RESUMEN
BACKGROUND: TH2 cells were thought to be a pivotal factor for initiation of the autoimmune blistering disease pemphigus. However, the role of other T-cell subsets in pemphigus pathogenesis remained unclear. OBJECTIVE: We aimed to characterize the exact phenotype of T cells responsible for the development of pemphigus. METHODS: Whole transcriptome shotgun sequencing was performed to determine differential gene expression in pemphigus lesions and skin of healthy individuals. The cutaneous cytokine signature was further evaluated by real-time quantitative PCR. In peripheral blood, the distribution of TH cell and folliclular helper (TFH) cell subsets was analyzed by flow cytometry. Finally, the capacity of TH and TFH cell subsets to induce desmoglein (Dsg)-specific autoantibodies by memory B cells was evaluated in coculture experiments. RESULTS: Transcriptome analysis of skin samples identified an IL-17A-dominated immune signature in patients with pemphigus, and Kyoto Encyclopedia of Genes and Genomes pathway analysis confirmed the dominance of the IL-17A signaling pathway. Increased expression of IL17A and associated cytokines was also detected by real-time quantitative PCR comparing lesional with perilesional or healthy skin. Interestingly, utilization of flow cytometry showed that patients with active pemphigus had elevated levels of circulating IL-17+, TH17, TFH17, and TFH17.1 cells. Notably, levels of TH17 and TFH17 cells correlated with levels of Dsg-specific CD19+CD27+ memory B cells, and patients with acute pemphigus showed higher levels of Dsg3-autoreactive TFH17 cells. Coculture experiments revealed TFH17 cells as primarily responsible for inducing Dsg-specific autoantibody production by B cells. CONCLUSION: Our findings show that TFH17 cells are critically involved in the pathogenesis of pemphigus and offer novel targets for therapeutic intervention.
Asunto(s)
Autoanticuerpos/inmunología , Desmogleína 1/inmunología , Desmogleína 3/inmunología , Pénfigo/inmunología , Pénfigo/metabolismo , Células Th17/inmunología , Células Th17/metabolismo , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/metabolismo , Autoinmunidad , Linfocitos B/inmunología , Linfocitos B/metabolismo , Biomarcadores , Humanos , Inmunofenotipificación , Piel/inmunología , Piel/metabolismo , Piel/patología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells, which are characterized by their capability to suppress T-cell responses. While MDSCs have been traditionally associated with cancer diseases, their role as regulators of autoimmune diseases is emerging. Pemphigus is a chronic autoimmune blistering skin disease characterized by dysregulated T-cell responses and autoantibody production. The role of MDSCs in pemphigus disease has not been defined yet. The aim of this study was to characterize MDSCs in pemphigus patients and to dissect their relationship with CD4+ T-cell subsets and clinical disease assessments. For this purpose, we performed a cross-sectional analysis of 20 patients with pemphigus. Our results indicate that a population of CD66b+ CD11b+ polymorphonuclear-like MDSCs (PMN-MDSCs) is expanded in the peripheral blood mononuclear cell fraction of pemphigus patients compared to age-matched healthy donors. These PMN-MDSCs have the capability of suppressing allogeneic T-cell proliferation in vitro and show increased expression of characteristic effector molecules such as arginase I and interleukin-10. We further demonstrate that PMN-MDSCs are especially expanded in patients with active pemphigus, but not in patients in remission. Moreover, MDSC frequencies correlate with an increased Th2/Th1 cell ratio. In conclusion, the identification of a functional PMN-MDSC population suggests a possible role of these cells as regulators of Th cell responses in pemphigus.
