiation factor 88 (MyD88) is an important transduction protein in the Toll-like receptor signaling pathway. In this study, we identified the cDNA of the MpMyD88 gene in black carp. We found that MpMyD88 was widely distributed in the tissues tested and showed significant immune responses both in vitro and in vivo after stimulation with bacterial and pathogen-associated molecular patterns. After MpMyD88 overexpression/silencing, proinflame-matory cytokines (TNF-α, IFN-α, IL-6, and IL-8) also showed significant up-regulation/down-regulation. Moreover, we found that the antibacterial ability of cells over-expressing MpMyD88 was significantly stronger than that of control cells, while that of silenced MpMyD88 was significantly lower than that in control cells. Besides, we found that the overexpression of MpMyD88 significantly increased the activity of NF-κB. These results indicate that MpMyD88 plays an important role in the innate immune response.


Assuntos
Carpas/genética , Carpas/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Aeromonas hydrophila/fisiologia , Sequência de Aminoácidos , Animais , Citocinas/genética , Citocinas/imunologia , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Fator 88 de Diferenciação Mieloide/química , Filogenia , Alinhamento de Sequência/veterinária
4.
Fish Shellfish Immunol ; 94: 539-547, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31533084

RESUMO

Myeloid differentiation factor 88 (MyD88) is a key adaptor of Toll-like receptors (TLR), an important pattern recognition receptor of the innate immune system. To study the origin and evolution of the vertebrate TLR signaling pathway in innate immune systems, we analyzed the biological characteristics and functions of the MyD88 gene in Northeast Chinese lamprey (Lethenteron morii) using PCR amplification, real-time PCR analysis, dual luciferase reporter gene assay, immunofluorescence assay, and other methods. Bioinformatics analysis showed that LmMyD88 has a modular structure consisting of Toll/IL-1R domain (TIR) and death domain (DD), which is typical of the MyD88 family. A phylogenetic tree showed that the homology of LmMyD88 was consistent with the phylogenetic status of lampreys. Tissue expression analysis indicated that the mRNA expression was expressed in some normal tissues of larval and adult L. morii. Real-time PCR analysis showed that the expression of LmMyD88 in tissues, such as gill and kidney, of the adult increased significantly after infection by Pseudomonas aeruginosa. Subcellular localization results showed that LmMyD88 was expressed in the nucleus, cytoplasm, and other parts. A dual luciferase reporter assay indicated that LmMyD88 activated nuclear factor kappa B downstream of the TLR signaling pathway. This study suggested that LmMyD88 might play an important role in the innate immune signal transduction process of L. morii.


Assuntos
Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Perciformes/genética , Perciformes/imunologia , Sequência de Aminoácidos , Animais , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Lampreias , Fator 88 de Diferenciação Mieloide/química , Filogenia , Pseudomonas aeruginosa/fisiologia , Alinhamento de Sequência/veterinária
5.
Fish Shellfish Immunol ; 94: 58-65, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31470137

RESUMO

TIR domain-containing protein is an important member for some bacterial pathogens to subvert host defenses. Here we described a fish virulent Yersinia ruckeri SC09 strain that interfered directly with Toll-like receptor (TLR) function by a TIR-containing protein. Firstly, the novel TIR-containing protein was identified by bioinformatics analysis and named as TcpA. Secondly, the toxic effects of TcpA in fish was demonstrated in vivo challenge experiments through knockout mutant and complement mutant of tcpA gene. Thirdly, The study in vitro revealed that TcpA could down-regulate the expression and secretion of IL-6, IL-1ß and TNF-α. Finally, we demonstrated that TcpA could inhibit the TLR signaling pathway through interaction with myeloid differentiation factor 88 (MyD88) in experiments such as NF-κB dependent luciferase reporter system, co-immunoprecipitation, GST pull-down and yeast two-hybrid. The study revealed that TcpA was essential for virulence and was able to interact with the TIR adaptor protein MyD88 and inhibit the pre-inflammatory signal of immune cells and promote the intracellular survival of pathogenic Yersinia ruckeri SC09 strain. In conclusion, our results showed that TcpA acted as a new virulence factor in Y. ruckeri could suppress innate immune response and increase virulence by inhibiting TLR and MyD88-mediated specific signaling, highlighting a novel strategy for innate immune evasion in bacteria.


