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1.
Int J Mol Sci ; 22(6)2021 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799511

RESUMO

In uremic patients, high-density lipoprotein (HDL) loses its anti-inflammatory features and can even become pro-inflammatory due to an altered protein composition. In chronic kidney disease (CKD), impaired functions of polymorphonuclear leukocytes (PMNLs) contribute to inflammation and an increased risk of cardiovascular disease. This study investigated the effect of HDL from CKD and hemodialysis (HD) patients on the CD14 expression on PMNLs. HDL was isolated using a one-step density gradient centrifugation. Isolation of PMNLs was carried out by discontinuous Ficoll-Hypaque density gradient centrifugation. CD14 surface expression was quantified by flow cytometry. The activity of the small GTPase Rac1 was determined by means of an activation pull-down assay. HDL increased the CD14 surface expression on PMNLs. This effect was more pronounced for HDL isolated from uremic patients. The acute phase protein serum amyloid A (SAA) caused higher CD14 expression, while SAA as part of an HDL particle did not. Lipid raft disruption with methyl-ß-cyclodextrin led to a reduced CD14 expression in the absence and presence of HDL. HDL from healthy subjects but not from HD patients decreased the activity of Rac1. Considering the known anti-inflammatory effects of HDL, the finding that even HDL from healthy subjects increased the CD14 expression was unexpected. The pathophysiological relevance of this result needs further investigation.


Assuntos
Receptores de Lipopolissacarídeos/genética , Lipoproteínas HDL/farmacologia , Neutrófilos/efeitos dos fármacos , Insuficiência Renal Crônica/genética , Uremia/genética , Idoso , Estudos de Casos e Controles , Feminino , Humanos , Receptores de Lipopolissacarídeos/metabolismo , Lipoproteínas HDL/isolamento & purificação , Masculino , Microdomínios da Membrana/química , Microdomínios da Membrana/efeitos dos fármacos , Microdomínios da Membrana/metabolismo , Pessoa de Meia-Idade , Neutrófilos/metabolismo , Neutrófilos/patologia , Cultura Primária de Células , Diálise Renal , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/fisiopatologia , Insuficiência Renal Crônica/terapia , Proteína Amiloide A Sérica/genética , Proteína Amiloide A Sérica/metabolismo , Uremia/metabolismo , Uremia/fisiopatologia , Uremia/terapia , beta-Ciclodextrinas/farmacologia , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
2.
Int J Mol Sci ; 22(3)2021 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-33513808

RESUMO

There is little known about the effect of the periodontopathogen Filifactor alocis on macrophages as key cells of the innate immune defense in the periodontium. Therefore, the aim of the present study was to investigate the effect of F. alocis and additionally of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNFα) on visfatin and other pro-inflammatory and proteolytic molecules associated with periodontitis in human macrophages. The presence of macrophage markers CD14, CD86, CD68, and CD163 was examined in gingival biopsies from healthy individuals and periodontitis patients. Human macrophages were incubated with F. alocis and TNFα for up to 2 d. The effects of both stimulants on macrophages were determined by real-time PCR, ELISA, immunocytochemistry, and immunofluorescence. F. alocis was able to significantly stimulate the synthesis of visfatin by human macrophages using TLR2 and MAPK pathways. In addition to visfatin, F. alocis was also able to increase the synthesis of cyclooxygenase 2, TNFα, and matrix metalloproteinase 1. Like F. alocis, TNFα was also able to stimulate the production of these proinflammatory and proteolytic molecules. Our results highlight the pathogenetic role of F. alocis in periodontal diseases and also underline the involvement of visfatin in the aetiopathogenesis of periodontitis.


Assuntos
Clostridiales/imunologia , Gengiva/metabolismo , Macrófagos/metabolismo , Nicotinamida Fosforribosiltransferase/biossíntese , Periodontite/imunologia , Receptor 2 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Antígenos de Diferenciação Mielomonocítica/metabolismo , Antígeno B7-2/genética , Antígeno B7-2/metabolismo , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Gengiva/citologia , Gengiva/patologia , Humanos , Imuno-Histoquímica , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , Sistema de Sinalização das MAP Quinases/imunologia , Macrófagos/efeitos dos fármacos , Metaloproteinase 1 da Matriz/biossíntese , Metaloproteinase 1 da Matriz/genética , Metaloproteinase 1 da Matriz/metabolismo , Nicotinamida Fosforribosiltransferase/genética , Nicotinamida Fosforribosiltransferase/metabolismo , Periodontite/metabolismo , Periodontite/microbiologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
3.
Mem Inst Oswaldo Cruz ; 115: e200110, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33146244

RESUMO

We aimed to investigate the association of CD14 -260C/T (rs2569190) polymorphism and Chagas cardiomyopathy and the functional characteristics of CD14+ and CD14- monocytes upon infection with Trypanosoma cruzi. We observed an association between the T- genotype (absence of allele -260T) related to low CD14 expression and the dilated cardiomyopathy type of Chagas disease. Furthermore, we observed that CD14- monocytes showed a more activated profile upon in vitro infection with T. cruzi than CD14+ monocytes. Our findings suggest that T- genotype is associated with susceptibility to develop Chagas dilated cardiomyopathy, likely linked to the T. cruzi-induced inflammatory profile of CD14- monocytes.