Asunto(s)
Arginasa/metabolismo , Células Supresoras de Origen Mieloide/metabolismo , Pénfigo/metabolismo , Células TH1/metabolismo , Células Th2/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Cutaneous lichen planus (CLP) and psoriasis (PSO) are both common chronic inflammatory skin diseases for which development of new treatments requires the identification of key targets. While PSO is a typical Th17/IL-17-disorder, there is some evidence that Th1/IFN-É£ dominate the inflammatory process in CLP. Nonetheless, the immunopathogenesis of CLP is not fully explained and key immunological factors still have to be recognized. In this study, we compared the immune signature of CLP lesions with the well-characterized inflammation present in PSO skin. First, we analysed the histological and immunohistological characteristics of CLP and PSO. Second, we assessed the cytokine expression (IL1A, IL1B, IL4, IL6, IL8, IL10, IL17A, IL19, IL21, IL22, IL23A, IL13, IFNG, TNF, IL12A, IL12B and IL36G) of lesional skin of CLP with PSO by qPCR. Histology revealed a similar epidermal thickness in CLP and PSO. Immunohistochemically, both diseases presented with an inflammatory infiltrate mainly composed by CD3+ CD4+ T cells rather than CD3+ CD8+ . Importantly, mRNA analysis showed a distinct cytokine signature: while levels of IL12B, IL1A, IL6 and IL23 were similar between the two groups, the characteristic PSO-associated cytokines IL8, IL17A, IL22, IL19 and IL36G were expressed at very low levels in CLP. In contrast, CLP lesional skin was dominated by the expression of IFNG, IL21, IL4, IL12A and TNF. Immunohistochemistry confirmed the dominance of IL-21, IFN-É£ and also pSTAT1 in the dermal infiltrate of CLP, while IL-17A was more present in PSO. Collectively, this study improves our understanding of the immunological factors dominating CLP. The dominating cytokines and signalling proteins identified suggest that anti-cytokine therapeutics like JAK inhibitors may be beneficial in CLP.
Asunto(s)
Citocinas/genética , Liquen Plano/genética , Liquen Plano/inmunología , Psoriasis/genética , Psoriasis/inmunología , Adolescente , Adulto , Anciano , Linfocitos T CD4-Positivos/patología , Niño , Citocinas/metabolismo , Femenino , Expresión Génica , Humanos , Inmunohistoquímica , Interferón gamma/genética , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Interleucina-8/genética , Interleucinas/genética , Interleucinas/metabolismo , Janus Quinasa 1/antagonistas & inhibidores , Liquen Plano/tratamiento farmacológico , Liquen Plano/patología , Masculino , Persona de Mediana Edad , Psoriasis/patología , ARN Mensajero/metabolismo , Factor de Transcripción STAT1/metabolismo , Adulto JovenRESUMEN
Cryopyrin-associated periodic syndromes (CAPS) are characterized by recurrent episodes of systemic inflammation caused by mutations in the NLRP3 gene. Besides confirmed pathogenic NLRP3 mutations, patients with CAPS-like symptoms frequently show low penetrance variants in NLRP3. The disease relevance of these variants is inconsistent. In this study, we investigated if an inflammasome activation assay differentiates between patients with confirmed pathogenic CAPS mutations, patients with low penetrance NLRP3 variants (V198M and Q703K) and healthy controls. The release of mature IL-1ß, IL-18, and caspase-1 into cell culture supernatants after 4h of inflammasome stimulation was significantly increased in patients with confirmed pathogenic CAPS mutations compared to low penetrance NLRP3 variants and controls. IL-1ß secretion in CAPS patients correlated with disease severity. This inflammasome activation assay differentiates between autoinflammation patients with confirmed pathogenic CAPS mutations and patients with low penetrance NLRP3 variants, and points towards alternative pathophysiological mechanisms in low penetrance NLRP3 variants.
