RESUMO
Germinal centers (GCs) are some of the most important structures in the human immune system. As such, their cell types and functions have been thoroughly investigated. B cells, T cells, follicular dendritic cells (FDCs), and macrophages have widely been found to typically be aggregated in GCs. However, the amount of space occupied by each of these cell types has yet to be investigated. In this study, we conducted confocal laser-based 3D cell-volume quantification of typical GC cells under reactive conditions in lymphadenitis and investigated how volume proportions change during GC development. For this investigation, we used anti-CD3 (T cells), anti-CD20 and anti-Pax5 (B cells), anti-CD23 (FDCs), anti-CD68 (macrophages), and DAPI (nuclear staining). We detected average proportions of about 11% CD3, 9% CD20, 6% CD23, and 2% CD68 in the largest possible regions of interest within GCs. Interestingly, these values remained steady relatively independent of GC size. The remarkably low B cell proportion can be attributed to technical constraints given the use of the CD20 antibody in 3D. Applying the B cell marker Pax5, we found that about 44% of the volume was occupied by B cells after extrapolating the volume of B cell nuclei to that of whole B cells. We concluded that Pax5 is more suitable than anti-CD20 for 3D B cell quantification in GCs. The substantial unstained volume in GCs raises the question of whether other cell types fill these open spaces. Our 3D investigation enabled a unique morphological and volumetric evaluation of GC cells that balance their overall volumes in GCs.
Assuntos
Células Dendríticas Foliculares , Linfadenite , Humanos , Células Dendríticas Foliculares/metabolismo , Linfócitos T , Centro Germinativo , Macrófagos , Linfadenite/metabolismoRESUMO
INTRODUCTION: Cervical scrofulous lymphadenitis due to Mycobacterium avium complex (MAC) in immunocompetent adults is a rare disease. The presence of MAC infections demands meticulous clinical evaluation of patients along with detailed phenotypic and functional evaluation of their immune system including next-generation sequencing (NGS) analyses of target genes. METHODS: Exact clinical histories of the index patients both suffering from retromandibular/cervical scrofulous lymphadenitis were obtained along with phenotypic and functional immunological evaluations of leukocyte populations followed by targeted NGS-based sequencing of candidate genes. RESULTS: Immunological investigations showed normal serum immunoglobulin and complement levels, but lymphopenia, which was caused by significantly reduced CD3+CD4+CD45RO+ memory T-cell and CD19+ B-cell numbers. Despite normal T-cell proliferation to a number of accessory cell-dependent and -independent stimuli, the PBMC of both patients elaborated clearly reduced levels of a number of cytokines, including IFN-γ, IL-10, IL-12p70, IL-1α, IL-1ß, and TNF-α upon TCR-dependent T-cell stimulation with CD3-coated beads but also superantigens. The IFN-γ production deficiency was confirmed for CD3+CD4+ helper and CD4+CD8+ cytotoxic T cells on the single-cell level by multiparametric flow cytometry irrespective of whether PMA/ionomycin-stimulated whole blood cells or gradient-purified PBMC was analyzed. In the female patient L1, targeted NGS-based sequencing revealed a homozygous c.110T>C mutation in the interferon-γ receptor type 1 (IFNGR1) leading to significantly reduced receptor expression on both CD14+ monocytes and CD3+ T cells. Patient S2 presented with normal IFNGR1 expression on CD14+ monocytes but significantly reduced IFNGR1 expression on CD3+ T cells, despite the absence of detectable homozygous mutations in the IFNGR1 itself or disease-related target genes. Exogenous addition of increasing doses of IFN-γ resulted in proper upregulation of high-affinity FcγRI (CD64) on monocytes from patient S2, whereas monocytes from patient L1 showed only partial induction of CD64 expression after incubation with high doses of IFN-γ. CONCLUSION: A detailed phenotypic and functional immunological examination is urgently required to determine the cause of a clinically relevant immunodeficiency, despite detailed genetic analyses.