Assuntos
Cardiomiopatia Dilatada/genética , Cardiomiopatia Chagásica/genética , Receptores de Lipopolissacarídeos/genética , Doença de Chagas , Genótipo , Insuficiência Cardíaca , Humanos , Trypanosoma cruzi , Disfunção Ventricular Esquerda
4.
PLoS Pathog ; 16(8): e1008639, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32790743

RESUMO

Leptospirosis is a worldwide re-emerging zoonosis caused by pathogenic Leptospira spp. All vertebrate species can be infected; humans are sensitive hosts whereas other species, such as rodents, may become long-term renal carrier reservoirs. Upon infection, innate immune responses are initiated by recognition of Microbial Associated Molecular Patterns (MAMPs) by Pattern Recognition Receptors (PRRs). Among MAMPs, the lipopolysaccharide (LPS) is recognized by the Toll-Like-Receptor 4 (TLR4) and activates both the MyD88-dependent pathway at the plasma membrane and the TRIF-dependent pathway after TLR4 internalization. We previously showed that leptospiral LPS is not recognized by the human-TLR4, whereas it signals through mouse-TLR4 (mTLR4), which mediates mouse resistance to acute leptospirosis. However, although resistant, mice are known to be chronically infected by leptospires. Interestingly, the leptospiral LPS has low endotoxicity in mouse cells and is an agonist of TLR2, the sensor for bacterial lipoproteins. Here, we investigated the signaling properties of the leptospiral LPS in mouse macrophages. Using confocal microscopy and flow cytometry, we showed that the LPS of L. interrogans did not induce internalization of mTLR4, unlike the LPS of Escherichia coli. Consequently, the LPS failed to induce the production of the TRIF-dependent nitric oxide and RANTES, both important antimicrobial responses. Using shorter LPS and LPS devoid of TLR2 activity, we further found this mTLR4-TRIF escape to be dependent on both the co-purifying lipoproteins and the full-length O antigen. Furthermore, our data suggest that the O antigen could alter the binding of the leptospiral LPS to the co-receptor CD14 that is essential for TLR4-TRIF activation. Overall, we describe here a novel leptospiral immune escape mechanism from mouse macrophages and hypothesize that the LPS altered signaling could contribute to the stealthiness and chronicity of the leptospires in mice.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Leptospira/imunologia , Leptospirose/imunologia , Lipopolissacarídeos/metabolismo , Lipoproteínas/metabolismo , Antígenos O/metabolismo , Receptor 4 Toll-Like/fisiologia , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Citocinas/metabolismo , Feminino , Leptospirose/metabolismo , Leptospirose/microbiologia , Leptospirose/patologia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , Lipoproteínas/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/fisiologia , Antígenos O/genética , Transdução de Sinais , Receptor 2 Toll-Like/fisiologia
6.
World Neurosurg ; 138: 535-540.e8, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32113992

RESUMO

BACKGROUND: Cerebral cavernous malformations (CCMs) are vascular capillary anomalies with a dysfunctional endothelial adherent junction profile, depicting hemorrhage and epilepsy as the main clinical features. With the advent of an increasingly personalized medicine, better comprehension of genetic mechanisms behind CCM represents an important key in the management of the patients and risk rating in relatives. In this context, genetic factors that might influence clinical expressiveness of CCM need to be identified. CASE DESCRIPTION: A 33-year-old woman harboring multiple CCM lesions with a CCM1 mutational profile already being treated conservatively for a right mesial temporal lobe CCM presented with refractory seizures. Magnetic resonance imaging showed no bleeding in the lesion, and the patient was submitted to complete resection of the CCM. Histopathology of the CCM samples depicted an extensive inflammatory reaction and colocalization of CD20+ and CD68+ cells. Genetic analyses of the patient and her mother demonstrated a novel CCM1 (KRIT1) frameshift mutation (c.1661_1662insT; p.Leu554PhefsTer14). Furthermore, variants in CD14 (rs778588), TLR-4 (rs10759930), SOD2 (rs4880), APEX1 (rs1130409), and OGG1 (rs1052133), known as polymorphisms related to disease aggressiveness, were detected in the patient and not in her oligosymptomatic mother harboring the same CCM1 mutation. CONCLUSIONS: Heterogeneity of clinical manifestations among individuals with familial CCM with the same genotype adds mechanistic involvement of modifier factors as phenotypic markers. We describe a novel CCM1/KRIT1 familial mutation in which the coexistence of genetic variants in inflammation and oxidative stress may be related to variable expressiveness of the disease.


Assuntos
Mutação da Fase de Leitura , Hemangioma Cavernoso do Sistema Nervoso Central/genética , Proteína KRIT1/genética , Adulto , Anticonvulsivantes/uso terapêutico , Brasil , DNA Glicosilases/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Feminino , Genótipo , Hemangioma Cavernoso do Sistema Nervoso Central/complicações , Hemangioma Cavernoso do Sistema Nervoso Central/diagnóstico por imagem , Hemangioma Cavernoso do Sistema Nervoso Central/cirurgia , Humanos , Inflamação/genética , Receptores de Lipopolissacarídeos/genética , Imagem por Ressonância Magnética , Mães , Estresse Oxidativo/genética , Fenótipo , Convulsões/tratamento farmacológico , Convulsões/etiologia , Índice de Gravidade de Doença , Superóxido Dismutase/genética , Receptor 4 Toll-Like/genética
7.
Int J Mol Sci ; 21(4)2020 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-32070062