Assuntos
Evasão da Resposta Imune/genética , Imunidade Inata/genética , Fator 88 de Diferenciação Mieloide/genética , Receptores Toll-Like/genética , Fatores de Virulência/genética , Yersiniose/veterinária , Yersinia ruckeri/genética , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Transdução de Sinais/imunologia , Receptores Toll-Like/metabolismo , Fatores de Virulência/metabolismo , Yersiniose/genética , Yersiniose/imunologia
6.
Fish Shellfish Immunol ; 93: 308-312, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31352113

RESUMO

Initiation of the innate immune response requires recognition of pathogen-associated molecular patterns by pathogen recognition receptors such as Toll-like receptors (TLRs). MyD88 adaptor-like (Mal) is an adaptor that responds to TLR activation and acts as a bridging adaptor for MyD88. In the present study, the open reading frame of Mal was identified in orange-spotted grouper (Epinephelus coioides), and named EcMal. It contained 831 bp encoding 276 aa, and was encoded by a 1299 bp DNA sequence with three exons and two introns. EcMal and the Mal sequence of other species shared different degrees of sequence identity, and clustered into the same group. EcMal was distributed in all tissues tested in healthy grouper, with the highest expression level in the head kidney. After infection with Cryptocaryon irritans, the expression level of EcMal was up-regulated in the gill and spleen. In addition, EcMal exhibited global cytosolic and nucleus localization, and could significantly activate NF-κB activity in grouper spleen cells.


Assuntos
Bass/genética , Bass/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Sequência de Aminoácidos , Animais , Cilióforos/fisiologia , Infecções por Cilióforos/imunologia , Infecções por Cilióforos/veterinária , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Fator 88 de Diferenciação Mieloide/química , Filogenia , Alinhamento de Sequência/veterinária
7.
Vet Res ; 50(1): 53, 2019 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-31300043

RESUMO

Our previous studies demonstrated that matrine directly acts on the replication process of porcine reproductive and respiratory syndrome virus (PRRSV). Matrine inhibits viral replication and is also associated with the NF-κB signalling pathway. These results suggest that matrine has antiviral and anti-inflammatory effects. However, the specific anti-inflammatory mechanism of matrine is still unclear. In this study, we investigated the anti-IL-1ß mechanism of matrine, as IL-1ß is a major inflammatory cytokine, in porcine alveolar macrophages (PAMs) stimulated with 4 µg PRRSV 5'-untranslated region (UTR) RNA and 1 µg/mL LPS. After 5'UTR RNA and LPS co-stimulation of PAMs for 12 h, the expression of IL-1ß, IL-6, IL-8 and TNF-α was significantly increased. The results also showed that co-stimulation induced the expression of MyD88, and activated the NF-κB signalling pathway and NLRP3 inflammasome. Furthermore, matrine treatment downregulated MyD88, NLRP3 and caspase-1 expression, inhibited ASC speck formation, suppressed IκBα phosphorylation, and interfered with the translocation of NF-κB from the cytoplasm to the nucleus. These results suggest that matrine plays an important role in PAMs co-stimulated with PRRSV 5'UTR RNA and LPS via its effect on NF-κB and the NLRP3 inflammasome. These findings lay the foundation for the exploration of the clinical application of matrine in PRRSV disease.


Assuntos
Alcaloides/farmacologia , Inflamassomos/imunologia , Interleucina-1beta/metabolismo , Macrófagos/efeitos dos fármacos , NF-kappa B/imunologia , Quinolizinas/farmacologia , Animais , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , RNA Viral/genética , Transdução de Sinais/imunologia , Sus scrofa , Transfecção/veterinária
8.
Int J Mol Med ; 44(2): 479-490, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31173183