Assuntos
Linfadenite , Infecção por Mycobacterium avium-intracellulare , Adulto , Humanos , Feminino , Complexo Mycobacterium avium/genética , Complexo Mycobacterium avium/metabolismo , Leucócitos Mononucleares , Infecção por Mycobacterium avium-intracellulare/genética , Infecção por Mycobacterium avium-intracellulare/metabolismo , Citocinas/metabolismo , Linfadenite/metabolismoRESUMO
Classical Hodgkin lymphoma (cHL) is one of the most common malignant lymphomas in Western Europe. It is diagnosed on the basis of histological sections by pathologists using a light microscope. The tumor cells, the Hodgkin- and Reed Sternberg cells (HRS), are visualized by morphology and positive response for the CD30-antigen. The same antigen can also be detected by immunohistochemistry on a reactive counterpart, showing CD30+ cells in special immunoreactions, such as inflammations of lymph nodes (lymphadenitis). CD30+ cells in reactive and neoplastic conditions are surrounded by lymphocytes and histiocytes, forming a micromilieu that enables the survival of the tumor cells, as well as their reactive counterparts. This study deals with an investigation of CD30+-surrounding cells using a confocal laser technology, visualizing the contacts of reactive and neoplastic CD30+ cells with CD68+ macrophages and CD163+ macrophages as well as to PD1+ lymphocytes and B cells (CD20+). CD4 immunostains were not included, because CD4+ cells were too numerous for clear dissection of single cells. 3D images visualized the, so-called, connectomes. Clear differences in the number of contacts between CD30-reactive and neoplastic cells (HRS) with macrophages and B lymphocytes were visible. Lymphadenitis and Mixed Cellularity type of classical Hodgkin Lymphoma (cHL) differed in that Mixed Cellularity (MC) cHL had more connections to macrophages (CD163+) and lower number of connections to B cells (CD20+). The connectomes of both Hodgkin variants MCcHL and Nodular Sclerosis cHL (NScHL) mainly differed in the number of contacts to CD163+ macrophages, which was higher in MCcHL. Investigating the volumes of CD30+ -reactive and neoplastic cells, we found out that reactive cells showed lesser volumes, which correlated with the number of contacts. The comparison between 2D and 3D images, including 3D prints, demonstrated clear advantages of the 3D method. 3D images visualized significantly more and clearly defined intercellular contacts. Complicated cellular networks and their contacts became especially evident in volume and surface evaluations, as well as in 3D prints.
Assuntos
Conectoma , Doença de Hodgkin/metabolismo , Imageamento Tridimensional/métodos , Antígeno Ki-1/metabolismo , Linfócitos/metabolismo , Biomarcadores Tumorais/metabolismo , Humanos , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Inflamação , Linfonodos/patologia , Linfadenite/metabolismo , Linfoma/metabolismo , Microscopia Confocal , Células de Reed-Sternberg/metabolismo , Software , Microambiente TumoralRESUMO
Necrotizing lymphadenitis (NEL) is a self-limited systemic disease exhibiting characteristic clinical features. The pathogenesis of the disease remains unclear, but it may be associated with viral infection. In lymph nodes affected by this disease, innumerable plasmacytoid dendritic cells produce interferon-α when triggered by certain viral stimuli. IFN-α presents antigens causing the transformation of CD8+ cells into immunoblasts and apoptosis of CD4+ cells. From the perspective of innate immunity, UNC93B1, an endoplasmic reticulum (ER)-resident protein, associates more strongly with TLR9 than TLR7. Homeostasis is maintained under normal conditions. However, in NEL, TLR 7 was observed more than TLR 9, possibly because mutant type UNC93B1 associates more tightly with TLR7. The inhibitory effects against TLR7 by TLR9 were reported to disappear. It is likely that more TLR7 than TLR9 is transported from the ER to endolysosomes. In conclusion, overexpression of TLR7, an innate immune sensor of microbial single-stranded RNA, is inferred. Consequently, NEL may be induced.
Assuntos
Células Dendríticas/patologia , Linfadenite/patologia , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Células Dendríticas/metabolismo , Feminino , Humanos , Linfadenite/metabolismo , Masculino , Pessoa de Meia-Idade , Transporte Proteico , Receptor 7 Toll-Like/análise , Receptor Toll-Like 9/análise , Adulto JovemRESUMO
Autoinflammatory disorders of the innate immune system present with recurrent episodes of inflammation often beginning in early childhood. While there are now more than 30 genetically-defined hereditary fever disorders, many patients lack a clear diagnosis. Many pediatric patients are often grouped with patients with periodic fever, aphthous stomatitis, pharyngitis, and adenitis (PFAPA) syndrome despite failing to meet diagnostic criteria. Here, we categorize these patients as syndrome of undifferentiated recurrent fever (SURF), and identify the unique features which distinguish them from the PFAPA syndrome. SURF patients were more likely to report gastrointestinal symptoms of nausea, vomiting and abdominal pain, and experienced inconsistent responses to on-demand steroid therapy compared to PFAPA patients. For this previously undefined cohort, an optimal course of therapy remains uncertain, with medical and surgical therapies largely driven by parental preference. A subset of patients with SURF underwent tonsillectomy with complete resolution. Flow cytometric evaluation demonstrates leukocytic populations distinct from PFAPA patients, with reduced CD3+ T cell numbers. SURF patient tonsils were predominantly characterized by an IL-1 signature compared to PFAPA, even during the afebrile period. Peripheral blood signatures were similar between groups suggesting that PFAPA and SURF patient tonsils have localized, persistent inflammation, without clinical symptoms. These data suggest that SURF is a heterogenous syndrome on the autoinflammatory disease spectrum.