RESUMO

The majority of meningiomas are grade I, but some grade I tumours are clinically more aggressive. Recent advances in the genetic study of meningiomas has allowed investigation into the influence of genetics on the tumour microenvironment, which is important for tumorigenesis. We have established that the endpoint genotyping method Kompetitive Allele Specific PCR (KASP™) is a fast, reliable method for the screening of meningioma samples into different non-NF2 mutational groups using a standard real-time PCR instrument. This genotyping method and four-colour flow cytometry has enabled us to assess the variability in the largest immune cell infiltrate population, M2 macrophages (CD45+HLA-DR+CD14+CD163+) in 42 meningioma samples, and to suggest that underlying genetics is relevant. Further immunohistochemistry analysis comparing AKT1 E17K mutants to WHO grade I NF2-negative samples showed significantly lower levels of CD163-positive activated M2 macrophages in meningiomas with mutated AKT1 E17K, signifying a more immunosuppressive tumour microenvironment in NF2 meningiomas. Our data suggested that underlying tumour genetics play a part in the development of the immune composition of the tumour microenvironment. Stratifying meningiomas by mutational status and correlating this with their cellular composition will aid in the development of new immunotherapies for patients.


Assuntos
Macrófagos/metabolismo , Meningioma/genética , Proteínas Proto-Oncogênicas c-akt/genética , Microambiente Tumoral/genética , Alelos , Antígenos CD/genética , Antígenos de Diferenciação Mielomonocítica/genética , Linhagem da Célula/genética , Feminino , Genótipo , Antígenos HLA-DR/genética , Humanos , Antígenos Comuns de Leucócito/genética , Receptores de Lipopolissacarídeos/genética , Macrófagos/classificação , Macrófagos/patologia , Masculino , Meningioma/classificação , Meningioma/patologia , Pessoa de Meia-Idade , Mutação/genética , Neurofibromina 2/genética , Receptores de Superfície Celular/genética
8.
Eur J Immunol ; 50(5): 685-694, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32012247

RESUMO

Syncytin-1 is the envelope protein of the human endogenous retrovirus W (HERV-W). It has been related to multiple sclerosis (MS) but its role in cellular immunity and its pathogenic mechanism in the autoimmune context are not fully understood. We analyzed syncytin-1 levels in peripheral blood mononuclear cells (PBMC) subsets from healthy donors, MS patients in relapse or remission, and patients with acute infections by flow cytometry. PBMC cultures were also prepared to analyze protein expression kinetics. MS patients had higher levels of syncytin-1 levels than controls. We found that syncytin-1 is elevated in monocytes during MS relapses and infections. Cells expressing syncytin-1, including monocytes, T and B lymphocytes, and NKs presented mainly an activated phenotype and, upon stimulation with LPS, its levels increased rapidly on antigen-presenting cells. Syncytin-1 ligation promoted the activation of monocytes, as demonstrated by the upregulation of CD80 and the nonclassical subset CD14low CD16+ . Our results suggest an important role for syncytin-1 in the activation of leukocytes. Given that the expression of syncytin-1 is upregulated in MS patients, this protein might be contributing to the autoimmune cascade in the disease.


Assuntos
Retrovirus Endógenos/imunologia , Produtos do Gene env/genética , Monócitos/virologia , Esclerose Múltipla/genética , Esclerose Múltipla/virologia , Proteínas da Gravidez/genética , Adulto , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/virologia , Antígeno B7-1/genética , Antígeno B7-1/imunologia , Estudos de Casos e Controles , Retrovirus Endógenos/genética , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Regulação da Expressão Gênica , Produtos do Gene env/imunologia , Humanos , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/virologia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/imunologia , Lipopolissacarídeos/farmacologia , Masculino , Pessoa de Meia-Idade , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Proteínas da Gravidez/imunologia , Cultura Primária de Células , Receptores de IgG/genética , Receptores de IgG/imunologia , Recidiva , Indução de Remissão , Transdução de Sinais , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/virologia
9.
Genes (Basel) ; 11(1)2020 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-31963925

RESUMO

Immune response to infections has been shown to be mediated by genetic diversity in pattern recognition receptors, leading to disease tolerance or susceptibility. We elucidated naturally occurring variations within the bovine CD14 gene promoter in trypanosome-tolerant (N'Dama) and susceptible (White Fulani) cattle, with genomic and computational approaches. Blood samples were collected from White Fulani and N'Dama cattle, genomic DNA extracted and the entire promoter region of the CD14 gene amplified by PCR. We sequenced this region and performed in silico computation to identify SNP variants, transcription factor binding sites, as well as micro RNAs in the region. CD14 promoter sequences were compared with the reference bovine genome from the Ensembl database to identify various SNPs. Furthermore, we validated three selected N'Dama specific SNPs using custom Taqman SNP genotyping assay for genetic diversity. In all, we identified a total of 54 and 41 SNPs at the CD14 promoter for N'Dama and White Fulani respectively, including 13 unique SNPs present in N'Dama only. The significantly higher SNP density at the CD14 gene promoter region in N'Dama may be responsible for disease tolerance, possibly an evolutionary adaptation. Our genotype analysis of the three loci selected for validation show that mutant alleles (A/A, C/C, and A/A) were adaptation profiles within disease tolerant N'Dama. A similar observation was made for our haplotype analysis revealing that haplotypes H1 (ACA) and H2 (ACG) were significant combinations within the population. The SNP effect prediction revealed 101 and 89 new transcription factor binding sites in N'Dama and White Fulani, respectively. We conclude that disease tolerant N'Dama possessing higher SNP density at the CD14 gene promoter and the preponderance of mutant alleles potentially confirms the significance of this promoter in immune response, which is lacking in susceptible White Fulani. We, therefore, recommend further in vitro and in vivo study of this observation in infected animals, as the next step for understanding genetic diversity relating to varying disease phenotypes in both breeds.