RESUMO

Acute lung injury (ALI) is a critical syndrome that is associated with a high morbidity and mortality in patients. Sevoflurane has a lung protective effect in ALI as it reportedly has anti­inflammatory and apoptotic­regulating activity. However, the mechanism is still not entirely understood. The aim of the present study was to explore the effects of sevoflurane on lipopolysaccharide (LPS)­induced ALI in mice and the possible mechanisms involved. The results revealed that sevoflurane treatment improved LPS­induced lung injury, as evidenced by the reduction in mortality, lung permeability, lung wet/dry ratio and lung histopathological changes in mice. Total cell counts and the production of pro­inflammatory cytokines [tumor necrosis factor­α, interleukin (IL)­1ß and IL­6] in bronchoalveolar fluid were also decreased following treatment with sevoflurane. Additionally, LPS­triggered apoptosis in lung tissues, which was eliminated by sevoflurane. Furthermore, a miRCURY™ LNA array was employed to screen for differentially expressed microRNAs (miRs/miRNAs). Among these miRNAs, 6 were differentially expressed and were involved in the inflammatory response, but only miR­27a­3p (miR­27a) was regulated by sevoflurane. Subsequently, the present study investigated whether sevoflurane exerts its function through the modulation of miR­27a. The results demonstrated that the overexpression of miR­27a via an injection with agomiR­27a produced similar protections as sevoflurane, while the inhibition of miR­27a suppressed the lung protective effects of sevoflurane in ALI mice. In addition, the present study identified that miR­27a inhibited Toll­like receptor 4 (TLR4) by binding to its 3'­untranslated region. Western blot analysis demonstrated that sevoflurane may ameliorate the inflammatory response by blocking the miR­27a/TLR4/MyD88/NF­κB signaling pathway. The present results indicate that sevoflurane may be a viable therapeutic option in the treatment of patients with ALI.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Anti-Inflamatórios/uso terapêutico , MicroRNAs/imunologia , NF-kappa B/imunologia , Sevoflurano/uso terapêutico , Receptor 4 Toll-Like/imunologia , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/imunologia , Animais , Anti-Inflamatórios/farmacologia , Apoptose , Lipopolissacarídeos/imunologia , Masculino , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Fator 88 de Diferenciação Mieloide/imunologia , Sevoflurano/farmacologia , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
9.
Int J Med Microbiol ; 309(5): 307-318, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31178418

RESUMO

Mycobacterium abscessus (MAB) is an emerging, rapidly growing non-tuberculous Mycobacterium causing therapy-resistant pulmonary disease especially in patients with cystic fibrosis (CF). Smooth and rough colony type MAB can be isolated from infected patients whereby rough colony type MAB are more often associated with severe disease. Disease severity is also associated with an alternated type I interferon (IFN-I) response of the MAB-infected patients. However the relevance of this response for the outcome of MAB infection is still unknown. In this study, we analyzed the IFNß expression of murine macrophages infected with a MAB rough colony strain (MAB-R) isolated from a patient with progressive CF and compared it to macrophages infected with the MAB smooth colony type reference strain (MAB-S). We found that MAB-R infected macrophages expressed significantly more IFNß mRNA and protein than MAB-S infected macrophages. Higher IFNß induction by MAB-R was associated with higher TNF expression and intracellular killing while low IFNß induction was associated with lower TNF expression and persistence of MAB-S. IFNß induction was independent of the intracellular cGAS-STING recognition pathway. MAB appeared to be recognized extracellularly and induced IFNß expression via TLR2-TLR4-MyD88-TRIF-IRF3 dependent pathways. By using macrophages lacking the IFN-I receptor we demonstrate that MAB induced IFN-I response essentially contributed to restricting MAB-R and MAB-S infections by activating macrophage Nos2 expression and nitric oxide production. Thus IFN-I seem to influence the intrinsic ability of macrophages to control MAB infections. As MAB persists over long time periods in susceptible patients, our findings suggest that virulence of MAB strains is promoted by an insufficient IFN-I response of the host.