Assuntos
Febre/diagnóstico , Doenças Hereditárias Autoinflamatórias/diagnóstico , Inflamação/diagnóstico , Interleucina-1/metabolismo , Linfadenite/diagnóstico , Faringite/diagnóstico , Estomatite Aftosa/diagnóstico , Complexo CD3/metabolismo , Pré-Escolar , Feminino , Febre/metabolismo , Gastroenteropatias/diagnóstico , Gastroenteropatias/metabolismo , Doenças Hereditárias Autoinflamatórias/metabolismo , Humanos , Inflamação/metabolismo , Linfadenite/metabolismo , Masculino , Tonsila Palatina/metabolismo , Pediatria , Faringite/metabolismo , Estomatite Aftosa/metabolismo , Síndrome , Linfócitos T/metabolismo , Tonsilectomia/métodosRESUMO
Hassanzad M, Farnia P, Darougar S, Velayati AA. A novel evaluation of genetic polymorphism in BCG adenitis. Turk J Pediatr 2019; 61: 466-470. Bacillus Calmette-Guerin (BCG) is a live attenuated vaccine which has been used to prevent tuberculosis, according to the World Health Organization (WHO) recommendation in parts of the world with an incidence of tuberculosis infection more than 1%. The incidence of BCG adverse reactions differs between regions with regional lymphadenitis as the most common presentation. The aim of this study was to detect the impact of polymorphisms causing BCG lymphadenitis in children receiving BCG vaccination at birth. Eight healthy infants with BCG adenitis from 4 to 12 months old were enrolled. All these patients underwent a thorough physical examination, abdominopelvic ultrasound evaluation to detect distant lymphadenopathies and immunodeficiency screening tests for any possible underlying immunodeficiency disorders. Then genotyping for known mutations was performed using restriction fragments length polymorphism (PCR-RFLP) assays. Sequencing was performed for IL-12 Rß1, IFN-Ï receptor 1, IL-10, TNF-α and P2X7. The mean age of onset of the adenitis was 6.5 months. TNF-857, IL-12Rß1 705, IL-10 1082, and IFN-Ï- 56 single nucleotide polymorphisms (SNPs) were common in the children studied. The most frequent polymorphism found in the patients with BCG adenitis except one, was the P2X7 -762 polymorphism. To conclude, these polymorphisms are more common in some ethnic populations but not others and make the genetic basis of immunity to BCG strains and the occurrence of post-BCG lymphadenitis in otherwise healthy children.
Assuntos
Vacina BCG/efeitos adversos , Citocinas/genética , Linfadenite/genética , Polimorfismo Genético , Tuberculose/prevenção & controle , Vacinação/efeitos adversos , Citocinas/metabolismo , Feminino , Humanos , Lactente , Linfadenite/etiologia , Linfadenite/metabolismo , Masculino , Mutação , Tuberculose/diagnóstico , UltrassonografiaRESUMO
INTRODUCTION: PFAPA (periodic fever, aphthous stomatitis, pharyngitis and cervical adenitis) is the most frequent non-infectious cause of high fever observed among the European child population. While its cause is still not yet fully identified, PFAPA patients were previously shown to have altered tonsillar microbiome composition. Our study hypothesized that this is associated with a change in antimicrobial peptide (AMP) expression levels, as in the case of Crohn's disease which is another autoinflammatory disorder. METHODS AND MATERIALS: The tonsil specimens were isolated from seven patients with PFAPA syndrome, and six patients with group A beta-hemolytic streptococcal (GAßHS) recurrent tonsillitis. Tonsillar expression levels of human beta-defensin 1-2, cathelicidin, ribonuclease-7, and liver expressed antimicrobial peptide-1 were monitored by immunohistochemistry (IHC). Expression levels were scored using semi-quantitative analysis method and were statistically analyzed by Two-Way Repeated Measures Analysis of Variance test. RESULTS: Our results showed no significant difference in AMP expression levels between PFAPA and GAßHS patients. Immunolocalization of human beta-defensin 1 was different between the two groups; expressed at higher levels on tonsil surface epithelium (SE) than lymphoid interior (LI) in PFAPA patient group, while this was not evident in GAßHS patients group. CONCLUSIONS: Our results suggest that, PFAPA patients may be associated with altered AMP expression as in other autoinflammatory diseases. Future studies with subjects without any inflammatory condition are required for more precise conclusions.
Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Febre/metabolismo , Linfadenite/metabolismo , Tonsila Palatina/metabolismo , Faringite/metabolismo , Estomatite Aftosa/metabolismo , Tonsilite/metabolismo , Adolescente , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Pescoço , Ribonucleases/metabolismo , Streptococcus pyogenes , Síndrome , Tonsilite/microbiologia , beta-Defensinas/metabolismo , CatelicidinasRESUMO
OBJECTIVE: PFAPA syndrome is a clinical entity of unknown etiology which presents with periodic episodes of fever, aphthous stomatitis, tonsillitis or pharyngitis, and cervical adenitis. In this study we investigated DNA damage and the oxidative stress parameters in patients diagnosed with PFAPA, to elucidate the underlying pathophysiological mechanism of this syndrome. METHODS: Thirty-one patients diagnosed with PFAPA (Group 1), 22 patients diagnosed with normal tonsillitis or pharyngitis (Group 2), and 20 healthy volunteers (Group 3) were included in our study. Heparinized peripheral blood samples were drawn from all patients and volunteers. DNA damage was assessed by single cell alkaline electrophoresis assay in peripheral mononuclear leukocytes. Plasma levels of total antioxidant status (TAS) and total oxidative status (TOS) were determined by using a novel automated measurement method, and oxidative stress index (OSI) was calculated. RESULTS: DNA damage in the mononuclear leukocytes of Group 1 was significantly higher than that of Group 2 and Group 3. The oxidative stress parameters revealed that the TOS and OSI values of Group 1 were significantly higher than those of Group 2 and Group 3. TAS values of Group 1 were significantly lower than those of Group 2 and Group 3. Correlation analysis of Group 1 demonstrated a significant correlation between TOS, one of the oxidative stress parameters, and DNA damage. Correlations between DNA damage and C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) values were also significant. CONCLUSION: Our study indicated that both the inflammatory and the oxidative stress parameters were significantly increased in patients with PFAPA syndrome, accompanied by a significant positive correlation between DNA damage and oxidative stress.
Assuntos
Dano ao DNA , Febre/metabolismo , Linfadenite/metabolismo , Estresse Oxidativo , Periodicidade , Faringite/metabolismo , Estomatite Aftosa/metabolismo , Tonsilite/metabolismo , Sedimentação Sanguínea , Proteína C-Reativa/metabolismo , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudos Transversais , Feminino , Febre/genética , Humanos , Leucócitos Mononucleares/metabolismo , Linfadenite/genética , Masculino , Pescoço , Faringite/genética , Estomatite Aftosa/genética , Síndrome , Tonsilite/genéticaRESUMO
INTRODUCTION: Trans-1-amino-3-[(18)F]fluorocyclobutanecarboxylic acid (anti-[(18)F]FACBC) is a positron emission tomography (PET) tracer used to visualize prostate cancer (PCa). In this study, we investigated the differences in anti-[(18)F]FACBC accumulation between metastatic and inflamed lymph node (LN) lesions. METHODS: A PCa LN metastasis (PLM) model was developed by inoculating a rat PCa cell line, MAT-Ly-Lu-B2, into popliteal LNs of Copenhagen rats. Acute lymphadenitis (AL) was induced by injecting concanavalin A (Con A) into the hind footpad, and chronic lymphadenitis (CL) was induced by daily injection of Con A into the tissues surrounding the popliteal LNs for 2weeks. Main lesions of all animal models were established in lumbar and/or inguinal LNs. Biodistribution and dynamic PET imaging data were acquired after tracer injection. T2-weighted magnetic resonance (MR) images were registered with PET images. RESULTS: In the biodistribution study, the uptake ratios of PLM-to-lymphadenitis in lesional lumbar and inguinal LNs were 0.97-1.57 and 1.47-2.08 at 15 and 60min post-anti-[(18)F]FACBC injection respectively. In PET imaging, the lesional lumbar LNs of CL and PLM, but not of AL, were visualized on anti-[(18)F]FACBC-PET/MR fusion images without disturbance from radioactivity from urine, andãthe rank order of anti-[(18)F]FACBC accumulation at 50-60 post-injection in lesional lumbar LNs was PLM>CL>AL. CONCLUSIONS: Anti-[(18)F]FACBC accumulation in LNs with PLM was higher than that in inflamed LNs. ADVANCES IN KNOWLEDGE: The study showed that although low but significant levels of anti-[(18)F]FACBC uptake by chronic inflamed lesions might cause false-positives in anti-[(18)F]FACBC-PET in some PCa patients, uptake of the tracer at acutely inflamed sites was minimal. IMPLICATIONS FOR PATIENT CARE: The findings of this study suggest the potential of Anti-[(18)F]FACBC for distinguishing between tumors and acute inflammation in clinical practice.