Assuntos
Bovinos/genética , Predisposição Genética para Doença/genética , Receptores de Lipopolissacarídeos/genética , Adaptação Biológica/genética , Animais , Cruzamento , Doenças dos Bovinos/genética , Suscetibilidade a Doenças , Feminino , Genoma/genética , Genômica/métodos , Genótipo , Masculino , Nigéria , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas/genética , Infestações por Carrapato/genética , Trypanosoma/genética , Trypanosoma/patogenicidade
10.
Dev Comp Immunol ; 103: 103529, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31669309

RESUMO

The myeloid differentiation protein 2 (MD2)-related lipid-recognition (ML) proteins display diverse biological functions in host immunity and lipid metabolism by interacting with different lipids. Human MD2, an indispensable accessory protein in TLR4 signaling pathway, specifically recognizes lipopolysaccharides (LPS), thereby leading to the activation of TLR4 signaling pathway to produce many effectors that participate in inflammatory and immuneresponses against Gram-negative bacteria. Toll and immune deficiency (IMD) pathways are first characterized in Drosophila and are reportedly present in crustaceans, but the recognition and activation mechanism of these signaling pathways in crustaceans remains unclear. In the present study, a novel ML protein was characterized in mud crab (Scylla paramamosain) and designated as SpMD2. The complete SpMD2 cDNA sequence is 1114 bp long with a 465 bp open reading frame; it encodes a protein that contains 154 amino acids (aa). In the deduced protein, a signal peptide (1-21 aa residues) and a ML domain (43-151 aa residues) were predicted. SpMD2 shared a similar three-dimensional structure and a close evolutionary relationship with human MD2. SpMD2 was highly expressed in gills, hemocytes, intestine, and hepatopancreas and was upregulated in gills and hemocytes after challenges with bacteria, thereby suggesting its involvement in antibacterial defense. Western blot assay showed that SpMD2 possesses strong binding activities to different bacteria and two fungi. ELISA demonstrated that SpMD2 exhibits binding abilities to LPS, lipid A, peptidoglycan (PGN), and lipoteichoic acid (LTA). Its binding ability to LPS and lipid A were stronger than to PGN or LTA, implying that SpMD2 was an important LPS-binding protein in mud crab. Bacterial clearance assay revealed that the pre-incubation of Vibrio parahemolyticus with SpMD2 facilitates bacterial clearance in vivo and that knockdown of SpMD2 dramatically suppresses the bacterial clearance and decreases the expression of several antimicrobial peptides (AMPs). Furthermore, SpMD2 overexpression could enhance the promoter activity of SpALF2. These results revealed that SpMD2 affects bacterial clearance by regulating AMPs. Thus, by binding to LPS and by regulating AMPs, SpMD2 may function as a potential receptor, which is involved in the recognition and activation of a certain immune signaling pathway against Gram-negative bacteria. This study provides new insights into the diverse functions of ML proteins and into the antibacterial mechanisms of crustaceans.


Assuntos
Proteínas de Artrópodes/imunologia , Braquiúros/imunologia , Receptores de Lipopolissacarídeos/imunologia , Antígeno 96 de Linfócito/imunologia , Animais , Peptídeos Catiônicos Antimicrobianos/imunologia , Proteínas de Artrópodes/genética , Receptores de Lipopolissacarídeos/genética , Antígeno 96 de Linfócito/genética , Vibrioses/imunologia , Vibrioses/veterinária , Vibrio parahaemolyticus/imunologia
11.
J Immunol Res ; 2019: 1462098, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31815150

RESUMO

Diabetes mellitus, a metabolic disease characterized by hyperglycemia and poor glucose control, is a risk factor for Mycobacterium tuberculosis (M. tuberculosis) infection and the development of active tuberculosis. To evaluate whether M. tuberculosis infection susceptibility is associated with an intrinsic factor in monocytes from type 2 diabetes (T2D) patients or it is associated with hyperglycemia per se, we analyzed TLR-2 and TLR-4 expression by flow cytometry and the cytokines IL-1ß, IL-6, IL-8, IL-10, and TNF-α by cytometric bead array assays, either stimulated with TLR-2 and TLR-4 ligands or infected with M. tuberculosis in the whole blood from T2D patients (n = 43) and healthy subjects (n = 26) or in CD14+ monocytes from healthy subjects cultured in high glucose (HG) (30 mM). The intracellular growth of M. tuberculosis was evaluated by CFU counts at 0, 1, and 3 days in both monocytes from T2D patients and monocytes from healthy subjects cultured in HG. We did not find significant differences in TLR expression, cytokine production, or growth of M. tuberculosis in monocytes from T2D patients compared with those in monocytes from healthy subjects. Despite these results, in vitro assays of monocytes cultured with 30 mM glucose led to significantly increased TLR-2 and TLR-4 basal expression compared to those of monocytes cultured with 11 mM glucose (P < 0.05). Conversely, the production of IL-6 by TLR-2 ligand stimulation, of IL-1ß, IL-6, and IL-8 by TLR-4 ligand stimulation, and of IL-8 by M. tuberculosis infection significantly decreased in monocytes cultured in HG (P < 0.05). Additionally, the intracellular survival of M. tuberculosis increased in monocytes in HG after day 3 of culture (P < 0.05). In conclusion, HG decreased IL-8 production and the intracellular growth control of M. tuberculosis by monocytes, supporting the hypothesis that hyperglycemia plays an important role in the impaired immune responses to M. tuberculosis in patients with T2D.