Assuntos
Interferon beta/imunologia , Macrófagos/microbiologia , Mycobacterium abscessus/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Óxido Nítrico/imunologia , Receptor 2 Toll-Like/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Interações Hospedeiro-Patógeno/imunologia , Humanos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/microbiologia , Óxido Nítrico Sintase Tipo II/genética , Escarro/microbiologia
10.
J Immunol ; 202(10): 3020-3032, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30988118

RESUMO

The inflammatory response to infection or injury dramatically increases the hematopoietic demand on the bone marrow to replace effector leukocytes consumed in the inflammatory response. In the setting of infection, pathogen-associated molecular patterns induce emergency hematopoiesis, activating hematopoietic stem and progenitor cells to proliferate and produce progeny for accelerated myelopoiesis. Sterile tissue injury due to trauma also increases leukocyte demand; however, the effect of sterile tissue injury on hematopoiesis is not well described. We find that tissue injury alone induces emergency hematopoiesis in mice subjected to polytrauma. This process is driven by IL-1/MyD88-dependent production of G-CSF. G-CSF induces the expansion of hematopoietic progenitors, including hematopoietic stem cells and multipotent progenitors, and increases the frequency of myeloid-skewed progenitors. To our knowledge, these data provide the first comprehensive description of injury-induced emergency hematopoiesis and identify an IL-1/MyD88/G-CSF-dependent pathway as the key regulator of emergency hematopoiesis after injury.


Assuntos
Fator Estimulador de Colônias de Granulócitos/imunologia , Hematopoese/imunologia , Interleucina-1/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Ferimentos e Lesões/imunologia , Animais , Fator Estimulador de Colônias de Granulócitos/genética , Hematopoese/genética , Interleucina-1/genética , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Ferimentos e Lesões/genética , Ferimentos e Lesões/patologia
11.
J Immunol ; 202(10): 3041-3052, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30952815

RESUMO

Serotonin (5-hydroxytryptamine [5-HT]) is a key enteric signaling molecule that mediates various physiological processes in the gut. Enterochromaffin (EC) cells in the mucosal layer of the gut are the main source of 5-HT in the body and are situated in close proximity to the gut microbiota. In this study, we identify a pivotal role of TLR2 in 5-HT production in the gut. Antibiotic treatment reduces EC cell numbers and 5-HT levels in naive C57BL/6 mice, which is associated with downregulation of TLR2 expression but not TLR1 or TLR4. TLR2-deficient (Tlr2 -/-) and Myd88 -/- mice express lower EC cell numbers and 5-HT levels, whereas treatment with TLR2/1 agonist upregulates 5-HT production in irradiated C57BL/6 mice, which are reconstituted with Tlr2 -/- bone marrow cells, and in germ-free mice. Human EC cell line (BON-1 cells) release higher 5-HT upon TLR2/1 agonist via NF-κB pathway. Tlr2 -/- mice and anti-TLR2 Ab-treated mice infected with enteric parasite, Trichuris muris, exhibited attenuated 5-HT production, compared with infected wild-type mice. Moreover, excretory-secretory products from T. muris induce higher 5-HT production in BON-1 cells via TLR2 in a dose-dependent manner, whereby the effect of excretory-secretory products is abrogated by TLR2 antagonist. These findings not only suggest an important role of TLR2 in mucosal 5-HT production in the gut by resident microbiota as well as by a nematode parasite but also provide, to our knowledge, novel information on the potential benefits of targeting TLR2 in various gut disorders that exhibit aberrant 5-HT signaling.


Assuntos
Células Enterocromafins/imunologia , Serotonina/imunologia , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/imunologia , Tricuríase/imunologia , Trichuris/imunologia , Animais , Linhagem Celular , Células Enterocromafins/patologia , Microbioma Gastrointestinal/imunologia , Humanos , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Serotonina/genética , Transdução de Sinais/genética , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Tricuríase/genética , Tricuríase/patologia
12.
J Immunol ; 202(10): 3053-3064, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30979817