Assuntos
Ácidos Carboxílicos/metabolismo , Ciclobutanos/metabolismo , Linfadenite/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Animais , Ácidos Carboxílicos/farmacocinética , Linhagem Celular Tumoral , Ciclobutanos/farmacocinética , Linfadenite/diagnóstico por imagem , Metástase Linfática , Masculino , Tomografia por Emissão de Pósitrons , Neoplasias da Próstata/diagnóstico por imagem , Ratos , Distribuição TecidualRESUMO
OBJECTIVES: To evaluate whether dual-energy computed tomography (DECT)-derived iodine content and iodine overlay could differentiate between normal, inflammatory and metastatic squamous cell carcinoma (SCC) cervical lymph nodes. METHODS: This study was approved by the institutional review board. Sixteen patients with normal lymph nodes, 20 patients with enlarged nodes draining deep cervical inflammations and 23 patients with pathologically proved metastatic SCC nodes who underwent contrast enhanced DECT were retrospectively identified. Iodine content and overlay of 36 normal, 43 inflammatory and 52 metastatic lymph nodes were calculated using circular regions of interest and compared among the three groups. A receiver operating characteristic (ROC) curve was used to determine the sensitivity and specificity of iodine content and overlay for diagnosis of metastatic nodes. RESULTS: Iodine content (mg/ml) was significantly lower for metastatic lymph nodes (2.34 ± 0.45) than for normal (2.86 ± 0.37) and inflammatory (3.53 ± 0.56) lymph nodes, P < 0.0001. Iodine overlay (HU) was also significantly lower for metastatic lymph nodes (47 ± 11.6) than normal (57.4 ± 8.2) and inflammatory nodes (69.3 ± 11.5), P < 0.0001. The areas under the ROC curve for iodine content and iodine overlay were 0.923 and 0.896. CONCLUSIONS: DECT-derived iodine content and overlay differ significantly among normal, inflammatory and metastatic SCC cervical lymph nodes. KEY POINTS: ⢠Derived iodine content can be calculated from contrast-enhanced dual-energy CT. ⢠Derived iodine content and iodine overlay could help characterise cervical lymph nodes. ⢠Iodine parameters were significantly lower in metastatic lymph nodes than normal/inflammatory lymph nodes. ⢠Iodine content appears more sensitive than iodine overlay for lymph node characterisation.
Assuntos
Carcinoma de Células Escamosas/diagnóstico por imagem , Neoplasias de Cabeça e Pescoço/diagnóstico por imagem , Iodo/metabolismo , Linfadenite/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/secundário , Feminino , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Linfonodos/diagnóstico por imagem , Linfonodos/metabolismo , Linfadenite/metabolismo , Linfadenite/patologia , Metástase Linfática/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Pescoço/patologia , Variações Dependentes do Observador , Curva ROC , Estudos Retrospectivos , Sensibilidade e Especificidade , Tomografia Computadorizada por Raios X/estatística & dados numéricos , Adulto JovemRESUMO
OBJECTIVE: Periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis (PFAPA) syndrome is an autoinflammatory disease of unknown etiology that primarily affects preschool-aged children. PFAPA syndrome is characterized by recurrent attacks of fever and symptoms of inflammation consistent with the disease acronym. Since autoinflammatory diseases are, by definition, mediated by cells of the innate immune system, the aim of this study was to evaluate the functional features of neutrophils, the most abundant innate immune cell in the circulation, in children with PFAPA syndrome. METHODS: Blood polymorphonuclear leukocytes (PMNs), obtained from patients with PFAPA syndrome during both febrile and asymptomatic, afebrile phases of the disease, as well as from healthy children (afebrile controls) and children with fever and abdominal pain (febrile controls), were analyzed for 3 key neutrophil characteristics: 1) apoptosis (measured by annexin V/7-aminoactinomycin D staining), 2) production of reactive oxygen species (ROS) (measured by luminol/isoluminol-amplified chemiluminescence), and 3) priming status (measured as responsiveness to galectin-3 and up-regulation of CD11b). RESULTS: Compared to PMNs obtained from patients with PFAPA syndrome during an afebrile interval and those from febrile controls, PMNs obtained from patients during a PFAPA syndrome flare produced elevated levels of intracellular NADPH oxidase-derived ROS, had significantly diminished rates of spontaneous apoptosis, and displayed signatures of priming. In contrast, PMNs from afebrile patients with PFAPA syndrome had a significantly elevated rate of spontaneous apoptosis compared to PMNs from afebrile controls. CONCLUSION: These findings demonstrate that 3 key aspects of neutrophil innate immune function, namely, apoptosis, priming, and generation of an intracellular oxidative burst, are altered, most prominently during febrile attacks, in children with PFAPA syndrome.