Assuntos
Diabetes Mellitus Tipo 2/imunologia , Glucose/farmacologia , Hiperglicemia/imunologia , Monócitos/imunologia , Mycobacterium tuberculosis/crescimento & desenvolvimento , Tuberculose Pulmonar/imunologia , Adulto , Idoso , Estudos de Casos e Controles , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/microbiologia , Diabetes Mellitus Tipo 2/patologia , Feminino , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Glucose/metabolismo , Humanos , Hiperglicemia/metabolismo , Hiperglicemia/microbiologia , Hiperglicemia/patologia , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/imunologia , Lipopolissacarídeos/farmacologia , Lipoproteínas/farmacologia , Masculino , Pessoa de Meia-Idade , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Monócitos/microbiologia , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/patogenicidade , Cultura Primária de Células , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/imunologia , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Tuberculose Pulmonar/metabolismo , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
12.
Front Immunol ; 10: 2443, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31681320

RESUMO

Fracture repair is initiated by a multitude of immune cells and induction of an inflammatory cascade. Alterations in the early healing response due to an aged adaptive immune system leads to impaired bone repair, delayed healing or even formation of non-union. However, immuno-senescence is not limited to the adaptive immunity, but is also described for macrophages, main effector cells from the innate immune system. Beside regulation of pro- and anti-inflammatory signaling, macrophages contribute to angiogenesis and granulation tissue maturation. Thus, it seems likely that an altered macrophage function due to aging may affect bone repair at various stages and contribute to age related deficiencies in bone regeneration. To prove this hypothesis, we analyzed the expression of macrophage markers and angiogenic factors in the early bone hematoma derived from young and aged osteotomized Spraque Dawley rats. We detected an overall reduced expression of the monocyte/pan-macrophage markers CD14 and CD68 in aged rats. Furthermore, the analysis revealed an impaired expression of anti-inflammatory M2 macrophage markers in hematoma from aged animals that was connected to a diminished revascularization of the bone callus. To verify that the age related disturbed bone regeneration was due to a compromised macrophage function, CD14+ macrophage precursors were transplanted locally into the osteotomy gap of aged rats. Transplantation rescued bone regeneration partially after 6 weeks, demonstrated by a significantly induced deposition of new bone tissue, reduced fibrosis and significantly improved callus vascularization.


Assuntos
Envelhecimento/imunologia , Regeneração Óssea/imunologia , Fraturas Ósseas/imunologia , Macrófagos/imunologia , Cicatrização/imunologia , Fatores Etários , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Antígenos de Diferenciação Mielomonocítica/imunologia , Antígenos de Diferenciação Mielomonocítica/metabolismo , Biomarcadores/metabolismo , Regeneração Óssea/genética , Regeneração Óssea/fisiologia , Osso e Ossos/irrigação sanguínea , Osso e Ossos/imunologia , Osso e Ossos/lesões , Feminino , Expressão Gênica/genética , Expressão Gênica/imunologia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/imunologia , Receptores de Lipopolissacarídeos/metabolismo , Macrófagos/classificação , Macrófagos/metabolismo , Osteotomia , Ratos Sprague-Dawley , Cicatrização/genética
13.
Int J Mol Sci ; 20(20)2019 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-31658592

RESUMO

The microneme organelles of Toxoplasma gondii tachyzoites release protein complexes (MICs), including one composed of the transmembrane protein MIC6 plus MIC1 and MIC4. In this complex, carbohydrate recognition domains of MIC1 and MIC4 are exposed and interact with terminal sialic acid and galactose residues, respectively, of host cell glycans. Recently, we demonstrated that MIC1 and MIC4 binding to the N-glycans of Toll-like receptor (TLR) 2 and TLR4 on phagocytes triggers cell activation and pro-inflammatory cytokine production. Herein, we investigated the requirement for TLR2 heterodimerization and co-receptors in MIC-induced responses, as well as the signaling molecules involved. We used MICs to stimulate macrophages and HEK293T cells transfected with TLR2 and TLR1 or TLR6, both with or without the co-receptors CD14 and CD36. Then, the cell responses were analyzed, including nuclear factor-kappa B (NF-κB) activation and cytokine production, which showed that (1) only TLR2, among the studied factors, is crucial for MIC-induced cell activation; (2) TLR2 heterodimerization augments, but is not critical for, activation; (3) CD14 and CD36 enhance the response to MIC stimulus; and (4) MICs activate cells through a transforming growth factor beta-activated kinase 1 (TAK1)-, mammalian p38 mitogen-activated protein kinase (p38)-, and NF-κB-dependent pathway. Remarkably, among the studied factors, the interaction of MIC1 and MIC4 with TLR2 N-glycans is sufficient to induce cell activation, which promotes host protection against T. gondii infection.