RESUMO

Acute graft-versus-host disease (aGVHD) hinders the efficacy of allogeneic hematopoietic cell transplantation (HCT). Plasma levels of soluble membrane-bound ST2 (ST2) are elevated in human and murine aGVHD and correlated to type 1 T cells response. ST2 signals through the adapter protein MyD88. The role of MyD88 in T cells during aGVHD has yet to be elucidated. We found that knocking out MyD88 in the donor T cells protected against aGVHD independent of IL-1R and TLR4 signaling in two murine HCT models. This protection was entirely driven by MyD88-/- CD4 T cells. Transplanting donor MyD88-/- conventional T cells (Tcons) with wild-type (WT) or MyD88-/- regulatory T cells (Tregs) lowered aGVHD severity and mortality. Transcriptome analysis of sorted MyD88-/- CD4 T cells from the intestine 10 d post-HCT showed lower levels of Il1rl1 (gene of ST2), Ifng, Csf2, Stat5, Batf, and Jak2 Transplanting donor ST2-/- Tcons with WT or ST2-/- Tregs showed a similar phenotype with what we observed when using donor MyD88-/- Tcons. Decreased ST2 was confirmed at the protein level with less secretion of soluble ST2 and more expression of ST2 compared with WT T cells. Our data suggest that Treg suppression from lack of MyD88 signaling in donor Tcons during alloreactivity uses the ST2 but not the IL-1R or TLR4 pathways, and ST2 represents a potential aGVHD therapeutic target sparing Tregs.


Assuntos
Doença Enxerto-Hospedeiro/imunologia , Transplante de Células-Tronco Hematopoéticas , Proteína 1 Semelhante a Receptor de Interleucina-1/deficiência , Fator 88 de Diferenciação Mieloide/deficiência , Transdução de Sinais/imunologia , Linfócitos T Reguladores/imunologia , Doença Aguda , Animais , Doença Enxerto-Hospedeiro/genética , Doença Enxerto-Hospedeiro/patologia , Proteína 1 Semelhante a Receptor de Interleucina-1/imunologia , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/imunologia , Transdução de Sinais/genética , Linfócitos T Reguladores/patologia , Transplante Homólogo
13.
Leukemia ; 33(9): 2195-2207, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-30816327

RESUMO

Successful adoptive chimeric antigen receptor (CAR) T-cell therapies against hematological malignancies require CAR-T expansion and durable persistence following infusion. Balancing increased CAR-T potency with safety, including severe cytokine-release syndrome (sCRS) and neurotoxicity, warrants inclusion of safety mechanisms to control in vivo CAR-T activity. Here, we describe a novel CAR-T cell platform that utilizes expression of the toll-like receptor (TLR) adaptor molecule, MyD88, and tumor-necrosis factor family member, CD40 (MC), tethered to the CAR molecule through an intentionally inefficient 2A linker system, providing a constitutive signal that drives CAR-T survival, proliferation, and antitumor activity against CD19+ and CD123+ hematological cancers. Robust activity of MC-enhanced CAR-T cells was associated with cachexia in animal models that corresponded with high levels of human cytokine production. However, toxicity could be successfully resolved by using the inducible caspase-9 (iC9) safety switch to reduce serum cytokines, by administration of a neutralizing antibody against TNF-α, or by selecting "low" cytokine-producing CD8+ T cells, without loss of antitumor activity. Interestingly, high basal activity was essential for in vivo CAR-T expansion. This study shows that co-opting novel signaling elements (i.e., MyD88 and CD40) and development of a unique CAR-T architecture can drive T-cell proliferation in vivo to enhance CAR-T therapies.


Assuntos
Antígenos CD40/imunologia , Linfócitos T CD8-Positivos/imunologia , Neoplasias Hematológicas/imunologia , Neoplasias Hematológicas/terapia , Fator 88 de Diferenciação Mieloide/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos Quiméricos/imunologia , Animais , Antígenos CD19/imunologia , Proliferação de Células/efeitos dos fármacos , Células HEK293 , Humanos , Imunoterapia Adotiva/métodos , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos NOD , Transdução de Sinais/imunologia , Células THP-1
14.
J Immunol ; 202(9): 2737-2746, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30885957

RESUMO

Dimethyl fumarate (DMF) is a prescribed treatment for multiple sclerosis and has also been used to treat psoriasis. The electrophilicity of DMF suggests that its immunosuppressive activity is related to the covalent modification of cysteine residues in the human proteome. Nonetheless, our understanding of the proteins modified by DMF in human immune cells and the functional consequences of these reactions remains incomplete. In this study, we report that DMF inhibits human plasmacytoid dendritic cell function through a mechanism of action that is independent of the major electrophile sensor NRF2. Using chemical proteomics, we instead identify cysteine 13 of the innate immune kinase IRAK4 as a principal cellular target of DMF. We show that DMF blocks IRAK4-MyD88 interactions and IRAK4-mediated cytokine production in a cysteine 13-dependent manner. Our studies thus identify a proteomic hotspot for DMF action that constitutes a druggable protein-protein interface crucial for initiating innate immune responses.