Asunto(s)
Bioensayo/métodos , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Variación Genética , Mutación/genética , Adulto , Síndromes Periódicos Asociados a Criopirina/genética , Síndromes Periódicos Asociados a Criopirina/fisiopatología , Femenino , Humanos , Inflamasomas/genética , Inflamasomas/metabolismo , Masculino , Proteína con Dominio Pirina 3 de la Familia NLRRESUMEN
Pseudomonas aeruginosa persists in patients with cystic fibrosis (CF) and drives CF lung disease progression. P. aeruginosa potently activates the innate immune system, mainly mediated through pathogen-associated molecular patterns, such as flagellin. However, the host is unable to eradicate this flagellated bacterium efficiently. The underlying immunological mechanisms are incompletely understood. Myeloid-derived suppressor cells (MDSCs) are innate immune cells generated in cancer and proinflammatory microenvironments and are capable of suppressing T cell responses. We hypothesized that P. aeruginosa induces MDSCs to escape T cell immunity. In this article, we demonstrate that granulocytic MDSCs accumulate in CF patients chronically infected with P. aeruginosa and correlate with CF lung disease activity. Flagellated P. aeruginosa culture supernatants induced the generation of MDSCs, an effect that was 1) dose-dependently mimicked by purified flagellin protein, 2) significantly reduced using flagellin-deficient P. aeruginosa bacteria, and 3) corresponded to TLR5 expression on MDSCs in vitro and in vivo. Both purified flagellin and flagellated P. aeruginosa induced an MDSC phenotype distinct from that of the previously described MDSC-inducing cytokine GM-CSF, characterized by an upregulation of the chemokine receptor CXCR4 on the surface of MDSCs. Functionally, P. aeruginosa-infected CF patient ex vivo-isolated as well as flagellin or P. aeruginosa in vitro-generated MDSCs efficiently suppressed polyclonal T cell proliferation in a dose-dependent manner and modulated Th17 responses. These studies demonstrate that flagellin induces the generation of MDSCs and suggest that P. aeruginosa uses this mechanism to undermine T cell-mediated host defense in CF and other P. aeruginosa-associated chronic lung diseases.
Asunto(s)
Fibrosis Quística/complicaciones , Flagelina/inmunología , Evasión Inmune/inmunología , Tolerancia Inmunológica/inmunología , Células Mieloides/inmunología , Neumonía Bacteriana/inmunología , Infecciones por Pseudomonas/inmunología , Pseudomonas aeruginosa/patogenicidad , Adolescente , Adulto , Proteínas Bacterianas/genética , Células Cultivadas/inmunología , Medios de Cultivo Condicionados/farmacología , Fibrosis Quística/microbiología , Susceptibilidad a Enfermedades , Femenino , Flagelos/inmunología , Flagelos/fisiología , Flagelina/genética , Flagelina/farmacología , Regulación de la Expresión Génica/inmunología , Humanos , Inmunidad Innata , Pulmón/microbiología , Masculino , Células Mieloides/efectos de los fármacos , Mielopoyesis/inmunología , Neumonía Bacteriana/etiología , Neumonía Bacteriana/microbiología , Infecciones por Pseudomonas/etiología , Infecciones por Pseudomonas/microbiología , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/inmunología , Pseudomonas aeruginosa/aislamiento & purificación , Receptores CXCR4/biosíntesis , Receptores CXCR4/genética , Receptores CXCR4/inmunología , Subgrupos de Linfocitos T/inmunología , Células Th17/inmunología , Receptor Toll-Like 5/inmunología , Regulación hacia Arriba/inmunología , Adulto JovenRESUMEN
In acute myeloid leukemia (AML), the leukemia-initiating cell is found within the CD34(+)/CD38(-) cell compartment. Over the last years evidence grew that AML is initiated and propagated by leukemic stem cells (LSCs). Conceivably, these most immature leukemia cells are more resistant to therapy and subsequently initiate relapse. The authors studied 17 patients with childhood AML treated according to the AML-BFM 98/04 protocol. At diagnosis, the authors determined the characteristic immunophenotype of the leukemic cells by flow cytometry and investigated the expression of CD34, CD38, and CD45 to define a population of immunophenotypically immature cells (CD34(+)/CD38(-)/CD45(-/low)) enriched for LSCs in many cases of AML. The authors compared the fraction of this population of all