Assuntos
Moléculas de Adesão Celular/química , Moléculas de Adesão Celular/metabolismo , Dimerização , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Receptor 2 Toll-Like/química , Receptor 2 Toll-Like/metabolismo , Toxoplasma/metabolismo , Animais , Antígenos CD36/genética , Antígenos CD36/metabolismo , Citocinas/análise , Feminino , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Células RAW 264.7 , Transdução de Sinais , Receptor 1 Toll-Like/metabolismo , Receptor 6 Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
14.
PLoS One ; 14(9): e0222912, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31569199

RESUMO

To confirm that neoplastic monocyte-derived collagen- and fibronectin-producing fibrocytes induce bone marrow (BM) fibrosis in primary myelofibrosis (PMF), we injected PMF BM-derived fibrocyte-precursor CD14+/CD34- monocytes into the tail vein of NOD-SCID-γ (NSG) mice. PMF BM-derived CD14+/CD34- monocytes engrafted and induced a PMF-like phenotype with splenomegaly, myeloid hyperplasia with clusters of atypical megakaryocytes, persistence of the JAK2V617F mutation, and BM and spleen fibrosis. As control we used normal human BM-derived CD14+/CD34- monocytes. These monocytes also engrafted and gave rise to normal megakaryocytes that, like PMF CD14+/CD34--derived megakaryocytes, expressed HLA-ABC and human CD42b antigens. Using 2 clonogenic assays we confirmed that PMF and normal BM-derived CD14+/CD34- monocytes give rise to megakaryocyte colony-forming cells, suggesting that a subpopulation BM monocytes harbors megakaryocyte progenitor capacity. Taken together, our data suggest that PMF monocytes induce myelofibrosis-like phenotype in immunodeficient mice and that PMF and normal BM-derived CD14+/CD34- monocytes give rise to megakaryocyte progenitor cells.


Assuntos
Células da Medula Óssea/imunologia , Fibroblastos/imunologia , Hiperplasia/imunologia , Hospedeiro Imunocomprometido , Monócitos/imunologia , Mielofibrose Primária/imunologia , Esplenomegalia/imunologia , Transferência Adotiva , Animais , Antígenos CD34/genética , Antígenos CD34/imunologia , Células da Medula Óssea/patologia , Feminino , Fibroblastos/patologia , Fibroblastos/transplante , Expressão Gênica , Antígenos HLA/genética , Antígenos HLA/imunologia , Humanos , Hiperplasia/etiologia , Hiperplasia/genética , Hiperplasia/patologia , Janus Quinase 2/genética , Janus Quinase 2/imunologia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/imunologia , Megacariócitos/imunologia , Megacariócitos/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Monócitos/patologia , Monócitos/transplante , Mutação , Mielofibrose Primária/etiologia , Mielofibrose Primária/genética , Mielofibrose Primária/patologia , Esplenomegalia/etiologia , Esplenomegalia/genética , Esplenomegalia/patologia
15.
Hum Immunol ; 80(11): 930-936, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31474499

RESUMO

Variable immune response to external stimuli remains a major concern in sickle cell disease (SCD), with such responses predicted to be contributors to disease pathogenesis. Elucidating the diversity of host genes contributing to immune response would assist to clarify differing outcomes among and between disease groups. We hypothesize that there is a significant interethnic diversity in the CD14 (rs2569190), CD28 (rs35593994), CTLA-4 (rs5742909) and ICOS (rs4404254) gene polymorphisms among and between SCD groups. We genotyped single nucleotide polymorphisms of the 4 loci among African and African American SCD and control groups and between SCD groups. In all, 375 individuals from Mali (145 SCD and 230 controls) and 700 DNA samples from the United States (321 SCD and 379 controls) were subjected to a PCR-RFLP assay. We found no intraethnic difference in genotypic and allelic frequencies of the 4 loci among Africans and African Americans, potentially significant in disease association studies, including a similar observation for interethnic frequencies of CD28, CTLA-4 and ICOS genes, but not CD14. The CD14 (rs2569190) gene promoter demonstrated a significant difference (p < 0.02) between African and African American SCD groups, with the mutant variant (-159 T/T) more frequent (p < 0.0002) in African American SCD (38.9% versus 26.2%). The higher frequency of CD14 mutants among African Americans without an accompanying defect in CD28, CTLA-4 and ICOS diversity possibly indicates a defective innate response, driven by CD14, is untethered to downstream T cell differentiation or effector function. Additionally, we show that CD28 (rs35593994) mutant variants have no impact on T cell differentiation, as the ICOS gene provides an alternative pathway to override this impairment. We conclude that in spite of the defect in CD14, T cell selection and differentiation is unimpeded and a robust adaptive immune response initiated.


Assuntos
Afro-Americanos , Anemia Falciforme/genética , Genótipo , Receptores de Lipopolissacarídeos/genética , Regiões Promotoras Genéticas/genética , Adolescente , Adulto , Anemia Falciforme/epidemiologia , Antígenos CD28/genética , Antígeno CTLA-4/genética , Criança , Pré-Escolar , Feminino , Frequência do Gene , Estudos de Associação Genética , Humanos , Proteína Coestimuladora de Linfócitos T Induzíveis/genética , Lactente , Louisiana/epidemiologia , Masculino , Mali/epidemiologia , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Adulto Jovem
16.
Biochem Biophys Res Commun ; 519(4): 909-915, 2019 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-31563324