Assuntos
Células Dendríticas/imunologia , Fumarato de Dimetilo/farmacologia , Imunidade Inata/efeitos dos fármacos , Quinases Associadas a Receptores de Interleucina-1/imunologia , Complexos Multiproteicos/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Plasmócitos/imunologia , Transdução de Sinais/efeitos dos fármacos , Adulto , Citocinas/imunologia , Feminino , Humanos , Pessoa de Meia-Idade
15.
J Immunol ; 202(8): 2266-2275, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30842274

RESUMO

It is not clear how hepatitis B virus (HBV) modulates host immunity during chronic infection. In addition to the key mediators of inflammatory response in viral infection, monocytes also express a high-level IFN-stimulated gene, CH25H, upon response to IFN-α exerting an antiviral effect. In this study, the mechanism by which HBV manipulates IFN signaling in human monocytes was investigated. We observed that monocytes from chronic hepatitis B patients express lower levels of IFN signaling/stimulated genes and higher levels of inflammatory cytokines compared with healthy donors. HBV induces monocyte production of inflammatory cytokines via TLR2/MyD88/NF-κB signaling and STAT1-Ser727 phosphorylation and inhibits IFN-α-induced stat1, stat2, and ch25h expression through the inhibition of STAT1-Tyr701 phosphorylation and in an IL-10-dependent, partially autocrine manner. Further, we found that enhancement of STAT1 activity with a small molecule (2-NP) rescued HBV-mediated inhibition of IFN signaling and counteracted the induction of inflammatory cytokines. In conclusion, HBV contributes to the monocyte inflammatory response but inhibits their IFN-α/ß responsiveness to impair antiviral innate immunity. These effects are mediated via differential phosphorylation of Tyr701 and Ser727 of STAT1.


Assuntos
Vírus da Hepatite B/imunologia , Hepatite B/imunologia , Imunidade Inata , Monócitos/imunologia , Fator de Transcrição STAT1/imunologia , Transdução de Sinais/imunologia , Células Hep G2 , Hepatite B/patologia , Humanos , Interleucina-10/imunologia , Monócitos/patologia , Fator 88 de Diferenciação Mieloide/imunologia , NF-kappa B/imunologia , Fosforilação/imunologia , Fator de Transcrição STAT2/imunologia , Receptor 2 Toll-Like/imunologia
16.
J Immunol ; 202(8): 2384-2396, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30787108

RESUMO

MyD88 adaptor-like (Mal) protein is the most polymorphic of the four key adaptor proteins involved in TLR signaling. TLRs play a critical role in the recognition and immune response to pathogens through activation of the prototypic inflammatory transcription factor NF-κB. The study of single nucleotide polymorphisms in TLRs, adaptors, and signaling mediators has provided key insights into the function of the corresponding genes but also into the susceptibility to infectious diseases in humans. In this study, we have analyzed the immune response of mice carrying the human Mal-D96N genetic variation that has previously been proposed to confer protection against septic shock. We have found that Mal-D96N macrophages display reduced cytokine expression in response to TLR4 and TLR2 ligand challenge. Mal-D96N macrophages also display reduced MAPK activation, NF-κB transactivation, and delayed NF-κB nuclear translocation, presumably via delayed kinetics of Mal interaction with MyD88 following LPS stimulation. Importantly, Mal-D96N genetic variation confers a physiological protective phenotype to in vivo models of LPS-, Escherichia coli-, and influenza A virus-induced hyperinflammatory disease in a gene dosage-dependent manner. Together, these results highlight the critical role Mal plays in regulating optimal TLR-induced inflammatory signaling pathways and suggest the potential therapeutic advantages of targeting the Mal D96 signaling nexus.