myeloid cells at diagnosis with event-free survival. Kaplan-Meier analysis revealed significant higher event free survival of patients with low CD34(+)/CD38(-)/CD45(-/low) cell proportion (<0.68%) compared to patients with high burden of this population (>0.83%; log-rank P < .04). This correlation was not found for the total number of CD34(+) cells. This is the first study to show that a higher proportion of immature CD34(+)/CD38(-)/CD45(-/low) blasts at diagnosis correlates with unfavorable prognosis in childhood AML. The results suggest that a large CD34(+)/CD38(-)/CD45(-/low) population reflects a higher fraction of LSCs, leading to increased chemotherapy resistance and elevated relapse rate. Thus the initial frequency of CD34(+)/CD38(-)/CD45(-/low) cells may serve as a prognostic marker in pediatric AML. Future treatment in childhood AML should specifically target this immature population as well as the mature blast population.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Leucemia Mieloide Aguda/diagnóstico , Neoplasia Residual/diagnóstico , Células Madre Neoplásicas/patología , ADP-Ribosil Ciclasa 1/metabolismo , Adolescente , Antígenos CD34/metabolismo , Niño , Preescolar , Femenino , Citometría de Flujo , Humanos , Inmunofenotipificación , Lactante , Leucemia Mieloide Aguda/clasificación , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/metabolismo , Antígenos Comunes de Leucocito/metabolismo , Masculino , Pronóstico , Tasa de SupervivenciaRESUMEN
Myeloid-derived suppressor cells (MDSC) represent an innate immune cell subset capable of suppressing T-cell responses in cancer and chronic inflammation. While the effect of MDSC on T cells has been defined thoroughly, the reciprocal impact of T cells on MDSC homeostasis remains poorly understood. Therefore, we comprehensively analyzed the effect of different T-cell subsets on the generation and survival of human MDSC. Using an in vitro MDSC generation assay, we demonstrate that unstimulated CD4+, but not CD8+ T cells, induce polymorphonuclear MDSC (PMN-MDSC) from CD33+ myeloid cells. This effect was dependent on direct cell-cell contact and required TNF-α signaling. Soluble TNF-α was dispensable for PMN-MDSC generation, suggesting that transmembrane TNF-α is involved in that trans-cellular process. Stimulated human CD3+ T cells delayed the apoptosis of PMN-MDSC, which was independent of TNF-α signaling or direct cell-cell contact, but was recapitulated by IL-2. Taken together, our study shows that human T cells modulate MDSC generation and survival through two distinct mechanisms and thereby fine-tune the homeostasis of human MDSC in a regulated manner.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Células Mieloides/inmunología , Células Supresoras de Origen Mieloide/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Apoptosis/inmunología , Células Cultivadas , Homeostasis/inmunología , Humanos , Interleucina-2/inmunología , Neoplasias/inmunología , Lectina 3 Similar a Ig de Unión al Ácido Siálico/inmunología , Transducción de Señal/inmunologíaRESUMEN
Staphylococcus aureus is one of the major human bacterial pathogens causing a broad spectrum of serious infections. Myeloid-derived suppressor cells (MDSC) represent an innate immune cell subset capable of regulating host-pathogen interactions, yet their role in the pathogenesis of S. aureus infections remains incompletely defined. The aim of this study was to determine the influence of different S. aureus strains and associated virulence factors on human MDSC generation. Using an in vitro MDSC generation assay we demonstrate that low concentrations of supernatants of different S. aureus strains led to an induction of functional MDSC, whereas increased concentrations, conversely, reduced MDSC numbers. The concentration-dependent reduction of MDSC correlated with T cell proliferation and cytotoxicity. Several findings supported a role for staphylococcal enterotoxins in modulating MDSC generation. Staphylococcal enterotoxins recapitulated concentration-dependent MDSC induction and inhibition, T cell proliferation and cytotoxicity, while an enterotoxin-deficient S. aureus strain largely failed to alter MDSC. Taken together, we identified staphylococcal enterotoxins as main modulators of MDSC generation. The inhibition of MDSC generation by staphylococcal enterotoxins might represent a novel therapeutic target in S. aureus infections and beyond in non-infectious conditions, such as cancer.