Assuntos
Febre/metabolismo , Linfadenite/metabolismo , Neutrófilos/metabolismo , Faringite/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Estomatite Aftosa/metabolismo , Proteínas de Fase Aguda/imunologia , Proteínas de Fase Aguda/metabolismo , Criança , Pré-Escolar , Feminino , Febre/imunologia , Humanos , Imunidade Inata/imunologia , Lactente , Linfadenite/imunologia , Masculino , Neutrófilos/imunologia , Faringite/imunologia , Estomatite Aftosa/imunologiaRESUMO
Dendritic cell (DC) transmigration across the lymphatic endothelium is critical for the initiation and sustenance of immune responses. Under noninflammatory conditions, DC transit across the lymphatic endothelial cell (LEC) has been shown to be integrin independent. In contrast, there is increasing evidence for the participation of integrins and their ligands in DC transit across lymphatic endothelium under inflammation. In this sense, we describe the formation of ICAM-1 (CD54)-enriched three-dimensional structures on LEC/DC contacts, as these DCs adhere to inflamed skin lymphatic vessels and transmigrate into them. In vitro imaging revealed that under inflammation ICAM-1 accumulated on microvilli projections surrounding 60% of adhered DCs. In contrast, these structures were scarcely formed in noninflammatory conditions. Furthermore, ICAM-1-enriched microvilli were important in promoting DC transendothelial migration and DC crawling over the LEC surface. Microvilli formation was dependent on the presence of ß-integrins on the DC side and on integrin conformational affinity to ligand. Finally, we observed that LEC microvilli structures appeared in close vicinity of CCL21 depots and that their assembly was partially inhibited by CCL21-neutralizing antibodies. Therefore, under inflammatory conditions, integrin ligands form three-dimensional membrane projections around DCs. These structures offer docking sites for DC transit from the tissue toward the lymphatic vessel lumen.
Assuntos
Movimento Celular/fisiologia , Células Dendríticas/citologia , Dermatite/patologia , Células Endoteliais/citologia , Linfadenite/patologia , Vasos Linfáticos/citologia , Animais , Adesão Celular/fisiologia , Comunicação Celular/fisiologia , Diferenciação Celular/fisiologia , Células Dendríticas/metabolismo , Dermatite/metabolismo , Derme/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Humanos , Imageamento Tridimensional/métodos , Integrinas/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Linfadenite/metabolismo , Vasos Linfáticos/fisiologia , Antígeno-1 Associado à Função Linfocitária/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microvilosidades/fisiologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
BACKGROUND: The exact pathogenesis of the pediatric disorder periodic fever, aphthous stomatitis, pharyngitis, cervical adenitis (PFAPA) syndrome is unknown. OBJECTIVES: We hypothesized that PFAPA might be due to dysregulated monocyte IL-1ß production linked to genetic variants in proinflammatory genes. METHODS: Fifteen patients with PFAPA syndrome were studied during and outside a febrile episode. Hematologic profile, inflammatory markers, and cytokine levels were measured in the blood. The capacity of LPS-stimulated PBMCs and monocytes to secrete IL-1ß was assessed by using ELISA, and active IL-1ß secretion was visualized by means of Western blotting. Real-time quantitative PCR was performed to assess cytokine gene expression. DNA was screened for variants of the MEFV, TNFRSF1A, MVK, and NLRP3 genes in a total of 57 patients with PFAPA syndrome. RESULTS: During a febrile attack, patients with PFAPA syndrome revealed significantly increased neutrophil counts, erythrocyte sedimentation rates, and C-reactive protein, serum amyloid A, myeloid-related protein 8/14, and S100A12 levels compared with those seen outside attacks. Stimulated PBMCs secreted significantly more IL-1ß during an attack (during a febrile episode, 575 ± 88 pg/mL; outside a febrile episode, 235 ± 56 pg/mL; P < .001), and this was in the mature active p17 form. IL-1ß secretion was inhibited by ZYVAD, a caspase inhibitor. Similar results were found for stimulated monocytes (during a febrile episode, 743 ± 183 pg/mL; outside a febrile episode, 227 ± 92 pg/mL; P < .05). Genotyping identified variants in 15 of 57 patients, with 12 NLRP3 variants, 1 TNFRSF1A variant, 4 MEFV variants, and 1 MVK variant. CONCLUSION: Our data strongly suggest that IL-1ß monocyte production is dysregulated in patients with PFAPA syndrome. Approximately 20% of them were found to have NLRP3 variants, suggesting that inflammasome-related genes might be involved in this autoinflammatory syndrome.