RESUMO

1,25-Dihydroxyvitamin D3 or 1,25(OH)2D3 is known to play an important role in the differentiation of human myeloid cells. However, the molecular mechanism underlying the 1,25(OH)2D3-mediated differentiation of human myeloid cells is incompletely understood. Here, we report that 1,25(OH)2D3 induces differentiation of human myeloid cell lines such as U937 and THP-1 cells via the mammalian target of rapamycin (mTOR) signaling pathway. Both the expression of the differentiation marker CD14 and activation of the mTOR signaling pathway were induced by 1,25(OH)2D3 in phorbol 12-myristate 13-acetate (PMA)-differentiated U937 and THP-1 cells. The 1,25(OH)2D3-induced expression of CD14 in PMA-differentiated U937 and THP-1 cells was prevented by mTOR inhibitors, PP242 and Torin1. The 1,25(OH)2D3-induced morphological changes as characteristics of differentiated myeloid cells were also reversed after PP242 and Torin1 treatment. Silencing of either regulatory-associated protein of mTOR (Raptor) or rapamycin-insensitive companion of mTOR (Rictor) in PMA-differentiated THP-1 cells with small-interfering RNA resulted in the inhibition of CD14 expression and morphological changes induced by 1,25(OH)2D3, indicating that both mTORC1 and mTORC2 were important for the differentiation of myeloid THP-1 cells. Previous studies have shown that phosphatidic acid (PA) maintains the stability of the mTOR complex. Here we found that the attenuation of PA production with 1-butanol or a PLD inhibitor prevented the 1,25(OH)2D3-induced upregulation of CD14. Taken together, our results show that 1,25(OH)2D3 enhances the differentiation of human myeloid cells through the mTOR signaling pathway.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Células Mieloides/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Vitamina D/análogos & derivados , Expressão Gênica/efeitos dos fármacos , Humanos , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Células Mieloides/metabolismo , Células Mieloides/patologia , Ácidos Fosfatídicos/metabolismo , Células THP-1 , Acetato de Tetradecanoilforbol/farmacologia , Células U937 , Vitamina D/farmacologia , Vitaminas/farmacologia
17.
BMC Med Genomics ; 12(1): 130, 2019 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-31519222

RESUMO

BACKGROUND: Pneumonia, sepsis, meningitis, and empyema due to Streptococcus pneumoniae is a major cause of morbidity and mortality. We provide a systemic overview of genetic variants associated with susceptibility, phenotype and outcome of community acquired pneumococcal pneumonia (CAP) and invasive pneumococcal disease (IPD). METHODS: We searched PubMed for studies on the influence of host genetics on susceptibility, phenotype, and outcome of CAP and IPD between Jan 1, 1983 and Jul 4, 2018. We listed methodological characteristics and when genetic data was available we calculated effect sizes. We used fixed or random effect models to calculate pooled effect sizes in the meta-analysis. RESULTS: We identified 1219 studies of which 60 studies involving 15,358 patients were included. Twenty-five studies (42%) focused on susceptibility, 8 (13%) on outcome, 1 (2%) on disease phenotype, and 26 (43%) on multiple categories. We identified five studies with a hypothesis free approach of which one resulted in one genome wide significant association in a gene coding for lincRNA with pneumococcal disease susceptibility. We performed 17 meta-analyses of which two susceptibility polymorphisms had a significant overall effect size: variant alleles of MBL2 (odds ratio [OR] 1·67, 95% confidence interval [CI] 1·04-2·69) and a variant in CD14 (OR 1·77, 95% CI 1·18-2·66) and none of the outcome polymorphisms. CONCLUSIONS: Studies have identified several host genetics factors influencing risk of pneumococcal disease, but many result in non-reproducible findings due to methodological limitations. Uniform case definitions and pooling of data is necessary to obtain more robust findings.


Assuntos
Infecções Pneumocócicas/genética , Polimorfismo Genético , Suscetibilidade a Doenças , Humanos , Receptores de Lipopolissacarídeos/genética , Lectina de Ligação a Manose/genética , Razão de Chances , Fenótipo , Infecções Pneumocócicas/microbiologia , Infecções Pneumocócicas/patologia , RNA Longo não Codificante/metabolismo , Fatores de Risco
18.
Adv Clin Exp Med ; 28(8): 1087-1094, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31403268

RESUMO

BACKGROUND: There is evidence that suggests variation in gene encoding pattern recognition receptors, the essential components of innate immunity, might be associated with atopic diseases. However, results have been inconclusive. OBJECTIVES: The aim of the study was to determine the individual associations and possible interactive effects of the CD14 (cluster of differentiation 14), TLR4 (toll-like receptor 4) and TLR2 (toll-like receptor 2) polymorphisms on allergic diseases. MATERIAL AND METHODS: The CD14 C-159T, TLR4 +896A/G and TLR2 A-16934T polymorphisms were identified in 115 children aged from 6 to 17 years. All subjects were selected using a detailed questionnaire which included questions on symptoms and each one underwent skin prick testing. All single-nucleotide polymorphisms (SNPs) were determined using real-time polymerase chain reaction (PCR) assays. RESULTS: There was no statistically significant correlation between the 3 polymorphisms (CD14 C-159T, TLR4 +896A/G and TLR2/-16934A/T) and either asthma, allergic rhinitis or atopy. We observed that children who were heterozygous or homozygous for both the CD14/-159T and TLR2/-16934A alleles had a 4-fold lower risk for asthma than children who were carriers of the T allele of CD14 but non-carriers of the A allele of TLR2, and an almost 3-fold lower risk for asthma when compared to all other groups. Concerning allergic rhinitis, a similar trend was observed. In addition, the presence of at least 1 A allele in the TLR2/-16934 polymorphism reduced the risk for asthma and allergic rhinitis, but only in children who were homozygous for the common A allele in the TLR4 +896 polymorphism. CONCLUSIONS: Our study supports the idea that the CD14, TLR2 and TLR4 polymorphisms may not be directly involved in the development of atopic diseases. However, our results suggest that their impact on the risk of asthma and allergic rhinitis might be modulated by gene-gene interactions.