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Enterotoxinas/inmunología , Células Supresoras de Origen Mieloide/efectos de los fármacos , Células Supresoras de Origen Mieloide/inmunología , Staphylococcus aureus/inmunología , Células Cultivadas , Enterotoxinas/metabolismo , Humanos , Evasión Inmune , Tolerancia Inmunológica , Modelos Teóricos , Infecciones Estafilocócicas/inmunología , Infecciones Estafilocócicas/patología , Staphylococcus aureus/metabolismoRESUMEN
BACKGROUND: Psoriasis is an inflammatory skin disease with aberrant keratinocyte proliferation, presumably as a result of immune cell activation. Th17 cytokines like IL-17A and IL-22 are critically implicated in epidermal thickening, altered keratinocyte differentiation and production of innate factors such as antimicrobial peptides. Psoriasis treatment options include modern targeted therapies using anti-cytokine antibodies and traditional non-targeted treatments like anthralin (dithranol). While the mode of action of anti-cytokine antibodies is defined, the effects of topical anthralin on psoriatic skin are not fully understood. OBJECTIVE: This study aims to unravel the direct effects of anthralin on keratinocyte proliferation, differentiation and production of psoriasis-associated factors. METHODS: We tested the effects of anthralin on cell proliferation, cytokeratin expression and changes in the expression of antimicrobial peptides using primary keratinocytes and 3D psoriasis tissue models with and without stimulation of the psoriasis-promoting cytokines IL-17A and IL-22. Moreover, we compared the findings derived from monolayer and multilayer cultures to data derived from lesional skin of patients with psoriasis before and under treatment with anthralin. RESULTS: Our study shows that anthralin directly induces cell apoptosis in vitro in monolayer cultures but not in 3D psoriasis tissue models treated with IL-17A and IL-22. Yet, keratinocyte proliferation as determined by Ki-67 staining is impaired by anthralin in vivo. In lesional skin but not in 3D psoriasis tissue models anthralin rapidly normalizes cytokeratin (CK)16 expression. Furthermore, anthralin directly inhibits DEFB4 expression in vitro and in vivo, while other antimicrobial peptides and cytokines studied like IL-6 and IL-8 are regulated differently in vitro and in vivo. CONCLUSIONS: Our results show that anthralin directly regulates DEFB4A expression. However, its beneficial effects on psoriasis cannot be explained by direct effects on keratinocyte differentiation or cytokine expression.
Asunto(s)
Antralina/farmacología , Fármacos Dermatológicos/farmacología , Queratina-16/metabolismo , Queratinocitos/efectos de los fármacos , Psoriasis/tratamiento farmacológico , beta-Defensinas/metabolismo , Administración Cutánea , Antralina/uso terapéutico , Apoptosis/efectos de los fármacos , Biopsia , Diferenciación Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Fármacos Dermatológicos/uso terapéutico , Técnica del Anticuerpo Fluorescente , Humanos , Interleucina-17/metabolismo , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Interleucinas/metabolismo , Queratina-10/metabolismo , Queratina-5/metabolismo , Queratinocitos/metabolismo , Antígeno Ki-67/metabolismo , Psoriasis/patología , Piel/citología , Piel/efectos de los fármacos , Piel/patología , Técnicas de Cultivo de Tejidos/métodos , Interleucina-22RESUMEN
Myeloid-derived suppressor cells (MDSCs) are key regulators of adaptive immunity by suppressing T-cell functions. However, their potential action on or interaction with B cells remained poorly understood. Here we demonstrate that human polymorphonuclear MDSCs differentially modulate B-cell function by suppressing B-cell proliferation and antibody production. We further demonstrate that this MDSC-mediated effect is cell contact dependent and involves established mediators such as arginase-1, nitric oxide (NO), reactive oxygen species (ROS) as well as B-cell death. Collectively, our studies provide novel evidence that human MDSCs modulate B cells, which could have future implications for immunotherapy approaches.