Assuntos
Febre/metabolismo , Interleucina-1beta/biossíntese , Linfadenite/metabolismo , Monócitos/metabolismo , Faringite/metabolismo , Estomatite Aftosa/metabolismo , Adolescente , Adulto , Idoso , Criança , Feminino , Febre/genética , Febre/imunologia , Variação Genética , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Mediadores da Inflamação/sangue , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Contagem de Leucócitos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/imunologia , Linfadenite/genética , Linfadenite/imunologia , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Neutrófilos , Faringite/genética , Faringite/imunologia , Estomatite Aftosa/genética , Estomatite Aftosa/imunologia , Síndrome , Adulto JovemRESUMO
Common dendritic cell progenitors (CDPs) in the bone marrow (BM) regenerate dendritic cells (DCs) in lymphoid and nonlymphoid tissues. How the dissemination of progenitor-derived DCs to peripheral tissues is regulated on need remains elusive. Microbes are sensed by pathogen recognition receptors such as Toll-like receptors (TLRs). We found that CDPs in the BM express TLR2, TLR4, and TLR9. On TLR stimulation, CDPs down-regulated CXCR4, the nonredundant chemokine receptor for their BM retention, up-regulated CCR7, and migrated to lymph nodes (LNs). When TLR agonists were injected locally, CDPs preferentially gave rise to DCs in inflamed LNs in expense of noninflamed LNs and the BM, but they did not alter their lineage differentiation and proliferative activity. Consequently, BM DC progenitors can sense TLR agonists and, via regulation of CXCR4 and CCR7, support the replenishment of DCs in reactive LNs. This mechanism likely developed to support DC homeostasis on specific need at sites of inflammation.
Assuntos
Movimento Celular , Células Dendríticas/imunologia , Interações Hospedeiro-Patógeno , Linfadenite/imunologia , Células-Tronco/imunologia , Receptores Toll-Like/fisiologia , Animais , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Células da Medula Óssea/fisiologia , Movimento Celular/imunologia , Movimento Celular/fisiologia , Fenômenos Fisiológicos Celulares/genética , Células Cultivadas , Células Dendríticas/metabolismo , Células Dendríticas/fisiologia , Feminino , Interações Hospedeiro-Patógeno/genética , Inflamação/imunologia , Linfonodos/imunologia , Linfonodos/metabolismo , Linfonodos/patologia , Linfadenite/genética , Linfadenite/metabolismo , Linfadenite/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células-Tronco/metabolismo , Células-Tronco/fisiologia , Receptores Toll-Like/metabolismoAssuntos
Carcinoma Pulmonar de Células não Pequenas/secundário , Epitélio/patologia , Queratinas/metabolismo , Neoplasias Pulmonares/patologia , Linfonodos/patologia , Adulto , Biomarcadores Tumorais/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Fatores de Confusão Epidemiológicos , Diagnóstico Diferencial , Epitélio/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Linfonodos/metabolismo , Linfadenite/diagnóstico , Linfadenite/metabolismo , Metástase Linfática , Biópsia de Linfonodo Sentinela/métodosRESUMO
The syndrome of periodic fever, aphthous stomatitis, pharyngitis, and cervical adenitis (PFAPA) is the most common periodic fever disease in children. However, the pathogenesis is unknown. Using a systems biology approach we analyzed blood samples from PFAPA patients whose genetic testing excluded hereditary periodic fevers (HPFs), and from healthy children and pediatric HPF patients. Gene expression profiling could clearly distinguish PFAPA flares from asymptomatic intervals, HPF flares, and healthy controls. During PFAPA attacks, complement (C1QB, C2, SERPING1), IL-1-related (IL-1B, IL-1RN, CASP1, IL18RAP), and IFN-induced (AIM2, IP-10/CXCL10) genes were significantly overexpressed, but T cell-associated transcripts (CD3, CD8B) were down-regulated. On the protein level, PFAPA flares were accompanied by significantly increased serum levels of chemokines for activated T lymphocytes (IP-10/CXCL10, MIG/CXCL9), G-CSF, and proinflammatory cytokines (IL-18, IL-6). PFAPA flares also manifested a relative lymphopenia. Activated CD4(+)/CD25(+) T-lymphocyte counts correlated negatively with serum concentrations of IP-10/CXCL10, whereas CD4(+)/HLA-DR(+) T lymphocyte counts correlated positively with serum concentrations of the counterregulatory IL-1 receptor antagonist. Based on the evidence for IL-1ß activation in PFAPA flares, we treated five PFAPA patients with a recombinant IL-1 receptor antagonist. All patients showed a prompt clinical and IP-10/CXCL10 response. Our data suggest an environmentally triggered activation of complement and IL-1ß/-18 during PFAPA flares, with induction of Th1-chemokines and subsequent retention of activated T cells in peripheral tissues. IL-1 inhibition may thus be beneficial for treatment of PFAPA attacks, with IP-10/CXCL10 serving as a potential biomarker.