Assuntos
Lipopolissacarídeos/toxicidade , Sistema de Sinalização das MAP Quinases , Macrófagos/imunologia , Mutação de Sentido Incorreto , Fator 88 de Diferenciação Mieloide , Polimorfismo de Nucleotídeo Único , Receptores Toll-Like , Substituição de Aminoácidos , Animais , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/imunologia , Camundongos , Camundongos Mutantes , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Receptores Toll-Like/genética , Receptores Toll-Like/imunologia
17.
Fish Shellfish Immunol ; 87: 829-838, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30790663

RESUMO

Myeloid differentiation factor 88 (MyD88) and tumor necrosis factor receptor-associated factor 6 (TRAF6) are two critical signal transducers in toll-like receptor (TLR) pathway. In the present study, we identified and characterized the homologues of MyD88 and TRAF6 in Qihe crucian carp Carassius auratus, termed as CaMyD88 and CaTRAF6, respectively, and examined their roles during pathogenic infection. Full-length cDNA of CaMyD88 was 2463 bp, including a 191 bp 5'-untranslated region (UTR), a 1417 bp 3'-UTR, and an 855 bp open reading frame (ORF) encoding for a putative protein with 284 amino acids. Full-length cDNA of CaTRAF6 was identified to be 2555 bp, consisting of a 52 bp 5'-UTR, an 871 bp 3'-UTR, and a 1632 bp ORF encoding a protein of 543 amino acids. Deduced amino acid sequences of CaMyD88 and CaTRAF6 contained the typical domains (CaMyD88: death domain and TIR domain; CaTRAF6: one RING-type zinc finger domain, two TRAF-type zinc finger domains, one coiled-coil region, and one conserved C-terminal meprin and TRAF homology domain) as in other fish. Quantitative Real-Time PCR (qRT-PCR) analysis revealed that both CaMyD88 and CaTRAF6 were ubiquitously expressed throughout the development stages and appeared to be developmentally regulated. In addition, CaMyD88 and CaTRAF6 had a broadly distribution of expression in all examined eleven tissues of healthy fish, although the transcript levels varied among the different tissues. Moreover, it was found that mRNA expressions of CaMyD88 and CaTRAF6 were generally up-regulated after stimulation by polyI:C, flagellin, and Aeromonas hydrophila in spite of the down-regulation appeared at some time points or tissues. These results indicated that CaMyD88 and CaTRAF6 play the critical roles in the immune defense of Qihe crucian carp against pathogenic invasion. The present findings will provide the valuable information for understanding the innate immune responses of Qihe crucian carp and contribute to develop the preventive way against pathogens.


Assuntos
Carpas/genética , Carpas/imunologia , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Aeromonas hydrophila/fisiologia , Animais , Clonagem Molecular , Proteínas de Peixes/química , Flagelina/farmacologia , Perfilação da Expressão Gênica/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Fator 88 de Diferenciação Mieloide/química , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/imunologia , Poli I-C/farmacologia , Fator 6 Associado a Receptor de TNF/química , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/imunologia
18.
J Periodontal Res ; 54(4): 396-404, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30793777

RESUMO

AIM: To explore the role of keratinocyte myeloid differentiation primary response 88 (MyD88) expression in the adhesion of Porphyromonas gingivalis to the cells and its subsequent invasion and intracellular survival. MATERIALS AND METHODS: Primary mouse keratinocytes from wild-type (WT) or Myd88-/- mice were infected with P gingivalis alone or co-infected with Fusobacterium nucleatum. Bacterial adhesion and invasion were measured using fluorescent microscopy and flow cytometry, and intracellular survival in keratinocytes was quantified by an antibiotic protection assay. Keratinocyte expression of antimicrobial peptides was measured by real-time PCR. RESULTS: In the absence of MyD88, P gingivalis adherence, invasion, and intracellular survival were enhanced compared with WT keratinocytes. The presence of F nucleatum during infection increased the adhesion of P gingivalis to WT keratinocytes but reduced the adhesion to Myd88-/- keratinocytes. Fusobacterium nucleatum improved mildly the invasion and survival of P gingivalis in both cell types. Baseline expression of beta-defensin 2, 3, 4 and RegIII-γ was elevated in Myd88-/- keratinocytes compared to WT cells; however, following infection beta-defensin expression was strongly induced in WT cells but decreased dramatically in the MyD88 deficient cells. CONCLUSION: In the absence of MyD88 expression, P gingivalis adhesion to keratinocytes is improved, and invasion and intracellular survival are increased. Furthermore, keratinocyte infection by P gingivalis induces antimicrobial peptide expression in a MyD88-dependent manner. Thus, MyD88 plays a key role in the interaction between P gingivalis and keratinocytes.