Asunto(s)
Linfocitos B/inmunología , Linfocitos B/metabolismo , Comunicación Celular/inmunología , Inmunomodulación , Células Supresoras de Origen Mieloide/inmunología , Células Supresoras de Origen Mieloide/metabolismo , Formación de Anticuerpos/inmunología , Arginasa/metabolismo , Biomarcadores , Células Cultivadas , Humanos , Inmunoglobulina M/inmunología , Activación de Linfocitos/inmunología , Neutrófilos/inmunología , Neutrófilos/metabolismo , Óxido Nítrico/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Myeloid-derived suppressor cells (MDSCs) are innate immune cells characterized by their ability to suppress T-cell responses. Recently, we demonstrated that the human-pathogenic fungi Candida albicans and Aspergillus fumigatus induced a distinct subset of neutrophilic MDSCs. To dissect Candida-mediated MDSC induction in more depth, we studied the relative efficacy of different pathogenic non-albicans Candida species to induce and functionally modulate neutrophilic MDSCs, including C. glabrata, C. parapsilosis, C. dubliniensis, and C. krusei. Our data demonstrate that the extent of MDSC generation is largely dependent on the Candida species with MDSCs induced by C. krusei and C. glabrata showing a higher suppressive activity compared to MDSCs induced by C. albicans. In summary, these studies show that fungal MDSC induction is differentially regulated at the species level and differentially affects effector T-cell responses.
RESUMEN
Despite continuous contact with fungi, immunocompetent individuals rarely develop pro-inflammatory antifungal immune responses. The underlying tolerogenic mechanisms are incompletely understood. Using both mouse models and human patients, we show that infection with the human pathogenic fungi Aspergillus fumigatus and Candida albicans induces a distinct subset of neutrophilic myeloid-derived suppressor cells (MDSCs), which functionally suppress T and NK cell responses. Mechanistically, pathogenic fungi induce neutrophilic MDSCs through the pattern recognition receptor Dectin-1 and its downstream adaptor protein CARD9. Fungal MDSC induction is further dependent on pathways downstream of Dectin-1 signaling, notably reactive oxygen species (ROS) generation as well as caspase-8 activity and interleukin-1 (IL-1) production. Additionally, exogenous IL-1ß induces MDSCs to comparable levels observed during C. albicans infection. Adoptive transfer and survival experiments show that MDSCs are protective during invasive C. albicans infection, but not A. fumigatus infection. These studies define an innate immune mechanism by which pathogenic fungi regulate host defense.
Asunto(s)
Aspergillus fumigatus/inmunología , Candida albicans/inmunología , Interacciones Huésped-Patógeno , Tolerancia Inmunológica , Neutrófilos/inmunología , Animales , Aspergilosis/inmunología , Aspergilosis/microbiología , Proteínas Adaptadoras de Señalización CARD/metabolismo , Candidiasis/inmunología , Candidiasis/microbiología , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , Lectinas Tipo C/metabolismo , Ratones , Transducción de SeñalRESUMEN
Prognosis for children with acute lymphoblastic leukemia (ALL) has considerably improved, yet relapse still occurs in a significant proportion of patients. Conceivably, the most immature leukemia cells may be more resistant to therapy and initiate relapse. We studied 42 patients with childhood ALL treated according to the ALL-BFM 2000 protocol. At diagnosis, we determined the characteristic immunophenotype of the leukemic cells by flow cytometry and also investigated the expression of CD34 and CD38 to define a population of immunophenotypically immature cells (CD34(+)/CD38(-)). We then studied levels of minimal residual disease (MRD) after induction therapy (day 33) and after consolidation therapy (week 12). We found a significant, increasing correlation between the prevalence of CD34(+)/CD38(-) cells at diagnosis and MRD levels at day 33 and week 12. Our results suggest that the initial frequency of CD34(+)/CD38(-) cells may serve as a prognostic marker in pediatric ALL.