Assuntos
Febre/imunologia , Imunidade Inata , Interleucina-1/imunologia , Linfadenite/imunologia , Ativação Linfocitária/imunologia , Faringite/imunologia , Células Th1/imunologia , Adolescente , Criança , Pré-Escolar , Citocinas/imunologia , Citocinas/metabolismo , Feminino , Febre/metabolismo , Febre/terapia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/imunologia , Humanos , Interleucina-1/antagonistas & inibidores , Interleucina-1/metabolismo , Linfadenite/metabolismo , Linfadenite/terapia , Masculino , Faringite/metabolismo , Faringite/terapia , Estomatite Aftosa/imunologia , Estomatite Aftosa/metabolismo , Estomatite Aftosa/terapiaRESUMO
Fas-associated death domain protein is a key component of the extrinsic apoptotic pathway. In addition, in animal models, Fas-associated death domain protein phosphorylation at serine 194 has been shown to affect cell proliferation, especially in T lymphocytes. The importance of Fas-associated death domain protein phosphorylation at serine 194 for the proliferation of B lymphocytes, however, is uncertain. Here we show in reactive lymph nodes that serine 194 phosphorylated Fas-associated death domain protein is expressed predominantly in the dark (proliferative) zone of germinal centers. In B-cell non-Hodgkin lymphoma cell lines, serine 194 phosphorylated Fas-associated death domain protein levels are substantially higher in highly proliferating cells and lower in serum-starved cells. We also used immunohistochemical analysis to assess Fas-associated death domain protein phosphorylation at serine 194 expression in 122 B-cell non-Hodgkin-type lymphomas. The mean percentage of serine 194 phosphorylated Fas-associated death domain protein positive tumor cells was 81% in Burkitt lymphoma, 41% in diffuse large B-cell lymphoma, 18% in follicular lymphoma, 18% in plasma cell myeloma, 12% in extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue, 11% in mantle cell lymphoma, and 2% in chronic lymphocytic leukemia/small lymphocytic lymphoma (P < .0001, Kruskal-Wallis test). Furthermore, in chronic lymphocytic leukemia/small lymphocytic lymphoma, serine 194 phosphorylated Fas-associated death domain protein was detected predominantly in proliferation centers. In the entire study group, the percentage of cells positive for serine 194 phosphorylated Fas-associated death domain protein correlated significantly with the proliferation index Ki-67 (Spearman R = 0.9, P < .0001). These data provide evidence that serine 194 phosphorylated Fas-associated death domain protein is involved in the proliferation of normal and neoplastic B cells and has features of a novel proliferation marker.
Assuntos
Ciclo Celular/fisiologia , Proteína de Domínio de Morte Associada a Fas/metabolismo , Linfoma de Células B/patologia , Serina/metabolismo , Biomarcadores Tumorais/metabolismo , Contagem de Células , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Proliferação de Células , Centro Germinativo/metabolismo , Centro Germinativo/patologia , Humanos , Imuno-Histoquímica , Antígeno Ki-67/metabolismo , Linfonodos/metabolismo , Linfonodos/patologia , Linfadenite/metabolismo , Linfadenite/patologia , Linfoma de Células B/metabolismo , Fosforilação , Tonsilite/metabolismo , Tonsilite/patologiaRESUMO
A 25-year-old man had complained of sudden fever spikes for two years and his blood tests were within the normal range. In 1993, a surgical biopsy of swollen left inguinal lymph nodes was negative for malignancy, but showed reactive lymphadenitis and widespread sinus histiocytosis. A concomitant needle biopsy of the periaortic lymph nodes and a bone marrow aspirate were also negative. In 1994, after an emergency hospital admission because of a sport-related thoracic trauma, a right inguinal lymph node biopsy demonstrated Hodgkin's lymphoma Stage IVB (scleronodular mixed cell subtype). Although it was improved by chemotherapy, the disease suddenly relapsed, and a further lymph node biopsy was performed in 1998 confirming the same diagnosis. Despite further treatment, the patient died of septic shock in 2004, at the age of 38 years. Retrospective analysis of the various specimens showed intracellular heavy metal nanoparticles within lymph node, bone marrow, and liver samples by field emission gun environmental scanning electron microscopy and energy dispersive spectroscopy. Heavy metals from environmental pollution may accumulate in sites far from the entry route and, in genetically conditioned individuals with tissue specificity, may act as cofactors for chronic inflammation or even malignant transformation. The present anecdotal report highlights the need for further pathologic ultrastructural investigations using serial samples and the possible role of intracellular nanoparticles in human disease.