Assuntos
Asma , Epistasia Genética , Imunidade Inata , Polimorfismo de Nucleotídeo Único , Adolescente , Alelos , Asma/genética , Asma/imunologia , Criança , Predisposição Genética para Doença , Humanos , Receptores de Lipopolissacarídeos/genética , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética
19.
J Alzheimers Dis ; 71(3): 751-761, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31450497

RESUMO

BACKGROUND: Identifying the mechanisms involved in the pathogenesis of Alzheimer's disease (AD) remains crucially important. Chronic age-related low-grade inflammation is considered to be one such mechanism, although its causes are unclear. Lipopolysaccharide (LPS)-type endotoxins, a major component of the outer membrane of Gram-negative bacteria, are known as potent pro-inflammatory molecules. Therefore, we hypothesized that greater exposure to circulating LPS, potentially mediated by the inflammatory pathway, would be a key step of the onset of AD. OBJECTIVE: The aim of this study was to investigate the link between plasma endotoxin-exposure, inflammation, and AD. METHODS: Applying a nested case-control design, we evaluated the associations among baseline plasma endotoxin-exposure (assessed by measuring LPS-binding protein (LBP) and soluble cluster of differentiation-14 (sCD14) levels), inflammation (assessed by measuring interleukin-6 (IL6) levels), and the odds of developing AD over 12 years. Selected from a population-based cohort, 212 incident cases of AD were matched with 424 controls without dementia with regard to age, gender, and education level. RESULTS: After adjusting for a large set of confounders, including the use of anti-inflammatory drugs, only higher LBP levels were significantly associated with a 30% higher odds of developing AD over 12 years (OR 1.30, 95% CIs [1.07-1.59]), regardless of IL6 levels. CONCLUSION: This large case-control study provides preliminary results concerning plasma endotoxin-exposure among the elderly and suggests that higher LBP levels, an acute-phase reactant involved in the pro-inflammatory response to LPS, are associated with higher odds of developing AD.


Assuntos
Proteínas da Fase Aguda/genética , Doença de Alzheimer/epidemiologia , Doença de Alzheimer/genética , Proteínas de Transporte/genética , Receptores de Lipopolissacarídeos/genética , Glicoproteínas de Membrana/genética , Idoso , Idoso de 80 Anos ou mais , Proteínas de Transporte/sangue , Estudos de Casos e Controles , Feminino , França/epidemiologia , Humanos , Incidência , Vida Independente , Interleucina-6/sangue , Receptores de Lipopolissacarídeos/sangue , Masculino , Glicoproteínas de Membrana/sangue , Projetos Piloto , População Urbana
20.
Hum Immunol ; 80(10): 855-862, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31285077

RESUMO

High mortality in pregnant women is a characteristic of hepatitis E virus (HEV) infection. Role of monocytes/T cells in HEV infection during pregnancy is still unclear. We compared CD14+monocytes and CD4+T cells by flow-cytometry in hepatitis-E patients including 13 pregnant (Antenatal care, ANC), 25 non-ANC patients and respective controls (12 and 20). Non-ANC-patients showed significantly higher frequency of monocytes with increased expression of CD80, CD86 and HLA-DR than control individuals (p < 0.001). Healthy pregnancy was associated with increased frequency of monocytes with higher CD80 expression and lower levels of HLA-DR (p < 0.05) compared to non-ANC controls. ANC-patients exhibited elevated levels of monocytes (p < 0.01) with higher expression of CD80 (p < 0.001) and reduced levels of HLA-DR and CD86 (p < 0.05) when compared with non-ANC patients. TLR2 and TLR4 surface expression on monocytes was higher in non-ANC-patients (p < 0.00) and lower in the ANC-patients (p < 0.01). Healthy-ANCs exhibited lower TLR4 expression on monocytes (p < 0.05). HEV infection did not change the frequency of CD4+ and CD4+CD28+T cells in patients' group (p > 0.05). Compared to respective controls, CD137+ and CD152+CD4+T cells were higher (p < 0.05) in both patients' categories. Higher levels of CD152+CD4+T cells (p < 0.001) was noted in healthy pregnant women. Among patients' groups, the CD4+T cells and their subpopulation were not different (p > 0.05). We found higher and reduced levels of circulating inflammatory cytokines (IL12, TNFα, IL6 and IL8; miliplex-assay) in non-ANC and ANC-patients respectively. In conclusion, on contrary to the classical activation of CD14+monocytes in the non-ANC-patients, impaired response was evident in the ANC-patients while the CD4+T cell populations were similar in the patient groups.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Vírus da Hepatite E/imunologia , Hepatite E/genética , Monócitos/imunologia , Fenótipo , Adulto , Antígeno B7-1/genética , Antígeno B7-1/metabolismo , Antígeno B7-2/genética , Antígeno B7-2/metabolismo , Citocinas/sangue , Feminino , Antígenos HLA-DR/genética , Antígenos HLA-DR/metabolismo , Hepatite E/patologia , Humanos , Índia , Receptores de Lipopolissacarídeos/genética , Receptores de Lipopolissacarídeos/metabolismo , Masculino , Gravidez , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Adulto Jovem
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