Assuntos
Infecções por Bacteroidaceae/imunologia , Queratinócitos/microbiologia , Fator 88 de Diferenciação Mieloide/imunologia , Porphyromonas gingivalis , Animais , Peptídeos Catiônicos Antimicrobianos/imunologia , Aderência Bacteriana , Fusobacterium nucleatum , Queratinócitos/imunologia , Camundongos , Camundongos Knockout
19.
PLoS One ; 14(2): e0212236, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30794604

RESUMO

Viral-bacterial coinfections, such as with influenza A virus and Streptococcus pneumoniae (S.p.), are known to cause severe pneumonia. It is well known that the host response has an important role in disease. Interleukin-1ß (IL-1ß) is an important immune signaling cytokine responsible for inflammation and has been previously shown to contribute to disease severity in numerous infections. Other studies in mice indicate that IL-1ß levels are dramatically elevated during IAV-S.p. coinfection. However, the regulation of IL-1ß during coinfection is unknown. Here, we report the NLRP3 inflammasome is the major inflammasome regulating IL-1ß activation during coinfection. Furthermore, elevated IL-1ß mRNA expression is due to enhanced TLR2-MYD88 signaling, which increases the amount of pro-IL-1ß substrate for the inflammasome to process. Finally, NLRP3 and high IL-1ß levels were associated with increased bacterial load in the brain. Our results show the NLRP3 inflammasome is not protective during IAV-S.p. coinfection.


Assuntos
Coinfecção/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Interleucina-1beta/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Infecções por Orthomyxoviridae/imunologia , Infecções Pneumocócicas/imunologia , Transdução de Sinais/imunologia , Streptococcus pneumoniae/imunologia , Receptor 2 Toll-Like/imunologia , Animais , Encéfalo , Linhagem Celular , Embrião de Galinha , Coinfecção/genética , Coinfecção/patologia , Interleucina-1beta/genética , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/patologia , Infecções Pneumocócicas/genética , Infecções Pneumocócicas/patologia , Transdução de Sinais/genética , Receptor 2 Toll-Like/genética
20.
Viruses ; 11(1)2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30650519

RESUMO

Respiratory syncytial virus (RSV) is a major cause of respiratory infectious disease in infants and young children. Dendritic cells (DCs) and macrophages (MACs) are known to play important roles in RSV recognition, and in the production of type I interferons (IFNs) and pro-inflammatory cytokine in RSV infection. Toll-like receptor 7 (TLR7), myeloid differentiation primary response 88 (MyD88), and mitochondrial antiviral-signaling protein (MAVS) are known to be important for the RSV sensing pathway in DCs and MACs. However, despite the critical roles of type I IFNs in the anti-RSV immune response, the pattern recognition receptors (PRRs) that are required for RSV sensing in DCs and MACs remain unclear. Here, we investigate the pathway activated by RSV A2 strain infection using an IFN-ß/YFP reporter mouse model to visualize IFN-ß-producing cells and in vitro RSV infection in bone marrow-derived DCs (BM-DCs) and macrophages (BM-DMs). We present our finding that MyD88, but not TLR7, are important for RSV recognition and type I IFN and pro-inflammatory production in DCs and MACs. MAVS-deficient BM-DCs and BM-DMs show impaired induction of IFN-ß production upon RSV stimulation, and this effect is RSV replication-dependent. Our study provides information on cell type-specific PRR requirements in innate immune responses against RSV infection.


Assuntos
Células Dendríticas/imunologia , Imunidade Inata , Interferon beta/imunologia , Macrófagos/imunologia , Vírus Sincicial Respiratório Humano/imunologia , Transdução de Sinais , Animais , Células Cultivadas , Células Dendríticas/virologia , Macrófagos/virologia , Glicoproteínas de Membrana/imunologia , Camundongos , Fator 88 de Diferenciação Mieloide/imunologia , Receptores de Reconhecimento de Padrão , Receptor 7 Toll-Like/imunologia , Replicação Viral
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